Interleukin 6 Influences Germinal Center Development and Antibody Production via a Contribution of C3 Complement Component

PubMed Central

Kopf, Manfred; Herren, Suzanne; Wiles, Michael V.; Pepys, Mark B.; Kosco-Vilbois, Marie H.

1998-01-01

Mice rendered deficient for interleukin (IL) 6 by gene targeting were evaluated for their response to T cell–dependent antigens. Antigen-specific immunoglobulin (Ig)M levels were unaffected whereas all IgG isotypes showed varying degrees of alteration. Germinal center reactions occurred but remained physically smaller in comparison to those in the wild-type mice. This concurred with the observations that molecules involved in initial signaling events leading to germinal center formation were not altered (e.g., B7.2, CD40 and tumor necrosis factor R1). T cell priming was not impaired nor was a gross imbalance of T helper cell (Th) 1 versus Th2 cytokines observed. However, B7.1 molecules, absent from wild-type counterparts, were detected on germinal center B cells isolated from the deficient mice suggesting a modification of costimulatory signaling. A second alteration involved impaired de novo synthesis of C3 both in serum and germinal center cells from IL-6–deficient mice. Indeed, C3 provided an essential stimulatory signal for wild-type germinal center cells as both monoclonal antibodies that interrupted C3-CD21 interactions and sheep anti–mouse C3 antibodies caused a significant decrease in antigen-specific antibody production. In addition, germinal center cells isolated from C3–deficient mice produced a similar defect in isotype production. Low density cells with dendritic morphology were the local source of IL-6 and not the germinal center lymphocytes. Adding IL-6 in vitro to IL-6–deficient germinal center cells stimulated cell cycle progression and increased levels of antibody production. These findings reveal that the germinal center produces and uses molecules of the innate immune system, evolutionarily pirating them in order to optimally generate high affinity antibody responses. PMID:9815267

The unique stem cell system of the immortal larva of the human parasite Echinococcus multilocularis

PubMed Central

2014-01-01

Background It is believed that in tapeworms a separate population of undifferentiated cells, the germinative cells, is the only source of cell proliferation throughout the life cycle (similar to the neoblasts of free living flatworms). In Echinococcus multilocularis, the metacestode larval stage has a unique development, growing continuously like a mass of vesicles that infiltrate the tissues of the intermediate host, generating multiple protoscoleces by asexual budding. This unique proliferation potential indicates the existence of stem cells that are totipotent and have the ability for extensive self-renewal. Results We show that only the germinative cells proliferate in the larval vesicles and in primary cell cultures that undergo complete vesicle regeneration, by using a combination of morphological criteria and by developing molecular markers of differentiated cell types. The germinative cells are homogeneous in morphology but heterogeneous at the molecular level, since only sub-populations express homologs of the post-transcriptional regulators nanos and argonaute. Important differences are observed between the expression patterns of selected neoblast marker genes of other flatworms and the E. multilocularis germinative cells, including widespread expression in E. multilocularis of some genes that are neoblast-specific in planarians. Hydroxyurea treatment results in the depletion of germinative cells in larval vesicles, and after recovery following hydroxyurea treatment, surviving proliferating cells grow as patches that suggest extensive self-renewal potential for individual germinative cells. Conclusions In E. multilocularis metacestodes, the germinative cells are the only proliferating cells, presumably driving the continuous growth of the larval vesicles. However, the existence of sub-populations of the germinative cells is strongly supported by our data. Although the germinative cells are very similar to the neoblasts of other flatworms in function and in undifferentiated morphology, their unique gene expression pattern and the evolutionary loss of conserved stem cells regulators suggest that important differences in their physiology exist, which could be related to the unique biology of E. multilocularis larvae. PMID:24602211

The unique stem cell system of the immortal larva of the human parasite Echinococcus multilocularis.

PubMed

Koziol, Uriel; Rauschendorfer, Theresa; Zanon Rodríguez, Luis; Krohne, Georg; Brehm, Klaus

2014-03-06

It is believed that in tapeworms a separate population of undifferentiated cells, the germinative cells, is the only source of cell proliferation throughout the life cycle (similar to the neoblasts of free living flatworms). In Echinococcus multilocularis, the metacestode larval stage has a unique development, growing continuously like a mass of vesicles that infiltrate the tissues of the intermediate host, generating multiple protoscoleces by asexual budding. This unique proliferation potential indicates the existence of stem cells that are totipotent and have the ability for extensive self-renewal. We show that only the germinative cells proliferate in the larval vesicles and in primary cell cultures that undergo complete vesicle regeneration, by using a combination of morphological criteria and by developing molecular markers of differentiated cell types. The germinative cells are homogeneous in morphology but heterogeneous at the molecular level, since only sub-populations express homologs of the post-transcriptional regulators nanos and argonaute. Important differences are observed between the expression patterns of selected neoblast marker genes of other flatworms and the E. multilocularis germinative cells, including widespread expression in E. multilocularis of some genes that are neoblast-specific in planarians. Hydroxyurea treatment results in the depletion of germinative cells in larval vesicles, and after recovery following hydroxyurea treatment, surviving proliferating cells grow as patches that suggest extensive self-renewal potential for individual germinative cells. In E. multilocularis metacestodes, the germinative cells are the only proliferating cells, presumably driving the continuous growth of the larval vesicles. However, the existence of sub-populations of the germinative cells is strongly supported by our data. Although the germinative cells are very similar to the neoblasts of other flatworms in function and in undifferentiated morphology, their unique gene expression pattern and the evolutionary loss of conserved stem cells regulators suggest that important differences in their physiology exist, which could be related to the unique biology of E. multilocularis larvae.

EGF-mediated EGFR/ERK signaling pathway promotes germinative cell proliferation in Echinococcus multilocularis that contributes to larval growth and development.

PubMed

Cheng, Zhe; Liu, Fan; Li, Xiu; Dai, Mengya; Wu, Jianjian; Guo, Xinrui; Tian, Huimin; Heng, Zhijie; Lu, Ying; Chai, Xiaoli; Wang, Yanhai

2017-02-01

Larvae of the tapeworm E. multilocularis cause alveolar echinococcosis (AE), one of the most lethal helminthic infections in humans. A population of stem cell-like cells, the germinative cells, is considered to drive the larval growth and development within the host. The molecular mechanisms controlling the behavior of germinative cells are largely unknown. Using in vitro cultivation systems we show here that the EGFR/ERK signaling in the parasite can promote germinative cell proliferation in response to addition of human EGF, resulting in stimulated growth and development of the metacestode larvae. Inhibition of the signaling by either the EGFR inhibitors CI-1033 and BIBW2992 or the MEK/ERK inhibitor U0126 impairs germinative cell proliferation and larval growth. These data demonstrate the contribution of EGF-mediated EGFR/ERK signaling to the regulation of germinative cells in E. multilocularis, and suggest the EGFR/ERK signaling as a potential therapeutic target for AE and perhaps other human cestodiasis.

Bacillus anthracis spores germinate extracellularly at air-liquid interface in an in vitro lung model under serum-free conditions

DOE PAGES

Powell, Joshua D.; Hutchison, Janine R.; Hess, Becky M.; ...

2015-07-30

Aims: To better understand the parameters that govern spore dissemination after lung exposure using in vitro cell systems. Methods and Results: We evaluated the kinetics of uptake, germination and proliferation of B. anthracis Sterne spores in association with human primary lung epithelial cells, Calu-3, and A549 cell lines. We also analyzed the influence of various cell culture media formulations related to spore germination. Conclusions: We found negligible spore uptake by epithelial cells, but germination and proliferation of spores in the extracellular environment was evident, and was appreciably higher in A549 and Calu-3 cultures than in primary epithelial cells. Additionally, ourmore » results revealed spores in association with primary cells submerged in cell culture media germinated 1 h« less

Constitutive CD40L Expression on B Cells Prematurely Terminates Germinal Center Response and Leads to Augmented Plasma Cell Production in T Cell Areas

PubMed Central

Bolduc, Anna; Long, Eugene; Stapler, Dale; Cascalho, Marilia; Tsubata, Takeshi; Koni, Pandelakis A.; Shimoda, Michiko

2013-01-01

CD40/CD40L engagement is essential to T cell-dependent B cell proliferation and differentiation. However, the precise role of CD40 signaling through cognate T–B interaction in the generation of germinal center and memory B cells is still incompletely understood. To address this issue, a B cell-specific CD40L transgene (CD40LBTg) was introduced into mice with B cell-restricted MHC class II deficiency. Using this mouse model, we show that constitutive CD40L expression on B cells alone could not induce germinal center differentiation of MHC class II-deficient B cells after immunization with T cell-dependent Ag. Thus, some other MHC class II-dependent T cell-derived signals are essential for the generation of germinal center B cells in response to T cell-dependent Ag. In fact, CD40LBTg mice generated a complex Ag-specific IgG1 response, which was greatly enhanced in early, but reduced in late, primary response compared with control mice. We also found that the frequency of Ag-specific germinal center B cells in CD40LBTg mice was abruptly reduced 1 wk after immunization. As a result, the numbers of Ag-specific IgG1 long-lived plasma cells and memory B cells were reduced. By histology, large numbers of Ag-specific plasma cells were found in T cell areas adjacent to Ag-specific germinal centers of CD40LBTg mice, temporarily during the second week of primary response. These results indicate that CD40L expression on B cells prematurely terminated their ongoing germinal center response and produced plasma cells. Our results support the notion that CD40 signaling is an active termination signal for germinal center reaction. PMID:20505142

In situ dynamics of cyst and vegetative cell populations of the toxic dinoflagellate Alexandrium catenella in Ago Bay, central Japan

PubMed Central

Ishikawa, Akira; Hattori, Mayuko; Ishii, Ken-Ichiro; Kulis, David M.; Anderson, Donald M.; Imai, Ichiro

2014-01-01

Temporal changes in the in situ germination flux of cysts and the abundance of vegetative cells of the toxic dinoflagellate Alexandrium catenella were investigated in Ago Bay, central Japan from July 2003 to December 2004. The in situ germination flux (cells m−2 day−1) was measured using ‘plankton emergence trap/chambers (PET chambers)’. Germination of the cysts in the sediments occurred continuously during the study, ranging from 52 to 1753 cells m−2 day−1, with no temporal trend. This germination pattern appeared to be promoted by a short mandatory dormancy period for newly formed cysts coupled with a broad temperature window for germination. For the vegetative populations, high abundances (>105 cells m−2) were recorded in the water column from spring to summer and from autumn to early winter. The size of the vegetative populations did not correlate with the cyst germination flux but rather larger vegetative populations were often observed when the water temperature was around 20°C, indicating that bloom development was mainly regulated by the temperature. Nonetheless, the continuous germination pattern of A. catenella is advantageous enabling the germinated cells to immediately exploit favorable bloom conditions. PMID:25221373

EGF-mediated EGFR/ERK signaling pathway promotes germinative cell proliferation in Echinococcus multilocularis that contributes to larval growth and development

PubMed Central

Li, Xiu; Dai, Mengya; Wu, Jianjian; Guo, Xinrui; Tian, Huimin; Heng, Zhijie; Lu, Ying; Chai, Xiaoli

2017-01-01

Background Larvae of the tapeworm E. multilocularis cause alveolar echinococcosis (AE), one of the most lethal helminthic infections in humans. A population of stem cell-like cells, the germinative cells, is considered to drive the larval growth and development within the host. The molecular mechanisms controlling the behavior of germinative cells are largely unknown. Methodology/Principal findings Using in vitro cultivation systems we show here that the EGFR/ERK signaling in the parasite can promote germinative cell proliferation in response to addition of human EGF, resulting in stimulated growth and development of the metacestode larvae. Inhibition of the signaling by either the EGFR inhibitors CI-1033 and BIBW2992 or the MEK/ERK inhibitor U0126 impairs germinative cell proliferation and larval growth. Conclusions/Significance These data demonstrate the contribution of EGF-mediated EGFR/ERK signaling to the regulation of germinative cells in E. multilocularis, and suggest the EGFR/ERK signaling as a potential therapeutic target for AE and perhaps other human cestodiasis. PMID:28241017

α-Xylosidase plays essential roles in xyloglucan remodelling, maintenance of cell wall integrity, and seed germination in Arabidopsis thaliana

PubMed Central

Shigeyama, Takuma; Watanabe, Asuka; Tokuchi, Konatsu; Toh, Shigeo; Sakurai, Naoki; Shibuya, Naoto; Kawakami, Naoto

2016-01-01

Regulation and maintenance of cell wall physical properties are crucial for plant growth and environmental response. In the germination process, hypocotyl cell expansion and endosperm weakening are prerequisites for dicot seeds to complete germination. We have identified the Arabidopsis mutant thermoinhibition-resistant germination 1 (trg1), which has reduced seed dormancy and insensitivity to unfavourable conditions for germination owing to a loss-of-function mutation of TRG1/XYL1, which encodes an α-xylosidase. Compared to those of wild type, the elongating stem of trg1 showed significantly lower viscoelasticity, and the fruit epidermal cells were longitudinally shorter and horizontally enlarged. Actively growing tissues of trg1 over-accumulated free xyloglucan oligosaccharides (XGOs), and the seed cell wall had xyloglucan with a greatly reduced molecular weight. These observations suggest that XGOs reduce xyloglucan size by serving as an acceptor in transglycosylation and eventually enhancing cell wall loosening. TRG1/XYL1 gene expression was abundant in growing wild-type organs and tissues but relatively low in cells at most actively elongating part of the tissues, suggesting that α-xylosidase contributes to maintaining the mechanical integrity of the primary cell wall in the growing and pre-growing tissues. In germinating seeds of trg1, expression of genes encoding specific abscisic acid and gibberellin metabolism enzymes was altered in accordance with the aberrant germination phenotype. Thus, cell wall integrity could affect seed germination not only directly through the physical properties of the cell wall but also indirectly through the regulation of hormone gene expression. PMID:27605715

The extent of clonal structure in different lymphoid organs

PubMed Central

1992-01-01

To gain insight into the clonal organization of lymphoid organs, we studied the distribution in situ of donor-derived cells in near- physiological chimeras. We introduced RT7b fetal liver cells into nonirradiated congenic RT7a neonatal rats. The chimerism 6-20 wk after injection ranged from 0.3 to 20%. The numbers of cell clones simultaneously contributing to cell generation in a particular histological feature were deduced from the variance in donor cell distribution. In bone marrow and thymus, donor-derived lymphoid cells were found scattered among host cells, indicating a high mobility of cells. In bone marrow, donor cells were evenly distributed over the entire marrow, even at low chimerism. This indicates that leukopoiesis is maintained by the proliferation of many clones. In the thymus, the various lobules showed different quantities of donor-derived lymphoid cells. Mathematical analysis of these differences indicated that 17-18 cell division cycles occur in the cortex. In spleen, the distribution of donor-derived cells over the germinal centers indicated that 5 d after antigenic stimulation, germinal centers develop oligoclonally. The main conclusions of this work are that (a) bone marrow and thymus are highly polyclonal; (b) 17-18 divisions occur between prothymocyte and mature T cell; and (c) lymphoid cells disperse rapidly while proliferating and differentiating. PMID:1569396

DOE Office of Scientific and Technical Information (OSTI.GOV)

Powell, Joshua D.; Hutchison, Janine R.; Hess, Becky M.

Aims: To better understand the parameters that govern spore dissemination after lung exposure using in vitro cell systems. Methods and Results: We evaluated the kinetics of uptake, germination and proliferation of B. anthracis Sterne spores in association with human primary lung epithelial cells, Calu-3, and A549 cell lines. We also analyzed the influence of various cell culture media formulations related to spore germination. Conclusions: We found negligible spore uptake by epithelial cells, but germination and proliferation of spores in the extracellular environment was evident, and was appreciably higher in A549 and Calu-3 cultures than in primary epithelial cells. Additionally, ourmore » results revealed spores in association with primary cells submerged in cell culture media germinated 1 h« less

Disappearance and reappearance of high endothelial venules and immigrating lymphocytes in lymph nodes deprived of afferent lymphatic vessels: a possible regulatory role of macrophages in lymphocyte migration.

PubMed

Hendriks, H R; Eestermans, I L

1983-08-01

Interruption of the afferent lymphatic vessels of the popliteal lymph node resulted in the disappearance of high endothelial venules (HEV) and immigrating lymphocytes within 3 weeks. HEV showed several characteristic morphological changes: the endothelial cells became flattened and less pyroninophilic, the chromatine became condensed and protein synthetizing and secretory cell organelles became scarce. At the same time the number of macrophages in the lymph node was severely reduced. Injection of sheep red blood cells into such lymph nodes, 6 weeks after operation, resulted in reappearance of HEV and immigrating lymphocytes, and development of many plasma cells and some germinal centres. Injection of lipopolysaccharide into the operated lymph nodes resulted in the appearance of many plasma cells and a few poorly developed germinal centres; HEV and immigrating lymphocytes, however, remained almost absent. The results show a relationship between the immigration of lymphocytes and the activity of the endothelial cells in the HEV. The activation of the latter may occur by mediators released by antigen-stimulated macrophages and T cells. Moreover, the morphological features of the HEV are independent of the presence of recirculating lymphocytes.

α-Xylosidase plays essential roles in xyloglucan remodelling, maintenance of cell wall integrity, and seed germination in Arabidopsis thaliana.

PubMed

Shigeyama, Takuma; Watanabe, Asuka; Tokuchi, Konatsu; Toh, Shigeo; Sakurai, Naoki; Shibuya, Naoto; Kawakami, Naoto

2016-10-01

Regulation and maintenance of cell wall physical properties are crucial for plant growth and environmental response. In the germination process, hypocotyl cell expansion and endosperm weakening are prerequisites for dicot seeds to complete germination. We have identified the Arabidopsis mutant thermoinhibition-resistant germination 1 (trg1), which has reduced seed dormancy and insensitivity to unfavourable conditions for germination owing to a loss-of-function mutation of TRG1/XYL1, which encodes an α-xylosidase. Compared to those of wild type, the elongating stem of trg1 showed significantly lower viscoelasticity, and the fruit epidermal cells were longitudinally shorter and horizontally enlarged. Actively growing tissues of trg1 over-accumulated free xyloglucan oligosaccharides (XGOs), and the seed cell wall had xyloglucan with a greatly reduced molecular weight. These observations suggest that XGOs reduce xyloglucan size by serving as an acceptor in transglycosylation and eventually enhancing cell wall loosening. TRG1/XYL1 gene expression was abundant in growing wild-type organs and tissues but relatively low in cells at most actively elongating part of the tissues, suggesting that α-xylosidase contributes to maintaining the mechanical integrity of the primary cell wall in the growing and pre-growing tissues. In germinating seeds of trg1, expression of genes encoding specific abscisic acid and gibberellin metabolism enzymes was altered in accordance with the aberrant germination phenotype. Thus, cell wall integrity could affect seed germination not only directly through the physical properties of the cell wall but also indirectly through the regulation of hormone gene expression. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Cell division versus cell elongation: the control of radicle elongation during thermoinhibition of Tagetes minuta achenes.

PubMed

Taylor, Nicky J; Hills, Paul N; van Staden, Johannes

2007-12-01

Endogenous embryo factors, which act mainly in the radicle, prevent germination in Tagetes minuta at high temperatures. These factors act to prevent cell elongation, which is critical for radicle protrusion under optimal conditions. Once the radicle has emerged both cell elongation and cell division are required for post-germination growth. Germination can be induced at high temperatures by fusicoccin, which rapidly stimulates cell elongation. In addition, priming seeds at 25 degrees C on polyethylene glycol (PEG) 6000 and mannitol could also induce germination on water at 36 degrees C, indicating that priming prevents radicle protrusion at a point subsequent to the point of control in thermoinhibited achenes. Flow cytometry studies revealed that DNA synthesis occurs during thermoinhibition and the inhibition of DNA synthesis during this process inhibits subsequent germination on water under optimal conditions, suggesting a protective role for DNA synthesis in thermoinhibited achenes of T. minuta.

Immunological aspects in chronic lymphocytic leukemia (CLL) development.

PubMed

García-Muñoz, Ricardo; Galiacho, Verónica Roldan; Llorente, Luis

2012-07-01

Chronic lymphocytic leukemia (CLL) is unique among B cell malignancies in that the malignant clones can be featured either somatically mutated or unmutated IGVH genes. CLL cells that express unmutated immunoglobulin variable domains likely underwent final development prior to their entry into the germinal center, whereas those that express mutated variable domains likely transited through the germinal center and then underwent final development. Regardless, the cellular origin of CLL remains unknown. The aim of this review is to summarize immunological aspects involved in this process and to provide insights about the complex biology and pathogenesis of this disease. We propose a mechanistic hypothesis to explain the origin of B-CLL clones into our current picture of normal B cell development. In particular, we suggest that unmutated CLL arises from normal B cells with self-reactivity for apoptotic bodies that have undergone receptor editing, CD5 expression, and anergic processes in the bone marrow. Similarly, mutated CLL would arise from cells that, while acquiring self-reactivity for autoantigens-including apoptotic bodies-in germinal centers, are also still subject to tolerization mechanisms, including receptor editing and anergy. We believe that CLL is a proliferation of B lymphocytes selected during clonal expansion through multiple encounters with (auto)antigens, despite the fact that they differ in their state of activation and maturation. Autoantigens and microbial pathogens activate BCR signaling and promote tolerogenic mechanisms such as receptor editing/revision, anergy, CD5+ expression, and somatic hypermutation in CLL B cells. The result of these tolerogenic mechanisms is the survival of CLL B cell clones with similar surface markers and homogeneous gene expression signatures. We suggest that both immunophenotypic surface markers and homogenous gene expression might represent the evidence of several attempts to re-educate self-reactive B cells.

Effect of Sodium Nitrite, Sodium Chloride, and Sodium Nitrate on Germination and Outgrowth of Anaerobic Spores1

PubMed Central

Duncan, Charles L.; Foster, E. M.

1968-01-01

The effects of meat-curing agents on germination and outgrowth of putrefactive anaerobe 3679h (PA 3679h) spores were studied in microcultures. Nitrite concentrations up to 0.06% at pH 6.0 or between 0.8 and 1% at pH 7.0 allowed emergence and elongation of vegetative cells but blocked cell division. The newly emerged cells then lysed. With more than 0.06% nitrite at pH 6.0 or more than 0.8 to 1% at pH 7.0, the spores lost refractility and swelled, but vegetative cells did not emerge. Even as much as 4% nitrite failed to prevent germination (complete loss of refractility) and swelling of the spores. Sodium chloride concentrations above 6% prevented complete germination (i.e., the spores retained a refractile core). In the presence of 3 to 6% sodium chloride, most of the spores germinated and produced vegetative cells, but cell division was often blocked. Sodium nitrate had no apparent effect on germination and outgrowth at concentrations up to 2%. Images Fig. 1 Fig. 2 Fig. 3 PMID:5645423

Changes in cell wall polysaccharide composition, gene transcription and alternative splicing in germinating barley embryos.

PubMed

Zhang, Qisen; Zhang, Xiaoqi; Pettolino, Filomena; Zhou, Gaofeng; Li, Chengdao

2016-02-01

Barley (Hordeum vulgare L.) seed germination initiates many important biological processes such as DNA, membrane and mitochondrial repairs. However, little is known on cell wall modifications in germinating embryos. We have investigated cell wall polysaccharide composition change, gene transcription and alternative splicing events in four barley varieties at 24h and 48 h germination. Cell wall components in germinating barley embryos changed rapidly, with increases in cellulose and (1,3)(1,4)-β-D-glucan (20-100%) within 24h, but decreases in heteroxylan and arabinan (3-50%). There were also significant changes in the levels of type I arabinogalactans and heteromannans. Alternative splicing played very important roles in cell wall modifications. At least 22 cell wall transcripts were detected to undergo either alternative 3' splicing, alternative 5' splicing or intron retention type of alternative splicing. These genes coded enzymes catalyzing synthesis and degradation of cellulose, heteroxylan, (1,3)(1,4)-β-D-glucan and other cell wall polymers. Furthermore, transcriptional regulation also played very important roles in cell wall modifications. Transcript levels of primary wall cellulase synthase, heteroxylan synthesizing and nucleotide sugar inter-conversion genes were very high in germinating embryos. At least 50 cell wall genes changed transcript levels significantly. Expression patterns of many cell wall genes coincided with changes in polysaccharide composition. Our data showed that cell wall polysaccharide metabolism was very active in germinating barley embryos, which was regulated at both transcriptional and post-transcriptional levels. Copyright © 2015 Elsevier GmbH. All rights reserved.

The maize pentatricopeptide repeat gene empty pericarp4 (emp4) is required for proper cellular development in vegetative tissues.

PubMed

Gabotti, Damiano; Caporali, Elisabetta; Manzotti, Priscilla; Persico, Martina; Vigani, Gianpiero; Consonni, Gabriella

2014-06-01

The empty pericarp4 (emp4) gene encodes a mitochondrion-targeted pentatricopeptide repeat (ppr) protein that is involved in the regulation of mitochondrial gene expression and is required for seed development. In homozygous mutant emp4-1 kernels the endosperm is drastically reduced and the embryo is retarded in its development and unable to germinate. With the aim of investigating the role of emp4 during post-germinative development, homozygous mutant seedlings were obtained by cultivation of excised immature embryos on a synthetic medium. In the mutants both germination frequency as well as the proportion of seedlings reaching the first and second leaf stages were reduced. The anatomy of the leaf blades and the root cortex was not affected by the mutation, however severe alterations such as the presence of empty cells or cells containing poorly organized organelles, were observed. Moreover both mitochondria and chloroplast functionality was impaired in the mutants. Our hypothesis is that mitochondrial impairment, the primary effect of the mutation, causes secondary effects on the development of other cellular organelles. Ultra-structural features of mutant leaf blade mesophyll cells are reminiscent of cells undergoing senescence. Interestingly, both structural and functional damage was less severe in seedlings grown in total darkness compared with those exposed to light, thus suggesting that the effects of the mutation are enhanced by the presence of light. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Structural Features of Sugars That Trigger or Support Conidial Germination in the Filamentous Fungus Aspergillus niger

PubMed Central

Hayer, Kimran; Stratford, Malcolm

2013-01-01

The asexual spores (conidia) of Aspergillus niger germinate to produce hyphae under appropriate conditions. Germination is initiated by conidial swelling and mobilization of internal carbon and energy stores, followed by polarization and emergence of a hyphal germ tube. The effects of different pyranose sugars, all analogues of d-glucose, on the germination of A. niger conidia were explored, and we define germination as the transition from a dormant conidium into a germling. Within germination, we distinguish two distinct stages, the initial swelling of the conidium and subsequent polarized growth. The stage of conidial swelling requires a germination trigger, which we define as a compound that is sensed by the conidium and which leads to catabolism of d-trehalose and isotropic growth. Sugars that triggered germination and outgrowth included d-glucose, d-mannose, and d-xylose. Sugars that triggered germination but did not support subsequent outgrowth included d-tagatose, d-lyxose, and 2-deoxy-d-glucose. Nontriggering sugars included d-galactose, l-glucose, and d-arabinose. Certain nontriggering sugars, including d-galactose, supported outgrowth if added in the presence of a complementary triggering sugar. This division of functions indicates that sugars are involved in two separate events in germination, triggering and subsequent outgrowth, and the structural features of sugars that support each, both, or none of these events are discussed. We also present data on the uptake of sugars during the germination process and discuss possible mechanisms of triggering in the absence of apparent sugar uptake during the initial swelling of conidia. PMID:23995938

Structural features of sugars that trigger or support conidial germination in the filamentous fungus Aspergillus niger.

PubMed

Hayer, Kimran; Stratford, Malcolm; Archer, David B

2013-11-01

The asexual spores (conidia) of Aspergillus niger germinate to produce hyphae under appropriate conditions. Germination is initiated by conidial swelling and mobilization of internal carbon and energy stores, followed by polarization and emergence of a hyphal germ tube. The effects of different pyranose sugars, all analogues of d-glucose, on the germination of A. niger conidia were explored, and we define germination as the transition from a dormant conidium into a germling. Within germination, we distinguish two distinct stages, the initial swelling of the conidium and subsequent polarized growth. The stage of conidial swelling requires a germination trigger, which we define as a compound that is sensed by the conidium and which leads to catabolism of d-trehalose and isotropic growth. Sugars that triggered germination and outgrowth included d-glucose, d-mannose, and d-xylose. Sugars that triggered germination but did not support subsequent outgrowth included d-tagatose, d-lyxose, and 2-deoxy-d-glucose. Nontriggering sugars included d-galactose, l-glucose, and d-arabinose. Certain nontriggering sugars, including d-galactose, supported outgrowth if added in the presence of a complementary triggering sugar. This division of functions indicates that sugars are involved in two separate events in germination, triggering and subsequent outgrowth, and the structural features of sugars that support each, both, or none of these events are discussed. We also present data on the uptake of sugars during the germination process and discuss possible mechanisms of triggering in the absence of apparent sugar uptake during the initial swelling of conidia.

Seed Hydropriming and Smoke Water Significantly Improve Low-Temperature Germination of Lupinus angustifolius L.

PubMed

Płażek, Agnieszka; Dubert, Franciszek; Kopeć, Przemysław; Dziurka, Michał; Kalandyk, Agnieszka; Pastuszak, Jakub; Wolko, Bogdan

2018-03-26

Seed imbibition under cold temperature is dangerous when dry seeds have relatively low water content. The aim of this study was to investigate germination of 20 lines/cultivars of narrow-leaf lupine at 7 °C (cold) and 13 °C (control) under the influence of smoke water and following seed hydropriming for 3 h at 20 °C. The efficacy of individual treatments was examined with regard to seed protection during low-temperature germination. Based on seed germination, vigour at cold was evaluated four days after sowing by means of hypocotyl length, the studied lines/cultivars were divided into three groups with low, high and very high germination rates. Germination vigour correlated with cell membrane permeability, dehydrogenase activity and abscisic acid (ABA) content and was analysed in the seeds one day after sowing. Gibberellin content did not correlate with germination vigour. The seeds of weakly germinating lines/cultivars had the highest cell permeability and ABA content as well as the lowest amylolytic activity at both studied temperatures. Additionally, the vigour of weakly germinating seeds at 7 °C correlated with dehydrogenase activity. Three-hour hydropriming was the most effective for seed germination under cold due to reduced cell membrane permeability and ABA level. Stimulating effects of smoke water on germination under cold could be explained by enhanced dehydrogenase activity.

Complexes of D-type cyclins with CDKs during maize germination

PubMed Central

Vázquez-Ramos, Jorge M.

2013-01-01

The importance of cell proliferation in plant growth and development has been well documented. The majority of studies on basic cell cycle mechanisms in plants have been at the level of gene expression and much less knowledge has accumulated in terms of protein interactions and activation. Two key proteins, cyclins and cyclin-dependent kinases (CDKs) are fundamental for cell cycle regulation and advancement. Our aim has been to understand the role of D-type cyclins and type A and B CDKs in the cell cycle taking place during a developmental process such as maize seed germination. Results indicate that three maize D-type cyclins—D2;2, D4;2, and D5;3—(G1-S cyclins by definition) bind and activate two different types of CDK—A and B1;1—in a differential way during germination. Whereas CDKA–D-type cyclin complexes are more active at early germination times than at later times, it was surprising to observe that CDKB1;1, a supposedly G2-M kinase, bound in a differential way to all D-type cyclins tested during germination. Binding to cyclin D2;2 was detectable at all germination times, forming a complex with kinase activity, whereas binding to D4;2 and D5;3 was more variable; in particular, D5;3 was only detected at late germination times. Results are discussed in terms of cell cycle advancement and its importance for seed germination. PMID:24127516

Testing Nucleoside Analogues as Inhibitors of Bacillus anthracis Spore Germination In Vitro and in Macrophage Cell Culture ▿

PubMed Central

Alvarez, Zadkiel; Lee, Kyungae; Abel-Santos, Ernesto

2010-01-01

Bacillus anthracis, the etiological agent of anthrax, has a dormant stage in its life cycle known as the endospore. When conditions become favorable, spores germinate and transform into vegetative bacteria. In inhalational anthrax, the most fatal manifestation of the disease, spores enter the organism through the respiratory tract and germinate in phagosomes of alveolar macrophages. Germinated cells can then produce toxins and establish infection. Thus, germination is a crucial step for the initiation of pathogenesis. B. anthracis spore germination is activated by a wide variety of amino acids and purine nucleosides. Inosine and l-alanine are the two most potent nutrient germinants in vitro. Recent studies have shown that germination can be hindered by isomers or structural analogues of germinants. 6-Thioguanosine (6-TG), a guanosine analogue, is able to inhibit germination and prevent B. anthracis toxin-mediated necrosis in murine macrophages. In this study, we screened 46 different nucleoside analogues as activators or inhibitors of B. anthracis spore germination in vitro. These compounds were also tested for their ability to protect the macrophage cell line J774a.1 from B. anthracis cytotoxicity. Structure-activity relationship analysis of activators and inhibitors clarified the binding mechanisms of nucleosides to B. anthracis spores. In contrast, no structure-activity relationships were apparent for compounds that protected macrophages from B. anthracis-mediated killing. However, multiple inhibitors additively protected macrophages from B. anthracis. PMID:20921305

Evaluation of pathogenicity of Candida albicans in germination-ready states using a silkworm infection model.

PubMed

Matsumoto, Haruhito; Nagao, Jun-ichi; Cho, Tamaki; Kodama, Jun

2013-01-01

We previously developed an N-acetyl-D-glucosamine (GlcNAc) medium which induces Candida albicans to undergo a yeast-to-hyphal transition through a cAMP-PKA pathway. Microarray analysis demonstrated that 18 genes, including ALS3 that encodes a cell wall adhesion, were upregulated by 30-min incubation of yeast cells at 37°C in the GlcNAc medium. To investigate the differences between morphological transition and morphotype in C. albicans as a consequence of infection, this study utilized a silkworm infection model as an invertebrate mini-host. We prepared 3 different conditions of C. albicans cells in vitro by changing the incubation times in the GlcNAc medium: yeast-form cells at 0 min (Y0 cells), yeast-form cells in germination-ready state at 60 min (Y60 cells), and hyphal cells at 120 min (H120 cells), and compared their pathogenicities. We performed the infection study at various temperatures to find temperature-dependent virulence factors in vivo. Y60 cells in germination-ready state in the GlcNAc medium showed higher pathogenicity in vivo compared to Y0 and H120 cells at 30°C. Y60 cells proliferated in silkworms 24 h post-injection at 30°C, whereas the other 2 cell types did not. In vitro analysis demonstrated that Y60 cells, but not Y0 cells, germinated in the silkworm hemolymph at 30°C. However, Y0 and Y60 cells showed a similar degree of germination in the silkworm hemolymph at 37°C, although no significant difference in silkworm survival after infection with each cell type was observed at 37°C. These results suggested that the germination-ready state induced by the GlcNAc medium contributed to virulence in the silkworm.

Histochemical studies on protease formation in the cotyledons of germinating bean seeds.

PubMed

Yomo, H; Taylor, M P

1973-03-01

Protease formation in Phaseolus vulgaris L. cotyledons during seed germination was studied histochemically using a gelatin-film-substrate method. Protease activity can be detected by this method on the 5th day of germination, at approximately the same time that a rapid increase of activity was observed by a test-tube assay with casein as a substrate. At the early stage of germination, protease activity was observed throughout the cotyledon except in two or three cell layers below the cotyledon surface and in several cell layers around the vascular bundles. A highly active cell layer surrounding the protease-inactive cells near the vascular bundles is suggested to be a source of the protease.

Chloroplast Growth and Replication in Germinating Spinach Cotyledons following Massive γ-Irradiation of the Seed

PubMed Central

Rose, Ray; Possingham, John

1976-01-01

Spinach seeds (Spinacia oleracea L.) given massive doses of γ-irradiation (500 krad) germinate and form a seedling with two green cotyledons and a radicle, but develop no further. Irradiated cotyledons show no increase in cell number or total DNA over a 7-day period in the light, while in control cotyledons there is a small increase in cell number and large increases in total DNA and chloroplast number. The chloroplasts of irradiated cotyledons are delayed in their division, become greatly enlarged and contain large amounts of starch. The whole population of chloroplasts subsequently undergoes a wave of division. The daughter chloroplasts show normal thylakoid development, but have some abnormal structural features caused by the radiation stress. Information on the effect of X-irradiation, ultraviolet irradiation, and 5-fluorodeoxyuridine on chloroplast replication and on chloroplast and nuclear DNA synthesis was obtained from cultured spinach leaf discs. It appears that chloroplast replication is more resistant to ionizing radiation than cell division and can proceed in the absence of nuclear DNA synthesis and greatly reduced chloroplast DNA synthesis. Images PMID:16659421

Human CD30+ B cells represent a unique subset related to Hodgkin lymphoma cells.

PubMed

Weniger, Marc A; Tiacci, Enrico; Schneider, Stefanie; Arnolds, Judith; Rüschenbaum, Sabrina; Duppach, Janine; Seifert, Marc; Döring, Claudia; Hansmann, Martin-Leo; Küppers, Ralf

2018-06-11

Very few B cells in germinal centers (GCs) and extrafollicular (EF) regions of lymph nodes express CD30. Their specific features and relationship to CD30-expressing Hodgkin and Reed/Sternberg (HRS) cells of Hodgkin lymphoma are unclear but highly relevant, because numerous patients with lymphoma are currently treated with an anti-CD30 immunotoxin. We performed a comprehensive analysis of human CD30+ B cells. Phenotypic and IgV gene analyses indicated that CD30+ GC B lymphocytes represent typical GC B cells, and that CD30+ EF B cells are mostly post-GC B cells. The transcriptomes of CD30+ GC and EF B cells largely overlapped, sharing a strong MYC signature, but were strikingly different from conventional GC B cells and memory B and plasma cells, respectively. CD30+ GC B cells represent MYC+ centrocytes redifferentiating into centroblasts; CD30+ EF B cells represent active, proliferating memory B cells. HRS cells shared typical transcriptome patterns with CD30+ B cells, suggesting that they originate from these lymphocytes or acquire their characteristic features during lymphomagenesis. By comparing HRS to normal CD30+ B cells we redefined aberrant and disease-specific features of HRS cells. A remarkable downregulation of genes regulating genomic stability and cytokinesis in HRS cells may explain their genomic instability and multinuclearity.

Cytological and Proteomic Analyses of Osmunda cinnamomea Germinating Spores Reveal Characteristics of Fern Spore Germination and Rhizoid Tip Growth.

PubMed

Suo, Jinwei; Zhao, Qi; Zhang, Zhengxiu; Chen, Sixue; Cao, Jian'guo; Liu, Guanjun; Wei, Xing; Wang, Tai; Yang, Chuanping; Dai, Shaojun

2015-09-01

Fern spore is a good single-cell model for studying the sophisticated molecular networks in asymmetric cell division, differentiation, and polar growth. Osmunda cinnamomea L. var. asiatica is one of the oldest fern species with typical separate-growing trophophyll and sporophyll. The chlorophyllous spores generated from sporophyll can germinate without dormancy. In this study, the spore ultrastructure, antioxidant enzyme activities, as well as protein and gene expression patterns were analyzed in the course of spore germination at five typical stages (i.e. mature spores, rehydrated spores, double-celled spores, germinated spores, and spores with protonemal cells). Proteomic analysis revealed 113 differentially expressed proteins, which were mainly involved in photosynthesis, reserve mobilization, energy supplying, protein synthesis and turnover, reactive oxygen species scavenging, signaling, and cell structure modulation. The presence of multiple proteoforms of 25 differentially expressed proteins implies that post-translational modification may play important roles in spore germination. The dynamic patterns of proteins and their encoding genes exhibited specific characteristics in the processes of cell division and rhizoid tip growth, which include heterotrophic and autotrophic metabolisms, de novo protein synthesis and active protein turnover, reactive oxygen species and hormone (brassinosteroid and ethylene) signaling, and vesicle trafficking and cytoskeleton dynamic. In addition, the function skew of proteins in fern spores highlights the unique and common mechanisms when compared with evolutionarily divergent spermatophyte pollen. These findings provide an improved understanding of the typical single-celled asymmetric division and polar growth during fern spore germination. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

THE CELLULAR ORIGIN OF HUMAN IMMUNOGLOBULINS (γ2, γ1M, γ1A)

PubMed Central

Mellors, Robert C.; Korngold, Leonhard

1963-01-01

A study was made of the cellular origin of human immunoglobulins (γ2, γ1M, γ1A). The results indicated that two closely related families of cells form immunoglobulins in human lymphoid tissue: germinal (reticular) centers and plasma cells. Thus their cellular origin in addition to their known antigenic relations further justifies placing the immunoglobulins in one family of proteins. Immunoglobulins were also formed to a small extent in primitive reticular cells which resembled those of germinal centers but were separated from them. Possibly such cells were undergoing transition to the much more numerous plasma cells with which they were commonly associated. The mantles of small lymphocytes which surrounded germinal centers did not contain detectable quantities of immunoglobulins. While in general only one type of immunoglobulin was present in an individual cell or germinal center, γ2- and γ1M-globulin were identified on occasion in the same plasma cell and germinal center. A peculiarity of the fetal thymus gland was the presence of immunoglobulin, mainly γ1M, in a small number of cells of small and intermediate size and primitive reticular appearance and in Hassall's corpuscles. PMID:14077999

Re-analysis of protein data reveals the germination pathway and up accumulation mechanism of cell wall hydrolases during the radicle protrusion step of seed germination in Podophyllum hexandrum- a high altitude plant

PubMed Central

Dogra, Vivek; Bagler, Ganesh; Sreenivasulu, Yelam

2015-01-01

Podophyllum hexandrum Royle is an important high-altitude plant of Himalayas with immense medicinal value. Earlier, it was reported that the cell wall hydrolases were up accumulated during radicle protrusion step of Podophyllum seed germination. In the present study, Podophyllum seed Germination protein interaction Network (PGN) was constructed by using the differentially accumulated protein (DAP) data set of Podophyllum during the radicle protrusion step of seed germination, with reference to Arabidopsis protein–protein interaction network (AtPIN). The developed PGN is comprised of a giant cluster with 1028 proteins having 10,519 interactions and a few small clusters with relevant gene ontological signatures. In this analysis, a germination pathway related cluster which is also central to the topology and information dynamics of PGN was obtained with a set of 60 key proteins. Among these, eight proteins which are known to be involved in signaling, metabolism, protein modification, cell wall modification, and cell cycle regulation processes were found commonly highlighted in both the proteomic and interactome analysis. The systems-level analysis of PGN identified the key proteins involved in radicle protrusion step of seed germination in Podophyllum. PMID:26579141

ENZYMES OF GLUCOSE AND PYRUVATE CATABOLISM IN CELLS, SPORES, AND GERMINATED SPORES OF CLOSTRIDIUM BOTULINUM1

PubMed Central

Simmons, Richard J.; Costilow, Ralph N.

1962-01-01

Simmons, R. J. (Michigan State University, East Lansing), and R. N. Costilow. Enzymes of glucose and pyruvate catabolism in cells, spores, and germinated spores of Clostridium botulinum. J. Bacteriol. 84:1274–1281. 1962.—An investigation was made of the enzymes of vegetative cells, spores, and germinated spores of Clostridium botulinum 62-A to elucidate a pathway of glucose metabolism. Manometric studies were conducted with intact cells, and various enzymes and enzyme systems were assayed in cell-free and spore-free extracts by use of spectrophotometric and colorimetric procedures. Glucose fermentation was found to be inducible; glucokinase was the controlling enzyme. All other enzymes of the Embden-Meyerhof-Parnas (EMP) pathway were found in both induced and non-induced cells, but they were in relatively low concentrations in the latter. This, plus the fact that no glucose-6-phosphate dehydrogenase was detected, led to the conclusion that glucose is catabolized primarily by the EMP system. A number of glycolytic enzymes were also found in extracts of spores and germinated spores of this organism, but the activities were extremely low as compared with activities in cell extracts. A phosphoroclastic-type reaction was readily demonstrated in both glucose-adapted and non-adapted cells, but not in spores and germinated spores. However, both acetokinase and phosphotransacetylase, as well as coenzyme A transphorase, were detected in spores and germinated-spore extracts, although at very low activity levels as compared with cell extracts. The specific activity of diaphorase in spore extracts was about one-half that of corresponding cell extracts, and the activity of reduced diphosphopyridine nucleotide (DPNH) oxidase was actually higher in the spore extracts. In addition, the DPNH oxidase in spore extracts was considerably more heat-stable than that in extracts of cells or germinated spores. PMID:13977433

A Waking Review: Old and Novel Insights into the Spore Germination in Streptomyces.

PubMed

Bobek, Jan; Šmídová, Klára; Čihák, Matouš

2017-01-01

The complex development undergone by Streptomyces encompasses transitions from vegetative mycelial forms to reproductive aerial hyphae that differentiate into chains of single-celled spores. Whereas their mycelial life - connected with spore formation and antibiotic production - is deeply investigated, spore germination as the counterpoint in their life cycle has received much less attention. Still, germination represents a system of transformation from metabolic zero point to a new living lap. There are several aspects of germination that may attract our attention: (1) Dormant spores are strikingly well-prepared for the future metabolic restart; they possess stable transcriptome, hydrolytic enzymes, chaperones, and other required macromolecules stabilized in a trehalose milieu; (2) Germination itself is a specific sequence of events leading to a complete morphological remodeling that include spore swelling, cell wall reconstruction, and eventually germ tube emergences; (3) Still not fully unveiled are the strategies that enable the process, including a single cell's signal transduction and gene expression control, as well as intercellular communication and the probability of germination across the whole population. This review summarizes our current knowledge about the germination process in Streptomyces , while focusing on the aforementioned points.

Effect of sublethal heat treatment on the later stage of germination-to-outgrowth of Clostridium perfringens spores.

PubMed

Sakanoue, Hideyo; Yasugi, Mayo; Miyake, Masami

2018-05-04

Sublethal heating of spores has long been known to stimulate or activate germination, but the underlying mechanisms are not yet fully understood. In this study, we visualized the entire germination-to-outgrowth process of spores from an anaerobic sporeformer, C. perfringens, at single-cell resolution. Quantitative analysis revealed that sublethal heating significantly reduced the time from completion of germination to the beginning of the first cell division. The results indicate that sublethal heating of C. perfringens spores not only sensitizes the responsiveness of germinant receptors but also directly or indirectly facilitates multiple steps during the bacterial regrowth process. © 2018 The Societies and John Wiley & Sons Australia, Ltd.

Gibberellic Acid-Stimulated Arabidopsis6 Serves as an Integrator of Gibberellin, Abscisic Acid, and Glucose Signaling during Seed Germination in Arabidopsis.

PubMed

Zhong, Chunmei; Xu, Hao; Ye, Siting; Wang, Shiyi; Li, Lingfei; Zhang, Shengchun; Wang, Xiaojing

2015-11-01

The DELLA protein REPRESSOR OF ga1-3-LIKE2 (RGL2) plays an important role in seed germination under different conditions through a number of transcription factors. However, the functions of the structural genes associated with RGL2-regulated germination are less defined. Here, we report the role of an Arabidopsis (Arabidopsis thaliana) cell wall-localized protein, Gibberellic Acid-Stimulated Arabidopsis6 (AtGASA6), in functionally linking RGL2 and a cell wall loosening expansin protein (Arabidopsis expansin A1 [AtEXPA1]), resulting in the control of embryonic axis elongation and seed germination. AtGASA6-overexpressing seeds showed precocious germination, whereas transfer DNA and RNA interference mutant seeds displayed delayed seed germination under abscisic acid, paclobutrazol, and glucose (Glc) stress conditions. The differences in germination rates resulted from corresponding variation in cell elongation in the hypocotyl-radicle transition region of the embryonic axis. AtGASA6 was down-regulated by RGL2, GLUCOSE INSENSITIVE2, and ABSCISIC ACID-INSENSITIVE5 genes, and loss of AtGASA6 expression in the gasa6 mutant reversed the insensitivity shown by the rgl2 mutant to paclobutrazol and the gin2 mutant to Glc-induced stress, suggesting that it is involved in regulating both the gibberellin and Glc signaling pathways. Furthermore, it was found that the promotion of seed germination and length of embryonic axis by AtGASA6 resulted from a promotion of cell elongation at the embryonic axis mediated by AtEXPA1. Taken together, the data indicate that AtGASA6 links RGL2 and AtEXPA1 functions and plays a role as an integrator of gibberellin, abscisic acid, and Glc signaling, resulting in the regulation of seed germination through a promotion of cell elongation. © 2015 American Society of Plant Biologists. All Rights Reserved.

Gibberellic Acid-Stimulated Arabidopsis6 Serves as an Integrator of Gibberellin, Abscisic Acid, and Glucose Signaling during Seed Germination in Arabidopsis1[OPEN

PubMed Central

Zhong, Chunmei; Xu, Hao; Ye, Siting; Wang, Shiyi; Li, Lingfei; Zhang, Shengchun; Wang, Xiaojing

2015-01-01

The DELLA protein REPRESSOR OF ga1-3-LIKE2 (RGL2) plays an important role in seed germination under different conditions through a number of transcription factors. However, the functions of the structural genes associated with RGL2-regulated germination are less defined. Here, we report the role of an Arabidopsis (Arabidopsis thaliana) cell wall-localized protein, Gibberellic Acid-Stimulated Arabidopsis6 (AtGASA6), in functionally linking RGL2 and a cell wall loosening expansin protein (Arabidopsis expansin A1 [AtEXPA1]), resulting in the control of embryonic axis elongation and seed germination. AtGASA6-overexpressing seeds showed precocious germination, whereas transfer DNA and RNA interference mutant seeds displayed delayed seed germination under abscisic acid, paclobutrazol, and glucose (Glc) stress conditions. The differences in germination rates resulted from corresponding variation in cell elongation in the hypocotyl-radicle transition region of the embryonic axis. AtGASA6 was down-regulated by RGL2, GLUCOSE INSENSITIVE2, and ABSCISIC ACID-INSENSITIVE5 genes, and loss of AtGASA6 expression in the gasa6 mutant reversed the insensitivity shown by the rgl2 mutant to paclobutrazol and the gin2 mutant to Glc-induced stress, suggesting that it is involved in regulating both the gibberellin and Glc signaling pathways. Furthermore, it was found that the promotion of seed germination and length of embryonic axis by AtGASA6 resulted from a promotion of cell elongation at the embryonic axis mediated by AtEXPA1. Taken together, the data indicate that AtGASA6 links RGL2 and AtEXPA1 functions and plays a role as an integrator of gibberellin, abscisic acid, and Glc signaling, resulting in the regulation of seed germination through a promotion of cell elongation. PMID:26400990

Redox Changes During the Cell Cycle in the Embryonic Root Meristem of Arabidopsis thaliana.

PubMed

de Simone, Ambra; Hubbard, Rachel; de la Torre, Natanael Viñegra; Velappan, Yazhini; Wilson, Michael; Considine, Michael J; Soppe, Wim J J; Foyer, Christine H

2017-12-20

The aim of this study was to characterize redox changes in the nuclei and cytosol occurring during the mitotic cell cycle in the embryonic roots of germinating Arabidopsis seedlings, and to determine how redox cycling was modified in mutants with a decreased capacity for ascorbate synthesis. Using an in vivo reduction-oxidation (redox) reporter (roGFP2), we show that transient oxidation of the cytosol and the nuclei occurred at G1 in the synchronized dividing cells of the Arabidopsis root apical meristem, with reduction at G2 and mitosis. This redox cycle was absent from low ascorbate mutants in which nuclei were significantly more oxidized than controls. The cell cycle-dependent increase in nuclear size was impaired in the ascorbate-deficient mutants, which had fewer cells per unit area in the root proliferation zone. The transcript profile of the dry seeds and size of the imbibed seeds was strongly influenced by low ascorbate but germination, dormancy release and seed aging characteristics were unaffected. These data demonstrate the presence of a redox cycle within the plant cell cycle and that the redox state of the nuclei is an important factor in cell cycle progression. Controlled oxidation is a key feature of the early stages of the plant cell cycle. However, sustained mild oxidation restricts nuclear functions and impairs progression through the cell cycle leading to fewer cells in the root apical meristem. Antioxid. Redox Signal. 27, 1505-1519.

HRS1 acts as a negative regulator of abscisic acid signaling to promote timely germination of Arabidopsis seeds.

PubMed

Wu, Chongming; Feng, Juanjuan; Wang, Ran; Liu, Hong; Yang, Huixia; Rodriguez, Pedro L; Qin, Huanju; Liu, Xin; Wang, Daowen

2012-01-01

In this work, we conducted functional analysis of Arabidopsis HRS1 gene in order to provide new insights into the mechanisms governing seed germination. Compared with wild type (WT) control, HRS1 knockout mutant (hrs1-1) exhibited significant germination delays on either normal medium or those supplemented with abscisic acid (ABA) or sodium chloride (NaCl), with the magnitude of the delay being substantially larger on the latter media. The hypersensitivity of hrs1-1 germination to ABA and NaCl required ABI3, ABI4 and ABI5, and was aggravated in the double mutant hrs1-1abi1-2 and triple mutant hrs1-1hab1-1abi1-2, indicating that HRS1 acts as a negative regulator of ABA signaling during seed germination. Consistent with this notion, HRS1 expression was found in the embryo axis, and was regulated both temporally and spatially, during seed germination. Further analysis showed that the delay of hrs1-1 germination under normal conditions was associated with reduction in the elongation of the cells located in the lower hypocotyl (LH) and transition zone (TZ) of embryo axis. Interestingly, the germination rate of hrs1-1 was more severely reduced by the inhibitor of cell elongation, and more significantly decreased by the suppressors of plasmalemma H(+)-ATPase activity, than that of WT control. The plasmalemma H(+)-ATPase activity in the germinating seeds of hrs1-1 was substantially lower than that exhibited by WT control, and fusicoccin, an activator of this pump, corrected the transient germination delay of hrs1-1. Together, our data suggest that HRS1 may be needed for suppressing ABA signaling in germinating embryo axis, which promotes the timely germination of Arabidopsis seeds probably by facilitating the proper function of plasmalemma H(+)-ATPase and the efficient elongation of LH and TZ cells.

HRS1 Acts as a Negative Regulator of Abscisic Acid Signaling to Promote Timely Germination of Arabidopsis Seeds

PubMed Central

Wang, Ran; Liu, Hong; Yang, Huixia; Rodriguez, Pedro L.; Qin, Huanju; Liu, Xin; Wang, Daowen

2012-01-01

In this work, we conducted functional analysis of Arabidopsis HRS1 gene in order to provide new insights into the mechanisms governing seed germination. Compared with wild type (WT) control, HRS1 knockout mutant (hrs1-1) exhibited significant germination delays on either normal medium or those supplemented with abscisic acid (ABA) or sodium chloride (NaCl), with the magnitude of the delay being substantially larger on the latter media. The hypersensitivity of hrs1-1 germination to ABA and NaCl required ABI3, ABI4 and ABI5, and was aggravated in the double mutant hrs1-1abi1-2 and triple mutant hrs1-1hab1-1abi1-2, indicating that HRS1 acts as a negative regulator of ABA signaling during seed germination. Consistent with this notion, HRS1 expression was found in the embryo axis, and was regulated both temporally and spatially, during seed germination. Further analysis showed that the delay of hrs1-1 germination under normal conditions was associated with reduction in the elongation of the cells located in the lower hypocotyl (LH) and transition zone (TZ) of embryo axis. Interestingly, the germination rate of hrs1-1 was more severely reduced by the inhibitor of cell elongation, and more significantly decreased by the suppressors of plasmalemma H+-ATPase activity, than that of WT control. The plasmalemma H+-ATPase activity in the germinating seeds of hrs1-1 was substantially lower than that exhibited by WT control, and fusicoccin, an activator of this pump, corrected the transient germination delay of hrs1-1. Together, our data suggest that HRS1 may be needed for suppressing ABA signaling in germinating embryo axis, which promotes the timely germination of Arabidopsis seeds probably by facilitating the proper function of plasmalemma H+-ATPase and the efficient elongation of LH and TZ cells. PMID:22545134

Cell of origin associated classification of B-cell malignancies by gene signatures of the normal B-cell hierarchy.

PubMed

Johnsen, Hans Erik; Bergkvist, Kim Steve; Schmitz, Alexander; Kjeldsen, Malene Krag; Hansen, Steen Møller; Gaihede, Michael; Nørgaard, Martin Agge; Bæch, John; Grønholdt, Marie-Louise; Jensen, Frank Svendsen; Johansen, Preben; Bødker, Julie Støve; Bøgsted, Martin; Dybkær, Karen

2014-06-01

Recent findings have suggested biological classification of B-cell malignancies as exemplified by the "activated B-cell-like" (ABC), the "germinal-center B-cell-like" (GCB) and primary mediastinal B-cell lymphoma (PMBL) subtypes of diffuse large B-cell lymphoma and "recurrent translocation and cyclin D" (TC) classification of multiple myeloma. Biological classification of B-cell derived cancers may be refined by a direct and systematic strategy where identification and characterization of normal B-cell differentiation subsets are used to define the cancer cell of origin phenotype. Here we propose a strategy combining multiparametric flow cytometry, global gene expression profiling and biostatistical modeling to generate B-cell subset specific gene signatures from sorted normal human immature, naive, germinal centrocytes and centroblasts, post-germinal memory B-cells, plasmablasts and plasma cells from available lymphoid tissues including lymph nodes, tonsils, thymus, peripheral blood and bone marrow. This strategy will provide an accurate image of the stage of differentiation, which prospectively can be used to classify any B-cell malignancy and eventually purify tumor cells. This report briefly describes the current models of the normal B-cell subset differentiation in multiple tissues and the pathogenesis of malignancies originating from the normal germinal B-cell hierarchy.

In vitro study on effect of germinated wheat on human breast cancer cells

USDA-ARS?s Scientific Manuscript database

This research investigated the possible anti-cancer effects of germinated wheat flours (GWF) on cell growth and apoptosis of human breast cancer cells. In a series of in vitro experiments, estrogen receptor-positive (MCF-7) and negative (MDA-MB-231) cells were cultured and treated with GWF that wer...

Physico-chemical factors influencing spore germination in cyanobacterium Fischerella muscicola.

PubMed

Mishra, Biranchi N; Kaushik, Manish S; Abraham, Gerard; Singh, Pawan K

2018-06-19

Spore (akinete) formation in the heterocystous and branched filamentous cyanobacterium Fischerella muscicola involves a significant increase in cell size and formation of several endospores in each of the cells. In present study, the physico-chemical factors (pH, light sources, nutrient deficiency, nitrogen sources, carbon sources, and growth hormones) affecting the germination of spores of F. muscicola were examined. Increase in spore germination frequency was detected above pH 8 with maximum germination (46.04%) recorded at pH 9, whereas a significant decrease in germination was observed at pH 6 when compared to control (pH 7.6). Spore germination was not observed at pH 5. Among light sources germination frequency followed the following order, that is, red light (39.9%) > white light (33.8%) > yellow light (3.4%) > green light (1.3%) whereas germination did not take place in dark and blue light. Ammonium chloride (NH 4 Cl) supported maximum (99.5%) germination frequency followed by calcium nitrate (Ca(NO 3 ) 2 ), potassium nitrate (KNO 3 ), and minimum germination was observed in urea. Nutrient (phosphorus, calcium, and magnesium) deficiency significantly enhanced the germination frequency with maximum increase in magnesium (Mg) deficient condition. Further, supplementation of carbon sources (glucose, fructose, and sodium acetate) and growth hormones (IAA and GA) also enhanced the germination frequency in this cyanobacterium. Therefore, it may be concluded that, those factors supporting higher germination frequency could be considered for successful production and use of this cyanobacterium in biofertilizer and other algal production technologies. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Reorganization of plasma membrane lipid domains during conidial germination.

PubMed

Santos, Filipa C; Fernandes, Andreia S; Antunes, Catarina A C; Moreira, Filipe P; Videira, Arnaldo; Marinho, H Susana; de Almeida, Rodrigo F M

2017-02-01

Neurospora crassa, a filamentous fungus, in the unicellular conidial stage has ideal features to study sphingolipid (SL)-enriched domains, which are implicated in fundamental cellular processes ranging from antifungal resistance to apoptosis. Several changes in lipid metabolism and in the membrane composition of N. crassa occur during spore germination. However, the biophysical impact of those changes is unknown. Thus, a biophysical study of N. crassa plasma membrane, particularly SL-enriched domains, and their dynamics along conidial germination is prompted. Two N. crassa strains, wild-type (WT) and slime, which is devoid of cell wall, were studied. Conidial growth of N. crassa WT from a dormancy state to an exponential phase was accompanied by membrane reorganization, namely an increase of membrane fluidity, occurring faster in a supplemented medium than in Vogel's minimal medium. Gel-like domains, likely enriched in SLs, were found in both N. crassa strains, but were particularly compact, rigid and abundant in the case of slime cells, even more than in budding yeast Saccharomyces cerevisiae. In N. crassa, our results suggest that the melting of SL-enriched domains occurs near growth temperature (30°C) for WT, but at higher temperatures for slime. Regarding biophysical properties strongly affected by ergosterol, the plasma membrane of slime conidia lays in between those of N. crassa WT and S. cerevisiae cells. The differences in biophysical properties found in this work, and the relationships established between membrane lipid composition and dynamics, give new insights about the plasma membrane organization and structure of N. crassa strains during conidial growth. Copyright © 2016 Elsevier B.V. All rights reserved.

Knockin' on pollen's door: live cell imaging of early polarization events in germinating Arabidopsis pollen

PubMed Central

Vogler, Frank; Konrad, Sebastian S. A.; Sprunck, Stefanie

2015-01-01

Pollen tubes are an excellent system for studying the cellular dynamics and complex signaling pathways that coordinate polarized tip growth. Although several signaling mechanisms acting in the tip-growing pollen tube have been described, our knowledge on the subcellular and molecular events during pollen germination and growth site selection at the pollen plasma membrane is rather scarce. To simultaneously track germinating pollen from up to 12 genetically different plants we developed an inexpensive and easy mounting technique, suitable for every standard microscope setup. We performed high magnification live-cell imaging during Arabidopsis pollen activation, germination, and the establishment of pollen tube tip growth by using fluorescent marker lines labeling either the pollen cytoplasm, vesicles, the actin cytoskeleton or the sperm cell nuclei and membranes. Our studies revealed distinctive vesicle and F-actin polarization during pollen activation and characteristic growth kinetics during pollen germination and pollen tube formation. Initially, the germinating Arabidopsis pollen tube grows slowly and forms a uniform roundish bulge, followed by a transition phase with vesicles heavily accumulating at the growth site before switching to rapid tip growth. Furthermore, we found the two sperm cells to be transported into the pollen tube after the phase of rapid tip growth has been initiated. The method presented here is suitable to quantitatively study subcellular events during Arabidopsis pollen germination and growth, and for the detailed analysis of pollen mutants with respect to pollen polarization, bulging, or growth site selection at the pollen plasma membrane. PMID:25954283

Immunolocalization and Changes of Hydroxyproline-Rich Glycoproteins During Symbiotic Germination of Dendrobium officinale.

PubMed

Li, Yuan-Yuan; Chen, Xiao-Mei; Zhang, Ying; Cho, Yu-Hsiu; Wang, Ai-Rong; Yeung, Edward C; Zeng, Xu; Guo, Shun-Xing; Lee, Yung-I

2018-01-01

Hydroxyproline-rich glycoproteins (HRGPs) are abundant cell wall components involved in mycorrhizal symbiosis, but little is known about their function in orchid mycorrhizal association. To gain further insight into the role of HRGPs in orchid symbiosis, the location and function of HRGPs were investigated during symbiotic germination of Dendrobium officinale . The presence of JIM11 epitope in developing protocorms was determined using immunodot blots and immunohistochemical staining procedures. Real-time PCR was also employed to verify the expression patterns of genes coding for extensin-like genes selected from the transcriptomic database. The importance of HRGPs in symbiotic germination was further investigated using 3,4-dehydro-L-proline (3,4-DHP), an inhibitor of HRGP biosynthesis. In symbiotic cultures, immunodot blots of JIM11 signals were moderate in mature seeds, and the signals became stronger in swollen embryos. After germination, signal intensities decreased in developing protocorms. In contrast, in asymbiotic cultures, JIM11 signals were much lower as compared with those stages in symbiotic cultures. Immunofluorescence staining enabled the visualization of JIM11 epitope in mature embryo and protocorm cells. Positive signals were initially localized in the larger cells near the basal (suspensor) end of uninfected embryos, marking the future colonization site of fungal hyphae. After 1 week of inoculation, the basal end of embryos had been colonized, and a strong signal was detected mostly at the mid- and basal regions of the enlarging protocorm. As protocorm development progressed, the signal was concentrated in the colonized cells at the basal end. In colonized cells, signals were present in the walls and intracellularly associated with hyphae and the pelotons. The precise localization of JIM11 epitope is further examined by immunogold labeling. In the colonized cells, gold particles were found mainly in the cell wall and the interfacial matrix near the fungal cell wall. Four extensin-like genes were verified to be highly up-regulated in symbiotically germinated protocorms as compared to asymbiotically germinated ones. The 3,4-DHP treatment inhibited the accumulation of HRGPs and symbiotic seed germination. In these protocorms, fungal hyphae could be found throughout the protocorms. Our results indicate that HRGPs play an important role in symbiotic germination. They can serve as markers for fungal colonization, establishing a symbiotic compartment and constraining fungal colonization inside the basal cells of protocorms.

Immunolocalization and Changes of Hydroxyproline-Rich Glycoproteins During Symbiotic Germination of Dendrobium officinale

PubMed Central

Li, Yuan-Yuan; Chen, Xiao-Mei; Zhang, Ying; Cho, Yu-Hsiu; Wang, Ai-Rong; Yeung, Edward C.; Zeng, Xu; Guo, Shun-Xing; Lee, Yung-I

2018-01-01

Hydroxyproline-rich glycoproteins (HRGPs) are abundant cell wall components involved in mycorrhizal symbiosis, but little is known about their function in orchid mycorrhizal association. To gain further insight into the role of HRGPs in orchid symbiosis, the location and function of HRGPs were investigated during symbiotic germination of Dendrobium officinale. The presence of JIM11 epitope in developing protocorms was determined using immunodot blots and immunohistochemical staining procedures. Real-time PCR was also employed to verify the expression patterns of genes coding for extensin-like genes selected from the transcriptomic database. The importance of HRGPs in symbiotic germination was further investigated using 3,4-dehydro-L-proline (3,4-DHP), an inhibitor of HRGP biosynthesis. In symbiotic cultures, immunodot blots of JIM11 signals were moderate in mature seeds, and the signals became stronger in swollen embryos. After germination, signal intensities decreased in developing protocorms. In contrast, in asymbiotic cultures, JIM11 signals were much lower as compared with those stages in symbiotic cultures. Immunofluorescence staining enabled the visualization of JIM11 epitope in mature embryo and protocorm cells. Positive signals were initially localized in the larger cells near the basal (suspensor) end of uninfected embryos, marking the future colonization site of fungal hyphae. After 1 week of inoculation, the basal end of embryos had been colonized, and a strong signal was detected mostly at the mid- and basal regions of the enlarging protocorm. As protocorm development progressed, the signal was concentrated in the colonized cells at the basal end. In colonized cells, signals were present in the walls and intracellularly associated with hyphae and the pelotons. The precise localization of JIM11 epitope is further examined by immunogold labeling. In the colonized cells, gold particles were found mainly in the cell wall and the interfacial matrix near the fungal cell wall. Four extensin-like genes were verified to be highly up-regulated in symbiotically germinated protocorms as compared to asymbiotically germinated ones. The 3,4-DHP treatment inhibited the accumulation of HRGPs and symbiotic seed germination. In these protocorms, fungal hyphae could be found throughout the protocorms. Our results indicate that HRGPs play an important role in symbiotic germination. They can serve as markers for fungal colonization, establishing a symbiotic compartment and constraining fungal colonization inside the basal cells of protocorms. PMID:29922306

In Vivo Cell Wall Loosening by Hydroxyl Radicals during Cress Seed Germination and Elongation Growth1[W][OA

PubMed Central

Müller, Kerstin; Linkies, Ada; Vreeburg, Robert A.M.; Fry, Stephen C.; Krieger-Liszkay, Anja; Leubner-Metzger, Gerhard

2009-01-01

Loosening of cell walls is an important developmental process in key stages of the plant life cycle, including seed germination, elongation growth, and fruit ripening. Here, we report direct in vivo evidence for hydroxyl radical (·OH)-mediated cell wall loosening during plant seed germination and seedling growth. We used electron paramagnetic resonance spectroscopy to show that ·OH is generated in the cell wall during radicle elongation and weakening of the endosperm of cress (Lepidium sativum; Brassicaceae) seeds. Endosperm weakening precedes radicle emergence, as demonstrated by direct biomechanical measurements. By 3H fingerprinting, we showed that wall polysaccharides are oxidized in vivo by the developmentally regulated action of apoplastic ·OH in radicles and endosperm caps: the production and action of ·OH increased during endosperm weakening and radicle elongation and were inhibited by the germination-inhibiting hormone abscisic acid. Both effects were reversed by gibberellin. Distinct and tissue-specific target sites of ·OH attack on polysaccharides were evident. In vivo ·OH attack on cell wall polysaccharides were evident not only in germinating seeds but also in elongating maize (Zea mays; Poaceae) seedling coleoptiles. We conclude that plant cell wall loosening by ·OH is a controlled action of this type of reactive oxygen species. PMID:19493972

TLR4 signaling augments B lymphocyte migration and overcomes the restriction that limits access to germinal center dark zones

PubMed Central

Hwang, Il-Young; Park, Chung; Harrison, Kathleen

2009-01-01

B lymphocyte–intrinsic Toll-like receptor (TLR) signals amplify humoral immunity and can exacerbate autoimmune diseases. We identify a new mechanism by which TLR signals may contribute to autoimmunity and chronic inflammation. We show that TLR4 signaling enhances B lymphocyte trafficking into lymph nodes (LNs), induces B lymphocyte clustering and interactions within LN follicles, leads to sustained in vivo B cell proliferation, overcomes the restriction that limits the access of nonantigen-activated B cells to germinal center dark zones, and enhances the generation of memory and plasma cells. Intravital microscopy and in vivo tracking studies of B cells transferred to recipient mice revealed that TLR4-activated, but not nonstimulated, B cells accumulated within the dark zones of preexisting germinal centers even when transferred with antigen-specific B cells. The TLR4-activated cells persist much better than nonstimulated cells, expanding both within the memory and plasma cell compartments. TLR-mediated activation of B cells may help to feed and stabilize the spontaneous and ectopic germinal centers that are so commonly found in autoimmune individuals and that accompany chronic inflammation. PMID:19917774

T regulatory cells participate in the control of germinal centre reactions

PubMed Central

Alexander, Carla-Maria; Tygrett, Lorraine T; Boyden, Alexander W; Wolniak, Kristy L; Legge, Kevin L; Waldschmidt, Thomas J

2011-01-01

Germinal centre (GC) reactions are central features of T-cell-driven B-cell responses, and the site where antibody-producing cells and memory B cells are generated. Within GCs, a range of complex cellular and molecular events occur which are critical for the generation of high affinity antibodies. These processes require exquisite regulation not only to ensure the production of desired antibodies, but to minimize unwanted autoreactive or low affinity antibodies. To assess whether T regulatory (Treg) cells participate in the control of GC responses, immunized mice were treated with an anti-glucocorticoid-induced tumour necrosis factor receptor-related protein (GITR) monoclonal antibody (mAb) to disrupt Treg-cell activity. In anti-GITR-treated mice, the GC B-cell pool was significantly larger compared with control-treated animals, with switched GC B cells composing an abnormally high proportion of the response. Dysregulated GCs were also observed regardless of strain, T helper type 1 or 2 polarizing antigens, and were also seen after anti-CD25 mAb treatment. Within the spleens of immunized mice, CXCR5+ and CCR7− Treg cells were documented by flow cytometry and Foxp3+ cells were found within GCs using immunohistology. Final studies demonstrated administration of either anti-transforming growth factor-β or anti-interleukin-10 receptor blocking mAb to likewise result in dysregulated GCs, suggesting that generation of inducible Treg cells is important in controlling the GC response. Taken together, these findings indicate that Treg cells contribute to the overall size and quality of the humoral response by controlling homeostasis within GCs. PMID:21635248

Expression of Immunoglobulin Receptors with Distinctive Features Indicating Antigen Selection by Marginal Zone B Cells from Human Spleen

PubMed Central

Colombo, Monica; Cutrona, Giovanna; Reverberi, Daniele; Bruno, Silvia; Ghiotto, Fabio; Tenca, Claudya; Stamatopoulos, Kostas; Hadzidimitriou, Anastasia; Ceccarelli, Jenny; Salvi, Sandra; Boccardo, Simona; Calevo, Maria Grazia; De Santanna, Amleto; Truini, Mauro; Fais, Franco; Ferrarini, Manlio

2013-01-01

Marginal zone (MZ) B cells, identified as surface (s)IgMhighsIgDlowCD23low/−CD21+CD38− B cells, were purified from human spleens, and the features of their V(D)J gene rearrangements were investigated and compared with those of germinal center (GC), follicular mantle (FM) and switched memory (SM) B cells. Most MZ B cells were CD27+ and exhibited somatic hypermutations (SHM), although to a lower extent than SM B cells. Moreover, among MZ B-cell rearrangements, recurrent sequences were observed, some of which displayed intraclonal diversification. The same diversifying sequences were detected in very low numbers in GC and FM B cells and only when a highly sensitive, gene-specific polymerase chain reaction was used. This result indicates that MZ B cells could expand and diversify in situ and also suggested the presence of a number of activation-induced cytidine deaminase (AID)-expressing B cells in the MZ. The notion of antigen-driven expansion/selection in situ is further supported by the VH CDR3 features of MZ B cells with highly conserved amino acids at specific positions and by the finding of shared (“stereotyped”) sequences in two different spleens. Collectively, the data are consistent with the notion that MZ B cells are a special subset selected by in situ antigenic stimuli. PMID:23877718

Osmoconditioning prevents the onset of microtubular cytoskeleton and activation of cell cycle and is detrimental for germination of Jatropha curcas L. seeds.

PubMed

de Brito, C D; Loureiro, M B; Ribeiro, P R; Vasconcelos, P C T; Fernandez, L G; de Castro, R D

2016-11-01

Jatropha curcas is an oilseed crop renowned for its tolerance to a diverse range of environmental stresses. In Brazil, this species is grown in semiarid regions where crop establishment requires a better understanding of the mechanisms underlying appropriate seed, seedling and plant behaviour under water restriction conditions. In this context, the objective of this study was to investigate the physiological and cytological profiles of J. curcas seeds in response to imbibition in water (control) and in polyethylene glycol solution (osmoticum). Seed germinability and reactivation of cell cycle events were assessed by means of different germination parameters and immunohistochemical detection of tubulin and microtubules, i.e. tubulin accumulation and microtubular cytoskeleton configurations in water imbibed seeds (control) and in seeds imbibed in the osmoticum. Immunohistochemical analysis revealed increasing accumulation of tubulin and appearance of microtubular cytoskeleton in seed embryo radicles imbibed in water from 48 h onwards. Mitotic microtubules were only visible in seeds imbibed in water, after radicle protrusion, as an indication of cell cycle reactivation and cell proliferation, with subsequent root development. Imbibition in osmoticum prevented accumulation of microtubules, i.e. activation of cell cycle, therefore germination could not be resumed. Osmoconditioned seeds were able to survive re-drying and could resume germination after re-imbibition in water, however, with lower germination performance, possibly due to acquisition of secondary dormancy. This study provides important insights into understanding of the physiological aspects of J. curcas seed germination in response to water restriction conditions. © 2016 German Botanical Society and The Royal Botanical Society of the Netherlands.

Up-regulating the abscisic acid inactivation gene ZmABA8ox1b contributes to seed germination heterosis by promoting cell expansion.

PubMed

Li, Yangyang; Wang, Cheng; Liu, Xinye; Song, Jian; Li, Hongjian; Sui, Zhipeng; Zhang, Ming; Fang, Shuang; Chu, Jinfang; Xin, Mingming; Xie, Chaojie; Zhang, Yirong; Sun, Qixin; Ni, Zhongfu

2016-04-01

Heterosis has been widely used in agriculture, but the underlying molecular principles are still largely unknown. During seed germination, we observed that maize (Zea mays) hybrid B73/Mo17 was less sensitive than its parental inbred lines to exogenous abscisic acid (ABA), and endogenous ABA content in hybrid embryos decreased more rapidly than in the parental inbred lines. ZmABA8ox1b, an ABA inactivation gene, was consistently more highly up-regulated in hybrid B73/Mo17 than in its parental inbred lines at early stages of seed germination. Moreover, ectopic expression of ZmABA8ox1b obviously promoted seed germination in Arabidopsis Remarkably, microscopic observation revealed that cell expansion played a major role in the ABA-mediated maize seed germination heterosis, which could be attributed to the altered expression of cell wall-related genes. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Portable Diagnostics and Rapid Germination

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dunn, Zachary Spencer

In the Bioenergy and Defense Department of Sandia National Laboratories, characterization of the BaDx (Bacillus anthracis diagnostic cartridge) was performed and rapid germination chemistry was investigated. BaDx was tested with complex sample matrixes inoculated with Bacillus anthracis, and the trials proved that BaDx will detect Bacillus anthracis in a variety of the medium, such as dirt, serum, blood, milk, and horse fluids. The dimensions of the device were altered to accommodate an E. coli or Listeria lateral flow immunoassay, and using a laser printer, BaDx devices were manufactured to identify E. coli and Listeria. Initial testing with E. coli versionsmore » of BaDx indicate that the device will be viable as a portable diagnostic cartridge. The device would be more effective with faster bacteria germination; hence studies were performed the use of rapid germination chemistry. Trials with calcium dipicolinic acid displayed increased cell germination, as shown by control studies using a microplate reader. Upon lyophilization the rapid germination chemistry failed to change growth patterns, indicating that the calcium dipicolinic acid was not solubilized under the conditions tested. Although incompatible with the portable diagnostic device, the experiments proved that the rapid germination chemistry was effective in increasing cell germination.« less

Germination traits explain soil seed persistence across species: the case of Mediterranean annual plants in cereal fields

PubMed Central

Saatkamp, Arne; Affre, Laurence; Dutoit, Thierry; Poschlod, Peter

2011-01-01

Background and Aims Seed persistence in the soil under field conditions is an important issue for the maintenance of local plant populations and the restoration of plant communities, increasingly so in the light of rapidly changing land use and climate change. Whereas processes important for dispersal in space are well known, knowledge of processes governing dispersal in time is still limited. Data for morphological seed traits such as size have given contradictory results for prediction of soil seed persistence or cover only a few species. There have been few experimental studies on the role of germination traits in determining soil seed persistence, while none has studied their predictive value consistently across species. Delayed germination, as well as light requirements for germination, have been suggested to contribute to the formation of persistent seed banks. Moreover, diurnally fluctuating temperatures can influence the timing of germination and are therefore linked to seed bank persistence. Methods The role of germination speed measured by T50 (days to germination of 50 % of all germinated seeds), light requirement and reaction to diurnally fluctuating temperatures in determining seed persistence in the soil was evaluated using an experimental comparative data set of 25 annual cereal weed species. Key Results It is shown that light requirements and slow germination are important features to maintain seeds ungerminated just after entering the soil, and hence influence survival of seeds in the soil. However, the detection of low diurnally fluctuating temperatures enhances soil seed bank persistence by limiting germination. Our data further suggest that the effect of diurnally fluctuating temperatures, as measured on seeds after dispersal and dry storage, is increasingly important to prevent fatal germination after longer burial periods. Conclusions These results underline the functional role of delayed germination and light for survival of seeds in the soil and hence their importance for shaping the first part of the seed decay curve. Our analyses highlight the detection of diurnally fluctuating temperatures as a third mechanism to achieve higher soil seed persistence after burial which interacts strongly with season. We therefore advocate focusing future research on mechanisms that favour soil seed persistence after longer burial times and moving from studies of morphological features to exploration of germination traits such as reaction to diurnally fluctuating temperatures. PMID:21224268

Calcineurin Orchestrates Lateral Transfer of Aspergillus fumigatus during Macrophage Cell Death.

PubMed

Shah, Anand; Kannambath, Shichina; Herbst, Susanne; Rogers, Andrew; Soresi, Simona; Carby, Martin; Reed, Anna; Mostowy, Serge; Fisher, Matthew C; Shaunak, Sunil; Armstrong-James, Darius P

2016-11-01

Pulmonary aspergillosis is a lethal mold infection in the immunocompromised host. Understanding initial control of infection and how this is altered in the immunocompromised host are key goals for comprehension of the pathogenesis of pulmonary aspergillosis. To characterize the outcome of human macrophage infection with Aspergillus fumigatus and how this is altered in transplant recipients on calcineurin inhibitor immunosuppressants. We defined the outcome of human macrophage infection with A. fumigatus, as well as the impact of calcineurin inhibitors, through a combination of single-cell fluorescence imaging, transcriptomics, proteomics, and in vivo studies. Macrophage phagocytosis of A. fumigatus enabled control of 90% of fungal germination. However, fungal germination in the late phagosome led to macrophage necrosis. During programmed necroptosis, we observed frequent cell-cell transfer of A. fumigatus between macrophages, which assists subsequent control of germination in recipient macrophages. Lateral transfer occurred through actin-dependent exocytosis of the late endosome in a vasodilator-stimulated phosphoprotein envelope. Its relevance to the control of fungal germination was also shown by direct visualization in our zebrafish aspergillosis model in vivo. The calcineurin inhibitor FK506 (tacrolimus) reduced cell death and lateral transfer in vitro by 50%. This resulted in uncontrolled fungal germination in macrophages and also resulted in hyphal escape. These observations identify programmed, necrosis-dependent lateral transfer of A. fumigatus between macrophages as an important host strategy for controlling fungal germination. This process is critically dependent on calcineurin. Our studies provide fundamental insights into the pathogenesis of pulmonary aspergillosis in the immunocompromised host.

Salt Induces Features of a Dormancy-Like State in Seeds of Eutrema (Thellungiella) salsugineum, a Halophytic Relative of Arabidopsis

PubMed Central

Kazachkova, Yana; Khan, Asif; Acuña, Tania; López-Díaz, Isabel; Carrera, Esther; Khozin-Goldberg, Inna; Fait, Aaron; Barak, Simon

2016-01-01

The salinization of land is a major factor limiting crop production worldwide. Halophytes adapted to high levels of salinity are likely to possess useful genes for improving crop tolerance to salt stress. In addition, halophytes could provide a food source on marginal lands. However, despite halophytes being salt-tolerant plants, the seeds of several halophytic species will not germinate on saline soils. Yet, little is understood regarding biochemical and gene expression changes underlying salt-mediated inhibition of halophyte seed germination. We have used the halophytic Arabidopsis relative model system, Eutrema (Thellungiella) salsugineum to explore salt-mediated inhibition of germination. We show that E. salsugineum seed germination is inhibited by salt to a far greater extent than in Arabidopsis, and that this inhibition is in response to the osmotic component of salt exposure. E. salsugineum seeds remain viable even when germination is completely inhibited, and germination resumes once seeds are transferred to non-saline conditions. Moreover, removal of the seed coat from salt-treated seeds allows embryos to germinate on salt-containing medium. Mobilization of seed storage reserves is restricted in salt-treated seeds, while many germination-associated metabolic changes are arrested or progress to a lower extent. Salt-exposed seeds are further characterized by a reduced GA/ABA ratio and increased expression of the germination repressor genes, RGL2, ABI5, and DOG1. Furthermore, a salt-mediated increase in expression of a LATE EMBRYOGENESIS ABUNDANT gene and accretion of metabolites involved in osmoprotection indicates induction of processes associated with stress tolerance, and accumulation of easily mobilized carbon reserves. Overall, our results suggest that salt inhibits E. salsugineum seed germination by inducing a seed state with molecular features of dormancy while a physical constraint to radicle emergence is provided by the seed coat layers. This seed state could facilitate survival on saline soils until a rain event(s) increases soil water potential indicating favorable conditions for seed germination and establishment of salt-tolerant E. salsugineum seedlings. PMID:27536302

Effect of animal sera on Bacillus anthracis Sterne spore germination and vegetative cell growth.

PubMed

Bensman, M D; Mackie, R S; Minter, Z A; Gutting, B W

2012-08-01

 The aims of this work were to investigate the effects of sera on B. anthracis Sterne germination and growth. Sera examined included human, monkey and rabbit sera, as well as sera from eight other species.  Standard dilution plate assay (with and without heat kill) was used as a measure of germination, and spectroscopy was used to measure growth. In addition, a Coulter Counter particle counter was used to monitor germination and growth based on bacterial size. Spores germinated best in foetal bovine and monkey sera, moderately with human sera and showed limited germination in the presence of rabbit or rat sera. Vegetative bacteria grew best in foetal bovine sera and moderately in rabbit sera. Human and monkey sera supported little growth of vegetative bacteria.  The data suggested sera can have a significant impact on germination and growth of Sterne bacteria.  These data should be considered when conducting in vitro cell culture studies and may aid in interpreting in vivo infection studies. © 2012 The Authors Journal of Applied Microbiology © 2012 The Society for Applied Microbiology.

AID-dependent activation of a MYC transgene induces multiple myeloma in a conditional mouse model of post-germinal center malignancies

PubMed Central

Chesi, Marta; Robbiani, Davide F.; Sebag, Michael; Chng, Wee Joo; Affer, Maurizio; Tiedemann, Rodger; Valdez, Riccardo; Palmer, Stephen E.; Haas, Stephanie S.; Stewart, A. Keith; Fonseca, Rafael; Kremer, Richard; Cattoretti, Giorgio; Bergsagel, P. Leif

2008-01-01

Summary By misdirecting the activity of Activation-Induced Deaminase (AID) to a conditional MYC transgene, we have achieved sporadic, AID-dependent MYC activation in germinal center B-cells of Vk*MYC mice. Whereas control C57BL/6 mice develop benign monoclonal gammopathy with age, all Vk*MYC mice progress to an indolent multiple myeloma associated with the biological and clinical features highly characteristic of the human disease. Furthermore, antigen-dependent myeloma could be induced by immunization with a T-dependent antigen. Consistent with these findings in mice, more frequent MYC rearrangements, elevated levels of MYC mRNA and MYC target genes distinguish human patients with multiple myeloma from individuals with monoclonal gammopathy, implicating a causal role for MYC in the progression of monoclonal gammopathy to multiple myeloma in man. PMID:18242516

Germinated wheat: Phytochemical composition and mixing characteristics

USDA-ARS?s Scientific Manuscript database

Germinated grain recently attracts interest due to its beneficial effect on human health. In this research, whole wheat flour samples obtained after three days and five days of germination were analyzed for biochemical components, mixing quality, and effects on human breast cancer cells. Germinati...

The Cysteine-Rich Domain of the Macrophage Mannose Receptor Is a Multispecific Lectin That Recognizes Chondroitin Sulfates a and B and Sulfated Oligosaccharides of Blood Group Lewisa and Lewisx Types in Addition to the Sulfated N-Glycans of Lutropin

PubMed Central

Leteux, Christine; Chai, Wengang; Loveless, R. Wendy; Yuen, Chun-Ting; Uhlin-Hansen, Lars; Combarnous, Yves; Jankovic, Mila; Maric, Svetlana C.; Misulovin, Ziva; Nussenzweig, Michel C.; Ten Feizi

2000-01-01

The mannose receptor (MR) is an endocytic protein on macrophages and dendritic cells, as well as on hepatic endothelial, kidney mesangial, tracheal smooth muscle, and retinal pigment epithelial cells. The extracellular portion contains two types of carbohydrate-recognition domain (CRD): eight membrane-proximal C-type CRDs and a membrane-distal cysteine-rich domain (Cys-MR). The former bind mannose-, N-acetylglucosamine-, and fucose-terminating oligosaccharides, and may be important in innate immunity towards microbial pathogens, and in antigen trapping for processing and presentation in adaptive immunity. Cys-MR binds to the sulfated carbohydrate chains of pituitary hormones and may have a role in hormonal clearance. A second feature of Cys-MR is binding to macrophages in marginal zones of the spleen, and to B cell areas in germinal centers which may help direct MR-bearing cells toward germinal centers during the immune response. Here we describe two novel classes of carbohydrate ligand for Cys-MR: chondroitin-4 sulfate chains of the type found on proteoglycans produced by cells of the immune system, and sulfated blood group chains. We further demonstrate that Cys-MR interacts with cells in the spleen via the binding site for sulfated carbohydrates. Our data suggest that the three classes of sulfated carbohydrate ligands may variously regulate the trafficking and function of MR-bearing cells. PMID:10748230

Granulocyte-macrophage colony-stimulating factor responses of oral epithelial cells to Candida albicans.

PubMed

Dongari-Bagtzoglou, A; Kashleva, H

2003-06-01

Candida albicans is the principal fungal species responsible for oropharyngeal candidiasis, the most frequent opportunistic infection associated with immune deficiencies. Cytokines, such as granulocyte-macrophage colony-stimulating factor (GM-CSF), are important in the generation of effective immunity to C. albicans. The purposes of this investigation were to determine whether C. albicans triggers secretion of GM-CSF by oral epithelial cells in vitro and to investigate mechanisms of host cell-fungal interactions that trigger such responses. Oral epithelial cell lines as well as primary oral mucosal epithelial cells were challenged with stationary phase viable C. albicans, added to human cell cultures at varying yeast:oral cell ratios. Yeast were allowed to germinate for up to 48 h and supernatants were analyzed for GM-CSF by ELISA. Fixed organisms, germination-deficient mutants and separation of yeast from epithelial cells using cell culture inserts were used to assess the effects of viability, germination and physical contact, respectively, on the GM-CSF responses of these cells. Two out of three cell lines and three out of six primary cultures responded to C. albicans with an increase in GM-CSF secretion. GM-CSF responses were contact-dependent, strain-dependent, required yeast viability and were optimal when the yeast germinated into hyphae.

Functional and Structural Characterization of a Receptor-Like Kinase Involved in Germination and Cell Expansion in Arabidopsis.

PubMed

Wu, Zhen; Liang, Shan; Song, Wen; Lin, Guangzhong; Wang, Weiguang; Zhang, Heqiao; Han, Zhifu; Chai, Jijie

2017-01-01

Leucine-rich repeat receptor-like kinases (LRR-RLKs) are widespread in different plant species and play important roles in growth and development. Germination inhibition is vital for the completion of seed maturation and cell expansion is a fundamental cellular process driving plant growth. Here, we report genetic and structural characterizations of a functionally uncharacterized LRR-RLK, named GRACE (Germination Repression and Cell Expansion receptor-like kinase). Overexpression of GRACE in Arabidopsis exhibited delayed germination, enlarged cotyledons, rosette leaves and stubbier petioles. Conversely, these phenotypes were reversed in the T-DNA insertion knock-down mutant grace-1 plants. A crystal structure of the extracellular domain of GRACE (GRACE-LRR) determined at the resolution of 3.0 Å revealed that GRACE-LRR assumed a right-handed super-helical structure with an island domain (ID). Structural comparison showed that structure of the ID in GRACE-LRR is strikingly different from those observed in other LRR-RLKs. This structural observation implies that GRACE might perceive a new ligand for signaling. Collectively, our data support roles of GRACE in repressing seed germination and promoting cell expansion of Arabidopsis , presumably by perception of unknown ligand(s).

Functional and Structural Characterization of a Receptor-Like Kinase Involved in Germination and Cell Expansion in Arabidopsis

PubMed Central

Wu, Zhen; Liang, Shan; Song, Wen; Lin, Guangzhong; Wang, Weiguang; Zhang, Heqiao; Han, Zhifu; Chai, Jijie

2017-01-01

Leucine-rich repeat receptor-like kinases (LRR-RLKs) are widespread in different plant species and play important roles in growth and development. Germination inhibition is vital for the completion of seed maturation and cell expansion is a fundamental cellular process driving plant growth. Here, we report genetic and structural characterizations of a functionally uncharacterized LRR-RLK, named GRACE (Germination Repression and Cell Expansion receptor-like kinase). Overexpression of GRACE in Arabidopsis exhibited delayed germination, enlarged cotyledons, rosette leaves and stubbier petioles. Conversely, these phenotypes were reversed in the T-DNA insertion knock-down mutant grace-1 plants. A crystal structure of the extracellular domain of GRACE (GRACE-LRR) determined at the resolution of 3.0 Å revealed that GRACE-LRR assumed a right-handed super-helical structure with an island domain (ID). Structural comparison showed that structure of the ID in GRACE-LRR is strikingly different from those observed in other LRR-RLKs. This structural observation implies that GRACE might perceive a new ligand for signaling. Collectively, our data support roles of GRACE in repressing seed germination and promoting cell expansion of Arabidopsis, presumably by perception of unknown ligand(s). PMID:29213277

RNA-seq analysis identifies potential modulators of gravity response in spores of Ceratopteris (Parkeriaceae): evidence for modulation by calcium pumps and apyrase activity.

PubMed

Bushart, Thomas J; Cannon, Ashley E; Ul Haque, Aeraj; San Miguel, Phillip; Mostajeran, Kathy; Clark, Gregory B; Porterfield, D Marshall; Roux, Stanley J

2013-01-01

Gravity regulates the magnitude and direction of a trans-cell calcium current in germinating spores of Ceratopteris richardii. Blocking this current with nifedipine blocks the spore's downward polarity alignment, a polarization that is fixed by gravity ∼10 h after light induces the spores to germinate. RNA-seq analysis at 10 h was used to identify genes potentially important for the gravity response. The data set will be valuable for other developmental and phylogenetic studies. De novo Newbler assembly of 958 527 reads from Roche 454 sequencing was executed. The sequences were identified and analyzed using in silico methods. The roles of endomembrane Ca(2+)-ATPase pumps and apyrases in the gravity response were further tested using pharmacological agents. Transcripts related to calcium signaling and ethylene biosynthesis were identified as notable constituents of the transcriptome. Inhibiting the activity of endomembrane Ca(2+)-ATPase pumps with 2,5-di-(t-butyl)-1,4-hydroquinone diminished the trans-cell current, but increased the orientation of the polar axis to gravity. The effects of applied nucleotides and purinoceptor antagonists gave novel evidence implicating extracellular nucleotides as regulators of the gravity response in these fern spores. In addition to revealing general features of the transcriptome of germinating spores, the results highlight a number of calcium-responsive and light-receptive transcripts. Pharmacologic assays indicate endomembrane Ca(2+)-ATPases and extracellular nucleotides may play regulatory roles in the gravity response of Ceratopteris spores.

Phosphate limitation induces sporulation in the chytridiomycete Blastocladiella emersonii.

PubMed

Bongiorno, Vagner Alexandre; Ferreira da Cruz, Angela; Nunis da Silva, Antonio; Corrêa, Luiz Carlos

2012-09-01

The cell cycle is controlled by numerous mechanisms that ensure correct cell division. If growth is not possible, cells may eventually promote autophagy, differentiation, or apoptosis. Microorganisms interrupt their growth and differentiate under general nutrient limitation. We analyzed the effects of phosphate limitation on growth and sporulation in the chytridiomycete Blastocladiella emersonii using kinetic data, phase-contrast, and laser confocal microscopy. Under phosphate limitation, zoospores germinated and subsequently formed 2-4 spores, regardless of the nutritional content of the medium. The removal of phosphate at any time during growth induced sporulation of vegetative cells. If phosphate was later added to the same cultures, growth was restored if the cells were not yet committed to sporulation. The cycles of addition and withdrawal of phosphate from growth medium resulted in cycles of germination-growth, germination-sporulation, or germination-growth-sporulation. These results show that phosphate limitation is sufficient to interrupt cell growth and to induce complete sporulation in B. emersonii. We concluded that the determination of growth or sporulation in this microorganism is linked to phosphate availability when other nutrients are not limiting. This result provides a new tool for the dissection of nutrient-energy and signal pathways in cell growth and differentiation.

HFR1 Sequesters PIF1 to Govern the Transcriptional Network Underlying Light-Initiated Seed Germination in Arabidopsis[C][W][OPEN

PubMed Central

Shi, Hui; Zhong, Shangwei; Mo, Xiaorong; Liu, Na; Nezames, Cynthia D.; Deng, Xing Wang

2013-01-01

Seed germination is the first step for seed plants to initiate a new life cycle. Light plays a predominant role in promoting seed germination, where the initial phase is mediated by photoreceptor phytochrome B (phyB). Previous studies showed that PHYTOCHROME-INTERACTING FACTOR1 (PIF1) represses seed germination downstream of phyB. Here, we identify a positive regulator of phyB-dependent seed germination, LONG HYPOCOTYL IN FAR-RED1 (HFR1). HFR1 blocks PIF1 transcriptional activity by forming a heterodimer with PIF1 that prevents PIF1 from binding to DNA. Our whole-genomic analysis shows that HFR1 and PIF1 oppositely mediate the light-regulated transcriptome in imbibed seeds. Through the HFR1–PIF1 module, light regulates expression of numerous genes involved in cell wall loosening, cell division, and hormone pathways to initiate seed germination. The functionally antagonistic HFR1–PIF1 pair constructs a fail-safe mechanism for fine-tuning seed germination during low-level illumination, ensuring a rapid response to favorable environmental changes. This study identifies the HFR1–PIF1 pair as a central module directing the whole genomic transcriptional network to rapidly initiate light-induced seed germination. PMID:24179122

Hormonal profile and the role of cell expansion in the germination control of Cerrado biome palm seeds.

PubMed

Dias, Daiane Souza; Ribeiro, Leonardo Monteiro; Lopes, Paulo Sérgio Nascimento; Munné-Bosch, Sergi; Garcia, Queila Souza

2017-09-01

Little information is currently available concerning the mechanisms controlling palm seed germination. We compared the anatomical and physiological aspects of seeds of two neotropical palm species showing different levels of dormancy. The seeds of Attalea vitrivir and Butia capitata were evaluated for the endogenous contents of hormones (ABA, GAs, CKs, BRs, IAA, JA, SA and the ethylene precursor ACC) in their cotyledonary petiole and operculum (structures involved in germination control), the force necessary to displace the operculum, endo-β-mannanase activities, and embryo cell elongation. The analyses were carried out on with intact dry and imbibed seeds as well as with seeds with the operculum mechanically removed, 2, 5 and 10 days after sowing. The germinabilities of the intact seeds of A. vitrivir and B. capitata were 68% and 3%, respectively; the removal of the operculum increased germination to more than 90% in both species. Reductions of ABA and increases in GAs contents coincided with cell elongation, although there is no evidence that hormonal balance and endo-β-mannanase activity are involved in operculum weakening. The ratio between the embryo length and the force required for operculum displacement (EL/OF) was found to be 1.9 times greater in A. vitrivir than in B. capitata, which means that very small elongations in each cell would be sufficient to promote germination, resulting in a lower level of dormancy in the former species. EL/OF and cell growth control are therefore important for defining dormancy level in palm seeds. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Chapter 26. Seed germination

Treesearch

Kent R. Jorgensen; G. Richard Wilson

2004-01-01

Seed germination represents the means for survival and spread of many plants (McDonough 1977). Germination consists of three overlapping processes: (1) absorption of water, mainly by imbibition, causing swelling of the seed; (2) concurrent enzymatic activity and increased respiration and assimilation rates; and (3) cell enlargement and divisions resulting in emergence...

Sulfinylated Azadecalins act as functional mimics of a pollen germination stimulant in Arabidopsis pistils

PubMed Central

Qin, Yuan; Wysocki, Ronald J; Somogyi, Arpad; Feinstein, Yelena; Franco, Jessica Y; Tsukamoto, Tatsuya; Dunatunga, Damayanthi; Levy, Clara; Smith, Steven; Simpson, Robert; Gang, David; Johnson, Mark A; Palanivelu, Ravishankar

2011-01-01

SUMMARY Polarized cell elongation is triggered by small molecule cues during development of diverse organisms. During plant reproduction, pollen interactions with the stigma result in the polar outgrowth of a pollen tube, which delivers sperm cells to the female gametophyte to effect double fertilization. In many plants, pistils stimulate pollen germination. However, in Arabidopsis, the effect of pistils on pollen germination and the pistil factors that stimulate pollen germination remain poorly characterized. Here, we demonstrate that stigma, style, and ovules in Arabidopsis pistils stimulate pollen germination. We isolated an Arabidopsis pistil extract fraction that stimulates Arabidopsis pollen germination, and employed ultrahigh resolution ESI FT-ICR and MS/MS techniques to accurately determine the mass (202.126 daltons) of a compound that is specifically present in this pistil extract fraction. Using the molecular formula (C10H19NOS) and tandem mass spectral fragmentation patterns of the m/z (mass to charge ratio) 202.126 ion, we postulated chemical structures, devised protocols, synthesized N-Methanesulfinyl 1- and 2-azadecalins that are close structural mimics of the m/z 202.126 ion, and showed that they are sufficient to stimulate Arabidopsis pollen germination in vitro (30 µM stimulated ~50% germination) and elicit accession-specific response. Although N-Methanesulfinyl 2-azadecalin stimulated pollen germination in three species of Lineage I of Brassicaceae, it did not induce a germination response in Sisymbrium irio (Lineage II of Brassicaceae) and tobacco, indicating that activity of the compound is not random. Our results show that Arabidopsis pistils promote germination by producing azadecalin-like molecules to ensure rapid fertilization by the appropriate pollen. PMID:21801250

The distinctive germinal center phase of IgE+ B lymphocytes limits their contribution to the classical memory response

USDA-ARS?s Scientific Manuscript database

The mechanisms involved in the maintenance of memory IgE responses are poorly understood, and the role played by germinal center (GC) IgE+ cells in memory responses is particularly unclear. IgE B cell differentiation is characterized by a transient GC phase, a bias towards the plasma cell (PC) fate,...

Leveraging a high resolution microfluidic assay reveals insights into pathogenic fungal spore germination

PubMed Central

Barkal, Layla J.; Walsh, Naomi M.; Botts, Michael R.; Beebe, David J.; Hull, Christina M.

2016-01-01

Germination of spores into actively growing cells is a process essential for survival and pathogenesis of many microbes. Molecular mechanisms governing germination, however, are poorly understood in part because few tools exist for evaluating and interrogating the process. Here, we introduce an assay that leverages developments in microfluidic technology and image processing to quantitatively measure germination with unprecedented resolution, assessing both individual cells and the population as a whole. Using spores from Cryptococcus neoformans, a leading cause of fatal fungal disease in humans, we developed a platform to evaluate spores as they undergo morphological changes during differentiation into vegetatively growing yeast. The assay uses pipet-accessible microdevices that can be arrayed for efficient testing of diverse microenvironmental variables, including temperature and nutrients. We discovered that temperature influences germination rate, a carbon source alone is sufficient to induce germination, and the addition of a nitrogen source sustains it. Using this information, we optimized the assay for use with fungal growth inhibitors to pinpoint stages of germination inhibition. Unexpectedly, the clinical antifungal drugs amphotericin B and fluconazole did not significantly alter the process or timing of the transition from spore to yeast, indicating that vegetative growth and germination are distinct processes in C. neoformans. Finally, we used the high temporal resolution of the assay to determine the precise defect in a slow-germination mutant. Combining advances in microfluidics with a robust fungal molecular genetic system allowed us to identify and alter key temporal, morphological, and molecular events that occur during fungal germination. PMID:27026574

Nivolumab With or Without Varlilumab in Treating Patients With Relapsed or Refractory Aggressive B-cell Lymphomas

ClinicalTrials.gov

2018-06-11

ALK-Positive Large B-Cell Lymphoma; Atypical Burkitt/Burkitt-Like Lymphoma; Burkitt-Like Lymphoma With 11q Aberration; Diffuse Large B-Cell Lymphoma Activated B-Cell Type; Diffuse Large B-Cell Lymphoma Associated With Chronic Inflammation; Diffuse Large B-Cell Lymphoma Germinal Center B-Cell Type; Diffuse Large B-Cell Lymphoma, Not Otherwise Specified; EBV-Positive Diffuse Large B-Cell Lymphoma, Not Otherwise Specified; EBV-Positive Mucocutaneous Ulcer; High-Grade B-Cell Lymphoma With MYC, BCL2, and BCL6 Rearrangements; Human Herpesvirus 8-Positive Neoplastic Cells Present; Intravascular Large B-Cell Lymphoma; Large B-Cell Lymphoma With IRF4 Rearrangement; Plasmablastic Lymphoma; Primary Cutaneous Diffuse Large B-Cell Lymphoma; Primary Cutaneous Diffuse Large B-Cell Lymphoma, Leg Type; Primary Diffuse Large B-Cell Lymphoma of the Central Nervous System; Primary Effusion Lymphoma; Recurrent B-Cell Lymphoma, Unclassifiable, With Features Intermediate Between Diffuse Large B-Cell Lymphoma and Classic Hodgkin Lymphoma; Recurrent Burkitt Lymphoma; Recurrent Diffuse Large B-Cell Lymphoma; Recurrent Lymphomatoid Granulomatosis; Recurrent Primary Mediastinal (Thymic) Large B-Cell Cell Lymphoma; Refractory B-Cell Lymphoma, Unclassifiable, With Features Intermediate Between Diffuse Large B-Cell Lymphoma and Classic Hodgkin Lymphoma; Refractory Burkitt Lymphoma; Refractory Diffuse Large B-Cell Lymphoma; Refractory Primary Mediastinal (Thymic) Large B-Cell Cell Lymphoma; Small Intestinal High Grade B-Cell Lymphoma, Not Otherwise Specified; T-Cell/Histiocyte-Rich Large B-Cell Lymphoma

B cell immunopoiesis: visualizing the impact of CD40 engagement on the course of T cell-independent immune responses in an Ig transgenic system.

PubMed

Erickson, L D; Vogel, L A; Cascalho, M; Wong, J; Wabl, M; Durell, B G; Noelle, R J

2000-11-01

This study tracks the fate of antigen-reactive B cells through follicular and extrafollicular responses and addresses the function of CD40 in these processes. The unique feature of this system is the use of transgenic B cells in which the heavy chain locus has been altered by site-directed insertion of a rearranged V(H) DJ(H) exon such that they are able to clonally expand, isotype-switch and follow a normal course of differentiation upon immunization. These Ig transgenic B cells when adoptively transferred into non-transgenic (Tg) mice in measured amounts expanded and differentiated distinctively in response to T cell-independent (TI) or T cell-dependent (TD) antigens. The capacity of these Tg B cells to faithfully recapitulate the humoral immune response to TI and TD antigens provides the means to track clonal B cell behavior in vivo. Challenge with TI antigen in the presence of agonistic anti-CD40 mAb resulted in well-defined alterations of the TI response. In vivo triggering of Tg B cells with TI antigen and CD40 caused an increase in the levels IgG produced and a broadening of the Ig isotype profile, characteristics which partially mimic TD responses. Although some TD characteristics were induced by TI antigen and CD40 triggering, the Tg B cells failed to acquire a germinal center phenotype and failed to generate a memory response. Therefore, TD-like immunity can be only partially reconstituted with CD40 agonists and TI antigens, suggesting that there are additional signals required for germinal center formation and development of memory.

Specific role of LeMAN2 in the control of seed germination exposed by overexpression of the LeMAN3 gene in tomato plants.

PubMed

Belotserkovsky, Harel; Berger, Yael; Shahar, Ron; Wolf, Shmuel

2007-12-01

Endo-beta-mannanase is one of the key enzymes involved in the hydrolysis of the mannan-rich cell walls of tomato (Solanum lycopersicon) seeds. Two isoforms of endo-beta-mannanase have been characterized in tomato seeds: LeMAN2 is active in the micropylar area prior to germination and LeMAN1 is active after germination in all endosperm cells surrounding the cotyledons. To explore whether general mannanase activity in the endosperm cap is sufficient to promote germination, the gene encoding LeMAN3 was inserted into transgenic tomato plants under the control of a CaMV-35S promoter. Expression of LeMAN3 was evident in the endosperm cap and in the lateral endosperm of the transgenic seeds 10 min after imbibition. An activity test indicated increased activity of endo-beta-mannanase in the transgenic lines relative to the control line in all seed parts, during the first 20 h of imbibition. However, overexpression of LeMAN3 in transgenic seeds inhibited seed germination at both optimal and suboptimal temperatures. Detailed RT-PCR analyses revealed the transcription patterns of the genes encoding the various mannanase isoforms, and indicated a delay in LeMAN2 transcription in the endosperm cap of the transgenic seeds. Interestingly, tissue-print assays indicated similar mannanase activity in the micropylar areas for both transgenic and control seeds. These results indicate that overexpression of active endo-beta-mannanase in the endosperm cap is not sufficient to enable hydrolysis of the cell walls or to promote germination of tomato seeds. Cell-wall hydrolysis in these endosperm cells is under tight control and requires the specific activity of LeMAN2.

Influence of antiretroviral therapy on programmed death-1 (CD279) expression on T cells in lymph nodes of human immunodeficiency virus-infected individuals.

PubMed

Ehrhard, Simone; Wernli, Marion; Dürmüller, Ursula; Battegay, Manuel; Gudat, Fred; Erb, Peter

2009-10-01

Human immunodeficiency virus infection leads to T-cell exhaustion and involution of lymphoid tissue. Recently, the programmed death-1 pathway was found to be crucial for virus-specific T-cell exhaustion during human immunodeficiency virus infection. Programmed death-1 expression was elevated on human immunodeficiency virus-specific peripheral blood CD8+ and CD4+ T cells and correlated with disease severity. During human immunodeficiency infection, lymphoid tissue acts as a major viral reservoir and is an important site for viral replication, but it is also essential for regulatory processes important for immune recovery. We compared programmed death-1 expression in 2 consecutive inguinal lymph nodes of 14 patients, excised before antiretroviral therapy (antiretroviral therapy as of 1997-1999) and 16 to 20 months under antiretroviral therapy. In analogy to lymph nodes of human immunodeficiency virus-negative individuals, in all treated patients, the germinal center area decreased, whereas the number of germinal centers did not significantly change. Programmed death-1 expression was mostly found in germinal centers. The absolute extent of programmed death 1 expression per section was not significantly altered after antiretroviral therapy resulting in a significant-relative increase of programmed death 1 per shrunken germinal center. In colocalization studies, CD45R0+ cells that include helper/inducer T cells strongly expressed programmed death-1 before and during therapy, whereas CD8+ T cells, fewer in numbers, showed a weak expression for programmed death-1. Thus, although antiretroviral therapy seems to reduce the number of programmed death-1-positive CD8+ T lymphocytes within germinal centers, it does not down-regulate programmed death-1 expression on the helper/inducer T-cell subset that may remain exhausted and therefore unable to trigger immune recovery.

Lectin histochemistry as a tool to identify apoptotic cells in the seminiferous epithelium of Syrian hamster (Mesocricetus auratus) subjected to short photoperiod.

PubMed

Seco-Rovira, V; Beltrán-Frutos, E; Ferrer, C; Sánchez-Huertas, M M; Madrid, J F; Saez, F J; Pastor, L M

2013-12-01

Lectins have been widely used to study the pattern of cellular glycoconjugates in numerous species. In the process of cellular apoptosis, it has been observed that changes occur in the membrane sugar sequences of these apoptotic cells. The aim of our work was to identify which lectins, out of an extensive battery of the same (PNA, SBA, HPA, LTA, Con-A, UEA-I, WGA, DBA, MAA, GNA, AAA, SNA), show affinity for germinal cells in apoptosis, at what stage of cell death they do so and in which germinal cell types they can be detected. For this, we studied testis sections during testicular regression in Syrian hamster (Mesocricetus auratus) subjected to short photoperiod. Several lectins showed an affinity for the glycoconjugate residues of germ cells in apoptosis: Gal β1,3-GalNAcα1, α-d-mannose, N-acetylgalactosamine and l-fucose. Furthermore, lectin specificity was observed for some specific germinal cells and in certain stages of apoptosis. It was also observed that one of these lectins (PNA) showed affinity for Sertoli cells undergoing apoptosis. Therefore, we conclude that the use of lectin histochemistry could be a very useful tool for studying apoptosis in the seminiferous epithelium because of the specificity shown towards germinal cells in pathological or experimentally induced epithelial depletion models. © 2013 Blackwell Verlag GmbH.

Consequences of neonatal thymectomy in New Zealand black mice

PubMed Central

East, June; de Sousa, Maria A. B.; Parrott, Delphine M. V.; Jaquet, H.

1967-01-01

The possible role of the thymus in autoimmune disease was studied by comparing the effects of neonatal thymectomy on New Zealand Black (NZB) mice (which develop a Coombs positive haemolytic anaemia) and on C3H/Bi, F1 (C57BL × C3H/Bi), C57BL and TO mice. The neonatally thymectomized NZB mice, in common with those of other strains, showed lethal wasting, a lymphocyte deficit in their lymphoid organs and blood, their packed cell volume was reduced and some had liver lesions associated with the hepatotrophic virus MHV-1. As in C3H/Bi and hybrid mice, thymectomy had little effect on the levels of immunoglobulins (IgG, IgA, IgM) present in their sera. Removing the thymus from NZB mice did not prevent and could precipitate Coombs positive reactions; thymectomized mice of the other strains were Coombs negative. Although germinal centres develop and plasma cells occur in the lymphoid organs of most thymectomized mice, the striking feature of the NZB mice was the large number, size and activity of the germinal centres that persisted after thymectomy. ImagesFig. 3Fig. 4 PMID:4166241

Histone Arginine Methylation by PRMT7 Controls Germinal Center Formation via Regulating Bcl6 Transcription.

PubMed

Ying, Zhengzhou; Mei, Mei; Zhang, Peizhun; Liu, Chunyi; He, Huacheng; Gao, Fei; Bao, Shilai

2015-08-15

B cells are the center of humoral immunity and produce Abs to protect against foreign Ags. B cell defects lead to diseases such as leukemia and lymphomas. Histone arginine methylation is important for regulating gene activation and silencing in cells. Although the process commonly exists in mammalian cells, its roles in B cells are unknown. To explore the effects of aberrant histone arginine methylation on B cells, we generated mice with a B cell-specific knockout of PRMT7, a member of the methyltransferases that mediate arginine methylation of histones. In this article, we showed that the loss of PRMT7 led to decreased mature marginal zone B cells and increased follicular B cells and promoted germinal center formation after immunization. Furthermore, mice lacking PRMT7 expression in B cells secreted low levels of IgG1 and IgA. Abnormal expression of germinal center genes (i.e., Bcl6, Prdm1, and Irf4) was detected in conditional knockout mice. By overexpressing PRMT7 in the Raji and A20 cell lines derived from B cell lymphomas, we validated the fact that PRMT7 negatively regulated Bcl6 expression. Using chromatin immunoprecipitation-PCR, we found that PRMT7 could recruit H4R3me1 and symmetric H4R3me2 to the Bcl6 promoter. These results provide evidence for the important roles played by PRMT7 in germinal center formation. Copyright © 2015 by The American Association of Immunologists, Inc.

Uptake of and Resistance to the Antibiotic Berberine by Individual Dormant, Germinating and Outgrowing Bacillus Spores as Monitored by Laser Tweezers Raman Spectroscopy

PubMed Central

Wang, Shiwei; Yu, Jing; Suvira, Milomir; Setlow, Peter; Li, Yong-qing

2015-01-01

Berberine, an alkaloid originally extracted from the plant Coptis chinensis and other herb plants, has been used as a pharmacological substance for many years. The therapeutic effect of berberine has been attributed to its interaction with nucleic acids and blocking cell division. However, levels of berberine entering individual microbial cells minimal for growth inhibition and its effects on bacterial spores have not been determined. In this work the kinetics and levels of berberine accumulation by individual dormant and germinated spores were measured by laser tweezers Raman spectroscopy and differential interference and fluorescence microscopy, and effects of berberine on spore germination and outgrowth and spore and growing cell viability were determined. The major conclusions from this work are that: (1) colony formation from B. subtilis spores was blocked ~ 99% by 25 μg/mL berberine plus 20 μg/mL INF55 (a multidrug resistance pump inhibitor); (2) 200 μg/mL berberine had no effect on B. subtilis spore germination with L-valine, but spore outgrowth was completely blocked; (3) berberine levels accumulated in single spores germinating with ≥ 25 μg/mL berberine were > 10 mg/mL; (4) fluorescence microscopy showed that germinated spores accumulated high-levels of berberine primarily in the spore core, while dormant spores accumulated very low berberine levels primarily in spore coats; and (5) during germination, uptake of berberine began at the time of commitment (T1) and reached a maximum after the completion of CaDPA release (Trelease) and spore cortex lysis (Tlysis). PMID:26636757

Increased germination and growth rates of pea and Zucchini seed by FSG plasma

NASA Astrophysics Data System (ADS)

Khatami, Shohreh; Ahmadinia, Arash

2018-04-01

Recently, cold atmospheric plasma (CAP) with the unique bio-disinfection features is used in various fields of industry, medicine, and agriculture. The main objectives of this work were to design FSG plasma (a semi-automatic device) and investigate the effect of the cold plasma in the enhancement of the Pea and Zucchini seed germination. Plasma irradiation time was studied to obtain a proper condition for the germination enhancement of seeds. The growth rate was calculated by measuring length of root and stem and dry weight of plants treated by plasma. To investigate drought resistance of plants, all treated and untreated samples were kept in darkness without water for 48 h. From the experimental results, it could be confirmed both drought resistance and germination of seedlings increased after plasma was applied to seeds at 30 s, while seeds treated whiten 60 s showed a decrease in both germination rate and seedling growth.

Dead Pericarps of Dry Fruits Function as Long-Term Storage for Active Hydrolytic Enzymes and Other Substances That Affect Germination and Microbial Growth

PubMed Central

Godwin, James; Raviv, Buzi; Grafi, Gideon

2017-01-01

It is commonly assumed that dead pericarps of dry indehiscent fruits have evolved to provide an additional physical layer for embryo protection and as a means for long distance dispersal. The pericarps of dry fruits undergo programmed cell death (PCD) during maturation whereby most macromolecules such DNA, RNA, and proteins are thought to be degraded and their constituents remobilized to filial tissues such as embryo and endosperm. We wanted to test the hypothesis that the dead pericarp represents an elaborated layer that is capable of storing active proteins and other substances for increasing survival rate of germinating seeds. Using in gel assays we found that dead pericarps of both dehiscent and indehiscent dry fruits of various plant species including Arabidopsis thaliana and Sinapis alba release upon hydration multiple active hydrolytic enzymes that can persist in an active form for decades, including nucleases, proteases, and chitinases. Proteomic analysis of indehiscent pericarp of S. alba revealed multiple proteins released upon hydration, among them proteases and chitinases, as well as proteins involved in reactive oxygen species (ROS) detoxification and cell wall modification. Pericarps appear to function also as a nutritional element-rich storage for nitrate, potassium, phosphorus, sulfur, and others. Sinapis alba dehiscent and indehiscent pericarps possess germination inhibitory substances as well as substances that promote microbial growth. Collectively, our study explored previously unknown features of the dead pericarp acting also as a reservoir of biological active proteins, and other substances capable of “engineering” the microenvironment for the benefit of the embryo. PMID:29257090

Infectious mononucleosis lymphoadenitis showing histologic findings indistinguishable from toxoplasma lymphadenitis. A report of three cases.

PubMed

Kojima, Masaru; Kashimura, Makoto; Itoh, Hideaki; Noro, Masahiro; Matsuda, Hazuki; Tsukamoto, Norihumi; Akikusa, Bunshiro; Masawa, Nobuhide; Morita, Yukio

2010-06-15

Lymph node lesions in infectious mononucleosis (IM) show a marked histologic diversity. We report here three cases of IM lymphadenitis with histologic findings indistinguishable from those of toxoplasmic lymphadenitis. The histologic findings of the three cases presented here showed a histologic triad of toxoplasmic lymphadenitis, including (i) numerous lymphoid follicles with hyperplastic germinal centers; (ii) small clusters or single epithelioid histiocytes; and (iii) multiple foci of monocytoid B-cells. Moreover, all three lesions contained isolated or small clusters of epithelioid histiocytes within the hyperplastic germinal centers and the periphery of lymphoid follicles, which are the most specific histologic findings of toxoplasmic lymphadenitis. However, serologic findings confirmed EBV infection in all three cases. On in situ hybridization, numerous Epstein-Barr virus (EBV)-encoded small RNA (EBER)-positive cells were demonstrated in the germinal center, as well as in interfollicular areas in all three cases. Toxoplasmosis gondii infection was excluded in at least one case, based on serologic findings. Polymerase chain reaction analysis also demonstrated that there was no T. gondii DNA in the remaining two cases. Two of our three cases showed atypical clinical presentations, including an absence of atypical lymphocytosis in peripheral blood in two cases, age more than 30 years, and an absence of systemic symptoms in one case. It appears that previous descriptions emphasize the differential diagnostic problems between IM lymphadenitis and malignant lymphomas. However, from a therapeutic perspective, it is important to discriminate IM lymphadenitis from toxoplasmic lymphadenitis particularly in patients showing atypical clinical features. 2010 Elsevier GmbH. All rights reserved.

[Morphofunctional organization of reserve stem cells providing for asexual and sexual reproduction of invertebrates].

PubMed

Isaeva, V V; Akhmadieva, A V; Aleksandriova, Ia N; Shukaliuk, A I

2009-01-01

Published and original data indicating evolutionary conservation of the morphofunctional organization of reserve stem cells providing for asexual and sexual reproduction of invertebrates are reviewed. Stem cells were studied in representatives of five animal types: archeocytes in sponge Oscarella malakhovi (Porifera), large interstitial cells in colonial hydroid Obelia longissima (Cnidaria), neoblasts in an asexual race of planarian Girardia tigrina (Platyhelmintes), stem cells in colonial rhizocephalans Peltogasterella gracilis, Polyascus polygenea, and Thylacoplethus isaevae (Arthropoda), and colonial ascidian Botryllus tuberatus (Chordata). Stem cells in animals of such diverse taxa feature the presence of germinal granules, are positive for proliferating cell nuclear antigen, demonstrate alkaline phosphatase activity (at marker of embryonic stem cells and primary germ cells in vertebrates), and rhizocephalan stem cells express the vasa-like gene (such genes are expressed in germline cells of different metazoans). The self-renewing pool of stem cells is the cellular basis of the reproductive strategy including sexual and asexual reproduction.

Primary cutaneous B-cell lymphoma is associated with somatically hypermutated immunoglobulin variable genes and frequent use of VH1-69 and VH4-59 segments.

PubMed

Perez, M; Pacchiarotti, A; Frontani, M; Pescarmona, E; Caprini, E; Lombardo, G A; Russo, G; Faraggiana, T

2010-03-01

Accurate assessment of the somatic mutational status of clonal immunoglobulin variable region (IgV) genes is relevant in elucidating tumour cell origin in B-cell lymphoma; virgin B cells bear unmutated IgV genes, while germinal centre and postfollicular B cells carry mutated IgV genes. Furthermore, biases in the IgV repertoire and distribution pattern of somatic mutations indicate a possible antigen role in the pathogenesis of B-cell malignancies. This work investigates the cellular origin and antigenic selection in primary cutaneous B-cell lymphoma (PCBCL). We analysed the nucleotide sequence of clonal IgV heavy-chain gene (IgVH) rearrangements in 51 cases of PCBCL (25 follicle centre, 19 marginal zone and seven diffuse large B-cell lymphoma, leg-type) and compared IgVH sequences with their closest germline segment in the GenBank database. Molecular data were then correlated with histopathological features. We showed that all but one of the 51 IgVH sequences analysed exhibited extensive somatic hypermutations. The detected mutation rate ranged from 1.6% to 21%, with a median rate of 9.8% and was independent of PCBCL histotype. Calculation of antigen-selection pressure showed that 39% of the mutated IgVH genes displayed a number of replacement mutations and silent mutations in a pattern consistent with antigenic selection. Furthermore, two segments, VH1-69 (12%) and VH4-59 (14%), were preferentially used in our case series. Data indicate that neoplastic B cells of PBCBL have experienced germinal centre reaction and also suggest that the involvement of IgVH genes is not entirely random in PCBCL and that common antigen epitopes could be pathologically relevant in cutaneous lymphomagenesis.

T Follicular Helper Cell-Germinal Center B Cell Interaction Strength Regulates Entry into Plasma Cell or Recycling Germinal Center Cell Fate.

PubMed

Ise, Wataru; Fujii, Kentaro; Shiroguchi, Katsuyuki; Ito, Ayako; Kometani, Kohei; Takeda, Kiyoshi; Kawakami, Eiryo; Yamashita, Kazuo; Suzuki, Kazuhiro; Okada, Takaharu; Kurosaki, Tomohiro

2018-04-17

Higher- or lower-affinity germinal center (GC) B cells are directed either to plasma cell or GC recycling, respectively; however, how commitment to the plasma cell fate takes place is unclear. We found that a population of light zone (LZ) GC cells, Bcl6 lo CD69 hi expressing a transcription factor IRF4 and higher-affinity B cell receptors (BCRs) or Bcl6 hi CD69 hi with lower-affinity BCRs, favored the plasma cell or recycling GC cell fate, respectively. Mechanistically, CD40 acted as a dose-dependent regulator for Bcl6 lo CD69 hi cell formation. Furthermore, we found that expression of intercellular adhesion molecule 1 (ICAM-1) and signaling lymphocytic activation molecule (SLAM) in Bcl6 lo CD69 hi cells was higher than in Bcl6 hi CD69 hi cells, thereby affording more stable T follicular helper (Tfh)-GC B cell contacts. These data support a model whereby commitment to the plasma cell begins in the GC and suggest that stability of Tfh-GC B cell contacts is key for plasma cell-prone GC cell formation. Copyright © 2018. Published by Elsevier Inc.

Pre-germinated brown rice could enhance maternal mental health and immunity during lactation.

PubMed

Sakamoto, Shigeko; Hayashi, Takashi; Hayashi, Keiko; Murai, Fumie; Hori, Miyo; Kimoto, Koichi; Murakami, Kazuo

2007-10-01

Rice is a dietary staple worldwide, especially pre-germinated brown rice has recently been widely served in Japan because of its abundant nutrition. Relationship between lactation and pre-germinated brown rice has attracted interest in terms of mental health and immunity. To demonstrate that Japanese foods are beneficial for psychosomatic health, the effects of pre-germinated brown rice on the mental status and immunological features during lactation were investigated. Forty-one breast-feeding mothers were recruited, and randomly divided into two groups. One group took pre-germinated brown rice and the other white rice (control) as their staple diet for 2 weeks. The Profile of Mood States (POMS) and salivary amylase activity as psychological indices and secretory IgA (s-IgA) and lactoferrin (LTF) in breast milk as immunological indices were determined before and after dietary intervention, and changes were investigated. In the psychological assessment, the scores of depression, anger-hostility, and fatigue were decreased on POMS analysis in the pre-germinated brown rice diet group, resulting in a significant decrease in total mood disturbance (TMD). The salivary amylase activity measurement suggested that resistance to stress was increased in the pre-germinated brown rice diet group. On the immunological assessment, the s-IgA level was significantly increased in the pre-germinated brown rice diet group. We have shown that pre-germinated brown rice may have beneficial effects on psychosomatic health.

Differential Effects of Carbohydrates on Arabidopsis Pollen Germination.

PubMed

Hirsche, Jörg; García Fernández, José M; Stabentheiner, Edith; Großkinsky, Dominik K; Roitsch, Thomas

2017-04-01

Pollen germination as a crucial process in plant development strongly depends on the accessibility of carbon as energy source. Carbohydrates, however, function not only as a primary energy source, but also as important signaling components. In a comprehensive study, we analyzed various aspects of the impact of 32 different sugars on in vitro germination of Arabidopsis pollen comprising about 150 variations of individual sugars and combinations. Twenty-six structurally different mono-, di- and oligosaccharides, and sugar analogs were initially tested for their ability to support pollen germination. Whereas several di- and oligosaccharides supported pollen germination, hexoses such as glucose, fructose and mannose did not support and even considerably inhibited pollen germination when added to germination-supporting medium. Complementary experiments using glucose analogs with varying functional features, the hexokinase inhibitor mannoheptulose and the glucose-insensitive hexokinase-deficient Arabidopsis mutant gin2-1 suggested that mannose- and glucose-mediated inhibition of sucrose-supported pollen germination depends partially on hexokinase signaling. The results suggest that, in addition to their role as energy source, sugars act as signaling molecules differentially regulating the complex process of pollen germination depending on their structural properties. Thus, a sugar-dependent multilayer regulation of Arabidopsis pollen germination is supported, which makes this approach a valuable experimental system for future studies addressing sugar sensing and signaling. © The Author 2017. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Aspergillus Cell Wall Melanin Blocks LC3-Associated Phagocytosis to Promote Pathogenicity.

PubMed

Akoumianaki, Tonia; Kyrmizi, Irene; Valsecchi, Isabel; Gresnigt, Mark S; Samonis, George; Drakos, Elias; Boumpas, Dimitrios; Muszkieta, Laetitia; Prevost, Marie-Christine; Kontoyiannis, Dimitrios P; Chavakis, Triantafyllos; Netea, Mihai G; van de Veerdonk, Frank L; Brakhage, Axel A; El-Benna, Jamel; Beauvais, Anne; Latge, Jean-Paul; Chamilos, Georgios

2016-01-13

Concealing pathogen-associated molecular patterns (PAMPs) is a principal strategy used by fungi to avoid immune recognition. Surface exposure of PAMPs during germination can leave the pathogen vulnerable. Accordingly, β-glucan surface exposure during Aspergillus fumigatus germination activates an Atg5-dependent autophagy pathway termed LC3-associated phagocytosis (LAP), which promotes fungal killing. We found that LAP activation also requires the genetic, biochemical or biological (germination) removal of A. fumigatus cell wall melanin. The attenuated virulence of melanin-deficient A. fumigatus is restored in Atg5-deficient macrophages and in mice upon conditional inactivation of Atg5 in hematopoietic cells. Mechanistically, Aspergillus melanin inhibits NADPH oxidase-dependent activation of LAP by excluding the p22phox subunit from the phagosome. Thus, two events that occur concomitantly during germination of airborne fungi, surface exposure of PAMPs and melanin removal, are necessary for LAP activation and fungal killing. LAP blockade is a general property of melanin pigments, a finding with broad physiological implications. Copyright © 2016 Elsevier Inc. All rights reserved.

Submergence Tolerance and Germination Dynamics of Roegneria nutans Seeds in Water-Level Fluctuation Zones with Different Water Rhythms in the Three Gorges Reservoir

PubMed Central

Zeng, Bo; Pan, Xiaojiao; Su, Xiaolei

2016-01-01

The Three Gorges Dam features two water-level fluctuation zones (WLFZs): the preupland drawdown zone (PU-DZ) and the preriparian drawdown zone (PR-DZ). To investigate the vegetation potential of Roegneria nutans in WLFZs, we compared the submergence tolerance and germination dynamics in the natural riparian zone (NRZ), PU-DZ and PR-DZ. We found that the NRZ seeds maintained an 81.3% intactness rate and >91% germination rate. The final seed germination rate and germination dynamics were consistent with those of the controls. Meanwhile, the PU-DZ seeds submerged at 5 m, 10 m, 15 m, and 20 m exhibited intactness rates of 70.5%, 79.95%, 40.75%, and 39.87%, respectively, and >75% germination. Furthermore, the PR-DZ seeds exhibited intactness rates of 22.44%, 61.13%, 81.87%, and 15.36% at 5 m, 10 m, 15 m, and 17 m, respectively, and 80% germination. The germination rates of the intact seeds submerged >10 m were >80%. Finally, the intact seeds germinated quickly in all WLFZs. The high proportion of intact seeds, rapid germination capacity, and high germination rate permit R. nutans seeds to adapt to the complicated water rhythms of the PU-DZ and PR-DZ and indicate the potential for their use in vegetation restoration and recovery. Thus, perennial seeds can be used for vegetation restoration in the WLFZs of large reservoirs and in other regions with water rhythms similar to the Three Gorges Reservoir. PMID:27031104

Mulberry cells in the thyroid: warthin-finkeldey-like cells in hashimoto thyroiditis-associated lymphoma.

PubMed

Lapadat, Razvan; Nam, Moon Woo; Mehrotra, Swati; Velankar, Milind; Pambuccian, Stefan E

2017-03-01

Warthin-Finkeldey type giant cells were first described in autopsies performed on young children who died during the highly lethal measles epidemic in Palermo during the winter of 1908. The cells had 8-15 nuclei without identifiable cytoplasm within the germinal centers of lymphoid organs resembling megakaryocytes. We describe a case of Hashimoto thyroiditis with an enlarging substernal throid mass. The resection specimen contained many Warthin-Finkeldey-Like Cells (WFLC) in an extranodal marginal zone lymphoma (MALT type) with focal transformation to diffuse large B-cell lymphoma. The WFLC showed nuclear features similar to those of neighboring follicular dendritic cells (FDCs), favoring the hypothesis that these cells might be the product of fusion of FDCs. This is supported by immunostaining results and the occurrence of similar cells in follicular dendritic cell sarcomas and in "dysplastic" FDCs in hyaline vascular type Castleman disease, a possible precursor of follicular dendritic cell tumors. Diagn. Cytopathol. 2017;45:212-216. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Nuclear dynamics during ascospore germination in Sordaria macrospora.

PubMed

Teichert, Ines

2017-01-01

The ascomycete Sordaria macrospora has a long history as a model organism for studying fungal sexual development. Starting from an ascospore, sexual fruiting bodies (perithecia) develop within seven days and discharge new ascospores. Sexual development has been studied in detail, revealing genes required for perithecium formation and ascospore germination. However, the germination process per se has not yet been examined. Here I analyze nuclear dynamics during ascospore germination using a fluorescently labeled histone. Live-cell imaging revealed that nuclei are transported into germination vesicles that form on one side of the spore. Polar growth is established from these vesicles. Copyright © 2016 Elsevier Inc. All rights reserved.

[Focal lymphoid hyperplasia (pseudolymphoma) of the terminal ileum in adults].

PubMed

Molas, G; Potet, F; Nogig, P

1985-01-01

We report two cases of focal lymphoid hyperplasia (FLH) of terminal ileum in adult patients. Both cases showed identical morphological findings. The first was discovered during cholecystectomy in a 75-year-old woman who complained mild non-specific abdominal discomfort. The second was manifested by right lower quadrant abdominal pain in a 32-year-old man. The surgical specimens revealed a thickened wall, a narrowed lumen and multiple ulcerations. The histologic features were small cell, well differentiated lymphocyte infiltration, with several follicles showing large germinal centers; regional lymph nodes revealed a conspicuous reactive size enlargement. Further clinical investigations revealed no other abnormalities. Clinical course showed benign evolution after 6 and 3 years of respective follow-up. FLH should be differentiated from terminal ileum inflammatory and infectious diseases. It can be differentiated from Crohn's disease by the absence of characteristic histological features; from Yersinia infection by the absence of significant rates of specific serum antibodies. Moreover, FLH can be differentiated from malignant lymphoma by the presence of follicles and enlarged germinal centers and by the long-term benign evolution. The nature of FLH in terminal ileum, as well as those of the stomach and colo-rectum is still to be determined. Several hypothesis are proposed: reactive, benign neoplastic, or prelymphomatous lesion?

The immune responses in CD40-deficient mice: impaired immunoglobulin class switching and germinal center formation.

PubMed

Kawabe, T; Naka, T; Yoshida, K; Tanaka, T; Fujiwara, H; Suematsu, S; Yoshida, N; Kishimoto, T; Kikutani, H

1994-06-01

An engagement of CD40 with CD40 ligand (CD40L) expressed on activated T cells is known to provide an essential costimulatory signal to B cells in vitro. To investigate the role of CD40 in in vivo immune responses, CD40-deficient mice were generated by gene targeting. The significant reduction of CD23 expression on mature B cells and relatively decreased number of IgM bright and IgD dull B cells were observed in the mutant mice. The mutant mice mounted IgM responses but no IgG, IgA, and IgE responses to thymus-dependent (TD) antigens. However, IgG as well as IgM responses to thymus-independent (TI) antigens were normal. Furthermore, the germinal center formation was defective in the mutant mice. These results suggest that CD40 is essential for T cell-dependent immunoglobulin class switching and germinal center formation, but not for in vivo T cell-dependent IgM responses and T cell-independent antibody responses.

Determining the Origin of Human Germinal Center B Cell-Derived Malignancies.

PubMed

Seifert, Marc; Küppers, Ralf

2017-01-01

Most human B cell lymphomas originate from germinal center (GC) B cells. This is partly caused by the high proliferative activity of GC B cells and the remodeling processes acting at the immunoglobulin (Ig) loci of these cells, i.e., somatic hypermutation and class-switching. Mistargeting of these processes can cause chromosomal translocations, and the hypermutation machinery may also target non-Ig genes. As somatic hypermutation is exclusively active in GC B cells, the presence of somatic mutations in rearranged IgV genes is a standard criterium for a GC or post-GC B cell origin of lymphomas. Beyond this, ongoing somatic hypermutation during lymphoma clone expansion indicates that the lymphoma has an active GC B cell differentiation program. The proto-oncogene BCL6 is specifically expressed in GC B cells and also acquires somatic mutations as a physiological by-product of the somatic hypermutation process, albeit at a lower level than IgV genes. Thus, detection of BCL6 mutations is a further genetic trait of a GC experience of a B cell lymphoma. Typically, B cell lymphomas retain key features of their specific cells of origin, including a differentiation stage-specific gene expression pattern. This is at least partly due to genetic lesions, which "freeze" the lymphoma cells at the differentiation stage at which the transformation occurred. Therefore, identification of the normal B cell subset with the most similar gene expression pattern to a particular type of B cell lymphoma has been instrumental to deduce the precise cell of origin of lymphomas.We present here protocols to analyze human B cell lymphomas for a potential origin from GC B cells by determining the presence of mutations in rearranged IgV genes and the BCL6 gene, and by comparing the gene expression pattern of lymphoma cells with those of normal B cell subsets by genechip or RNA-sequencing analysis.

Dendritic Macromer Replaces Sutures in Cataract Surgery; Promising Polymer Therapy for Paralyzed Dogs; Compound in Smoke Provides the Spark for Germination

NASA Astrophysics Data System (ADS)

King, Angela G.

2005-03-01

This Report surveys articles of interest to chemists that have been recently published in other science journals. Topics surveyed include reports that hydrogel offers an option to suturing the eye; polyethylene glycol aids recovery from spinal injury; and a compound in smoke increases germination rates. See Featured Molecules .

Emerging concepts in T follicular helper cell responses to malaria.

PubMed

Hansen, Diana S; Obeng-Adjei, Nyamekye; Ly, Ann; Ioannidis, Lisa J; Crompton, Peter D

2017-02-01

Antibody responses to malaria and candidate malaria vaccines are short-lived in children, leaving them susceptible to repeated malaria episodes. Because T follicular helper (T FH ) cells provide critical help to B cells to generate long-lived antibody responses, they have become the focus of recent studies of Plasmodium-infected mice and humans. The emerging data converge on common themes, namely, that malaria-induced T H1 cytokines are associated with the activation of (i) T-like memory T FH cells with impaired B cell helper function, and (ii) pre-T FH cells that acquire Th1-like features (T-bet expression, IFN-γ production), which impede their differentiation into fully functional T FH cells, thus resulting in germinal center dysfunction and suboptimal antibody responses. Deeper knowledge of T FH cells in malaria could illuminate strategies to improve vaccines through modulating T FH cell responses. This review summarizes emerging concepts in T FH cell responses to malaria. Copyright © 2016. Published by Elsevier Ltd.

Polyamine biosynthesis during germination of yeast ascospores.

PubMed Central

Brawley, J V; Ferro, A J

1979-01-01

The role of the diamine putrescine during germination and outgrowth of ascospores of Saccharomyces cerevisiae was examined. Ornithine decarboxylase activity increased and declined rapidly during germination and outgrowth; peak activity was attained after the cells had proceeded through the G1 interval of the cell cycle, whereas minimal activity was present at the completion of the first cell division. alpha-Methylornithine inhibited both ornithine decarboxylase activity and the in vivo accumulation of putrescine. In the presence of alpha-methylornithireak dormancy and proceed through one cell division. Subsequent cellular growth, however, was retarded but not completely inhibited. The supplementation of Methylglyoxal bis(guanylhydrazone) to sporulation medium greatly inhibited this sexual process. These data suggest that the synthesis of putrescine is not required for the breaking of spore dormancy, but that polyamine biosynthesis may be essential for meiosis and sporulation. PMID:387744

Elongator promotes germination and early post-germination growth.

PubMed

Woloszynska, Magdalena; Gagliardi, Olimpia; Vandenbussche, Filip; Van Lijsebettens, Mieke

2018-01-02

The Elongator complex interacts with RNA polymerase II and via histone acetylation and DNA demethylation facilitates epigenetically the transcription of genes involved in diverse processes in plants, including growth, development, and immune response. Recently, we have shown that the Elongator complex promotes hypocotyl elongation and photomorphogenesis in Arabidopsis thaliana by regulating the photomorphogenesis and growth-related gene network that converges on genes implicated in cell wall biogenesis and hormone signaling. Here, we report that germination in the elo mutant was delayed by 6 h in the dark when compared to the wild type in a time lapse and germination assay. A number of germination-correlated genes were down-regulated in the elo transcriptome, suggesting a transcriptional regulation by Elongator. We also show that the hypocotyl elongation defect observed in the elo mutants in darkness originates very early in the post-germination development and is independent from the germination delay.

Decontamination Of Bacterial Spores by a Peptide-Mimic

DTIC Science & Technology

2006-11-01

consisting of a thin cell wall and the outer cortex. The cell wall guarantees the maintenance of cellular integrity after germination. Lytic- enzymes ...percent of the water content of the vegetative cell. The enzymes contained in the core become active on germination. All minerals (mainly Ca2+, Mn2+ and...such as amino acids and sugars, by enzymes , by high hydrostatic pressure and by some non-nutrient chemicals such as dodecylamine (see next section

The structural requirements for immunoglobulin aggregates to localize in germinal centres.

PubMed Central

Embling, P H; Evans, H; Guttierez, C; Holborow, E J; Johns, P; Johnson, P M; Papamichail, M; Stanworth, D R

1978-01-01

The capacity of non-heat-aggregated monoclonal human immunoglobulins of different classes, to localize in murine splenic germinal centres within 24 h of intravenous injection has been investigated. It has been shown that at least trimerization of polyclonal IgG must occur before any germinal centre trapping is manifest. Studies of complement fixation by these IgG preparations in vivo, together with studies of the germinal centre trapping of various monoclonal immunoglobulins, have indicated that the sole structural requirement for germinal centre localization of immunoglobulin aggregates is the ability to fix complement. Results suggest that immunoglobulin aggregates are transported to germinal centres via membrane C3 receptors of mobile cells, and then are released with loss of complement to become fixed to dendritic macrophages by a separate mechanism. PMID:363602

Different Modes of Hydrogen Peroxide Action During Seed Germination

PubMed Central

Wojtyla, Łukasz; Lechowska, Katarzyna; Kubala, Szymon; Garnczarska, Małgorzata

2016-01-01

Hydrogen peroxide was initially recognized as a toxic molecule that causes damage at different levels of cell organization and thus losses in cell viability. From the 1990s, the role of hydrogen peroxide as a signaling molecule in plants has also been discussed. The beneficial role of H2O2 as a central hub integrating signaling network in response to biotic and abiotic stress and during developmental processes is now well established. Seed germination is the most pivotal phase of the plant life cycle, affecting plant growth and productivity. The function of hydrogen peroxide in seed germination and seed aging has been illustrated in numerous studies; however, the exact role of this molecule remains unknown. This review evaluates evidence that shows that H2O2 functions as a signaling molecule in seed physiology in accordance with the known biology and biochemistry of H2O2. The importance of crosstalk between hydrogen peroxide and a number of signaling molecules, including plant phytohormones such as abscisic acid, gibberellins, and ethylene, and reactive molecules such as nitric oxide and hydrogen sulfide acting on cell communication and signaling during seed germination, is highlighted. The current study also focuses on the detrimental effects of H2O2 on seed biology, i.e., seed aging that leads to a loss of germination efficiency. The dual nature of hydrogen peroxide as a toxic molecule on one hand and as a signal molecule on the other is made possible through the precise spatial and temporal control of its production and degradation. Levels of hydrogen peroxide in germinating seeds and young seedlings can be modulated via pre-sowing seed priming/conditioning. This rather simple method is shown to be a valuable tool for improving seed quality and for enhancing seed stress tolerance during post-priming germination. In this review, we outline how seed priming/conditioning affects the integrative role of hydrogen peroxide in seed germination and aging. PMID:26870076

Reassessment of an Arabidopsis cell wall invertase inhibitor AtCIF1 reveals its role in seed germination and early seedling growth.

PubMed

Su, Tao; Wolf, Sebastian; Han, Mei; Zhao, Hongbo; Wei, Hongbin; Greiner, Steffen; Rausch, Thomas

2016-01-01

In higher plants, cell wall invertase (CWI) and vacuolar invertase (VI) are recognized as essential players in sugar metabolism and sugar signaling, thereby affecting source-sink interactions, plant development and responses to environmental cues. CWI and VI expression levels are transcriptionally controlled; however, both enzymes are also subject to posttranslational control by invertase inhibitor proteins. The physiological significances of inhibitor proteins during seed germination and early seedling development are not yet fully understood. Here, we demonstrate that the inhibitor isoform AtCIF1 impacted on seed germination and early seedling growth in Arabidopsis. The primary target of AtCIF1 was shown to be localized to the apoplast after expressing an AtCIF1 YFP-fusion construct in tobacco epidermis and transgenic Arabidopsis root. The analysis of expression patterns showed that AtCWI1 was co-expressed spatiotemporally with AtCIF1 within the early germinating seeds. Seed germination was observed to be accelerated independently of exogenous abscisic acid (ABA) in the AtCIF1 loss-of-function mutant cif1-1. This effect coincided with a drastic increase of CWI activity in cif1-1 mutant seeds by 24 h after the onset of germination, both in vitro and in planta. Accordingly, quantification of sugar content showed that hexose levels were significantly boosted in germinating seeds of the cif1-1 mutant. Further investigation of AtCIF1 overexpressors in Arabidopsis revealed a markedly suppressed CWI activity as well as delayed seed germination. Thus, we conclude that the posttranslational modulation of CWI activity by AtCIF1 helps to orchestrate seed germination and early seedling growth via fine-tuning sucrose hydrolysis and, possibly, sugar signaling.

Ultrastructural analysis of oocytes of Rhipicephalus (Boophilus) annulatus during postengorgement period as a tool to evaluate the cytotoxic effects of amitraz and deltamethrin on the germinative cells.

PubMed

Sreelekha, Kanapadinchareveetil; Chandrasekhar, Leena; Kartha, Harikumar S; Ravindran, Reghu; Juliet, Sanis; Ajithkumar, Karapparambu G; Nair, Suresh N; Ghosh, Srikanta

2017-11-30

The present study utilizes the ultrastructural analysis of the fully engorged female Rhipicephalus (Boophilus) annulatus ticks, as a tool to evaluate the cytotoxic potential of deltamethrin and amitraz on the germinative cells. The ultrastructural analysis of the ovary of the normal (untreated) R (B.) annulatus revealed, oocytes in different stages of development, attached to the ovary wall by pedicel cells. The attachment site of oocyte to the pedicel cell was characterized by indentations of the plasma membrane. The oocyte was bound by three cell membranes viz., plasma membrane, chorion and basal lamina. The stages of oocytes were differentiated ultrastructurally based on the features of their outer membrane and the number and size of lipid and yolk droplets. Detailed day wise analysis of ultrastructural changes in the ovary during the post-engorgement period revealed the occurrence of the degenerative changes from day five onwards. These appeared first in the oocytes followed by the germinal epithelium. The ovary of ticks treated with methanol (control), revealed similar topographies as that of a normal ovary except for the presence of very few oocytes with ring shaped nucleoli. Ultrastructurally, treatment with deltamethrin produced more prominent and extensive morphological alterations when compared to amitraz. In the case of ticks treated with amitraz, the oocytes of stage IV and V showed wavy and disrupted outer boundaries along with the loss of integrity of the yolk droplets. Uneven nuclear membranes of stage II oocytes and cristolysis of mitochondria of mature oocytes were the other changes noticed. Ticks treated with deltamethrin revealed prominent modifications such as, detachment of the basal lamina, wrinkled boundary, inconsistent nuclear membrane, ring shaped nucleoli and chromatin clumping in the case of the early stage oocytes (I and II), whereas swelling and cristolysis of mitochondria were seen in mature oocytes. The study further indicated that, in addition to the previous proven neurotoxic effects, these compounds act directly on the ovary of tick. Copyright © 2017 Elsevier B.V. All rights reserved.

Dynamic proteomics emphasizes the importance of selective mRNA translation and protein turnover during Arabidopsis seed germination.

PubMed

Galland, Marc; Huguet, Romain; Arc, Erwann; Cueff, Gwendal; Job, Dominique; Rajjou, Loïc

2014-01-01

During seed germination, the transition from a quiescent metabolic state in a dry mature seed to a proliferative metabolic state in a vigorous seedling is crucial for plant propagation as well as for optimizing crop yield. This work provides a detailed description of the dynamics of protein synthesis during the time course of germination, demonstrating that mRNA translation is both sequential and selective during this process. The complete inhibition of the germination process in the presence of the translation inhibitor cycloheximide established that mRNA translation is critical for Arabidopsis seed germination. However, the dynamics of protein turnover and the selectivity of protein synthesis (mRNA translation) during Arabidopsis seed germination have not been addressed yet. Based on our detailed knowledge of the Arabidopsis seed proteome, we have deepened our understanding of seed mRNA translation during germination by combining two-dimensional gel-based proteomics with dynamic radiolabeled proteomics using a radiolabeled amino acid precursor, namely [(35)S]-methionine, in order to highlight de novo protein synthesis, stability, and turnover. Our data confirm that during early imbibition, the Arabidopsis translatome keeps reflecting an embryonic maturation program until a certain developmental checkpoint. Furthermore, by dividing the seed germination time lapse into discrete time windows, we highlight precise and specific patterns of protein synthesis. These data refine and deepen our knowledge of the three classical phases of seed germination based on seed water uptake during imbibition and reveal that selective mRNA translation is a key feature of seed germination. Beyond the quantitative control of translational activity, both the selectivity of mRNA translation and protein turnover appear as specific regulatory systems, critical for timing the molecular events leading to successful germination and seedling establishment.

Dynamic Proteomics Emphasizes the Importance of Selective mRNA Translation and Protein Turnover during Arabidopsis Seed Germination*

PubMed Central

Galland, Marc; Huguet, Romain; Arc, Erwann; Cueff, Gwendal; Job, Dominique; Rajjou, Loïc

2014-01-01

During seed germination, the transition from a quiescent metabolic state in a dry mature seed to a proliferative metabolic state in a vigorous seedling is crucial for plant propagation as well as for optimizing crop yield. This work provides a detailed description of the dynamics of protein synthesis during the time course of germination, demonstrating that mRNA translation is both sequential and selective during this process. The complete inhibition of the germination process in the presence of the translation inhibitor cycloheximide established that mRNA translation is critical for Arabidopsis seed germination. However, the dynamics of protein turnover and the selectivity of protein synthesis (mRNA translation) during Arabidopsis seed germination have not been addressed yet. Based on our detailed knowledge of the Arabidopsis seed proteome, we have deepened our understanding of seed mRNA translation during germination by combining two-dimensional gel-based proteomics with dynamic radiolabeled proteomics using a radiolabeled amino acid precursor, namely [35S]-methionine, in order to highlight de novo protein synthesis, stability, and turnover. Our data confirm that during early imbibition, the Arabidopsis translatome keeps reflecting an embryonic maturation program until a certain developmental checkpoint. Furthermore, by dividing the seed germination time lapse into discrete time windows, we highlight precise and specific patterns of protein synthesis. These data refine and deepen our knowledge of the three classical phases of seed germination based on seed water uptake during imbibition and reveal that selective mRNA translation is a key feature of seed germination. Beyond the quantitative control of translational activity, both the selectivity of mRNA translation and protein turnover appear as specific regulatory systems, critical for timing the molecular events leading to successful germination and seedling establishment. PMID:24198433

Molecular IgV(H) analysis demonstrates highly somatic mutated B cells in synovialitis of osteoarthritis: a degenerative disease is associated with a specific, not locally generated immune response.

PubMed

Krenn, V; Hensel, F; Kim, H J; Souto Carneiro, M M; Starostik, P; Ristow, G; König, A; Vollmers, H P; Müller-Hermelink, H K

1999-11-01

In osteoarthritis (OA), the synovial tissue exhibits a nonfollicular inflammatory infiltration with a characteristic arrangement of lymphocytes and plasma cells. These arrangements are either small perivascular aggregates with plasma cells surrounding the lymphocytes or small groups of plasma cells, located in the vicinity of small blood vessels. These patterns suggest that B lymphocytes directly differentiate into plasma cells. To understand the B-cell response in OA, we analyzed the V(H) genes from B cells of synovial tissue of nine OA patients (average age, 71.5+/-10.5 years; six female and three male). V(H) gene repertoires were determined from RNA prepared from tissue cryosections and from DNA of single isolated B lymphocytes and plasma cells. The inflammatory infiltrate was analyzed immunohistochemically by detecting CD20, Ki-M4 (follicular dendritic cells), CD4, IgG, IgM, IgA, Ki-67, and by simultaneous demonstration of the plasma-cell-specific antigen CD138 (syndecan-1) and factor VIII. The molecular data demonstrate B cells with a high number of somatic mutations (average, 16.5 to 19.8), and high ratios of replacement to silent mutations in the small lymphocytic/plasmacellular aggregates of OA. In the tissue cryosections, the values of the sigmaR/sigmaS at the complementarity determining regions were 5.3 and 2.0 in the framework regions. For both the isolated B lymphocytes and plasma cells, the value of this ratio in the complementarity determining regions was 3.5. In the framework regions, the values of this ratio were 2.0 for the isolated B cells and 1.8 for the plasma cells. B lymphocytes and plasma cells exhibited a distribution not described thus far. Two patterns of B-cell distribution could be observed: (a) Centrally located CD20+ B and CD4+ and CD8+ T lymphocytes were surrounded directly by IgG (predominantly) or IgA and IgM plasma cells. No proliferating Ki-67-positive cells and no follicular dendritic cells (germinal centers) could be detected in the aggregates; (b) Plasma cells (predominantly IgG) were located directly near endothelial cells of small blood vessels. The finding of highly mutated V(H) genes in B lymphocytes and the characteristic arrangement of B lymphocytes and plasma cells suggests that B cells, which participate in OA synovialitis, have undergone germinal center reaction at different sites. This may explain the low inflammatory infiltration without germinal centers in OA, which is a feature of this primarily degenerative joint disease.

A soluble form of IL-13 receptor alpha 1 promotes IgG2a and IgG2b production by murine germinal center B cells.

PubMed

Poudrier, J; Graber, P; Herren, S; Gretener, D; Elson, G; Berney, C; Gauchat, J F; Kosco-Vilbois, M H

1999-08-01

A functional IL-13R involves at least two cell surface proteins, the IL-13R alpha 1 and IL-4R alpha. Using a soluble form of the murine IL-13R alpha 1 (sIL-13R), we reveal several novel features of this system. The sIL-13R promotes proliferation and augmentation of Ag-specific IgM, IgG2a, and IgG2b production by murine germinal center (GC) B cells in vitro. These effects were enhanced by CD40 signaling and were not inhibited by an anti-IL4R alpha mAb, a result suggesting other ligands. In GC cell cultures, sIL-13R also promoted IL-6 production, and interestingly, sIL-13R-induced IgG2a and IgG2b augmentation was absent in GC cells isolated from IL-6-deficient mice. Furthermore, the effects of the sIL-13R molecule were inhibited in the presence of an anti-IL-13 mAb, and preincubation of GC cells with IL-13 enhanced the sIL-13R-mediated effects. When sIL-13R was injected into mice, it served as an adjuvant-promoting production to varying degrees of IgM and IgG isotypes. We thus propose that IL-13R alpha 1 is a molecule involved in B cell differentiation, using a mechanism that may involve regulation of IL-6-responsive elements. Taken together, our data reveal previously unknown activities as well as suggest that the ligand for the sIL-13R might be a component of the IL-13R complex or a counterstructure yet to be defined.

Promotion of Testa Rupture during Garden Cress Germination Involves Seed Compartment-Specific Expression and Activity of Pectin Methylesterases1[OPEN

PubMed Central

Scheler, Claudia; Weitbrecht, Karin; Pearce, Simon P.; Hampstead, Anthony; Büttner-Mainik, Annette; Lee, Kieran J.D.; Voegele, Antje; Oracz, Krystyna; Dekkers, Bas J.W.; Wang, Xiaofeng; Wood, Andrew T.A.; Bentsink, Leónie; King, John R.; Knox, J. Paul; Holdsworth, Michael J.; Müller, Kerstin; Leubner-Metzger, Gerhard

2015-01-01

Pectin methylesterase (PME) controls the methylesterification status of pectins and thereby determines the biophysical properties of plant cell walls, which are important for tissue growth and weakening processes. We demonstrate here that tissue-specific and spatiotemporal alterations in cell wall pectin methylesterification occur during the germination of garden cress (Lepidium sativum). These cell wall changes are associated with characteristic expression patterns of PME genes and resultant enzyme activities in the key seed compartments CAP (micropylar endosperm) and RAD (radicle plus lower hypocotyl). Transcriptome and quantitative real-time reverse transcription-polymerase chain reaction analysis as well as PME enzyme activity measurements of separated seed compartments, including CAP and RAD, revealed distinct phases during germination. These were associated with hormonal and compartment-specific regulation of PME group 1, PME group 2, and PME inhibitor transcript expression and total PME activity. The regulatory patterns indicated a role for PME activity in testa rupture (TR). Consistent with a role for cell wall pectin methylesterification in TR, treatment of seeds with PME resulted in enhanced testa permeability and promoted TR. Mathematical modeling of transcript expression changes in germinating garden cress and Arabidopsis (Arabidopsis thaliana) seeds suggested that group 2 PMEs make a major contribution to the overall PME activity rather than acting as PME inhibitors. It is concluded that regulated changes in the degree of pectin methylesterification through CAP- and RAD-specific PME and PME inhibitor expression play a crucial role during Brassicaceae seed germination. PMID:25429110

A microfluidic device for real-time monitoring of Bacillus subtilis bacterial spores during germination based on non-specific physicochemical interactions on the nanoscale level.

PubMed

Zabrocka, L; Langer, K; Michalski, A; Kocik, J; Langer, J J

2015-01-07

A microfluidic device for studies on the germination of bacterial spores (e.g. Bacillus subtilis) based on non-specific interactions on the nanoscale is presented. A decrease in the population of spores during germination followed by the appearance of transition forms and an increase in the number of vegetative cells can be registered directly and simultaneously by using the microfluidic device, which is equipped with a conductive polymer layer (polyaniline) in the form of a nano-network. The lab-on-a-chip-type device, operating in a continuous flow regime, allows monitoring of germination of bacterial spores and analysis of the process in detail. The procedure is fast and accurate enough for quantitative real-time monitoring of the main steps of germination, including final transformation of the spores into vegetative cells. All of this is done without the use of biomarkers or any bio-specific materials, such as enzymes, antibodies and aptamers, and is simply based on an analysis of physicochemical interactions on the nanoscale level.

Unique Features of Fish Immune Repertoires: Particularities of Adaptive Immunity Within the Largest Group of Vertebrates

PubMed Central

Sunyer, Oriol J.

2016-01-01

Fishes (i.e., teleost fishes) are the largest group of vertebrates. Although their immune system is based on the fundamental receptors, pathways, and cell types found in all groups of vertebrates, fishes show a diversity of particular features that challenge some classical concepts of immunology. In this chapter, we discuss the particularities of fish immune repertoires from a comparative perspective. We examine how allelic exclusion can be achieved when multiple Ig loci are present, how isotypic diversity and functional specificity impact clonal complexity, how loss of the MHC class II molecules affects the cooperation between T and B cells, and how deep sequencing technologies bring new insights about somatic hypermutation in the absence of germinal centers. The unique coexistence of two distinct B-cell lineages respectively specialized in systemic and mucosal responses is also discussed. Finally, we try to show that the diverse adaptations of immune repertoires in teleosts can help in understanding how somatic adaptive mechanisms of immunity evolved in parallel in different lineages across vertebrates. PMID:26537384

Seed germination in parasitic plants: what insights can we expect from strigolactone research?

PubMed

Brun, Guillaume; Braem, Lukas; Thoiron, Séverine; Gevaert, Kris; Goormachtig, Sofie; Delavault, Philippe

2018-04-23

Obligate root-parasitic plants belonging to the Orobanchaceae family are deadly pests for major crops all over the world. Because these heterotrophic plants severely damage their hosts even before emerging from the soil, there is an unequivocal need to design early and efficient methods for their control. The germination process of these species has probably undergone numerous selective pressure events in the course of evolution, in that the perception of host-derived molecules is a necessary condition for seeds to germinate. Although most of these molecules belong to the strigolactones, structurally different molecules have been identified. Since strigolactones are also classified as novel plant hormones that regulate several physiological processes other than germination, the use of autotrophic model plant species has allowed the identification of many actors involved in the strigolactone biosynthesis, perception, and signal transduction pathways. Nevertheless, many questions remain to be answered regarding the germination process of parasitic plants. For instance, how did parasitic plants evolve to germinate in response to a wide variety of molecules, while autotrophic plants do not? What particular features are associated with their lack of spontaneous germination? In this review, we attempt to illustrate to what extent conclusions from research into strigolactones could be applied to better understand the biology of parasitic plants.

Ceratopteris richardii: a productive model for revealing secrets of signaling and development

NASA Technical Reports Server (NTRS)

Chatterjee, A.; Roux, S. J.

2000-01-01

Ceratopteris richardii is an aquatic fern grown in tropical and subtropical regions of the world. It is proven to be a productive model system for studies in the genetics, biochemistry, and cell biology of basic biologic processes that occur in early gametophytic development. It provides several advantages to biologists, especially those interested in gravitational biology, polarity development, and in the genetics of sexual development. It is easy to culture, has a relatively short life cycle, and offers an array of attractive features that facilitate genetic studies. The germination and early development of large populations of genetically identical spores are easy to synchronize, and both the direction of polarity development and cell-level gravity responses can be measured and readily manipulated within the first 24 h of spore development. Although there is no reliable transformation system available yet in Ceratopteris, recent studies suggest that the technique of RNA interference can be used to block translation of specific genes in a related fern, Marsilea, and current studies will soon reveal the applicability of this approach, as well as of other transformation approaches, in Ceratopteris. A recently completed expressed sequence tag (EST) sequencing project makes available the partial sequence of more than 2000 cDNAs, representing a significant percentage of the genes being expressed during the first 24 h of spore germination, when many developmentally interesting processes are occurring. A microarray of these ESTs is being constructed, so especially for those scientists interested in basic cellular phenomena that occur early in spore germination, the availability of the ESTs and of the microarray will make Ceratopteris an even more attractive model system.

Ceratopteris richardii: a productive model for revealing secrets of signaling and development.

PubMed

Chatterjee, A; Roux, S J

2000-09-01

Ceratopteris richardii is an aquatic fern grown in tropical and subtropical regions of the world. It is proven to be a productive model system for studies in the genetics, biochemistry, and cell biology of basic biologic processes that occur in early gametophytic development. It provides several advantages to biologists, especially those interested in gravitational biology, polarity development, and in the genetics of sexual development. It is easy to culture, has a relatively short life cycle, and offers an array of attractive features that facilitate genetic studies. The germination and early development of large populations of genetically identical spores are easy to synchronize, and both the direction of polarity development and cell-level gravity responses can be measured and readily manipulated within the first 24 h of spore development. Although there is no reliable transformation system available yet in Ceratopteris, recent studies suggest that the technique of RNA interference can be used to block translation of specific genes in a related fern, Marsilea, and current studies will soon reveal the applicability of this approach, as well as of other transformation approaches, in Ceratopteris. A recently completed expressed sequence tag (EST) sequencing project makes available the partial sequence of more than 2000 cDNAs, representing a significant percentage of the genes being expressed during the first 24 h of spore germination, when many developmentally interesting processes are occurring. A microarray of these ESTs is being constructed, so especially for those scientists interested in basic cellular phenomena that occur early in spore germination, the availability of the ESTs and of the microarray will make Ceratopteris an even more attractive model system.

Strategies of seedlings to overcome their sessile nature: auxin in mobility control.

PubMed

Žádníková, Petra; Smet, Dajo; Zhu, Qiang; Van Der Straeten, Dominique; Benková, Eva

2015-01-01

Plants are sessile organisms that are permanently restricted to their site of germination. To compensate for their lack of mobility, plants evolved unique mechanisms enabling them to rapidly react to ever changing environmental conditions and flexibly adapt their postembryonic developmental program. A prominent demonstration of this developmental plasticity is their ability to bend organs in order to reach the position most optimal for growth and utilization of light, nutrients, and other resources. Shortly after germination, dicotyledonous seedlings form a bended structure, the so-called apical hook, to protect the delicate shoot meristem and cotyledons from damage when penetrating through the soil. Upon perception of a light stimulus, the apical hook rapidly opens and the photomorphogenic developmental program is activated. After germination, plant organs are able to align their growth with the light source and adopt the most favorable orientation through bending, in a process named phototropism. On the other hand, when roots and shoots are diverted from their upright orientation, they immediately detect a change in the gravity vector and bend to maintain a vertical growth direction. Noteworthy, despite the diversity of external stimuli perceived by different plant organs, all plant tropic movements share a common mechanistic basis: differential cell growth. In our review, we will discuss the molecular principles underlying various tropic responses with the focus on mechanisms mediating the perception of external signals, transduction cascades and downstream responses that regulate differential cell growth and consequently, organ bending. In particular, we highlight common and specific features of regulatory pathways in control of the bending of organs and a role for the plant hormone auxin as a key regulatory component.

Acquisition of physical dormancy and ontogeny of the micropyle--water-gap complex in developing seeds of Geranium carolinianum (Geraniaceae).

PubMed

Gama-Arachchige, N S; Baskin, J M; Geneve, R L; Baskin, C C

2011-07-01

The 'hinged valve gap' has been previously identified as the initial site of water entry (i.e. water gap) in physically dormant (PY) seeds of Geranium carolinianum (Geraniaceae). However, neither the ontogeny of the hinged valve gap nor acquisition of PY by seeds of Geraniaceae has been studied previously. The aims of the present study were to investigate the physiological events related to acquisition of PY and the ontogeny of the hinged valve gap and seed coat of G. carolinianum. Seeds of G. carolinianum were studied from the ovule stage until dispersal. The developmental stages of acquisition of germinability, physiological maturity and PY were determined by seed measurement, germination and imbibition experiments using intact seeds and isolated embryos of both fresh and slow-dried seeds. Ontogeny of the seed coat and water gap was studied using light microscopy. Developing seeds achieved germinability, physiological maturity and PY on days 9, 14 and 20 after pollination (DAP), respectively. The critical moisture content of seeds on acquisition of PY was 11 %. Slow-drying caused the stage of acquisition of PY to shift from 20 to 13 DAP. Greater extent of cell division and differentiation at the micropyle, water gap and chalaza than at the rest of the seed coat resulted in particular anatomical features. Palisade and subpalisade cells of varying forms developed in these sites. A clear demarcation between the water gap and micropyle is not evident due to their close proximity. Acquisition of PY in seeds of G. carolinianum occurs after physiological maturity and is triggered by maturation drying. The micropyle and water gap cannot be considered as two separate entities, and thus it is more appropriate to consider them together as a 'micropyle--water-gap complex'.

Strategies of seedlings to overcome their sessile nature: auxin in mobility control

PubMed Central

Žádníková, Petra; Smet, Dajo; Zhu, Qiang; Straeten, Dominique Van Der; Benková, Eva

2015-01-01

Plants are sessile organisms that are permanently restricted to their site of germination. To compensate for their lack of mobility, plants evolved unique mechanisms enabling them to rapidly react to ever changing environmental conditions and flexibly adapt their postembryonic developmental program. A prominent demonstration of this developmental plasticity is their ability to bend organs in order to reach the position most optimal for growth and utilization of light, nutrients, and other resources. Shortly after germination, dicotyledonous seedlings form a bended structure, the so-called apical hook, to protect the delicate shoot meristem and cotyledons from damage when penetrating through the soil. Upon perception of a light stimulus, the apical hook rapidly opens and the photomorphogenic developmental program is activated. After germination, plant organs are able to align their growth with the light source and adopt the most favorable orientation through bending, in a process named phototropism. On the other hand, when roots and shoots are diverted from their upright orientation, they immediately detect a change in the gravity vector and bend to maintain a vertical growth direction. Noteworthy, despite the diversity of external stimuli perceived by different plant organs, all plant tropic movements share a common mechanistic basis: differential cell growth. In our review, we will discuss the molecular principles underlying various tropic responses with the focus on mechanisms mediating the perception of external signals, transduction cascades and downstream responses that regulate differential cell growth and consequently, organ bending. In particular, we highlight common and specific features of regulatory pathways in control of the bending of organs and a role for the plant hormone auxin as a key regulatory component. PMID:25926839

Phytotoxic effects of Sicyos deppei (Cucurbitaceae) in germinating tomato seeds.

PubMed

Lara-Núñez, Aurora; Sánchez-Nieto, Sobeida; Luisa Anaya, Ana; Cruz-Ortega, Rocio

2009-06-01

The phytotoxic effect of allelochemicals is referred to as allelochemical stress and it is considered a biotic stress. Sicyos deppei G. Don (Cucurbitaceae) is an allelopathic weed that causes phytotoxicity in Lycopersicon esculentum, delaying seed germination and severely inhibiting radicle growth. This paper reports in in vitro conditions, the effects of the aqueous leachate of S. deppei-throughout tomato germination times-on (1) the dynamics of starch and sugars metabolism, (2) activity and expression of the cell wall enzymes involved in endosperm weakening that allows the protrusion of the radicle, and (3) whether abscisic acid (ABA) is involved in this altered metabolic processes. Results showed that S. deppei leachate on tomato seed germination mainly caused: (1) delay in starch degradation as well as in sucrose hydrolysis; (2) lower activity of sucrose phosphate synthase, cell wall invertase, and alpha-amylase; being sucrose phosphate synthase (SPS) gene expression down-regulated, and the last two up regulated; (3) also, lower activity of endo beta-mannanase, beta-1,3 glucanase, alpha-galactosidase, and exo-polygalacturonase with altered gene expression; and (4) higher content of ABA during all times of germination. The phytotoxic effect of S. deppei aqueous leachate is because of the sum of many metabolic processes affected during tomato seed germination that finally is evidenced by a strong inhibition of radicle growth.

Vacuolar status and water relations in embryonic axes of recalcitrant Aesculus hippocastanum seeds during stratification and early germination.

PubMed

Obroucheva, Natalie V; Lityagina, Snezhana V; Novikova, Galina V; Sin'kevich, Irina A

2012-01-01

In tropical recalcitrant seeds, their rapid transition from shedding to germination at high hydration level is of physiological interest but difficult to study because of the time constraint. In recalcitrant horse chestnut seeds produced in central Russia, this transition is much longer and extends through dormancy and dormancy release. This extended time period permits studies of the water relations in embryonic axes during the long recalcitrant period in terms of vacuolar status and water transport. Horse chestnut (Aesculus hippocastanum) seeds sampled in Moscow were stratified in cold wet sand for 4 months. Vacuole presence and development in embryonic axes were examined by vital staining, light and electron microscopy. Aquaporins and vacuolar H(+)-ATPase were identified immunochemically. Water channel operation was tested by water inflow rate. Vacuolar acid invertase was estimated in terms of activity and electrophoretic properties. Throughout the long recalcitrant period after seed shedding, cells of embryonic axes maintained active vacuoles and a high water content. Preservation of enzyme machinery in vacuoles was evident from retention of invertase activity, substrate specificity, molecular mass and subunit composition. Plasmalemma and tonoplast aquaporins and the E subunit of vacuolar H(+)-ATPase were also present. In non-dormant seeds prior to growth initiation, vacuoles enlarged at first in hypocotyls, and then in radicles, with their biogenesis being similar. Vacuolation was accompanied by increasing invertase activity, leading to sugar accumulation and active osmotic functioning. After growth initiation, vacuole enlargement was favoured by enhanced water inflow through water channels formed by aquaporins. Maintenance of high water content and desiccation sensitivity, as well as preservation of active vacuoles in embryonic axes after shedding, can be considered a specific feature of recalcitrant seeds, overlooked when studying tropical recalcitrants due to the short duration. The retained physiological activity of vacuoles allows them to function rapidly as dormancy is lost and when external conditions permit. Cell vacuolation precedes cell elongation in both hypocotyl and radicle, and provides impetus for rapid germination.

CRISPR-Mediated Triple Knockout of SLAMF1, SLAMF5 and SLAMF6 Supports Positive Signaling Roles in NKT Cell Development.

PubMed

Huang, Bonnie; Gomez-Rodriguez, Julio; Preite, Silvia; Garrett, Lisa J; Harper, Ursula L; Schwartzberg, Pamela L

2016-01-01

The SLAM family receptors contribute to diverse aspects of lymphocyte biology and signal via the small adaptor molecule SAP. Mutations affecting SAP lead to X-linked lymphoproliferative syndrome Type 1, a severe immunodysregulation characterized by fulminant mononucleosis, dysgammaglobulinemia, and lymphoproliferation/lymphomas. Patients and mice having mutations affecting SAP also lack germinal centers due to a defect in T:B cell interactions and are devoid of invariant NKT (iNKT) cells. However, which and how SLAM family members contribute to these phenotypes remains uncertain. Three SLAM family members: SLAMF1, SLAMF5 and SLAMF6, are highly expressed on T follicular helper cells and germinal center B cells. SLAMF1 and SLAMF6 are also implicated in iNKT development. Although individual receptor knockout mice have limited iNKT and germinal center phenotypes compared to SAP knockout mice, the generation of multi-receptor knockout mice has been challenging, due to the genomic linkage of the genes encoding SLAM family members. Here, we used Cas9/CRISPR-based mutagenesis to generate mutations simultaneously in Slamf1, Slamf5 and Slamf6. Genetic disruption of all three receptors in triple-knockout mice (TKO) did not grossly affect conventional T or B cell development and led to mild defects in germinal center formation post-immunization. However, the TKO worsened defects in iNKT cells development seen in SLAMF6 single gene-targeted mice, supporting data on positive signaling and potential redundancy between these receptors.

Effects of UV-B radiation on the isoflavone accumulation and physiological-biochemical changes of soybean during germination: Physiological-biochemical change of germinated soybean induced by UV-B.

PubMed

Ma, Meng; Wang, Pei; Yang, Runqiang; Gu, Zhenxin

2018-06-01

In this study, the effects of UV-B radiation on the isoflavones accumulation, physiological and nutritional quality, water status, and characteristics of proteins in germinated soybeans were investigated. The results showed that isoflavones content in soybeans increased with appropriate intensity and time of UV-B radiation and decreased with excessive treatment. Fresh weight, length, free amino acids, reducing sugar contents and bulk water (T 23 ) in germinated soybeans decreased with increasing radiation time, indicating that UV-B inhibited the growth and nutrients metabolism of soybean during germination. Cell damage was detected in germinated soybeans with excessive UV-B radiation, as shown by the black spots in cotyledons and the increased intercellular water determined by LF-NMR. Germination resulted in an increase in random coil structures, while UV-B radiation induced no obvious changes in FT-IR spectrum and protein conformation of soybeans. Both UV-B radiation and germination caused the increase in soluble proteins, especially in 1.0-75.0 kDa fraction. Copyright © 2018 Elsevier Ltd. All rights reserved.

Effect of ferrous sulfate fortification in germinated brown rice on seed iron concentration and bioavailability.

PubMed

Wei, Yanyan; Shohag, M J I; Ying, Feng; Yang, Xiaoe; Wu, Chunyong; Wang, Yuyan

2013-06-01

The present study evaluated the effectiveness of germination and iron fortification on iron concentration and bioavailability of brown rice. Iron fortification during germination process with 0.05-2 g/L ferrous sulfate increased the iron concentration in germinated brown rice from 1.1 to 15.6 times than those in raw brown rice. Based on the recommended dietary allowance of iron, maximum germination rate and γ-aminobutyric acid, we recommend the brown rice fortified with 0.25 g/L FeSO(4) as a suitable fortification level to use in germination process. Iron fortification during the germination process has a positive effect on iron concentration and bioavailability. A significant difference was observed among the cultivars in respect to the capacity for iron accumulation and bioavailability. Germination alone could improve in vitro iron solubility, but had no effect on iron bioavailability in Caco-2 cell, the additional fortification process should be combined to get high amount of bioavailable iron from the brown rice. Copyright © 2012 Elsevier Ltd. All rights reserved.

Changes in the pattern of protein synthesis during zoospore germination in Blastocladiella emersonii.

PubMed Central

Silva, A M; Maia, J C; Juliani, M H

1987-01-01

Using two-dimensional gel electrophoresis, we analyzed the pattern of proteins synthesized during Blastocladiella emersonii zoospore germination in an inorganic solution, in both the presence and absence of actinomycin D. During the transition from zoospore to round cells (the first 25 min), essentially no qualitative differences were noticeable, indicating that the earliest stages of germination are entirely preprogrammed with stored RNA. Later in germination (after 25 min), however, changes in the pattern of protein synthesis were found. Some of these proteins (a total of 6 polypeptides) correspond possibly to a selective translation of stored messages, whereas the majority of the changed proteins (22 polypeptides) corresponds to newly synthesized mRNA. Thus, multiple levels of protein synthesis regulation seem to occur during zoospore germination, involving both transcriptional and translational controls. We also analyzed the pattern of protein synthesis during germination in a nutrient medium; synthesis of specific polypeptides occurred during late germination. During early germination posttranslational control was also observed, several labeled proteins from zoospores being specifically degraded or charge modified. Images PMID:3571161

Immunoarchitectural patterns in nodal marginal zone B-cell lymphoma: a study of 51 cases.

PubMed

Salama, Mohamed E; Lossos, Izidore S; Warnke, Roger A; Natkunam, Yasodha

2009-07-01

Nodal marginal zone lymphoma (NMZL) represents a rare and heterogeneous group that lacks markers specific for the diagnosis. We evaluated morphologic and immunoarchitectural features of 51 NMZLs, and the following immunostains were performed: CD20, CD21, CD23, CD5, CD3, CD43, CD10, Ki-67, BCL1, BCL2, BCL6, HGAL, and LMO2. Four immunoarchitectural patterns were evident: diffuse (38 [75%]), well-formed nodular/follicular (5 [10%]), interfollicular (7 [14%]), and perifollicular (1 [2%]). Additional features included a monocytoid component (36 [71%]), admixed large cells (20 [39%]), plasma cells (24 [47%]), compartmentalizing stromal sclerosis (13 [25%]), and prominent blood vessel sclerosis (10 [20%]). CD21 highlighted disrupted follicular dendritic cell meshwork in 35 (71%) of 49 cases, and CD43 coexpression was present in 10 (24%) of 42 cases. A panel of germinal center-associated markers was helpful in eliminating cases of diffuse follicle center lymphoma. Our results highlight the histologic and immunoarchitectural spectrum of NMZL and the usefulness of immunohistochemical analysis for CD43, CD23, CD21, BCL6, HGAL, and LMO2 in the diagnosis of NMZL.

Metal Tolerance Protein 8 Mediates Manganese Homeostasis and Iron Reallocation during Seed Development and Germination.

PubMed

Eroglu, Seckin; Giehl, Ricardo F H; Meier, Bastian; Takahashi, Michiko; Terada, Yasuko; Ignatyev, Konstantin; Andresen, Elisa; Küpper, Hendrik; Peiter, Edgar; von Wirén, Nicolaus

2017-07-01

Metal accumulation in seeds is a prerequisite for germination and establishment of plants but also for micronutrient delivery to humans. To investigate metal transport processes and their interactions in seeds, we focused on METAL TOLERANCE PROTEIN8 (MTP8), a tonoplast transporter of the manganese (Mn) subclade of cation diffusion facilitators, which in Arabidopsis ( Arabidopsis thaliana ) is expressed in embryos of seeds. The x-ray fluorescence imaging showed that expression of MTP8 was responsible for Mn localization in subepidermal cells on the abaxial side of the cotyledons and in cortical cells of the hypocotyl. Accordingly, under low Mn availability, MTP8 increased seed stores of Mn, required for efficient seed germination. In mutant embryos lacking expression of VACUOLAR IRON TRANSPORTER1 ( VIT1 ), MTP8 built up iron (Fe) hotspots in MTP8 -expressing cells types, suggesting that MTP8 transports Fe in addition to Mn. In mtp8 vit1 double mutant seeds, Mn and Fe were distributed in all cell types of the embryo. An Fe transport function of MTP8 was confirmed by its ability to complement Fe hypersensitivity of a yeast mutant defective in vacuolar Fe transport. Imbibing mtp8-1 mutant seeds in the presence of Mn or subjecting seeds to wet-dry cycles showed that MTP8 conferred Mn tolerance. During germination, MTP8 promoted reallocation of Fe from the vasculature. These results indicate that cell type-specific accumulation of Mn and Fe in seeds depends on MTP8 and that this transporter plays an important role in the generation of seed metal stores as well as for metal homeostasis and germination efficiency under challenging environmental conditions. © 2017 American Society of Plant Biologists. All Rights Reserved.

Metal Tolerance Protein 8 Mediates Manganese Homeostasis and Iron Reallocation during Seed Development and Germination1[OPEN

PubMed Central

Takahashi, Michiko; Terada, Yasuko

2017-01-01

Metal accumulation in seeds is a prerequisite for germination and establishment of plants but also for micronutrient delivery to humans. To investigate metal transport processes and their interactions in seeds, we focused on METAL TOLERANCE PROTEIN8 (MTP8), a tonoplast transporter of the manganese (Mn) subclade of cation diffusion facilitators, which in Arabidopsis (Arabidopsis thaliana) is expressed in embryos of seeds. The x-ray fluorescence imaging showed that expression of MTP8 was responsible for Mn localization in subepidermal cells on the abaxial side of the cotyledons and in cortical cells of the hypocotyl. Accordingly, under low Mn availability, MTP8 increased seed stores of Mn, required for efficient seed germination. In mutant embryos lacking expression of VACUOLAR IRON TRANSPORTER1 (VIT1), MTP8 built up iron (Fe) hotspots in MTP8-expressing cells types, suggesting that MTP8 transports Fe in addition to Mn. In mtp8 vit1 double mutant seeds, Mn and Fe were distributed in all cell types of the embryo. An Fe transport function of MTP8 was confirmed by its ability to complement Fe hypersensitivity of a yeast mutant defective in vacuolar Fe transport. Imbibing mtp8-1 mutant seeds in the presence of Mn or subjecting seeds to wet-dry cycles showed that MTP8 conferred Mn tolerance. During germination, MTP8 promoted reallocation of Fe from the vasculature. These results indicate that cell type-specific accumulation of Mn and Fe in seeds depends on MTP8 and that this transporter plays an important role in the generation of seed metal stores as well as for metal homeostasis and germination efficiency under challenging environmental conditions. PMID:28461400

Chemical sporulation and germination: cytoprotective nanocoating of individual mammalian cells with a degradable tannic acid-FeIII complex

NASA Astrophysics Data System (ADS)

Lee, Juno; Cho, Hyeoncheol; Choi, Jinsu; Kim, Doyeon; Hong, Daewha; Park, Ji Hun; Yang, Sung Ho; Choi, Insung S.

2015-11-01

Individual mammalian cells were coated with cytoprotective and degradable films by cytocompatible processes maintaining the cell viability. Three types of mammalian cells (HeLa, NIH 3T3, and Jurkat cells) were coated with a metal-organic complex of tannic acid (TA) and ferric ion, and the TA-FeIII nanocoat effectively protected the coated mammalian cells against UV-C irradiation and a toxic compound. More importantly, the cell proliferation was controlled by programmed formation and degradation of the TA-FeIII nanocoat, mimicking the sporulation and germination processes found in nature.Individual mammalian cells were coated with cytoprotective and degradable films by cytocompatible processes maintaining the cell viability. Three types of mammalian cells (HeLa, NIH 3T3, and Jurkat cells) were coated with a metal-organic complex of tannic acid (TA) and ferric ion, and the TA-FeIII nanocoat effectively protected the coated mammalian cells against UV-C irradiation and a toxic compound. More importantly, the cell proliferation was controlled by programmed formation and degradation of the TA-FeIII nanocoat, mimicking the sporulation and germination processes found in nature. Electronic supplementary information (ESI) available: Experimental details, LSCM images, and SEM and TEM images. See DOI: 10.1039/c5nr05573c

A direct observation technique for evaluating sclerotium germination by Macrophomina phaseolina and effects of biocontrol materials on survival of sclerotia in soil.

PubMed

Pratt, Robert G

2006-08-01

Germination of sclerotia of Macrophomina phaseolina was quantified by direct microscopic observation following application of experimental treatments in vitro and incubation of sclerotia in soil. To assay germination, pieces of agar containing sclerotia were macerated in dilute, liquid cornmeal agar on glass slides; thinly spread; and incubated in a saturated atmosphere for 18-22 h. Germinated sclerotia then were identified by morphological features of germ hyphae. Frequencies of germination were similar in three dilute agar media. Germination was not affected by air-drying sclerotia for 2 weeks, but it was significantly reduced after 4 weeks and greatly reduced or eliminated after 6 or 8 weeks. Survival of sclerotia for 14 days in soil was greatest at 50, 75, and 100% moisture-holding capacity, less at 0 and 25%, and least at 125% (flooded soil). Incorporation of ground poultry litter into soil at 5% by weight reduced survival of sclerotia after 13 days, and incorporation of litter at 10% nearly eliminated it. These results indicate that the direct-observation technique may be used to evaluate animal wastes and other agricultural byproducts for biocontrol activity against sclerotia of M. phaseolina in soil.

Affinity of antigen encounter and other early B-cell signals determine B-cell fate

PubMed Central

Benson, Micah J; Erickson, Loren D; Gleeson, Michael W; Noelle, Randolph J

2010-01-01

Three possible effector fates await the naïve follicular B cell following antigen stimulation in thymus-dependent reactions. Short-lived plasma cells produce an initial burst of germline-encoded protective antibodies, and long-lived plasma cells and memory B cells arise from the germinal center and function to enhance and sustain the humoral immune response. The inherent B-cell receptor affinity of naïve follicular B cells and the contribution of other early B-cell signals pre-determines the pattern of transcription factor expression and the differentiation path taken by these cells. High initial B-cell receptor affinity shunts naïve follicular B-cell clones towards the short-lived plasma cell fate, whereas modest-affinity clones are skewed towards a plasma cell fate and low-affinity clones are recruited into the germinal center and are selected for both long-lived plasma cells and memory B cell pathways. In the germinal center reaction, increased levels of the transcription factor interferon regulatory factor-4 drive the molecular program that dictates differentiation into the long-lived plasma cell phenotype but has no impact on the memory B cell compartment. We hypothesize that graded interferon regulatory factor-4 levels driven by signals to B cells, including B-cell receptor signal strength, are responsible for this branch point in the B-cell terminal differentiation pathway. PMID:17433651

Germinal center texture entropy as possible indicator of humoral immune response: immunophysiology viewpoint.

PubMed

Pantic, Igor; Pantic, Senka

2012-10-01

In this article, we present the results indicating that spleen germinal center (GC) texture entropy determined by gray-level co-occurrence matrix (GLCM) method is related to humoral immune response. Spleen tissue was obtained from eight outbred male short-haired guinea pigs previously immunized by sheep red blood cells (SRBC). A total of 312 images from 39 germinal centers (156 GC light zone images and 156 GC dark zone images) were acquired and analyzed by GLCM method. Angular second moment, contrast, correlation, entropy, and inverse difference moment were calculated for each image. Humoral immune response to SRBC was measured using T cell-dependent antibody response (TDAR) assay. Statistically highly significant negative correlation was detected between light zone entropy and the number of TDAR plaque-forming cells (r (s) = -0.86, p < 0.01). The entropy decreased as the plaque-forming cells increased and vice versa. A statistically significant negative correlation was also detected between dark zone entropy values and the number of plaque-forming cells (r (s) = -0.69, p < 0.05). Germinal center texture entropy may be a powerful indicator of humoral immune response. This study is one of the first to point out the potential scientific value of GLCM image texture analysis in lymphoid tissue cytoarchitecture evaluation. Lymphoid tissue texture analysis could become an important and affordable addition to the conventional immunophysiology techniques.

Human germinal center CD4+CD57+ T cells act differently on B cells than do classical T-helper cells.

PubMed

Bouzahzah, F; Bosseloir, A; Heinen, E; Simar, L J

1995-01-01

We have isolated two subtypes of helper T cells from human tonsils: CD4+CD57+ cells, mostly located in the germinal center (GC), and CD4+CD57- cells, distributed through the interfollicular areas but also present in the GC. In a functional study, we have compared the capacities of these T-cell subtypes to stimulate B cells in cocultures. In order to block T-cell proliferation while maintaining their activation level, we pretreated isolated T cells with mitomycin C prior to culture in the presence of B cells and added polyclonal activators such as PHA and Con A, combined or not with IL-2. Contrary to CD4+ CD57- cells, CD4+CD57+ cells did not markedly enhance B-cell proliferation. Even when sIgD.B cells typical of germinal center cells were tested, the CD4+CD57+ cells had no significant effect. This is in accordance with the location of these cells: They mainly occupy the light zones of the GC where few B cells divide. Even when added to preactivated, actively proliferating cells, CD4+CD57 cells failed to modulate B-cell multiplication. On the supernatants of B-cell-T-cell cocultures, we examined by the ELISA technique the effect of T cells on Ig synthesis. Contrary to CD57+ T cells, whose effect was strong, CD57- T cells weakly stimulated Ig synthesis. More IgM than IgG was generally found. Because CD57 antigen is a typical marker of natural killer cells, we tested the cytolytic activity of tonsillar CD4+CD57+ cells on K562 target cells. Unlike NK cells, neither CD4+CD57+ nor CD4+CD57- cells exhibit any cytotoxicity. Thus, germinal center CD4+CD57+ cells are not cytolytic and do not strongly stimulate either B-cell proliferation or Ig secretion. CD4+CD57- cells, however, enhance B-cell proliferation and differentiation, thus acting like the classical helper cells of the T-dependent areas.

Synergistic Anti-Tumor Activity of EZH2 Inhibitors and Glucocorticoid Receptor Agonists in Models of Germinal Center Non-Hodgkin Lymphomas

PubMed Central

Klaus, Christine R.; Wigle, Tim J.; Iwanowicz, Dorothy; Littlefield, Bruce A.; Porter-Scott, Margaret; Smith, Jesse J.; Moyer, Mikel P.; Copeland, Robert A.; Pollock, Roy M.; Kuntz, Kevin W.; Raimondi, Alejandra; Keilhack, Heike

2014-01-01

Patients with non-Hodgkin lymphoma (NHL) are treated today with a cocktail of drugs referred to as CHOP (Cyclophosphamide, Hydroxyldaunorubicin, Oncovin, and Prednisone). Subsets of patients with NHL of germinal center origin bear oncogenic mutations in the EZH2 histone methyltransferase. Clinical testing of the EZH2 inhibitor EPZ-6438 has recently begun in patients. We report here that combining EPZ-6438 with CHOP in preclinical cell culture and mouse models results in dramatic synergy for cell killing in EZH2 mutant germinal center NHL cells. Surprisingly, we observe that much of this synergy is due to Prednisolone – a glucocorticoid receptor agonist (GRag) component of CHOP. Dramatic synergy was observed when EPZ-6438 is combined with Prednisolone alone, and a similar effect was observed with Dexamethasone, another GRag. Remarkably, the anti-proliferative effect of the EPZ-6438+GRag combination extends beyond EZH2 mutant-bearing cells to more generally impact germinal center NHL. These preclinical data reveal an unanticipated biological intersection between GR-mediated gene regulation and EZH2-mediated chromatin remodeling. The data also suggest the possibility of a significant and practical benefit of combining EZH2 inhibitors and GRag that warrants further investigation in a clinical setting. PMID:25493630

Synergistic Anti-Tumor Activity of EZH2 Inhibitors and Glucocorticoid Receptor Agonists in Models of Germinal Center Non-Hodgkin Lymphomas.

PubMed

Knutson, Sarah K; Warholic, Natalie M; Johnston, L Danielle; Klaus, Christine R; Wigle, Tim J; Iwanowicz, Dorothy; Littlefield, Bruce A; Porter-Scott, Margaret; Smith, Jesse J; Moyer, Mikel P; Copeland, Robert A; Pollock, Roy M; Kuntz, Kevin W; Raimondi, Alejandra; Keilhack, Heike

2014-01-01

Patients with non-Hodgkin lymphoma (NHL) are treated today with a cocktail of drugs referred to as CHOP (Cyclophosphamide, Hydroxyldaunorubicin, Oncovin, and Prednisone). Subsets of patients with NHL of germinal center origin bear oncogenic mutations in the EZH2 histone methyltransferase. Clinical testing of the EZH2 inhibitor EPZ-6438 has recently begun in patients. We report here that combining EPZ-6438 with CHOP in preclinical cell culture and mouse models results in dramatic synergy for cell killing in EZH2 mutant germinal center NHL cells. Surprisingly, we observe that much of this synergy is due to Prednisolone - a glucocorticoid receptor agonist (GRag) component of CHOP. Dramatic synergy was observed when EPZ-6438 is combined with Prednisolone alone, and a similar effect was observed with Dexamethasone, another GRag. Remarkably, the anti-proliferative effect of the EPZ-6438+GRag combination extends beyond EZH2 mutant-bearing cells to more generally impact germinal center NHL. These preclinical data reveal an unanticipated biological intersection between GR-mediated gene regulation and EZH2-mediated chromatin remodeling. The data also suggest the possibility of a significant and practical benefit of combining EZH2 inhibitors and GRag that warrants further investigation in a clinical setting.

Proteomic Analysis of Lettuce Seed Germination and Thermoinhibition by Sampling of Individual Seeds at Germination and Removal of Storage Proteins by Polyethylene Glycol Fractionation1

PubMed Central

Song, Bin-Yan; Deng, Zhi-Jun; Wang, Yue; Liu, Shu-Jun; Møller, Ian Max; Song, Song-Quan

2015-01-01

Germination and thermoinhibition in lettuce (Lactuca sativa ‘Jianyexianfeng No. 1’) seeds were investigated by a proteomic comparison among dry seeds, germinated seeds at 15°C, at 15°C after imbibition at 25°C for 48 h, or at 25°C in KNO3 (all sampled individually at germination), and ungerminated seeds at 25°C, a thermoinhibitory temperature. Before two-dimensional gel electrophoresis analysis, storage proteins (greater than 50% of total extractable protein) were removed by polyethylene glycol precipitation, which significantly improved the detection of less abundant proteins on two-dimensional gels. A total of 108 protein spots were identified to change more than 2-fold (P < 0.05) in abundance in at least one germination treatment. Nineteen proteins increasing and one protein decreasing in abundance during germination had higher abundance in germinated 15°C, 15°C after imbibition at 25°C for 48 h, and 25°C in KNO3 seeds than in ungerminated 25°C seeds. Gene expression of 12 of those proteins correlated well with the protein accumulation. Methionine metabolism, ethylene production, lipid mobilization, cell elongation, and detoxification of aldehydes were revealed to be potentially related to lettuce seed germination and thermoinhibition. Accumulation of three proteins and expression of five genes participating in the mevalonate (MVA) pathway of isoprenoid biosynthesis correlated positively with seed germinability. Inhibition of this pathway by lovastatin delayed seed germination and increased the sensitivity of germination to abscisic acid. MVA pathway-derived products, cytokinins, partially reversed the lovastatin inhibition of germination and released seed thermoinhibition at 25°C. We conclude that the MVA pathway for isoprenoid biosynthesis is involved in lettuce seed germination and thermoinhibition. PMID:25736209

Oxytropism: a new twist in pollen tube orientation

NASA Technical Reports Server (NTRS)

Blasiak, J.; Mulcahy, D. L.; Musgrave, M.

2001-01-01

Chemical gradients and structural features within the pistil have been previously proposed as factors determining the directionality of pollen tube growth. In this study, we examine the behavior of pollen of eight species germinated in a dynamic oxygen gradient. While the germination rates of some species decreased directly with decreasing oxygen tension, other species showed no decrease in germination at oxygen tensions as low as 2 kPa. In one species, germination was consistently greater at decreased oxygen tensions than at ambient atmospheric levels. In three of the eight species tested, the developing pollen tube showed clear directional growth away from the more-oxygenated regions of the growth medium, while in one species growth was towards the more-oxygenated region. The remaining four species showed random tube growth. The pattern of oxytropic responses among the taxa suggests that this tropic behavior is both widespread and phylogenetically unpredictable.

Investigating the Detrimental Effects of Low Pressure Plasma Sterilization on the Survival of Bacillus subtilis Spores Using Live Cell Microscopy.

PubMed

Fuchs, Felix M; Raguse, Marina; Fiebrandt, Marcel; Madela, Kazimierz; Awakowicz, Peter; Laue, Michael; Stapelmann, Katharina; Moeller, Ralf

2017-11-30

Plasma sterilization is a promising alternative to conventional sterilization methods for industrial, clinical, and spaceflight purposes. Low pressure plasma (LPP) discharges contain a broad spectrum of active species, which lead to rapid microbial inactivation. To study the efficiency and mechanisms of sterilization by LPP, we use spores of the test organism Bacillus subtilis because of their extraordinary resistance against conventional sterilization procedures. We describe the production of B. subtilis spore monolayers, the sterilization process by low pressure plasma in a double inductively coupled plasma reactor, the characterization of spore morphology using scanning electron microscopy (SEM), and the analysis of germination and outgrowth of spores by live cell microscopy. A major target of plasma species is genomic material (DNA) and repair of plasma-induced DNA lesions upon spore revival is crucial for survival of the organism. Here, we study the germination capacity of spores and the role of DNA repair during spore germination and outgrowth after treatment with LPP by tracking fluorescently-labelled DNA repair proteins (RecA) with time-resolved confocal fluorescence microscopy. Treated and untreated spore monolayers are activated for germination and visualized with an inverted confocal live cell microscope over time to follow the reaction of individual spores. Our observations reveal that the fraction of germinating and outgrowing spores is dependent on the duration of LPP-treatment reaching a minimum after 120 s. RecA-YFP (yellow fluorescence protein) fluorescence was detected only in few spores and developed in all outgrowing cells with a slight elevation in LPP-treated spores. Moreover, some of the vegetative bacteria derived from LPP-treated spores showed an increase in cytoplasm and tended to lyse. The described methods for analysis of individual spores could be exemplary for the study of other aspects of spore germination and outgrowth.

The biomechanics of seed germination.

PubMed

Steinbrecher, Tina; Leubner-Metzger, Gerhard

2017-02-01

From a biomechanical perspective, the completion of seed (and fruit) germination depends on the balance of two opposing forces: the growth potential of the embryonic axis (radicle-hypocotyl growth zone) and the restraint of the seed-covering layers (endosperm, testa, and pericarp). The diverse seed tissues are composite materials which differ in their dynamic properties based on their distinct cell wall composition and water uptake capacities. The biomechanics of embryo cell growth during seed germination depend on irreversible cell wall loosening followed by water uptake due to the decreasing turgor, and this leads to embryo elongation and eventually radicle emergence. Endosperm weakening as a prerequisite for radicle emergence is a widespread phenomenon among angiosperms. Research into the biochemistry and biomechanics of endosperm weakening has demonstrated that the reduction in puncture force of a seed's micropylar endosperm is environmentally and hormonally regulated and involves tissue-specific expression of cell wall remodelling proteins such as expansins, diverse hydrolases, and the production of directly acting apoplastic reactive oxygen. The endosperm-weakening biomechanics and its underlying cell wall biochemistry differ between the micropylar (ME) and chalazal (CE) endosperm domains. In the ME, they involve cell wall loosening, cell separation, and programmed cell death to provide decreased and localized ME tissue resistance, autolysis, and finally the formation of an ME hole required for radicle emergence. Future work will further unravel the molecular mechanisms, environmental regulation, and evolution of the diverse biomechanical cell wall changes underpinning the control of germination by endosperm weakening. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

A study of the effects of micro-gravity on seed germination

NASA Technical Reports Server (NTRS)

Klein, Lynn Suzanne; Mckibben, Mark; Brain, David A.; Johnson, Theodore C.; Dannenberg, Konrad K.

1992-01-01

This study will identify characteristics of seed germination dependent upon gravity. To accomplish this objective, four different seed types will be germinated in space and then be compared to a control group germinated on Earth. Both the experimental and control groups will be analyzed on the cellular level for the size of cells, structural anomalies, and gravitational effects. The experiment will be conducted in a Get Away Special Canister (GAS Can no. 608) owned by the U.S. Space and Rocket Center and designed for students. The GAS Can will remain in the cargo bay of the Space Shuttle with minimal astronaut interaction.

Cold tolerance in rice germinating seeds revealed by deep RNAseq analysis of contrasting indica genotypes.

PubMed

Dametto, Andressa; Sperotto, Raul A; Adamski, Janete M; Blasi, Édina A R; Cargnelutti, Denise; de Oliveira, Luiz F V; Ricachenevsky, Felipe K; Fregonezi, Jeferson N; Mariath, Jorge E A; da Cruz, Renata P; Margis, Rogério; Fett, Janette P

2015-09-01

Rice productivity is largely affected by low temperature, which can be harmful throughout plant development, from germination to grain filling. Germination of indica rice cultivars under cold is slow and not uniform, resulting in irregular emergence and small plant population. To identify and characterize novel genes involved in cold tolerance during the germination stage, two indica rice genotypes (sister lines previously identified as cold-tolerant and cold-sensitive) were used in parallel transcriptomic analysis (RNAseq) under cold treatment (seeds germinating at 13 °C for 7 days). We detected 1,361 differentially expressed transcripts. Differences in gene expression found by RNAseq were confirmed for 11 selected genes using RT-qPCR. Biological processes enhanced in the cold-tolerant seedlings include: cell division and expansion (confirmed by anatomical sections of germinating seeds), cell wall integrity and extensibility, water uptake and membrane transport capacity, sucrose synthesis, generation of simple sugars, unsaturation of membrane fatty acids, wax biosynthesis, antioxidant capacity (confirmed by histochemical staining of H2O2), and hormone and Ca(2+)-signaling. The cold-sensitive seedlings respond to low temperature stress increasing synthesis of HSPs and dehydrins, along with enhanced ubiquitin/proteasome protein degradation pathway and polyamine biosynthesis. Our findings can be useful in future biotechnological approaches aiming to cold tolerance in indica rice. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

A Multifaceted Study of Scedosporium boydii Cell Wall Changes during Germination and Identification of GPI-Anchored Proteins

PubMed Central

Ghamrawi, Sarah; Gastebois, Amandine; Zykwinska, Agata; Vandeputte, Patrick; Marot, Agnès; Mabilleau, Guillaume; Cuenot, Stéphane; Bouchara, Jean-Philippe

2015-01-01

Scedosporium boydii is a pathogenic filamentous fungus that causes a wide range of human infections, notably respiratory infections in patients with cystic fibrosis. The development of new therapeutic strategies targeting S. boydii necessitates a better understanding of the physiology of this fungus and the identification of new molecular targets. In this work, we studied the conidium-to-germ tube transition using a variety of techniques including scanning and transmission electron microscopy, atomic force microscopy, two-phase partitioning, microelectrophoresis and cationized ferritin labeling, chemical force spectroscopy, lectin labeling, and nanoLC-MS/MS for cell wall GPI-anchored protein analysis. We demonstrated that the cell wall undergoes structural changes with germination accompanied with a lower hydrophobicity, electrostatic charge and binding capacity to cationized ferritin. Changes during germination also included a higher accessibility of some cell wall polysaccharides to lectins and less CH3/CH3 interactions (hydrophobic adhesion forces mainly due to glycoproteins). We also extracted and identified 20 GPI-anchored proteins from the cell wall of S. boydii, among which one was detected only in the conidial wall extract and 12 only in the mycelial wall extract. The identified sequences belonged to protein families involved in virulence in other fungi like Gelp/Gasp, Crhp, Bglp/Bgtp families and a superoxide dismutase. These results highlighted the cell wall remodeling during germination in S. boydii with the identification of a substantial number of cell wall GPI-anchored conidial or hyphal specific proteins, which provides a basis to investigate the role of these molecules in the host-pathogen interaction and fungal virulence. PMID:26038837

A Multifaceted Study of Scedosporium boydii Cell Wall Changes during Germination and Identification of GPI-Anchored Proteins.

PubMed

Ghamrawi, Sarah; Gastebois, Amandine; Zykwinska, Agata; Vandeputte, Patrick; Marot, Agnès; Mabilleau, Guillaume; Cuenot, Stéphane; Bouchara, Jean-Philippe

2015-01-01

Scedosporium boydii is a pathogenic filamentous fungus that causes a wide range of human infections, notably respiratory infections in patients with cystic fibrosis. The development of new therapeutic strategies targeting S. boydii necessitates a better understanding of the physiology of this fungus and the identification of new molecular targets. In this work, we studied the conidium-to-germ tube transition using a variety of techniques including scanning and transmission electron microscopy, atomic force microscopy, two-phase partitioning, microelectrophoresis and cationized ferritin labeling, chemical force spectroscopy, lectin labeling, and nanoLC-MS/MS for cell wall GPI-anchored protein analysis. We demonstrated that the cell wall undergoes structural changes with germination accompanied with a lower hydrophobicity, electrostatic charge and binding capacity to cationized ferritin. Changes during germination also included a higher accessibility of some cell wall polysaccharides to lectins and less CH3/CH3 interactions (hydrophobic adhesion forces mainly due to glycoproteins). We also extracted and identified 20 GPI-anchored proteins from the cell wall of S. boydii, among which one was detected only in the conidial wall extract and 12 only in the mycelial wall extract. The identified sequences belonged to protein families involved in virulence in other fungi like Gelp/Gasp, Crhp, Bglp/Bgtp families and a superoxide dismutase. These results highlighted the cell wall remodeling during germination in S. boydii with the identification of a substantial number of cell wall GPI-anchored conidial or hyphal specific proteins, which provides a basis to investigate the role of these molecules in the host-pathogen interaction and fungal virulence.

Noonan syndrome: Severe phenotype and PTPN11 mutations.

PubMed

Carrasco Salas, Pilar; Gómez-Molina, Gertrudis; Carreto-Alba, Páxedes; Granell-Escobar, Reyes; Vázquez-Rico, Ignacio; León-Justel, Antonio

2018-04-24

Noonan syndrome (NS) is a genetic disorder characterized by a wide range of distinctive features and health problems. It caused in 50% of cases by missense mutations in PTPN11 gene. It has been postulated that it is possible to predict the disease course based into the impact of mutations on the protein. We report two cases of severe NS phenotype including hydrops fetalis. PTPN11 gene was studied in germinal cells of both patients by sequencing. Two different mutations (p.Gly503Arg and p.Met504Val) was detected in PTPN11 gene. These mutations have been reported previously, and when they were germinal variants, patients presented classic NS, NS with other malignancies and recently, p.Gly503Arg has been also observed in a patient with severe NS and hydrops fetalis, as our cases. Therefore, these observations shade light on that it is not always possibly to determine the genotype-phenotype relation based into the impact of mutations on the protein in NS patients with PTPN11 mutations. Copyright © 2018 Elsevier España, S.L.U. All rights reserved.

Phenotype, effector function, and tissue localization of PD-1-expressing human follicular helper T cell subsets

PubMed Central

2011-01-01

Background It is well established that PD-1 is expressed by follicular T cells but its function in regulation of human T helper cells has been unclear. We investigated the expression modality and function of PD-1 expressed by human T cells specialized in helping B cells. Results We found that PD-1-expressing T cells are heterogeneous in PD-1 expression. We identified three different PD-1-expressing memory T cell subsets (i.e. PD-1low (+), PD-1medium (++), and PD-1high (+++) cells). PD-1+++ T cells expressed CXCR5 and CXCR4 and were localized in the rim of germinal centers. PD-1+ or PD-1++ cells expressed CCR7 and were present mainly in the T cell area or other parts of the B cell follicles. Utilizing a novel antigen density-dependent magnetic sorting (ADD-MS) method, we isolated the three T cell subsets for functional characterization. The germinal center-located PD-1+++ T cells were most efficient in helping B cells and in producing IL-21 and CXCL13. Other PD-1-expressing T cells, enriched with Th1 and Th17 cells, were less efficient than PD-1+++ T cells in these capacities. PD-1+++ T cells highly expressed Ki-67 and therefore appear active in cell activation and proliferation in vivo. IL-2 is a cytokine important for proliferation and survival of the PD-1+++ T cells. In contrast, IL-21, while a major effector cytokine produced by the PD-1-expressing T helper cells, had no function in generation, survival, or proliferation of the PD-1-expressing helper T cells at least in vitro. PD-1 triggering has a suppressive effect on the proliferation and B cell-helping function of PD-1+++ germinal center T cells. Conclusion Our results revealed the phenotype and effector function of PD-1-expressing T helper cell subsets and indicate that PD-1 restrains the B cell-helping function of germinal center-localized T cells to prevent excessive antibody response. PMID:21914188

Glycerol enhances fungal germination at the water‐activity limit for life

PubMed Central

Stevenson, Andrew; Hamill, Philip G.; Medina, Ángel; Kminek, Gerhard; Rummel, John D.; Dijksterhuis, Jan; Timson, David J.; Magan, Naresh; Leong, Su‐Lin L.

2016-01-01

Summary For the most‐extreme fungal xerophiles, metabolic activity and cell division typically halts between 0.700 and 0.640 water activity (approximately 70.0–64.0% relative humidity). Here, we investigate whether glycerol can enhance xerophile germination under acute water‐activity regimes, using an experimental system which represents the biophysical limit of Earth's biosphere. Spores from a variety of species, including Aspergillus penicillioides, Eurotium halophilicum, Xerochrysium xerophilum (formerly Chrysosporium xerophilum) and Xeromyces bisporus, were produced by cultures growing on media supplemented with glycerol (and contained up to 189 mg glycerol g dry spores−1). The ability of these spores to germinate, and the kinetics of germination, were then determined on a range of media designed to recreate stresses experienced in microbial habitats or anthropogenic systems (with water‐activities from 0.765 to 0.575). For A. penicillioides, Eurotium amstelodami, E. halophilicum, X. xerophilum and X. bisporus, germination occurred at lower water‐activities than previously recorded (0.640, 0.685, 0.651, 0.664 and 0.637 respectively). In addition, the kinetics of germination at low water‐activities were substantially faster than those reported previously. Extrapolations indicated theoretical water‐activity minima below these values; as low as 0.570 for A. penicillioides and X. bisporus. Glycerol is present at high concentrations (up to molar levels) in many types of microbial habitat. We discuss the likely role of glycerol in expanding the water‐activity limit for microbial cell function in relation to temporal constraints and location of the microbial cell or habitat. The findings reported here have also critical implications for understanding the extremes of Earth's biosphere; for understanding the potency of disease‐causing microorganisms; and in biotechnologies that operate at the limits of microbial function. PMID:27631633

Spore Heat Activation Requirements and Germination Responses Correlate with Sequences of Germinant Receptors and with the Presence of a Specific spoVA2mob Operon in Foodborne Strains of Bacillus subtilis.

PubMed

Krawczyk, Antonina O; de Jong, Anne; Omony, Jimmy; Holsappel, Siger; Wells-Bennik, Marjon H J; Kuipers, Oscar P; Eijlander, Robyn T

2017-04-01

Spore heat resistance, germination, and outgrowth are problematic bacterial properties compromising food safety and quality. Large interstrain variation in these properties makes prediction and control of spore behavior challenging. High-level heat resistance and slow germination of spores of some natural Bacillus subtilis isolates, encountered in foods, have been attributed to the occurrence of the spoVA 2mob operon carried on the Tn 1546 transposon. In this study, we further investigate the correlation between the presence of this operon in high-level-heat-resistant spores and their germination efficiencies before and after exposure to various sublethal heat treatments (heat activation, or HA), which are known to significantly improve spore responses to nutrient germinants. We show that high-level-heat-resistant spores harboring spoVA 2mob required higher HA temperatures for efficient germination than spores lacking spoVA 2mob The optimal spore HA requirements additionally depended on the nutrients used to trigger germination, l-alanine (l-Ala), or a mixture of l-asparagine, d-glucose, d-fructose, and K + (AGFK). The distinct HA requirements of these two spore germination pathways are likely related to differences in properties of specific germinant receptors. Moreover, spores that germinated inefficiently in AGFK contained specific changes in sequences of the GerB and GerK germinant receptors, which are involved in this germination response. In contrast, no relation was found between transcription levels of main germination genes and spore germination phenotypes. The findings presented in this study have great implications for practices in the food industry, where heat treatments are commonly used to inactivate pathogenic and spoilage microbes, including bacterial spore formers. IMPORTANCE This study describes a strong variation in spore germination capacities and requirements for a heat activation treatment, i.e., an exposure to sublethal heat that increases spore responsiveness to nutrient germination triggers, among 17 strains of B. subtilis , including 9 isolates from spoiled food products. Spores of industrial foodborne isolates exhibited, on average, less efficient and slower germination responses and required more severe heat activation than spores from other sources. High heat activation requirements and inefficient, slow germination correlated with elevated resistance of spores to heat and with specific genetic features, indicating a common genetic basis of these three phenotypic traits. Clearly, interstrain variation and numerous factors that shape spore germination behavior challenge standardization of methods to recover highly heat-resistant spores from the environment and have an impact on the efficacy of preservation techniques used by the food industry to control spores. Copyright © 2017 American Society for Microbiology.

Spore Heat Activation Requirements and Germination Responses Correlate with Sequences of Germinant Receptors and with the Presence of a Specific spoVA2mob Operon in Foodborne Strains of Bacillus subtilis

PubMed Central

Krawczyk, Antonina O.; de Jong, Anne; Omony, Jimmy; Holsappel, Siger; Wells-Bennik, Marjon H. J.; Eijlander, Robyn T.

2017-01-01

ABSTRACT Spore heat resistance, germination, and outgrowth are problematic bacterial properties compromising food safety and quality. Large interstrain variation in these properties makes prediction and control of spore behavior challenging. High-level heat resistance and slow germination of spores of some natural Bacillus subtilis isolates, encountered in foods, have been attributed to the occurrence of the spoVA2mob operon carried on the Tn1546 transposon. In this study, we further investigate the correlation between the presence of this operon in high-level-heat-resistant spores and their germination efficiencies before and after exposure to various sublethal heat treatments (heat activation, or HA), which are known to significantly improve spore responses to nutrient germinants. We show that high-level-heat-resistant spores harboring spoVA2mob required higher HA temperatures for efficient germination than spores lacking spoVA2mob. The optimal spore HA requirements additionally depended on the nutrients used to trigger germination, l-alanine (l-Ala), or a mixture of l-asparagine, d-glucose, d-fructose, and K+ (AGFK). The distinct HA requirements of these two spore germination pathways are likely related to differences in properties of specific germinant receptors. Moreover, spores that germinated inefficiently in AGFK contained specific changes in sequences of the GerB and GerK germinant receptors, which are involved in this germination response. In contrast, no relation was found between transcription levels of main germination genes and spore germination phenotypes. The findings presented in this study have great implications for practices in the food industry, where heat treatments are commonly used to inactivate pathogenic and spoilage microbes, including bacterial spore formers. IMPORTANCE This study describes a strong variation in spore germination capacities and requirements for a heat activation treatment, i.e., an exposure to sublethal heat that increases spore responsiveness to nutrient germination triggers, among 17 strains of B. subtilis, including 9 isolates from spoiled food products. Spores of industrial foodborne isolates exhibited, on average, less efficient and slower germination responses and required more severe heat activation than spores from other sources. High heat activation requirements and inefficient, slow germination correlated with elevated resistance of spores to heat and with specific genetic features, indicating a common genetic basis of these three phenotypic traits. Clearly, interstrain variation and numerous factors that shape spore germination behavior challenge standardization of methods to recover highly heat-resistant spores from the environment and have an impact on the efficacy of preservation techniques used by the food industry to control spores. PMID:28130296

Glutathione synthesis is essential for pollen germination in vitro

PubMed Central

2011-01-01

Background The antioxidant glutathione fulfills many important roles during plant development, growth and defense in the sporophyte, however the role of this important molecule in the gametophyte generation is largely unclear. Bioinformatic data indicate that critical control enzymes are negligibly transcribed in pollen and sperm cells. Therefore, we decided to investigate the role of glutathione synthesis for pollen germination in vitro in Arabidopsis thaliana accession Col-0 and in the glutathione deficient mutant pad2-1 and link it with glutathione status on the subcellular level. Results The depletion of glutathione by buthionine sulfoximine (BSO), an inhibitor of glutathione synthesis, reduced pollen germination rates to 2-5% compared to 71% germination in wildtype controls. The application of reduced glutathione (GSH), together with BSO, restored pollen germination and glutathione contents to control values, demonstrating that inhibition of glutathione synthesis is responsible for the decrease of pollen germination in vitro. The addition of indole-3-acetic acid (IAA) to media containing BSO restored pollen germination to control values, which demonstrated that glutathione depletion in pollen grains triggered disturbances in auxin metabolism which led to inhibition of pollen germination. Conclusions This study demonstrates that glutathione synthesis is essential for pollen germination in vitro and that glutathione depletion and auxin metabolism are linked in pollen germination and early elongation of the pollen tube, as IAA addition rescues glutathione deficient pollen. PMID:21439079

The orphan germinant receptor protein GerXAO (but not GerX3b) is essential for L-alanine induced germination in Clostridium botulinum Group II.

PubMed

Brunt, Jason; Carter, Andrew T; Pye, Hannah V; Peck, Michael W

2018-05-04

Clostridium botulinum is an anaerobic spore forming bacterium that produces the potent botulinum neurotoxin that causes a severe and fatal neuro-paralytic disease of humans and animals (botulism). C. botulinum Group II is a psychrotrophic saccharolytic bacterium that forms spores of moderate heat resistance and is a particular hazard in minimally heated chilled foods. Spore germination is a fundamental process that allows the spore to transition to a vegetative cell and typically involves a germinant receptor (GR) that responds to environmental signals. Analysis of C. botulinum Group II genomes shows they contain a single GR cluster (gerX3b), and an additional single gerA subunit (gerXAO). Spores of C. botulinum Group II strain Eklund 17B germinated in response to the addition of L-alanine, but did not germinate following the addition of exogenous Ca 2+ -DPA. Insertional inactivation experiments in this strain unexpectedly revealed that the orphan GR GerXAO is essential for L-alanine stimulated germination. GerX3bA and GerX3bC affected the germination rate but were unable to induce germination in the absence of GerXAO. No role could be identified for GerX3bB. This is the first study to identify the functional germination receptor of C. botulinum Group II.

DELAY OF GERMINATION 1 mediates a conserved coat-dormancy mechanism for the temperature- and gibberellin-dependent control of seed germination.

PubMed

Graeber, Kai; Linkies, Ada; Steinbrecher, Tina; Mummenhoff, Klaus; Tarkowská, Danuše; Turečková, Veronika; Ignatz, Michael; Sperber, Katja; Voegele, Antje; de Jong, Hans; Urbanová, Terezie; Strnad, Miroslav; Leubner-Metzger, Gerhard

2014-08-26

Seed germination is an important life-cycle transition because it determines subsequent plant survival and reproductive success. To detect optimal spatiotemporal conditions for germination, seeds act as sophisticated environmental sensors integrating information such as ambient temperature. Here we show that the delay of germination 1 (DOG1) gene, known for providing dormancy adaptation to distinct environments, determines the optimal temperature for seed germination. By reciprocal gene-swapping experiments between Brassicaceae species we show that the DOG1-mediated dormancy mechanism is conserved. Biomechanical analyses show that this mechanism regulates the material properties of the endosperm, a seed tissue layer acting as germination barrier to control coat dormancy. We found that DOG1 inhibits the expression of gibberellin (GA)-regulated genes encoding cell-wall remodeling proteins in a temperature-dependent manner. Furthermore we demonstrate that DOG1 causes temperature-dependent alterations in the seed GA metabolism. These alterations in hormone metabolism are brought about by the temperature-dependent differential expression of genes encoding key enzymes of the GA biosynthetic pathway. These effects of DOG1 lead to a temperature-dependent control of endosperm weakening and determine the optimal temperature for germination. The conserved DOG1-mediated coat-dormancy mechanism provides a highly adaptable temperature-sensing mechanism to control the timing of germination.

The evolution of pollen germination timing in flowering plants: Austrobaileya scandens (Austrobaileyaceae)

PubMed Central

Williams, Joseph H.

2012-01-01

Background and aims The pollination to fertilization process (progamic phase) is thought to have become greatly abbreviated with the origin of flowering plants. In order to understand what developmental mechanisms enabled the speeding of fertilization, comparative data are needed from across the group, especially from early-divergent lineages. I studied the pollen germination process of Austrobaileya scandens, a perennial vine endemic to the Wet Tropics area of northeastern Queensland, Australia, and a member of the ancient angiosperm lineage, Austrobaileyales. Methodology I used in vivo and in vitro hand pollinations and timed collections to study development from late pollen maturation to just after germination. Then I compared the contribution of pollen germination timing to progamic phase duration in 131 angiosperm species (65 families). Principal findings Mature pollen of Austrobaileya was bicellular, starchless and moderately dehydrated—water content was 31.5 % by weight and volume increased by 57.9 % upon hydration. A callose layer in the inner intine appeared only after pollination. In vivo pollen germination followed a logarithmic curve, rising from 28 % at 1 hour after pollination (hap) to 97 % at 12 hap (R2 = 0.98). Sufficient pollen germination to fertilize all ovules was predicted to have occurred within 62 min. Across angiosperms, pollen germination ranged from 1 min to >60 h long and required 8.3 ± 9.8 % of the total duration of the progamic phase. Significance Pollen of Austrobaileya has many plesiomorphic features that are thought to prolong germination. Yet its germination is quite fast for species with desiccation-tolerant pollen (range: <1 to 60 h). Austrobaileya and other early-divergent angiosperms have relatively rapid pollen germination and short progamic phases, comparable to those of many insect-pollinated monocots and eudicots. These results suggest that both the pollen germination and pollen tube growth periods were marked by acceleration of developmental processes early in angiosperm history. PMID:22567221

The evolution of pollen germination timing in flowering plants: Austrobaileya scandens (Austrobaileyaceae).

PubMed

Williams, Joseph H

2012-01-01

The pollination to fertilization process (progamic phase) is thought to have become greatly abbreviated with the origin of flowering plants. In order to understand what developmental mechanisms enabled the speeding of fertilization, comparative data are needed from across the group, especially from early-divergent lineages. I studied the pollen germination process of Austrobaileya scandens, a perennial vine endemic to the Wet Tropics area of northeastern Queensland, Australia, and a member of the ancient angiosperm lineage, Austrobaileyales. I used in vivo and in vitro hand pollinations and timed collections to study development from late pollen maturation to just after germination. Then I compared the contribution of pollen germination timing to progamic phase duration in 131 angiosperm species (65 families). Mature pollen of Austrobaileya was bicellular, starchless and moderately dehydrated-water content was 31.5 % by weight and volume increased by 57.9 % upon hydration. A callose layer in the inner intine appeared only after pollination. In vivo pollen germination followed a logarithmic curve, rising from 28 % at 1 hour after pollination (hap) to 97 % at 12 hap (R(2) = 0.98). Sufficient pollen germination to fertilize all ovules was predicted to have occurred within 62 min. Across angiosperms, pollen germination ranged from 1 min to >60 h long and required 8.3 ± 9.8 % of the total duration of the progamic phase. Pollen of Austrobaileya has many plesiomorphic features that are thought to prolong germination. Yet its germination is quite fast for species with desiccation-tolerant pollen (range: <1 to 60 h). Austrobaileya and other early-divergent angiosperms have relatively rapid pollen germination and short progamic phases, comparable to those of many insect-pollinated monocots and eudicots. These results suggest that both the pollen germination and pollen tube growth periods were marked by acceleration of developmental processes early in angiosperm history.

An immunocytochemical study of the germinal layer vasculature in the developing fetal brain using Ulex europaeus 1 lectin.

PubMed

Gould, S J; Howard, S

1988-10-01

The characteristics of the germinal matrix vasculature were studied in the developing fetal brain using immunocytochemical methods. A preliminary comparative immunocytochemical study was made on six fetal brains to compare endothelial staining by Ulex europaeus I lectin with that of antibody to Factor VIII related antigen. Ulex was found to stain germinal layer vessels better than Factor VIII related antigen. Subsequently, the germinal layers of a further 15 fetal and preterm infant brains ranging from 13 to 35 weeks' gestation were stained with Ulex europaeus I to demonstrate the vasculature. With increasing gestation, there was a gradual increase in vessel density, particularly of capillaries. This was not a uniform process. A plexus of capillaries was prominent immediately beneath the ependyma while the more central parts of the germinal matrix contained fewer, but often larger diameter, vessels. The variation in vessel density which was a feature of the later gestation brains may have implications for local blood flow and may be a factor in haemorrhage at this site.

Involvement of Alternative Splicing in Barley Seed Germination

PubMed Central

Zhang, Qisen; Zhang, Xiaoqi; Wang, Songbo; Tan, Cong; Zhou, Gaofeng; Li, Chengdao

2016-01-01

Seed germination activates many new biological processes including DNA, membrane and mitochondrial repairs and requires active protein synthesis and sufficient energy supply. Alternative splicing (AS) regulates many cellular processes including cell differentiation and environmental adaptations. However, limited information is available on the regulation of seed germination at post-transcriptional levels. We have conducted RNA-sequencing experiments to dissect AS events in barley seed germination. We identified between 552 and 669 common AS transcripts in germinating barley embryos from four barley varieties (Hordeum vulgare L. Bass, Baudin, Harrington and Stirling). Alternative 3’ splicing (34%-45%), intron retention (32%-34%) and alternative 5’ splicing (16%-21%) were three major AS events in germinating embryos. The AS transcripts were predominantly mapped onto ribosome, RNA transport machineries, spliceosome, plant hormone signal transduction, glycolysis, sugar and carbon metabolism pathways. Transcripts of these genes were also very abundant in the early stage of seed germination. Correlation analysis of gene expression showed that AS hormone responsive transcripts could also be co-expressed with genes responsible for protein biosynthesis and sugar metabolisms. Our RNA-sequencing data revealed that AS could play important roles in barley seed germination. PMID:27031341

Inhibitors of protein phosphorylation including the retinoblastoma protein induce germination of Candida albicans.

PubMed

Cho, T; Hamatake, H; Hagihara, Y; Kaminishi, H

2000-02-01

It has been previously shown that the induction of germination in Candida albicans occurs following its cessation of growth as a yeast. Similarly, mammalian cells undergo a differentiation process that is preceded by a growth cessation associated with a hypophosphorylation of proteins of the retinoblastoma gene family. It is postulated that a similar type of mechanism may be operative in C. albicans and protein phosphorylation inhibitors: forskolin (stimulates cyclic adenosine monophosphate production), okadaic acid (phosphatase inhibitor) and D-erythro-sphingosine (retinoblastoma protein phosphorylation inhibitor) have been used to further strengthen this hypothesis. Okadaic acid (1-1000 nM) and D-erythro-sphingosine (100 microM) significantly inhibited the growth of yeast cells of C. albicans. D-Erythro-sphingosine at 1000 microM was candidicidal. Forskolin did not significantly affect growth. Exponentially grown C. albicans pretreated with forskolin (10 microM), okadaic acid (1000 nM) or D-erythro-sphingosine (100 microM) readily germinated. In comparison, when these inhibitors were incorporated in the same medium, germination of exponentially grown cells did not occur. These results suggest that protein dephosphorylation may be necessary at an early stage of the yeast-hyphae transition in C. albicans.

Bicarbonate and amino acids are co-germinants for spores of Clostridium perfringens type A isolates carrying plasmid-borne enterotoxin gene.

PubMed

Alnoman, Maryam; Udompijitkul, Pathima; Banawas, Saeed; Sarker, Mahfuzur R

2018-02-01

Clostridium perfringens type A isolates carrying a chromosomal enterotoxin (cpe) gene (C-cpe) are generally linked to food poisoning, while isolates carrying cpe on a plasmid (P-cpe) are associated with non-food-borne gastrointestinal diseases. Both C-cpe and P-cpe isolates can form metabolically dormant spores, which through germination process return to actively growing cells to cause diseases. In our previous study, we showed that only 3 out of 20 amino acids (aa) in phosphate buffer (pH 7.0) triggered germination of spores of P-cpe isolates (P-cpe spores). We now found that 14 out of 20 individual aa tested induced germination of P-cpe spores in the presence of bicarbonate buffer (pH 7.0). However, no significant spore germination was observed with bicarbonate (pH 7.0) alone, indicating that aa and bicarbonate are co-germinants for P-cpe spores. P-cpe strain F4969 gerKC spores did not germinate, and gerAA spores germinated extremely poorly as compared to wild-type and gerKA spores with aa-bicarbonate (pH 7.0) co-germinants. The germination defects in gerKC and gerAA spores were partially restored by complementing gerKC or gerAA spores with wild-type gerKC or gerAA, respectively. Collectively, this study identified aa-bicarbonate as a novel nutrient germinant for P-cpe spores and provided evidence that GerKC and GerAA play major roles in aa-bicarbonate induced germination. Copyright © 2017 Elsevier Ltd. All rights reserved.

Lymph node toxoplasmosis. Follow-up of 237 histologically diagnosed and serologically verified cases.

PubMed

Miettinen, M; Saxén, L; Saxén, E

1980-01-01

The clinical features, histology and follow-up of lymph node toxoplasmosis are presented in the light of 237 histologically and serologically verified cases. Lymph node toxoplasmosis is a disease with mild symptoms, and in most patients the enlarged lymph nodes were the only sign. Three fourths of the patients were women and the majority were under 40 years of age. The clinical picture was not specific, but suggestive features included a relatively short history, presence of the nodes in the neck and relative lymphocytosis in peripheral blood. Histological changes in the lymph nodes were characteristic. The most important features were strong hyperplasia but preserved general structure with small groups of epithelioid cells both in the paracortical area and in the germinal centers. Strands of monocytoid cells were usually found. 80% of the cases with typical histology also had high antibody titers, and in more than 85% of the cases with high antibodies, the lymph nodes presented a typical picture of toxoplasmosis. The follow-up revealed that lymph node toxoplasmosis. The follow-up revealed that lymph node toxoplasmosis is a disease without complications, nor is there any connection with malignant lymphomas.

Epstein-Barr virus-infected B cells expanding in germinal centers of infectious mononucleosis patients do not participate in the germinal center reaction.

PubMed

Kurth, Julia; Hansmann, Martin-Leo; Rajewsky, Klaus; Küppers, Ralf

2003-04-15

To assess the impact of the germinal center (GC) reaction on viral spread in Epstein-Barr virus (EBV) infection, we isolated EBV(+) GC B cells from the tonsils of two infectious mononucleosis patients, sequenced their rearranged V genes, and determined expression of the EBV latency genes EBV nuclear antigen 2 and latent membrane protein 1. Most EBV(+) GC B cells belonged to clones of cells harboring somatically mutated V gene rearrangements. Ongoing somatic hypermutation, the hallmark of the GC reaction, was seen only in uninfected GC B cell clones, not in EBV(+) B cell clones. Thus, in infectious mononucleosis, GC and/or memory B cells are directly infected by EBV and expand without somatic hypermutation, whereas the GC passage of EBV-infected naive B cells does not contribute detectably to the generation of infected memory B cells, the main reservoir of EBV during persistence. Most, if not all, EBV-infected cells in GCs exhibited an unusual EBV gene expression pattern in that they were positive for EBV nuclear antigen 2 but negative for latent membrane protein 1. Although the three main types of EBV-associated B cell lymphomas (Burkitt's, Hodgkin's, and posttransplant lymphomas) presumably are derived from GC B cells, EBV(+) GC B cells resembling these EBV(+) GC B cell lymphomas in terms of EBV gene expression and somatic hypermutation pattern could not be identified.

Gravity and light control of the developmental polarity of regenerating protoplasts isolated from prothallial cells of the fern Ceratopteris richardii

NASA Technical Reports Server (NTRS)

Edwards, E. S.; Roux, S. J.

1998-01-01

A procedure has been developed for isolating protoplasts from prothalli of Ceratopteris richardii which can be cultured and are capable of regeneration. Protoplasts were isolated from 2-week-old gametophytes in a medium containing wall-digesting enzymes in 0.5 M sucrose, followed by purification of the released protoplasts by floating them up into a 0.5 M sorbitol layer. Regeneration occurred over a period of 10-24 days, and, under optimal osmotic conditions, followed the developmental pattern seen during spore germination, in that the first division gave rise to a primary rhizoid. Thus, prothallial protoplasts are comparable to germinating spores as suitable models for studies of developmental polarity in single cells. As in germinating spores, the polarity of development in regenerating protoplasts is influenced by the vectors of gravity and unilateral light. However, the relative influence of light in fixing this polarity is greater in regenerating protoplasts, while in germinating spores, the influence of gravity is greater.

Multiple Phosphatases Regulate Carbon Source-Dependent Germination and Primary Metabolism in Aspergillus nidulans

PubMed Central

de Assis, Leandro José; Ries, Laure Nicolas Annick; Savoldi, Marcela; Dinamarco, Taisa Magnani; Goldman, Gustavo Henrique; Brown, Neil Andrew

2015-01-01

Aspergillus nidulans is an important mold and a model system for the study of fungal cell biology. In addition, invasive A. nidulans pulmonary infections are common in humans with chronic granulomatous disease. The morphological and biochemical transition from dormant conidia into active, growing, filamentous hyphae requires the coordination of numerous biosynthetic, developmental, and metabolic processes. The present study exhibited the diversity of roles performed by seven phosphatases in regulating cell cycle, development, and metabolism in response to glucose and alternative carbon sources. The identified phosphatases highlighted the importance of several signaling pathways regulating filamentous growth, the action of the pyruvate dehydrogenase complex as a metabolic switch controlling carbon usage, and the identification of the key function performed by the α-ketoglutarate dehydrogenase during germination. These novel insights into the fundamental roles of numerous phosphatases in germination and carbon sensing have provided new avenues of research into the identification of inhibitors of fungal germination, with implications for the food, feed, and pharmaceutical industries. PMID:25762568

Nitric oxide is involved in light-specific responses of tomato during germination under normal and osmotic stress conditions

PubMed Central

Piterková, Jana; Luhová, Lenka; Hofman, Jakub; Turečková, Veronika; Novák, Ondřej; Petřivalský, Marek; Fellner, Martin

2012-01-01

Background and Aims Nitric oxide (NO) is involved in the signalling and regulation of plant growth and development and responses to biotic and abiotic stresses. The photoperiod-sensitive mutant 7B-1 in tomato (Solanum lycopersicum) showing abscisic acid (ABA) overproduction and blue light (BL)-specific tolerance to osmotic stress represents a valuable model to study the interaction between light, hormones and stress signalling. The role of NO as a regulator of seed germination and ABA-dependent responses to osmotic stress was explored in wild-type and 7B-1 tomato under white light (WL) and BL. Methods Germination data were obtained from the incubation of seeds on germinating media of different composition. Histochemical analysis of NO production in germinating seeds was performed by fluorescence microscopy using a cell-permeable NO probe, and endogenous ABA was analysed by mass spectrometry. Key Results The NO donor S-nitrosoglutathione stimulated seed germination, whereas the NO scavenger 2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (PTIO) had an inhibitory effect. Under WL in both genotypes, PTIO strongly suppressed germination stimulated by fluridone, an ABA inhibitor. The stimulatory effect of the NO donor was also observed under osmotic stress for 7B-1 seeds under WL and BL. Seed germination inhibited by osmotic stress was restored by fluridone under WL, but less so under BL, in both genotypes. This effect of fluridone was further modulated by the NO donor and NO scavenger, but only to a minor extent. Fluorescence microscopy using the cell-permeable NO probe DAF-FM DA (4-amino-5-methylamino-2′,7′-difluorofluorescein diacetate) revealed a higher level of NO in stressed 7B-1 compared with wild-type seeds. Conclusions As well as defective BL signalling, the differential NO-dependent responses of the 7B-1 mutant are probably associated with its high endogenous ABA concentration and related impact on hormonal cross-talk in germinating seeds. These data confirm that light-controlled seed germination and stress responses include NO-dependent signalling. PMID:22782244

Nitric oxide is involved in light-specific responses of tomato during germination under normal and osmotic stress conditions.

PubMed

Piterková, Jana; Luhová, Lenka; Hofman, Jakub; Turecková, Veronika; Novák, Ondrej; Petrivalsky, Marek; Fellner, Martin

2012-09-01

Nitric oxide (NO) is involved in the signalling and regulation of plant growth and development and responses to biotic and abiotic stresses. The photoperiod-sensitive mutant 7B-1 in tomato (Solanum lycopersicum) showing abscisic acid (ABA) overproduction and blue light (BL)-specific tolerance to osmotic stress represents a valuable model to study the interaction between light, hormones and stress signalling. The role of NO as a regulator of seed germination and ABA-dependent responses to osmotic stress was explored in wild-type and 7B-1 tomato under white light (WL) and BL. Germination data were obtained from the incubation of seeds on germinating media of different composition. Histochemical analysis of NO production in germinating seeds was performed by fluorescence microscopy using a cell-permeable NO probe, and endogenous ABA was analysed by mass spectrometry. The NO donor S-nitrosoglutathione stimulated seed germination, whereas the NO scavenger 2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (PTIO) had an inhibitory effect. Under WL in both genotypes, PTIO strongly suppressed germination stimulated by fluridone, an ABA inhibitor. The stimulatory effect of the NO donor was also observed under osmotic stress for 7B-1 seeds under WL and BL. Seed germination inhibited by osmotic stress was restored by fluridone under WL, but less so under BL, in both genotypes. This effect of fluridone was further modulated by the NO donor and NO scavenger, but only to a minor extent. Fluorescence microscopy using the cell-permeable NO probe DAF-FM DA (4-amino-5-methylamino-2',7'-difluorofluorescein diacetate) revealed a higher level of NO in stressed 7B-1 compared with wild-type seeds. As well as defective BL signalling, the differential NO-dependent responses of the 7B-1 mutant are probably associated with its high endogenous ABA concentration and related impact on hormonal cross-talk in germinating seeds. These data confirm that light-controlled seed germination and stress responses include NO-dependent signalling.

Inhibitory effect of indole analogs against Paenibacillus larvae, the causal agent of American foulbrood disease.

PubMed

Alvarado, Israel; Margotta, Joseph W; Aoki, Mai M; Flores, Fernando; Agudelo, Fresia; Michel, Guillermo; Elekonich, Michelle M; Abel-Santos, Ernesto

2017-09-01

Paenibacillus larvae, a Gram-positive bacterium, causes American foulbrood (AFB) in honey bee larvae (Apis mellifera Linnaeus [Hymenoptera: Apidae]). P. larvae spores exit dormancy in the gut of bee larvae, the germinated cells proliferate, and ultimately bacteremia kills the host. Hence, spore germination is a required step for establishing AFB disease. We previously found that P. larvae spores germinate in response to l-tyrosine plus uric acid in vitro. Additionally, we determined that indole and phenol blocked spore germination. In this work, we evaluated the antagonistic effect of 35 indole and phenol analogs and identified strong inhibitors of P. larvae spore germination in vitro. We further tested the most promising candidate, 5-chloroindole, and found that it significantly reduced bacterial proliferation. Finally, feeding artificial worker jelly containing anti-germination compounds to AFB-exposed larvae significantly decreased AFB infection in laboratory-reared honey bee larvae. Together, these results suggest that inhibitors of P. larvae spore germination could provide another method to control AFB. © The Authors 2017. Published by Oxford University Press on behalf of Entomological Society of America.

Analysis of α-glucosidase enzyme activity used in a rapid test for steam sterilization assurance.

PubMed

Setlow, B; Korza, G; Setlow, P

2016-05-01

This study was to determine the sources, location and identity of α-glucosidases in dormant/germinating/outgrowing spores and growing cells of Geobacillus stearothermophilus ATCC 7953, an enzymatic activity in spores used in rapid tests of steam sterilization. α-Glucosidase activity in spores and cells was determined measuring methylumbelliferyl-α-d-glucoside (α-MUG) or α-MUG-6-phosphate hydrolysis fluorometrically. While α-MUG-6-phosphate was not hydrolysed by cell or spore extracts, assays with α-MUG showed that: (1) the α-glucosidase activity was inside and outside spores, and the activity outside spores was largely removed by buffer washes or heat activation, whereas α-glucosidase activity was only inside vegetative cells; (2) most α-glucosidase activity in cells and spores was soluble; (3) Western blots and enzyme inhibition using an anti-α-glucosidase antiserum identified ≥2 α-glucosidases in spores and growing cells; (4) α-glucosidase-specific activities were similar in dormant, germinated and outgrowing spore and growing cell extracts; and (5) significant α-glucosidase was synthesized during spore germination and outgrowth and cell growth, this synthesis was not repressed by glucose nor induced by α-MUG, but glucose inhibited α-MUG uptake. α-MUG hydrolysis by G. stearothermophilus is by α-MUG uptake and hydrolysis by ≥2 α-glucosidases associated with dormant spores and synthesized by germinating and outgrowing spores. The enzyme activity observed by sterilization assurance assays appears likely to come from heat-stable enzyme in the spore core and enzyme(s) synthesized in spore outgrowth. The results of this work provide new insight into the science behind a rapid test for steam sterilization assurance. © 2016 The Society for Applied Microbiology.

Regulation of germinal center responses and B-cell memory by the chromatin modifier MOZ.

PubMed

Good-Jacobson, Kim L; Chen, Yunshun; Voss, Anne K; Smyth, Gordon K; Thomas, Tim; Tarlinton, David

2014-07-01

Memory B cells and long-lived bone marrow-resident plasma cells maintain humoral immunity. Little is known about the intrinsic mechanisms that are essential for forming memory B cells or endowing them with the ability to rapidly differentiate upon reexposure while maintaining the population over time. Histone modifications have been shown to regulate lymphocyte development, but their role in regulating differentiation and maintenance of B-cell subsets during an immune response is unclear. Using stage-specific deletion of monocytic leukemia zinc finger protein (MOZ), a histone acetyltransferase, we demonstrate that mutation of this chromatin modifier alters fate decisions in both primary and secondary responses. In the absence of MOZ, germinal center B cells were significantly impaired in their ability to generate dark zone centroblasts, with a concomitant decrease in both cell-cycle progression and BCL-6 expression. In contrast, there was increased differentiation to IgM and low-affinity IgG1(+) memory B cells. The lack of MOZ affected the functional outcome of humoral immune responses, with an increase in secondary germinal centers and a corresponding decrease in secondary high-affinity antibody-secreting cell formation. Therefore, these data provide strong evidence that manipulating epigenetic modifiers can regulate fate decisions during humoral responses, and thus could be targeted for therapeutic intervention.

Germinal mosaicism in Noonan syndrome: A family with two affected siblings of normal parents.

PubMed

Elalaoui, Siham Chafai; Kraoua, Lilia; Liger, Céline; Ratbi, Ilham; Cavé, Hélène; Sefiani, Abdelaziz

2010-11-01

Noonan syndrome (NS; OMIM 163950) is an autosomal dominant disorder with variable clinical expression and genetic heterogeneity. Clinical manifestations include characteristic facial features, short stature, and cardiac anomalies. Mutations in protein-tyrosine phosphatase, non-receptor-type 11 (PTPN11), encoding SHP-2, account for about half of NS patients. We report on a Moroccan family with two children with NS and apparently unaffected parents. The molecular studies showed the heterozygous mutation c.922A>G of PTPN11 gene in the two affected sibs. Neither the parents, nor the oldest brother carries this mutation in hematologic cells. The mutation was also absent in buccal epithelial cells and fingernails of both parents. We believe this is the first report of germ cell mosaicism in NS and suggest an empirical risk for recurrence of that is less than 1%. © 2010 Wiley-Liss, Inc.

Echinococcus granulosus equinus: variation in the germinal layer of murine hydatids and evidence of autophagy.

PubMed

Richards, K S; Arme, C; Bridges, J F

1984-08-01

The germinal layer of sterile 9-month-old murine peritoneal cysts of Echinococcus granulosus equinus shows interrelated variation in depth, tissue integrity, metabolic reserves and the number of autophagic lamellar bodies present. These features are similar in large and medium-sized cysts from the same host, whether occurring singly or within cyst masses. Deep germinal layers (greater than 16 micron) are lipid- and glycogen-rich and possess numerous autophagic vacuoles with 6 nm period lamellar stacks asymmetrically disposed peripherally; shallow layers (less than 12 micron), with indications of degeneration, have depleted metabolic reserves and fewer lamellar bodies. These bodies are formed by smooth endoplasmic reticulum encirclement of small glycogen masses followed by further sequestration, and eventually definition of glycogen particles may be lost. Autophagy of mitochondria and cytoplasmic vesicles also occurs. The presence of lysosomal enzymes within the layer suggests autolysosomal compartmentalization of excess substrate and effete material. Mucopolysaccharide bodies, containing material similar to that exocytosed to form the laminated layer matrix, occur and are formed from fusion and autophagy of Golgi-derived vesicles. These bodies may also develop peripheral 6 nm period lamellar stacks, but of limited depth. Mucopolysaccharide bodies are the dominant feature of the germinal layer of very small cyst-mass cysts in which laminated layer production is considered to be arrested. They thus represent a repository for the unreleased mucopolysaccharide material.

The use of cluster analysis for plant grouping by their tolerance to soil contamination with hydrocarbons at the germination stage.

PubMed

Potashev, Konstantin; Sharonova, Natalia; Breus, Irina

2014-07-01

Clustering was employed for the analysis of obtained experimental data set (42 plants in total) on seed germination in leached chernozem contaminated with kerosene. Among investigated plants were 31 cultivated plants from 11 families (27 species and 20 varieties) and 11 wild plant species from 7 families, 23 annual and 19 perennial/biannual plant species, 11 monocotyledonous and 31 dicotyledonous plants. Two-dimensional (two-parameter) clustering approach, allowing the estimation of tolerance of germinating seeds using a pair of independent parameters (С75%, V7%) was found to be most effective. These parameters characterized the ability of seeds to both withstand high concentrations of contaminants without the significant reduction of the germination, and maintain high germination rate within certain contaminant concentrations. The performed clustering revealed a number of plant features, which define the relation of a particular plant to a particular tolerance cluster; it has also demonstrated the possibility of generalizing the kerosene results for n-tridecane, which is one of the typical kerosene components. In contrast to the "manual" plant ranking based on the assessment of germination at discrete concentrations of the contaminant, the proposed clustering approach allowed a generalized characterization of the seed tolerance/sensitivity to hydrocarbon contaminants. Copyright © 2014 Elsevier B.V. All rights reserved.

Spred1, a negative regulator of Ras–MAPK–ERK, is enriched in CNS germinal zones, dampens NSC proliferation, and maintains ventricular zone structure

PubMed Central

Phoenix, Timothy N.; Temple, Sally

2010-01-01

Neural stem cells (NSCs) have great potential for self-renewal, which must be tightly regulated to generate appropriate cell numbers during development and to prevent tumor formation. The Ras–MAPK–ERK pathway affects mitogen-stimulated proliferation, and negative regulators are likely to be important for keeping self-renewal in check. Sprouty-related protein with an EVH1 domain (Spred1) is a recently discovered negative Ras–MAPK–ERK regulator linked to a neurofibromatosis 1 (NF-1)-like human syndrome; however, its role in CNS development has not been explored. We show that Spred1 is highly enriched in CNS germinal zones during neurogenesis. Spred1 knockdown increases NSC self-renewal and progenitor proliferation cell-autonomously, and overexpression causes premature differentiation. Surprisingly, Spred1 knockdown in vivo in the embryonic mouse forebrain frequently resulted in periventricular heterotopia, developmental abnormalities often associated with mutations in genes in the vesicular trafficking pathway that cause disruption of germinal zones and impair cell migration. In cortical progenitor cells, Spred1 localizes within distinct vesicles, indicating a potential role in transport. Spred1 knockdown gradually leads to disruption of the apical ventricular zone and loss of radial glia alignment. This impairs late neuronal migration, resulting in the formation of periventricular masses. Thus, Spred1 is critical for normal cortical development, as it modulates progenitor self-renewal/proliferation and helps maintain the integrity and organization of germinal zones. PMID:20047999

Coffee cysteine proteinases and related inhibitors with high expression during grain maturation and germination

PubMed Central

2012-01-01

Background Cysteine proteinases perform multiple functions in seeds, including participation in remodelling polypeptides and recycling amino acids during maturation and germination. Currently, few details exist concerning these genes and proteins in coffee. Furthermore, there is limited information on the cysteine proteinase inhibitors which influence the activities of these proteinases. Results Two cysteine proteinase (CP) and four cysteine proteinase inhibitor (CPI) gene sequences have been identified in coffee with significant expression during the maturation and germination of coffee grain. Detailed expression analysis of the cysteine proteinase genes CcCP1 and CcCP4 in Robusta using quantitative RT-PCR showed that these transcripts accumulate primarily during grain maturation and germination/post germination. The corresponding proteins were expressed in E. coli and purified, but only one, CcCP4, which has a KDDL/KDEL C-terminal sequence, was found to be active after a short acid treatment. QRT-PCR expression analysis of the four cysteine proteinase inhibitor genes in Robusta showed that CcCPI-1 is primarily expressed in developing and germinating grain and CcCPI-4 is very highly expressed during the late post germination period, as well as in mature, but not immature leaves. Transcripts corresponding to CcCPI-2 and CcCPI-3 were detected in most tissues examined at relatively similar, but generally low levels. Conclusions Several cysteine proteinase and cysteine proteinase inhibitor genes with strong, relatively specific expression during coffee grain maturation and germination are presented. The temporal expression of the CcCP1 gene suggests it is involved in modifying proteins during late grain maturation and germination. The expression pattern of CcCP4, and its close identity with KDEL containing CP proteins, implies this proteinase may play a role in protein and/or cell remodelling during late grain germination, and that it is likely to play a strong role in the programmed cell death associated with post-germination of the coffee grain. Expression analysis of the cysteine proteinase inhibitor genes suggests that CcCPI-1 could primarily be involved in modulating the activity of grain CP activity; while CcCPI-4 may play roles modulating grain CP activity and in the protection of the young coffee seedlings from insects and pathogens. CcCPI-2 and CcCPI-3, having lower and more widespread expression, could be more general "house-keeping" CPI genes. PMID:22380654

Identification of the seasonal conditions required for dormancy break of Persoonia longifolia (Proteaceae), a species with a woody indehiscent endocarp.

PubMed

Chia, K A; Sadler, R; Turner, S R; Baskin, C C

2016-08-01

The mechanisms involved in breaking seed dormancy in species with woody endocarps are poorly understood. In a landmark study examining the role of endocarps in regulating germination, our aim was to investigate the effects of the natural sequence of environmental conditions on dormancy break of a species with a woody endocarp (Persoonia longifolia). The role of the endocarp in germination was investigated through imbibition and endocarp removal germination tests. The use of burial to break dormancy was examined and results from these experiments were used to guide laboratory investigations into the use of wet/dry cycling and stratification to break dormancy. Endocarps were water-permeable. Germination increased from 0 to 92·5 % when endocarps were removed. During burial in the field and nursery, 41·6 and 63·7 % of the endocarps germinated, respectively, after 36 months. Ex situ post-burial germination was cyclical and highest after 30 months of burial (45·4 % nursery and 31·8 % field). Highest germination occurred in wet/dry trials when the dry summer was long (20 weeks), had fluctuating temperatures (30/50 °C) and two long (7 d) wet cycles and was followed by moist winters at 10/20 °C. A stratification trial found that highest germination occurred following incubation for 12 weeks at 30 °C (including 2 weeks moist) + 6 weeks moist at 8 °C then placement at 20/10 °C for germination. Summer conditions break physiological dormancy of the embryo and promote opening of the endocarp, allowing seeds to germinate during winter conditions. By closely monitoring the environment that endocarps are exposed to in nature, dormancy breaking mechanisms can be identified and used to improve germination. These results outline for the first time how dormancy and germination are regulated in a species with a hard woody endocarp, insights which will significantly improve our understanding of other species with similar reproductive features. © The Author 2016. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

DNA damage checkpoint kinase ATM regulates germination and maintains genome stability in seeds

PubMed Central

Waterworth, Wanda M.; Footitt, Steven; Bray, Clifford M.; Finch-Savage, William E.; West, Christopher E.

2016-01-01

Genome integrity is crucial for cellular survival and the faithful transmission of genetic information. The eukaryotic cellular response to DNA damage is orchestrated by the DNA damage checkpoint kinases ATAXIA TELANGIECTASIA MUTATED (ATM) and ATM AND RAD3-RELATED (ATR). Here we identify important physiological roles for these sensor kinases in control of seed germination. We demonstrate that double-strand breaks (DSBs) are rate-limiting for germination. We identify that desiccation tolerant seeds exhibit a striking transcriptional DSB damage response during germination, indicative of high levels of genotoxic stress, which is induced following maturation drying and quiescence. Mutant atr and atm seeds are highly resistant to aging, establishing ATM and ATR as determinants of seed viability. In response to aging, ATM delays germination, whereas atm mutant seeds germinate with extensive chromosomal abnormalities. This identifies ATM as a major factor that controls germination in aged seeds, integrating progression through germination with surveillance of genome integrity. Mechanistically, ATM functions through control of DNA replication in imbibing seeds. ATM signaling is mediated by transcriptional control of the cell cycle inhibitor SIAMESE-RELATED 5, an essential factor required for the aging-induced delay to germination. In the soil seed bank, seeds exhibit increased transcript levels of ATM and ATR, with changes in dormancy and germination potential modulated by environmental signals, including temperature and soil moisture. Collectively, our findings reveal physiological functions for these sensor kinases in linking genome integrity to germination, thereby influencing seed quality, crucial for plant survival in the natural environment and sustainable crop production. PMID:27503884

DNA damage checkpoint kinase ATM regulates germination and maintains genome stability in seeds.

PubMed

Waterworth, Wanda M; Footitt, Steven; Bray, Clifford M; Finch-Savage, William E; West, Christopher E

2016-08-23

Genome integrity is crucial for cellular survival and the faithful transmission of genetic information. The eukaryotic cellular response to DNA damage is orchestrated by the DNA damage checkpoint kinases ATAXIA TELANGIECTASIA MUTATED (ATM) and ATM AND RAD3-RELATED (ATR). Here we identify important physiological roles for these sensor kinases in control of seed germination. We demonstrate that double-strand breaks (DSBs) are rate-limiting for germination. We identify that desiccation tolerant seeds exhibit a striking transcriptional DSB damage response during germination, indicative of high levels of genotoxic stress, which is induced following maturation drying and quiescence. Mutant atr and atm seeds are highly resistant to aging, establishing ATM and ATR as determinants of seed viability. In response to aging, ATM delays germination, whereas atm mutant seeds germinate with extensive chromosomal abnormalities. This identifies ATM as a major factor that controls germination in aged seeds, integrating progression through germination with surveillance of genome integrity. Mechanistically, ATM functions through control of DNA replication in imbibing seeds. ATM signaling is mediated by transcriptional control of the cell cycle inhibitor SIAMESE-RELATED 5, an essential factor required for the aging-induced delay to germination. In the soil seed bank, seeds exhibit increased transcript levels of ATM and ATR, with changes in dormancy and germination potential modulated by environmental signals, including temperature and soil moisture. Collectively, our findings reveal physiological functions for these sensor kinases in linking genome integrity to germination, thereby influencing seed quality, crucial for plant survival in the natural environment and sustainable crop production.

The Microanatomic Segregation of Selection by Apoptosis in the Germinal Center

PubMed Central

Mayer, Christian T.; Gazumyan, Anna; Kara, Ervin E.; Gitlin, Alexander D.; Golijanin, Jovana; Viant, Charlotte; Pai, Joy; Oliveira, Thiago Y.; Wang, Qiao; Escolano, Amelia; Medina-Ramirez, Max; Sanders, Rogier W.; Nussenzweig, Michel C.

2018-01-01

B cells undergo rapid cell division and affinity maturation in anatomically distinct sites in lymphoid organs called germinal centers (GCs). Homeostasis is maintained in part by B-cell apoptosis. However, the precise contribution of apoptosis to GC biology and selection is not well defined. We developed apoptosis-indicator mice and used them to visualize, purify, and characterize dying GC B cells. Apoptosis is prevalent in the GC with up to half of all GC B cells dying every 6h. Moreover, programmed cell death is differentially regulated in the light zone (LZ) and the dark zone (DZ): LZ B cells die by default if they are not positively selected, whereas DZ cells die when their antigen receptors are damaged by activation-induced cytidine deaminase (AID). PMID:28935768

Ricin-B-lectin enhances microsporidia Nosema bombycis infection in BmN cells from silkworm Bombyx mori.

PubMed

Liu, Han; Li, Mingqian; Cai, Shunfeng; He, Xinyi; Shao, Yongqi; Lu, Xingmeng

2016-11-01

Nosema bombycis is an obligate intracellular parasitic fungus that utilizes a distinctive mechanism to infect Bombyx mori Spore germination can be used for host cell invasion; however, the detailed mechanism remains to be elucidated. The ricin-B-lectin (RBL) gene is significantly differentially regulated after N. bombycis spore germination, and NbRBL might play roles in spore germination and infection. In this study, the biological function of NbRBL was examined. Protein sequence analysis showed that NbRBL is a secreted protein that attaches to carbohydrates. The relative expression level of the NbRBL gene was low during the first 30 h post-infection (hpi) in BmN cells, and high expression was detected from 42 hpi. Gene cloning, prokaryotic expression, and antibody preparation for NbRBL were performed. NbRBL was detected in total and secreted proteins using western blot analysis. Subcellular localization analysis showed that NbRBL is an intracellular protein. Spore adherence and infection assays showed that NbRBL could enhance spore adhesion to BmN cells; the proliferative activities of BmN cells incubated with anti-NbRBL were higher than those in negative control groups after N. bombycis infection; and the treatment groups showed less damage from spore invasion. We therefore, propose that NbRBL is released during spore germination, enhances spore adhesion to BmN cells, and contributes to spore invasion. © The Author 2016. Published by Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

DELAY OF GERMINATION 1 mediates a conserved coat-dormancy mechanism for the temperature- and gibberellin-dependent control of seed germination

PubMed Central

Graeber, Kai; Linkies, Ada; Steinbrecher, Tina; Mummenhoff, Klaus; Tarkowská, Danuše; Turečková, Veronika; Ignatz, Michael; Sperber, Katja; Voegele, Antje; de Jong, Hans; Urbanová, Terezie; Strnad, Miroslav; Leubner-Metzger, Gerhard

2014-01-01

Seed germination is an important life-cycle transition because it determines subsequent plant survival and reproductive success. To detect optimal spatiotemporal conditions for germination, seeds act as sophisticated environmental sensors integrating information such as ambient temperature. Here we show that the DELAY OF GERMINATION 1 (DOG1) gene, known for providing dormancy adaptation to distinct environments, determines the optimal temperature for seed germination. By reciprocal gene-swapping experiments between Brassicaceae species we show that the DOG1-mediated dormancy mechanism is conserved. Biomechanical analyses show that this mechanism regulates the material properties of the endosperm, a seed tissue layer acting as germination barrier to control coat dormancy. We found that DOG1 inhibits the expression of gibberellin (GA)-regulated genes encoding cell-wall remodeling proteins in a temperature-dependent manner. Furthermore we demonstrate that DOG1 causes temperature-dependent alterations in the seed GA metabolism. These alterations in hormone metabolism are brought about by the temperature-dependent differential expression of genes encoding key enzymes of the GA biosynthetic pathway. These effects of DOG1 lead to a temperature-dependent control of endosperm weakening and determine the optimal temperature for germination. The conserved DOG1-mediated coat-dormancy mechanism provides a highly adaptable temperature-sensing mechanism to control the timing of germination. PMID:25114251

Gene expression at each stage of the life cycle of spore-forming bacteria: different sensitivity of transcription to antibiotics which act on DNA gyrase.

PubMed

Matsuda, M; Kameyama, T

1985-01-01

Effects of antibiotics acting on DNA gyrase, novobiocin and nalidixic acid on RNA synthesis during germination, vegetative growth and sporulation of Bacillus subtilis were examined. These drugs were relatively ineffective in inhibiting RNA synthesis of phase Gm 1 (5-15 min) during germination but effective in those of Gm 2 (15-40 min) and Gm 3 (40-60 min) during germination (Matsuda and Kameyama 1980). No distinguishable inhibitory effects of RNA synthesis in B. subtilis NOVr1TT mutant could be detected by novobiocin. RNA synthesis of Gm 2 and Gm 3 of this mutant was inhibited by nalidixic acid. When novobiocin was added to exponential vegetative cell or sporulating cell culture at T0 and T1 stage, the rate of RNA synthesis was inhibited by 80% for 5 min following addition of the drug. However, RNA synthesis after T2 of sporulation became resistant toward novobiocin, as was the case at an early stage of germination. RNA profiles from transcripts synthesized on administration of NOV suggested that the suppression of the synthesis of 23S and 16S rRNA is relatively greater than 4 to 5S RNA at the middle stage of germination and at vegetative growth stage in the presence of NOV. Our present data suggest that DNA gyrase is involved in the regulation of gene transcription during middle and late phases of germination, vegetative growth and T0 and T1 of sporulation. The transcription during early phase of germination and sporulation after T2 may proceed independently of this enzyme.

Why was the 2012 bloom so early? Using chilling and warming metrics to resolve interannual variability in the timing of Alexandrium fundyense bloom initiation

NASA Astrophysics Data System (ADS)

Fischer, A. D.; Anderson, D. M.; Moore, S.; Brosnahan, M.

2016-02-01

The Nauset Marsh System (NMS) on Cape Cod (MA, USA) has recurrent Alexandrium fundyense blooms that have caused nearly annual shellfishing closures due to paralytic shellfish poisoning. Blooms were observed over a multi-year period (2009-2015) to examine the effects of seasonal cooling and warming on the transition from resting cysts to vegetative cells in the plankton. The life cycle processes of cyst dormancy, germination, and vegetative cell growth are all uniquely sensitive to temperature, which can translate to changes in bloom initiation phenology. Bloom initiation (>100 cells/l-1) occurred as early as 14 February 2012, and as late as 15 April 2015. To quantitatively examine the mechanisms responsible for this two-month range, laboratory studies were performed. In experiments mimicking winter's onset, mature cysts were exposed to chilling temperatures (2-8°C), and at regular intervals the germination potential of cyst cohorts was evaluated. Next, in experiments mimicking a range of late-winter, early-spring temperatures, the time to germination was observed for cold-conditioned cysts. To account for the interannual temperature variability in the NMS and enable comparison to laboratory studies, we calculated growing degree-days and chilling-units, both metrics that tabulate accumulated temperature exposures. Here we pair laboratory studies with seven years of bloom data to present a conceptual model of three temperature-dependent phases of bloom initiation for A. fundyense: 1) Winter dormancy. As temperatures cool, cysts enter a state of dormancy during which germination is physiologically inhibited, until they experience a threshold of winter chilling. 2) Quiescence. Cysts are physiologically able to germinate, but require a specific amount of heat, oxygen, and light. 3) Growth. Germling cells transform to vegetative cells, which divide asexually as a function of heat to create the bloom. These results help to explain differences in bloom timing between years, and provide insights into potential responses of A. fundyense to climate change.

Expression of a Polygalacturonase Associated with Tomato Seed Germination1

PubMed Central

Sitrit, Yaron; Hadfield, Kristen A.; Bennett, Alan B.; Bradford, Kent J.; Downie, A. Bruce

1999-01-01

Radicle protrusion from tomato (Lycopersicon esculentum Mill.) seeds to complete germination requires weakening of the endosperm tissue opposite the radicle tip. In common with other cell wall disassembly processes in plants, polygalacturonases (PGs) may be involved. Only calcium-dependent exo-PG activity was detected in tomato seed protein extracts. Chromatographic profiles of a partially acid-hydrolyzed fraction of polygalacturonic acid further digested with seed extract were consistent with the presence of only calcium-dependent exo-PG activity. In addition, a transcript encoding a previously unknown PG was detected prior to the completion of germination. The mRNA, produced from a gene (LeXPG1) estimated by Southern analysis to be represented once in the genome, was also present in flowers (anthers) and in lower amounts in roots and stems. LeXPG1 mRNA abundance was low during seed development, increased during imbibition, and was even greater in seeds that had completed germination. Expression of LeXPG1 during germination predominates in the endosperm cap and radicle tip, and in the radicle appears as a distinct band possibly associated with vascular tissue differentiation. We suggest that PG is involved in cell wall loosening of the endosperm necessary for radicle protrusion from tomato seeds and in subsequent embryo and seedling growth. PMID:10517833

Proteomic Dissection of Seed Germination and Seedling Establishment in Brassica napus

PubMed Central

Gu, Jianwei; Chao, Hongbo; Gan, Lu; Guo, Liangxing; Zhang, Kai; Li, Yonghong; Wang, Hao; Raboanatahiry, Nadia; Li, Maoteng

2016-01-01

The success of seed germination and establishment of a normal seedling are key determinants of plant species propagation. At present, only a few studies have focused on the genetic control of seed germination by using a proteomic approach in Brassica napus. In the present study, the protein expression pattern of seed germination was investigated using differential fluorescence two-dimensional gel electrophoresis in B. napus. One hundred and thirteen differentially expressed proteins (DEPs) that were mainly involved in storage (23.4%), energy metabolism (18.9%), protein metabolism (16.2%), defense/disease (12.6%), seed maturation (11.7%), carbohydrate metabolism (4.5%), lipid metabolism (4.5%), amino acids metabolism (3.6%), cell growth/division (3.6%), and some unclear functions (2.7%) were observed by proteomic analysis. Seventeen genes corresponding to 11 DEPs were identified within or near the associated linkage disequilibrium regions related to seed germination and vigor quantitative traits reported in B. napus in previous studies. The expression pattern of proteins showed that heterotrophic metabolism could be activated in the process of seed germination and that the onset of defense mechanisms might start during seed germination. These findings will help generate a more in-depth understanding of the mobilization of seed storage reserves and regulation mechanisms of the germination process in B. napus. PMID:27822216

Effect of lipopeptides of antagonistic strains of Bacillus subtilis on the morphology and ultrastructure of the cucurbit fungal pathogen Podosphaera fusca.

PubMed

Romero, D; de Vicente, A; Olmos, J L; Dávila, J C; Pérez-García, A

2007-10-01

To analyse the morphological and ultrastructural effects of lipopeptides of cell-free liquid cultures from the antagonistic Bacillus subtilis strains, UMAF6614 and UMAF6639, on the cucurbit powdery mildew fungus, Podosphaera fusca, conidial germination. Butanolic extracts from cell-free supernatants of B. subtilis cultures were tested for their ability to arrest P. fusca conidial germination using the zucchini cotyledon disc method. Previously, the occurrence of lipopeptide antibiotics fengycin, iturin/bacillomycin and surfactin in the extracts was verified by diverse chromatographic approaches. Conidial germination was strongly reduced by antifungal extracts obtained from liquid cultures of both B. subtilis strains. Scanning electron microscopy analysis showed morphological damage in conidia characterized by the presence of large depressions and loss of turgidness. Transmission electron microscopy analysis revealed severe modifications in the plasma membrane and disorganization of the P. fusca cell cytoplasm. The lipopeptides produced by the two strains of B. subtilis are able to reduce cucurbit powdery mildew disease by arresting conidial germination, which seems to result from the induction of important cytological alterations. We elucidated the mechanisms employed by these antagonistic strains of B. subtilis to suppress cucurbit powdery mildew disease and delineate the ultrastructural damages responsible for their suppressive effect.

Prunus necrotic ringspot virus Early Invasion and Its Effects on Apricot Pollen Grain Performance.

PubMed

Amari, Khalid; Burgos, Lorenzo; Pallas, Vicente; Sanchez-Pina, María Amelia

2007-08-01

ABSTRACT The route of infection and the pattern of distribution of Prunus necrotic ringspot virus (PNRSV) in apricot pollen were studied. PNRSV was detected both within and on the surface of infected pollen grains. The virus invaded pollen during its early developmental stages, being detected in pollen mother cells. It was distributed uniformly within the cytoplasm of uni- and bicellular pollen grains and infected the generative cell. In mature pollen grains, characterized by their triangular shape, the virus was located mainly at the apertures, suggesting that PNRSV distribution follows the same pattern as the cellular components required for pollen tube germination and cell wall tube synthesis. PNRSV also was localized inside pollen tubes, especially in the growth zone. In vitro experiments demonstrated that infection with PNRSV decreases the germination percentage of pollen grains by more than half and delays the growth of pollen tubes by approximately 24 h. However, although PNRSV infection affected apricot pollen grain performance during germination, the presence of the virus did not completely prevent fertilization, because the infected apricot pollen tubes, once germinated, were able to reach the apricot embryo sacs, which, in the climatic conditions of southeastern Spain, mature later than in other climates. Thus, infected pollen still could play an important role in the vertical transmission of PNRSV in apricot.

How Germinal Centers Evolve Broadly Neutralizing Antibodies: the Breadth of the Follicular Helper T Cell Response

DOE PAGES

De Boer, Rob J.; Perelson, Alan S.

2017-09-06

Many HIV-1-infected patients evolve broadly neutralizing antibodies (bnAbs). This evolutionary process typically takes several years and is poorly understood as selection taking place in germinal centers occurs on the basis of antibody affinity. B cells with the highest-affinity receptors tend to acquire the most antigen from the follicular dendritic cell (FDC) network and present the highest density of cognate peptides to follicular helper T (Tfh) cells, which provide survival signals to the B cell. bnAbs are therefore expected to evolve only when the B cell lineage evolving breadth is consistently capturing and presenting more peptides to Tfh cells than othermore » lineages of more specific B cells. Here we develop mathematical models of Tfh cells in germinal centers to explicitly define the mechanisms of selection in this complex evolutionary process. Our results suggest that broadly reactive B cells presenting a high density of peptides bound to major histocompatibility complex class II molecules (pMHC) are readily outcompeted by B cells responding to lineages of HIV-1 that transiently dominate the within host viral population. Conversely, if broadly reactive B cells acquire a large variety of several HIV-1 proteins from the FDC network and present a high diversity of several pMHC, they can be rescued by a large fraction of the Tfh cell repertoire in the germinal center. Under such circumstances the evolution of bnAbs is much more consistent. Increasing either the magnitude of the Tfh cell response or the breadth of the Tfh cell repertoire markedly facilitates the evolution of bnAbs. Because both the magnitude and breadth can be increased by vaccination with several HIV-1 proteins, this calls for experimental testing. Many HIV-infected patients slowly evolve antibodies that can neutralize a large variety of viruses. Such broadly neutralizing antibodies (bnAbs) could in the future become therapeutic agents. bnAbs appear very late, and patients are typically not protected by them. At the moment, we fail to understand why this takes so long and how the immune system selects for broadly neutralizing capacity. Typically, antibodies are selected based on affinity and not on breadth. We developed mathematical models to study two different mechanisms by which the immune system can select for broadly neutralizing capacity. One of these is based upon the repertoire of different follicular helper T (Tfh) cells in germinal centers. In conclusion, we suggest that broadly reactive B cells may interact with a larger fraction of this repertoire and demonstrate that this would select for bnAbs. Intriguingly, this suggests that broadening the Tfh cell repertoire by vaccination may speed up the evolution of bnAbs.« less

How Germinal Centers Evolve Broadly Neutralizing Antibodies: the Breadth of the Follicular Helper T Cell Response

DOE Office of Scientific and Technical Information (OSTI.GOV)

De Boer, Rob J.; Perelson, Alan S.

Many HIV-1-infected patients evolve broadly neutralizing antibodies (bnAbs). This evolutionary process typically takes several years and is poorly understood as selection taking place in germinal centers occurs on the basis of antibody affinity. B cells with the highest-affinity receptors tend to acquire the most antigen from the follicular dendritic cell (FDC) network and present the highest density of cognate peptides to follicular helper T (Tfh) cells, which provide survival signals to the B cell. bnAbs are therefore expected to evolve only when the B cell lineage evolving breadth is consistently capturing and presenting more peptides to Tfh cells than othermore » lineages of more specific B cells. Here we develop mathematical models of Tfh cells in germinal centers to explicitly define the mechanisms of selection in this complex evolutionary process. Our results suggest that broadly reactive B cells presenting a high density of peptides bound to major histocompatibility complex class II molecules (pMHC) are readily outcompeted by B cells responding to lineages of HIV-1 that transiently dominate the within host viral population. Conversely, if broadly reactive B cells acquire a large variety of several HIV-1 proteins from the FDC network and present a high diversity of several pMHC, they can be rescued by a large fraction of the Tfh cell repertoire in the germinal center. Under such circumstances the evolution of bnAbs is much more consistent. Increasing either the magnitude of the Tfh cell response or the breadth of the Tfh cell repertoire markedly facilitates the evolution of bnAbs. Because both the magnitude and breadth can be increased by vaccination with several HIV-1 proteins, this calls for experimental testing. Many HIV-infected patients slowly evolve antibodies that can neutralize a large variety of viruses. Such broadly neutralizing antibodies (bnAbs) could in the future become therapeutic agents. bnAbs appear very late, and patients are typically not protected by them. At the moment, we fail to understand why this takes so long and how the immune system selects for broadly neutralizing capacity. Typically, antibodies are selected based on affinity and not on breadth. We developed mathematical models to study two different mechanisms by which the immune system can select for broadly neutralizing capacity. One of these is based upon the repertoire of different follicular helper T (Tfh) cells in germinal centers. In conclusion, we suggest that broadly reactive B cells may interact with a larger fraction of this repertoire and demonstrate that this would select for bnAbs. Intriguingly, this suggests that broadening the Tfh cell repertoire by vaccination may speed up the evolution of bnAbs.« less

BCL6 interacting corepressor contributes to germinal center T follicular helper cell formation and B cell helper function

PubMed Central

Yang, Jessica A.; Tubo, Noah J.; Gearhart, Micah D.; Bardwell, Vivian J.; Jenkins, Marc K.

2015-01-01

CD4+ germinal center (GC) T follicular helper (GC-Tfh) cells help B cells become long-lived plasma cells and memory cells. The transcriptional repressor BCL6 plays a key role in GC-Tfh formation by inhibiting the expression of genes that promote differentiation into other lineages. We determined whether BCOR, a component of a Polycomb repressive complex that interacts with the BCL6 BTB domain, influences GC-Tfh differentiation. T cell-targeted BCOR deficiency led to a substantial loss of peptide:MHCII-specific GC-Tfh cells following Listeria monocytogenes infection and a 2-fold decrease following immunization with a peptide in CFA. The reduction in GC-Tfh cells was associated with diminished plasma cell and GC B cell formation. Thus, T cell-expressed BCOR is critical for optimal GC-Tfh differentiation and humoral immunity. PMID:25964495

Using atmospheric plasma to increase wettability, imbibition and germination of physically dormant seeds of Mimosa Caesalpiniafolia.

PubMed

da Silva, A R M; Farias, M L; da Silva, D L S; Vitoriano, J O; de Sousa, R C; Alves-Junior, C

2017-09-01

In this study, we analyzed seed wettability as well as imbibition and germination after treatment with atmospheric pressure cold plasma (APCP) using dielectric barrier discharge (DBD) in seeds that have very low germination rates. To aid industrial applications, several seeds were simultaneously treated with plasma within a space between two coaxial glass tubes sandwiched by two metal mesh screens that produced high-voltage pulses at 17.5kV with a frequency of 990Hz. Three treatment times (3min, 9min and 15min) as well as untreated seeds were used to conduct the wettability, imbibition and germination tests. The wettability and imbibition were found to be directly related to the treatment duration, but saturation of the imbibition was found for treatment durations greater than 9min. Plasma treatment was also effective in improving germination, but shorter treatment duration presented greater germination. This apparent contradiction is explained by the cell damage caused by the increased exposure to plasma, as observed in other studies. The results suggest that there must be an optimal wettability and imbibition condition that ensures that excessive moisture does not harm the germination process. Copyright © 2017 Elsevier B.V. All rights reserved.

Early detection of germinated wheat grains using terahertz image and chemometrics

NASA Astrophysics Data System (ADS)

Jiang, Yuying; Ge, Hongyi; Lian, Feiyu; Zhang, Yuan; Xia, Shanhong

2016-02-01

In this paper, we propose a feasible tool that uses a terahertz (THz) imaging system for identifying wheat grains at different stages of germination. The THz spectra of the main changed components of wheat grains, maltose and starch, which were obtained by THz time spectroscopy, were distinctly different. Used for original data compression and feature extraction, principal component analysis (PCA) revealed the changes that occurred in the inner chemical structure during germination. Two thresholds, one indicating the start of the release of α-amylase and the second when it reaches the steady state, were obtained through the first five score images. Thus, the first five PCs were input for the partial least-squares regression (PLSR), least-squares support vector machine (LS-SVM), and back-propagation neural network (BPNN) models, which were used to classify seven different germination times between 0 and 48 h, with a prediction accuracy of 92.85%, 93.57%, and 90.71%, respectively. The experimental results indicated that the combination of THz imaging technology and chemometrics could be a new effective way to discriminate wheat grains at the early germination stage of approximately 6 h.

Effects of lithium chloride as a potential radioprotective agent on radiation response of DNA synthesis in mouse germinal cells.

PubMed

Bhattacharjee, D; Rajan, R; Krishnamoorthy, L; Singh, B B

1997-06-01

Mouse spermatogonial germ cells are highly sensitive to ionizing radiation. Lithium salts are reported to stimulate the postirradiation recovery of hematopoietic marrow cells. We have, therefore, examined whether administered lithium chloride (LiCl) would also be able to protect the mouse germinal cells against radiation injury. Taking DNA synthesis as an endpoint, our results show that the testicular DNA-specific activity in irradiated mice was higher by 61% on average when they had been pretreated with LiCl both 24 h and 1 h prior to gamma-irradiation (2.0 Gy). It was also observed that the DNA synthetic activity in the germinal cells fully recovered after LiCl pretreatment at doses of 40 mg per kg body weight prior to total body irradiation of 0.05-0.25 Gy, whereas at doses of 0.5-6.0 Gy, following the same procedure of LiCl pretreatment, only an incomplete recovery was observed. The dose reduction factor for LiCl is 1.84. The current findings indicate that pretreatment with LiCl provides considerable protection against radiation damage in mouse spermatogonia.

Fate of Salmonella enterica and Enterohemorrhagic Escherichia coli Cells Artificially Internalized into Vegetable Seeds during Germination.

PubMed

Liu, Da; Cui, Yue; Walcott, Ronald; Chen, Jinru

2018-01-01

Vegetable seeds contaminated with bacterial pathogens have been linked to fresh-produce-associated outbreaks of gastrointestinal infections. This study was undertaken to observe the physiological behavior of Salmonella enterica and enterohemorrhagic Escherichia coli (EHEC) cells artificially internalized into vegetable seeds during the germination process. Surface-decontaminated seeds of alfalfa, fenugreek, lettuce, and tomato were vacuum-infiltrated with four individual strains of Salmonella or EHEC. Contaminated seeds were germinated at 25°C for 9 days, and different sprout/seedling tissues were microbiologically analyzed every other day. The internalization of Salmonella and EHEC cells into vegetable seeds was confirmed by the absence of pathogens in seed-rinsing water and the presence of pathogens in seed homogenates after postinternalization seed surface decontamination. Results show that 317 (62%) and 343 (67%) of the 512 collected sprout/seedling tissue samples were positive for Salmonella and EHEC, respectively. The average Salmonella populations were significantly larger ( P < 0.05) than the EHEC populations. Significantly larger Salmonella populations were recovered from the cotyledon and seed coat tissues, followed by the root tissues, but the mean EHEC populations from all sampled tissue sections were statistically similar, except in pregerminated seeds. Three Salmonella and two EHEC strains had significantly larger cell populations on sprout/seedling tissues than other strains used in the study. Salmonella and EHEC populations from fenugreek and alfalfa tissues were significantly larger than those from tomato and lettuce tissues. The study showed the fate of internalized human pathogens on germinating vegetable seeds and sprout/seedling tissues and emphasized the importance of using pathogen-free seeds for sprout production. IMPORTANCE The internalization of microorganisms into vegetable seeds could occur naturally and represents a possible pathway of vegetable seed contamination by human pathogens. The present study investigated the ability of two important bacterial pathogens, Salmonella and enterohemorrhagic Escherichia coli (EHEC), when artificially internalized into vegetable seeds, to grow and disseminate along vegetable sprouts/seedlings during germination. The data from the study revealed that the pathogen cells artificially internalized into vegetable seeds caused the contamination of different tissues of sprouts/seedlings and that pathogen growth on germinating seeds is bacterial species and vegetable seed-type dependent. These results further stress the necessity of using pathogen-free vegetable seeds for edible sprout production. Copyright © 2017 American Society for Microbiology.

A light microscopy and ultrastructural study of the testes of tortoise Testudo graeca (Testudinidae).

PubMed

Ibargüengoytía, N R; Pastor, L M; Pallares, J

1999-04-01

Adult males of Testudo graeca were used to preliminarily study the light microscopic morphology and the ultrastructure of the testes. Spermiogenesis has shown the presence of some interspecific variations among Chelonia, while the general features of the testes and spermatocytes are morphologically similar to other reptilians. The male reproductive state observed in the months analysed has shown spermatogenesis recrudescence in spring, a complete germinal series in autumn and testicular regression in winter. The observation of ultrastructural features, characteristic of steroidogenic activity, suggests a synchrony in tubular and interstitial compartments in T. graeca, with little steroidogenic activity in winter and active synthesis in spring and autumn. In conclusion, the results of this histological study suggest a probable asynchrony between the male and female reproductive cycle in this species and show synchrony in the steroidogenic activity of Sertoli and Leydig cells.

Karyological observations

NASA Technical Reports Server (NTRS)

Krikorian, A. D.; O'Connor, S. A.

1984-01-01

Root tips prepared for metaphase chromosome analysis from seedlings germinated under microgravity on the Space Shuttle (oats and mung bean) or which were exposed to space flight as very young seedlings (sunflower) have been examined. Experimental constraints did not permit pre-fixation in space with a cytostatic agent but arrest was achieved in the first division cycle on Earth after recovery. The number of cells in division was significantly depressed in all three species. Several chromosomal abnormalities were encountered in flight material. Bridge formation was seen in sunflower, as was aneuploidy. Breakage and fracture of chromosomes was prevalent in oats. No aberrant features could be detected in the chromosomes of mung bean. These results, although preliminary, should serve to alert investigators of the need to assess carefully as many aspects of cell division in higher plants exposed to space flight conditions as possible.

Germination Potential of Dormant and Nondormant Arabidopsis Seeds Is Driven by Distinct Recruitment of Messenger RNAs to Polysomes

PubMed Central

Basbouss-Serhal, Isabelle; Soubigou-Taconnat, Ludivine; Bailly, Christophe; Leymarie, Juliette

2015-01-01

Dormancy is a complex evolutionary trait that temporally prevents seed germination, thus allowing seedling growth at a favorable season. High-throughput analyses of transcriptomes have led to significant progress in understanding the molecular regulation of this process, but the role of posttranscriptional mechanisms has received little attention. In this work, we have studied the dynamics of messenger RNA association with polysomes and compared the transcriptome with the translatome in dormant and nondormant seeds of Arabidopsis (Arabidopsis thaliana) during their imbibition at 25°C in darkness, a temperature preventing germination of dormant seeds only. DNA microarray analysis revealed that 4,670 and 7,028 transcripts were differentially abundant in dormant and nondormant seeds in the transcriptome and the translatome, respectively. We show that there is no correlation between transcriptome and translatome and that germination regulation is also largely translational, implying a selective and dynamic recruitment of messenger RNAs to polysomes in both dormant and nondormant seeds. The study of 5′ untranslated region features revealed that GC content and the number of upstream open reading frames could play a role in selective translation occurring during germination. Gene Ontology clustering showed that the functions of polysome-associated transcripts differed between dormant and nondormant seeds and revealed actors in seed dormancy and germination. In conclusion, our results demonstrate the essential role of selective polysome loading in this biological process. PMID:26019300

5′ to 3′ mRNA Decay Contributes to the Regulation of Arabidopsis Seed Germination by Dormancy1

PubMed Central

Basbouss-Serhal, Isabelle; Pateyron, Stéphanie; Cochet, Françoise

2017-01-01

The regulation of plant gene expression, necessary for development and adaptive responses, relies not only on RNA transcription but also on messenger RNA (mRNA) fate. To understand whether seed germination relies on the degradation of specific subsets of mRNA, we investigated whether the 5′ to 3′ RNA decay machinery participated in the regulation of this process. Arabidopsis (Arabidopsis thaliana) seeds of exoribonuclease4 (xrn4) and varicose (vcs) mutants displayed distinct dormancy phenotypes. Transcriptome analysis of xrn4-5 and vcs-8 mutant seeds allowed us to identify genes that are likely to play a role in the control of germination. Study of 5′ untranslated region features of these transcripts revealed that specific motifs, secondary energy, and GC content could play a role in their degradation by XRN4 and VCS, and Gene Ontology clustering revealed novel actors of seed dormancy and germination. Several specific transcripts identified as being putative targets of XRN4 and VCS in seeds (PECTIN LYASE-LIKE, ASPARTYL PROTEASE, DWD-HYPERSENSITIVE-TO-ABA3, and YELLOW STRIPE-LIKE5) were further studied by reverse genetics, and their functional roles in the germination process were confirmed by mutant analysis. These findings suggest that completion of germination and its regulation by dormancy also depend on the degradation of specific subsets of mRNA. PMID:28126845

Vacuolar status and water relations in embryonic axes of recalcitrant Aesculus hippocastanum seeds during stratification and early germination

PubMed Central

Obroucheva, Natalie V.; Lityagina, Snezhana V.; Novikova, Galina V.; Sin'kevich, Irina A.

2012-01-01

Backgrounds and aims In tropical recalcitrant seeds, their rapid transition from shedding to germination at high hydration level is of physiological interest but difficult to study because of the time constraint. In recalcitrant horse chestnut seeds produced in central Russia, this transition is much longer and extends through dormancy and dormancy release. This extended time period permits studies of the water relations in embryonic axes during the long recalcitrant period in terms of vacuolar status and water transport. Methodology Horse chestnut (Aesculus hippocastanum) seeds sampled in Moscow were stratified in cold wet sand for 4 months. Vacuole presence and development in embryonic axes were examined by vital staining, light and electron microscopy. Aquaporins and vacuolar H+-ATPase were identified immunochemically. Water channel operation was tested by water inflow rate. Vacuolar acid invertase was estimated in terms of activity and electrophoretic properties. Principal results Throughout the long recalcitrant period after seed shedding, cells of embryonic axes maintained active vacuoles and a high water content. Preservation of enzyme machinery in vacuoles was evident from retention of invertase activity, substrate specificity, molecular mass and subunit composition. Plasmalemma and tonoplast aquaporins and the E subunit of vacuolar H+-ATPase were also present. In non-dormant seeds prior to growth initiation, vacuoles enlarged at first in hypocotyls, and then in radicles, with their biogenesis being similar. Vacuolation was accompanied by increasing invertase activity, leading to sugar accumulation and active osmotic functioning. After growth initiation, vacuole enlargement was favoured by enhanced water inflow through water channels formed by aquaporins. Conclusions Maintenance of high water content and desiccation sensitivity, as well as preservation of active vacuoles in embryonic axes after shedding, can be considered a specific feature of recalcitrant seeds, overlooked when studying tropical recalcitrants due to the short duration. The retained physiological activity of vacuoles allows them to function rapidly as dormancy is lost and when external conditions permit. Cell vacuolation precedes cell elongation in both hypocotyl and radicle, and provides impetus for rapid germination. PMID:22593822

Aberrant Meiotic Modulation Partially Contributes to the Lower Germination Rate of Pollen Grains in Maize (Zea mays L.) Under Low Nitrogen Supply.

PubMed

Zheng, Hongyan; Wu, Huamao; Pan, Xiaoying; Jin, Weiwei; Li, Xuexian

2017-02-01

Pollen germination is an essential step towards successful pollination during maize reproduction. How low niutrogen (N) affects pollen germination remains an interesting biological question to be addressed. We found that only low N resulted in a significantly lower germination rate of pollen grains after 4 weeks of low N, phosphorus or potassium treatment in maize production. Importantly, cytological analysis showed 7-fold more micronuclei in male meiocytes under the low N treatment than in the control, indicating that the lower germination rate of pollen grains was partially due to numerous chromosome loss events resulting from preceding meiosis. The appearance of 10 bivalents in the control and low N cells at diakinesis suggested that chromosome pairing and recombination in meiosis I was not affected by low N. Further gene expression analysis revealed dramatic down-regulation of Nuclear Division Cycle 80 (Ndc80) and Regulator of Chromosome Condensation 1 (Rcc1-1) expression and up-regulation of Cell Division Cycle 20 (Cdc20-1) expression, although no significant difference in the expression level of kinetochore foundation proteins Centromeric Histone H3 (Cenh3) and Centromere Protein C (Cenpc) and cohesion regulators Recombination 8 (Rec8) and Shugoshin (Sgo1) was observed. Aberrant modulation of three key meiotic regulators presumably resulted in a high likelihood of erroneous chromosome segregation, as testified by pronounced lagging chromosomes at anaphase I or cell cycle disruption at meiosis II. Thus, we proposed a cytogenetic mechanism whereby low N affects male meiosis and causes a higher chromosome loss frequency and eventually a lower germination rate of pollen grains in a staple crop plant. © The Author 2016. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Glycerol enhances fungal germination at the water-activity limit for life.

PubMed

Stevenson, Andrew; Hamill, Philip G; Medina, Ángel; Kminek, Gerhard; Rummel, John D; Dijksterhuis, Jan; Timson, David J; Magan, Naresh; Leong, Su-Lin L; Hallsworth, John E

2017-03-01

For the most-extreme fungal xerophiles, metabolic activity and cell division typically halts between 0.700 and 0.640 water activity (approximately 70.0-64.0% relative humidity). Here, we investigate whether glycerol can enhance xerophile germination under acute water-activity regimes, using an experimental system which represents the biophysical limit of Earth's biosphere. Spores from a variety of species, including Aspergillus penicillioides, Eurotium halophilicum, Xerochrysium xerophilum (formerly Chrysosporium xerophilum) and Xeromyces bisporus, were produced by cultures growing on media supplemented with glycerol (and contained up to 189 mg glycerol g dry spores -1 ). The ability of these spores to germinate, and the kinetics of germination, were then determined on a range of media designed to recreate stresses experienced in microbial habitats or anthropogenic systems (with water-activities from 0.765 to 0.575). For A. penicillioides, Eurotium amstelodami, E. halophilicum, X. xerophilum and X. bisporus, germination occurred at lower water-activities than previously recorded (0.640, 0.685, 0.651, 0.664 and 0.637 respectively). In addition, the kinetics of germination at low water-activities were substantially faster than those reported previously. Extrapolations indicated theoretical water-activity minima below these values; as low as 0.570 for A. penicillioides and X. bisporus. Glycerol is present at high concentrations (up to molar levels) in many types of microbial habitat. We discuss the likely role of glycerol in expanding the water-activity limit for microbial cell function in relation to temporal constraints and location of the microbial cell or habitat. The findings reported here have also critical implications for understanding the extremes of Earth's biosphere; for understanding the potency of disease-causing microorganisms; and in biotechnologies that operate at the limits of microbial function. © 2016 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.

Effects of acidity on tree pollen germination and tube growth

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jacobson, J.S.; Van Rye, D.M.; Lassoie, J.P.

Several studies have indicated that pollen germination and tube growth are adversely affected by air pollutants. Pollutants may inhibit the function of pollen by reducing the number of pollen grains which germinate, by reducing the maximum length to which the pollen tubes grow, or by interfering with the formation of the generative cell. The paper reports on studies that are attempting to determine the effects acid rain may have on these crucial stages in the life histories of northeastern tree species. The first stage of this work assessed the effects of acidity in the growth medium on in vitro pollenmore » germination for four deciduous forest species common to central New York State, Betula lutea (yellow birch), B. lenta (black birch), Acer saccharum (sugar maple), and Cornus florida (flowering dogwood). Measurements were taken at the end of the growth period to determine the percentage of grains which had germinated, and to estimate the average tube length. To determine the effects of pollen on the growth medium, the pH of the germination drop was measured at the end of the growth period.« less

Acquisition of physical dormancy and ontogeny of the micropyle–water-gap complex in developing seeds of Geranium carolinianum (Geraniaceae)

PubMed Central

Gama-Arachchige, N. S.; Baskin, J. M.; Geneve, R. L.; Baskin, C. C.

2011-01-01

Background and Aims The ‘hinged valve gap’ has been previously identified as the initial site of water entry (i.e. water gap) in physically dormant (PY) seeds of Geranium carolinianum (Geraniaceae). However, neither the ontogeny of the hinged valve gap nor acquisition of PY by seeds of Geraniaceae has been studied previously. The aims of the present study were to investigate the physiological events related to acquisition of PY and the ontogeny of the hinged valve gap and seed coat of G. carolinianum. Methods Seeds of G. carolinianum were studied from the ovule stage until dispersal. The developmental stages of acquisition of germinability, physiological maturity and PY were determined by seed measurement, germination and imbibition experiments using intact seeds and isolated embryos of both fresh and slow-dried seeds. Ontogeny of the seed coat and water gap was studied using light microscopy. Key Results Developing seeds achieved germinability, physiological maturity and PY on days 9, 14 and 20 after pollination (DAP), respectively. The critical moisture content of seeds on acquisition of PY was 11 %. Slow-drying caused the stage of acquisition of PY to shift from 20 to 13 DAP. Greater extent of cell division and differentiation at the micropyle, water gap and chalaza than at the rest of the seed coat resulted in particular anatomical features. Palisade and subpalisade cells of varying forms developed in these sites. A clear demarcation between the water gap and micropyle is not evident due to their close proximity. Conclusions Acquisition of PY in seeds of G. carolinianum occurs after physiological maturity and is triggered by maturation drying. The micropyle and water gap cannot be considered as two separate entities, and thus it is more appropriate to consider them together as a ‘micropyle–water-gap complex’. PMID:21546433

Expression and inhibition of aquaporins in germinating Arabidopsis seeds.

PubMed

Vander Willigen, Clare; Postaire, Olivier; Tournaire-Roux, Colette; Boursiac, Yann; Maurel, Christophe

2006-09-01

Extensive and kinetically well-defined water exchanges occur during germination of seeds. A putative role for aquaporins in this process was investigated in Arabidopsis. Macro-arrays carrying aquaporin gene-specific tags and antibodies raised against aquaporin subclasses revealed two distinct aquaporin expression programs between dry seeds and young seedlings. High expression levels of a restricted number of tonoplast intrinsic protein (TIP) isoforms (TIP3;1 and/or TIP3;2, and TIP5;1) together with a low expression of all 13 plasma membrane aquaporin (PIP) isoforms was observed in dry and germinating materials. In contrast, prevalent expression of aquaporins of the TIP1, TIP2 and PIP subgroups was induced during seedling establishment. Mercury (5 microM HgCl(2)), a general blocker of aquaporins in various organisms, reduced the speed of seed germination and induced a true delay in maternal seed coat (testa) rupture and radicle emergence, by 8-9 and 25-30 h, respectively. Most importantly, mercury did not alter seed lot homogeneity nor the seed germination developmental sequence, and its effects were largely reversed by addition of 2 mM dithiothreitol, suggesting that these effects were primarily due to oxidation of cell components, possibly aquaporins, without irreversible alteration of cell integrity. Measurements of water uptake in control and mercury-treated seeds suggested that aquaporin functions are not involved in early seed imbibition (phase I) but would rather be associated with a delayed initiation of phase III, i.e. water uptake accompanying expansion and growth of the embryo. A possible role for aquaporins in germinating seeds and more generally in plant tissue growth is discussed.

Latrunculin B-induced plant dwarfism: Plant cell elongation is F-actin-dependent.

PubMed

Baluska, F; Jasik, J; Edelmann, H G; Salajová, T; Volkmann, D

2001-03-01

Marine macrolides latrunculins are highly specific toxins which effectively depolymerize actin filaments (generally F-actin) in all eukaryotic cells. We show that latrunculin B is effective on diverse cell types in higher plants and describe the use of this drug in probing F-actin-dependent growth and in plant development-related processes. In contrast to other eukaryotic organisms, cell divisions occurs in plant cells devoid of all actin filaments. However, the alignment of the division planes is often distorted. In addition to cell division, postembryonic development and morphogenesis also continue in the absence of F-actin. These experimental data suggest that F-actin is of little importance in the morphogenesis of higher plants, and that plants can develop more or less normally without F-actin. In contrast, F-actin turns out to be essential for cell elongation. When latrunculin B was added during germination, morphologically normal Arabidopsis and rye seedlings developed but, as a result of the absence of cell elongation, these were stunted, resembling either genetic dwarfs or environmental bonsai plants. In conclusion, F-actin is essential for the plant cell elongation, while this F-actin-dependent cell elongation is not an essential feature of plant-specific developmental programs.

Vacuolar biogenesis and aquaporin expression at early germination of broad bean seeds.

PubMed

Novikova, Galina V; Tournaire-Roux, Colette; Sinkevich, Irina A; Lityagina, Snejana V; Maurel, Christophe; Obroucheva, Natalie

2014-09-01

A key event in seed germination is water uptake-mediated growth initiation in embryonic axes. Vicia faba var. minor (broad bean) seeds were used for studying cell growth, vacuolar biogenesis, expression and function of tonoplast water channel proteins (aquaporins) in embryonic axes during seed imbibition, radicle emergence and growth. Hypocotyl and radicle basal cells showed vacuole restoration from protein storage vacuoles, whereas de novo vacuole formation from provacuoles was observed in cells newly produced by root meristem. cDNA fragments of seven novel aquaporin isoforms including five Tonoplast Intrinsic Proteins (TIP) from three sub-types were amplified by PCR. The expression was probed using q-RT-PCR and when possible with isoform-specific antibodies. Decreased expression of TIP3s was associated to the transformation of protein storage vacuoles to vacuoles, whereas enhanced expression of a TIP2 homologue was closely linked to the fast cell elongation. Water channel functioning checked by inhibitory test with mercuric chloride showed closed water channels prior to growth initiation and active water transport into elongating cells. The data point to a crucial role of tonoplast aquaporins during germination, especially during growth of embryonic axes, due to accelerated water uptake and vacuole enlargement resulting in rapid cell elongation. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

Clinical, biologic, and pathologic features in 157 patients with angioimmunoblastic T-cell lymphoma treated within the Groupe d'Etude des Lymphomes de l'Adulte (GELA) trials

PubMed Central

Mourad, Nathalie; Mounier, Nicolas; Brière, Josette; Raffoux, Emmanuel; Delmer, Alain; Feller, Alfred; Meijer, Chris J. L. M.; Emile, Jean-François; Bouabdallah, Réda; Bosly, André; Diebold, Jacques; Haioun, Corinne; Coiffier, Bertrand; Gisselbrecht, Christian

2008-01-01

To evaluate the prognostic significance of clinicobiologic and pathological features in angioimmunoblastic T-cell lymphoma (AITL), 157 AITL patients were retrieved from the GELA LNH87-LNH93 randomized clinical trials. One hundred forty-seven patients received a cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP)–like regimen with intensified courses in half of them. Histologically, 41 cases were classified as “rich in large cells” and 116 as “classic” (including 19 rich in epithelioid cells, 14 rich in clear cells, and 4 with hyperplastic germinal centers). Sixty-two cases were scored for CD10 and CXCL13 expression according to the abundance of positive lymphoid cells. Median age was 62 years, with 81% advanced stage, 72% B symptoms, 65% anemia, 50% hypergammaglobulinemia, and 66% elevated LDH. Overall 7-year survival was 30%. In multivariate analysis, only male sex (P = .004), mediastinal lymphadenopathy (P = .041), and anemia (P = .042) adversely affected overall survival. Increase in large cells and high level of CD10 and CXCL13 did not affect survival. Intensive regimen did not improve survival. In conclusion, AITL is a morphologically heterogeneous T-cell lymphoma commonly expressing CXCL13 and CD10 and carrying few prognostic factors. It portends a poor prognosis even when treated intensively. However, AITL is not always lethal with 30% of patients alive at 7 years. PMID:18292286

B cells expressing the transcription factor T-bet drive lupus-like autoimmunity

PubMed Central

Rubtsov, Anatoly V.; Thurman, Joshua M.; Mennona, Johanna M.; Kappler, John W.; Marrack, Philippa

2017-01-01

B cells contribute to multiple aspects of autoimmune disorders and may play a role in triggering disease. Thus, targeting B cells may be a promising strategy for treating autoimmune disorders. Better understanding of the B cell subsets that are responsible for the development of autoimmunity will be critical for developing efficient therapies. Here we have reported that B cells expressing the transcription factor T-bet promote the rapid appearance of autoantibodies and germinal centers in spontaneous murine models of systemic lupus erythematosus (SLE). Conditional deletion of T-bet from B cells impaired the formation of germinal centers and mitigated the development of kidney damage and rapid mortality in SLE mice. B cell–specific deletion of T-bet was also associated with lower activation of both B cells and T cells. Taken together, our results suggest that targeting T-bet–expressing B cells may be a potential target for therapy for autoimmune diseases. PMID:28240602

The impact of inducing germination of Bacillus anthracis and Bacillus thuringiensis spores on potential secondary decontamination strategies.

PubMed

Omotade, T O; Bernhards, R C; Klimko, C P; Matthews, M E; Hill, A J; Hunter, M S; Webster, W M; Bozue, J A; Welkos, S L; Cote, C K

2014-12-01

Decontamination and remediation of a site contaminated by the accidental or intentional release of fully virulent Bacillus anthracis spores are difficult, costly and potentially damaging to the environment. Development of novel decontamination strategies that have minimal environmental impacts remains a high priority. Although ungerminated spores are amongst the most resilient organisms known, once exposed to germinants, the germinating spores, in some cases, become susceptible to antimicrobial environments. We evaluated the concept that once germinated, B. anthracis spores would be less hazardous and significantly easier to remediate than ungerminated dormant spores. Through in vitro germination and sensitivity assays, we demonstrated that upon germination, B. anthracis Ames spores and Bacillus thuringiensis Al Hakam spores (serving as a surrogate for B. anthracis) become susceptible to environmental stressors. The majority of these germinated B. anthracis and B. thuringiensis spores were nonviable after exposure to a defined minimal germination-inducing solution for prolonged periods of time. Additionally, we examined the impact of potential secondary disinfectant strategies including bleach, hydrogen peroxide, formaldehyde and artificial UV-A, UV-B and UV-C radiation, employed after a 60-min germination-induction step. Each secondary disinfectant employs a unique mechanism of killing; as a result, germination-induction strategies are better suited for some secondary disinfectants than others. These results provide evidence that the deployment of an optimal combination strategy of germination-induction/secondary disinfection may be a promising aspect of wide-area decontamination following a B. anthracis contamination event. By inducing spores to germinate, our data confirm that the resulting cells exhibit sensitivities that can be leveraged when paired with certain decontamination measures. This increased susceptibility could be exploited to devise more efficient and safe decontamination measures and may obviate the need for more stringent methods that are currently in place. Published 2014. This article is a U.S. Government work and is in the public domain in the USA.

Real-time visualization of immune cell clearance of Aspergillus fumigatus spores and hyphae.

PubMed

Knox, Benjamin P; Huttenlocher, Anna; Keller, Nancy P

2017-08-01

Invasive aspergillosis (IA) is a disease of the immunocompromised host and generally caused by the opportunistic fungal pathogen Aspergillus fumigatus. While both host and fungal factors contribute to disease severity and outcome, there are fundamental features of IA development including fungal morphological transition from infectious conidia to tissue-penetrating hyphae as well as host defenses rooted in mechanisms of innate phagocyte function. Here we address recent advances in the field and use real-time in vivo imaging in the larval zebrafish to visually highlight conserved vertebrate innate immune behaviors including macrophage phagocytosis of conidia and neutrophil responses post-germination. Copyright © 2017 Elsevier Inc. All rights reserved.

Translatome profiling in dormant and nondormant sunflower (Helianthus annuus) seeds highlights post-transcriptional regulation of germination.

PubMed

Layat, Elodie; Leymarie, Juliette; El-Maarouf-Bouteau, Hayat; Caius, José; Langlade, Nicolas; Bailly, Christophe

2014-12-01

Seed dormancy, which blocks germination in apparently favourable conditions, is a key regulatory control point of plant population establishment. As germination requires de novo translation, its regulation by dormancy is likely to be related to the association of individual transcripts to polysomes. Here, the polysome-associated mRNAs, that is, the translatome, were fractionated and characterized with microarrays in dormant and nondormant sunflower (Helianthus annuus) embryos during their imbibition at 10°C, a temperature preventing germination of dormant embryos. Profiling of mRNAs in polysomal complexes revealed that the translatome differs between germinating and nongerminating embryos. Association of transcripts with polysomes reached a maximum after 15 h of imbibition; at this time-point 194 polysome-associated transcripts were specifically found in nondormant embryos and 47 in dormant embryos only. The proteins corresponding to the polysomal mRNAs in nondormant embryos appeared to be very pertinent for germination and were involved mainly in transport, regulation of transcription or cell wall modifications. This work demonstrates that seed germination results from a timely regulated and selective recruitment of mRNAs to polysomes, thus opening novel fields of investigation for the understanding of this developmental process. © 2014 The Authors. New Phytologist © 2014 New Phytologist Trust.

Nanopriming technology for enhancing germination and starch metabolism of aged rice seeds using phytosynthesized silver nanoparticles.

PubMed

Mahakham, Wuttipong; Sarmah, Ajit K; Maensiri, Santi; Theerakulpisut, Piyada

2017-08-15

Application of nanomaterials for agriculture is relatively new as compared to their use in biomedical and industrial sectors. In order to promote sustainable nanoagriculture, biocompatible silver nanoparticles (AgNPs) have been synthesized through green route using kaffir lime leaf extract for use as nanopriming agent for enhancing seed germination of rice aged seeds. Results of various characterization techniques showed the successful formation of AgNPs which were capped with phytochemicals present in the plant extract. Rice aged seeds primed with phytosynthesized AgNPs at 5 and 10 ppm significantly improved germination performance and seedling vigor compared to unprimed control, AgNO 3 priming, and conventional hydropriming. Nanopriming could enhance α-amylase activity, resulting in higher soluble sugar content for supporting seedlings growth. Furthermore, nanopriming stimulated the up-regulation of aquaporin genes in germinating seeds. Meanwhile, more ROS production was observed in germinating seeds of nanopriming treatment compared to unprimed control and other priming treatments, suggesting that both ROS and aquaporins play important roles in enhancing seed germination. Different mechanisms underlying nanopriming-induced seed germination were proposed, including creation of nanopores for enhanced water uptake, rebooting ROS/antioxidant systems in seeds, generation of hydroxyl radicals for cell wall loosening, and nanocatalyst for fastening starch hydrolysis.

CTCF orchestrates the germinal centre transcriptional program and prevents premature plasma cell differentiation

PubMed Central

Pérez-García, Arantxa; Marina-Zárate, Ester; Álvarez-Prado, Ángel F.; Ligos, Jose M.; Galjart, Niels; Ramiro, Almudena R.

2017-01-01

In germinal centres (GC) mature B cells undergo intense proliferation and immunoglobulin gene modification before they differentiate into memory B cells or long-lived plasma cells (PC). GC B-cell-to-PC transition involves a major transcriptional switch that promotes a halt in cell proliferation and the production of secreted immunoglobulins. Here we show that the CCCTC-binding factor (CTCF) is required for the GC reaction in vivo, whereas in vitro the requirement for CTCF is not universal and instead depends on the pathways used for B-cell activation. CTCF maintains the GC transcriptional programme, allows a high proliferation rate, and represses the expression of Blimp-1, the master regulator of PC differentiation. Restoration of Blimp-1 levels partially rescues the proliferation defect of CTCF-deficient B cells. Thus, our data reveal an essential function of CTCF in maintaining the GC transcriptional programme and preventing premature PC differentiation. PMID:28677680

Structure-activity relationship of karrikin germination stimulants.

PubMed

Flematti, Gavin R; Scaffidi, Adrian; Goddard-Borger, Ethan D; Heath, Charles H; Nelson, David C; Commander, Lucy E; Stick, Robert V; Dixon, Kingsley W; Smith, Steven M; Ghisalberti, Emilio L

2010-08-11

Karrikins (2H-furo[2,3-c]pyran-2-ones) are potent smoke-derived germination promoters for a diverse range of plant species but, to date, their mode of action remains unknown. This paper reports the structure-activity relationship of numerous karrikin analogues to increase understanding of the key structural features of the molecule that are required for biological activity. The results demonstrate that modification at the C5 position is preferred over modification at the C3, C4, or C7 positions for retaining the highest bioactivity.

Germination of Spores of Astrobiologically Relevant Bacillus Species in High-Salinity Environments.

PubMed

Nagler, Katja; Julius, Christina; Moeller, Ralf

2016-07-01

In times of increasing space exploration and search for extraterrestrial life, new questions and challenges for planetary protection, aiming to avoid forward contamination of different planets or moons with terrestrial life, are emerging. Spore-forming bacteria such as Bacillus species have a high contamination potential due to their spores' extreme resistance, enabling them to withstand space conditions. Spores require liquid water for their conversion into a growing cell (i.e., spore germination and subsequent growth). If present, water on extraterrestrial planets or moons is likely to be closely associated with salts (e.g., in salty oceans or brines), thus constituting high-salinity environments. Spores of Bacillus subtilis can germinate despite very high salt concentrations, although salt stress does exert negative effects on this process. In this study, germination and metabolic reactivation ("outgrowth") of spores of five astrobiologically relevant Bacillus species (B. megaterium, B. pumilus SAFR-032, B. nealsonii, B. mojavensis, and B. vallismortis) in high salinity (≤3.6 M NaCl) were investigated. Spores of different species exhibited different germination and outgrowth capabilities in high salinity, which strongly depended on germination conditions, especially the exact composition of the medium. In this context, a new "universal" germination trigger for Bacillus spores, named KAGE (KCl, L-alanine, D-glucose, ectoine), was identified, which will be very useful for future comparative germination and outgrowth studies on different Bacillus species. Overall, this study yielded interesting new insights on salt stress effects on spore germination and points out the difficulty of predicting the potential of spores to contaminate salty environments on extraterrestrial celestial bodies. Bacillus species-Spores-Germination-High salinity-Salt stress-NaCl-Inhibition. Astrobiology 16, 500-512.

Effects of nifedipine on gravi-dependent germination of moss spores

NASA Astrophysics Data System (ADS)

Khorkavtsiv, O. Y.; Demkiv, O. T.

Influence of gravity on germination of spores and dependence of the generation of a polar axis on a Ca2+ influx were investigated. The germination of spores does not depend on gravity but outgrowth polarity is controlled by light and gravity (Sytnik et al., 1989; Pundiak et al., 2001). We have shown that gravity determines the polarity of germination of spores and development of rhizoid and chloronemal outgrowths in both moss species -- Ceratodon purpureus and Pohlia nutans, the alignment of polar of germinating spores in C. purpureus, however, is less dependent on gravistimulus than in P. nutans. In 48 h after sowing onto culture medium+0,2% glucose in vertically oriented petri dishes in darkness spores of P. nutans germinated positively gravitropic rhizoid at the lower spore side and negatively gravitropic chloronema at the opposite one. The germination of C. purpureus spores is similar but the outgrowths show the lower level of alignment to the gravity vector than that of P. nutans, the dispersion of angles being 8,9 vs. 1,2 respectively. The cellular mechanism by which gravity acts remains unknown. The intracellular signaling Ca2+ ions play a crucial role in gravity perception and ability of a single cell to respond to gravity. We determined relative intensity of Ca2+ luminescence in the spores before their germination and at the early stages of outgrowth formation after treatment with the nifedipine and in a dependence on gravity vector. Gravity determined the position of outgrowth initiation zone and later on the growth direction of spore filaments. Treatment with nifedipine suppressed the gravity-directed calcium channel influx and distrupted polar growth of outgrowths. In experiments with calcium channel blocker sterilized spores were pregerminated on normal Knop's agar one day after were transferred to 50 μ M nifedipine just before emergence of the germ tube. After 48 h on nifedipine treatment, 50% spores did not germinate, 35% grew apolarily and in 15% of spores cell filaments oriented parallely with respect to the gravity vector. Results shown suggest that the endogenic competency of a single-cell spore is necessary condition of gravi- induced initiation of polar axis the competency being realized with Ca2+ movement. The highest level of Ca2+ luminescence was at the bottom of spores. In other sites of the spores the Ca2+ luminescence was about 20-fold lower than at the site of Ca2+ influx. In the 24 h after formation of first outgrowth the new site of Ca2+ influx appeared at the opposite site of spore and the second outgrowth arised. Consequently during the period of gravi-dependent spore development the newly top Ca2+ influx was repeatedly established. The direction of the Ca2+ ions influx correlated with re-orientation of spores with respect to the gravity vector. It is known that the nifedipine partially inhibits polar axis formation (Chatterjee et al., 2000) the latter being formed under the influence Ca2+ gradient (Cove, 2000). Thus, our results confirm that the fast change of Ca2+ influx probably is one of the earliest cell-level responses induced by gravity and it plays a key role in guiding polar events of germinating spores. This research was supported by NASA grant NN-09 (R).

Novel hydrated graphene ribbon unexpectedly promotes aged seed germination and root differentiation

NASA Astrophysics Data System (ADS)

Hu, Xiangang; Zhou, Qixing

2014-01-01

It is well known that graphene (G) induces nanotoxicity towards living organisms. Here, a novel and biocompatible hydrated graphene ribbon (HGR) unexpectedly promoted aged (two years) seed germination. HGR formed at the normal temperature and pressure (120 days hydration), presented 17.1% oxygen, 0.9% nitrogen groups, disorder-layer structure, with 0.38 nm thickness ribbon morphology. Interestingly, there were bulges around the edges of HGR. Compared to G and graphene oxide (GO), HGR increased seed germination by 15% root differentiation between 52 and 59% and enhanced resistance to oxidative stress. The metabonomics analysis discovered that HGR upregulated carbohydrate, amino acid, and fatty acids metabolism that determined secondary metabolism, nitrogen sequestration, cell membrane integrity, permeability, and oxidation resistance. Hexadecanoic acid as a biomarker promoted root differentiation and increased the germination rate. Our discovery is a novel HGR that promotes aged seed germination, illustrates metabolic specificity among graphene-based materials, and inspires innovative concepts in the regulation of seed development.

Physiological, cellular and molecular aspects of the desiccation tolerance in Anadenanthera colubrina seeds during germination.

PubMed

Castro, L E; Guimarães, C C; Faria, J M R

2017-11-01

During germination, orthodox seeds become gradually intolerant to desiccation, and for this reason, they are a good model for recalcitrance studies. In the present work, physiological, biochemical, and ultrastructural aspects of the desiccation tolerance were characterized during the germination process of Anadenanthera colubrina seeds. The seeds were imbibed during zero (control), 2, 8, 12 (no germinated seeds), and 18 hours (germinated seeds with 1 mm protruded radicle); then they were dried for 72 hours, rehydrated and evaluated for survivorship. Along the imbibition, cytometric and ultrastructural analysis were performed, besides the extraction of the heat-stable proteins. Posteriorly to imbibition and drying, the evaluation of ultrastructural damages was performed. Desiccation tolerance was fully lost after root protrusion. There was no increase in 4C DNA content after the loss of desiccation tolerance. Ultrastructural characteristics of cells from 1mm roots resembled those found in the recalcitrant seeds, in both hydrated and dehydrated states. The loss of desiccation tolerance coincided with the reduction of heat-stable proteins.

Seed Germination and Seedling Growth under Simulated Microgravity Causes Alterations in Plant Cell Proliferation and Ribosome Biogenesis

NASA Astrophysics Data System (ADS)

Matía, Isabel; van Loon, Jack W. A.; Carnero-Díaz, Eugénie; Marco, Roberto; Medina, Francisco Javier

2009-01-01

The study of the modifications induced by altered gravity in functions of plant cells is a valuable tool for the objective of the survival of terrestrial organisms in conditions different from those of the Earth. We have used the system "cell proliferation-ribosome biogenesis", two inter-related essential cellular processes, with the purpose of studying these modifications. Arabidopsis seedlings belonging to a transformed line containing the reporter gene GUS under the control of the promoter of the cyclin gene CYCB1, a cell cycle regulator, were grown in a Random Positioning Machine, a device known to accurately simulate microgravity. Samples were taken at 2, 4 and 8 days after germination and subjected to biometrical analysis and cellular morphometrical, ultrastructural and immunocytochemical studies in order to know the rates of cell proliferation and ribosome biogenesis, plus the estimation of the expression of the cyclin gene, as an indication of the state of cell cycle regulation. Our results show that cells divide more in simulated microgravity in a Random Positioning Machine than in control gravity, but the cell cycle appears significantly altered as early as 2 days after germination. Furthermore, higher proliferation is not accompanied by an increase in ribosome synthesis, as is the rule on Earth, but the functional markers of this process appear depleted in simulated microgravity-grown samples. Therefore, the alteration of the gravitational environmental conditions results in a considerable stress for plant cells, including those not specialized in gravity perception.

Hide, Keep Quiet, and Keep Low: Properties That Make Aspergillus fumigatus a Successful Lung Pathogen

PubMed Central

Escobar, Natalia; Ordonez, Soledad R.; Wösten, Han A. B.; Haas, Pieter-Jan A.; de Cock, Hans; Haagsman, Henk P.

2016-01-01

Representatives of the genus Aspergillus are opportunistic fungal pathogens. Their conidia can reach the alveoli by inhalation and can give rise to infections in immunocompromised individuals. Aspergillus fumigatus is the causal agent of invasive aspergillosis in nearly 90% of the cases. It is not yet well-established what makes this fungus more pathogenic than other aspergilli such as A. niger. Here, we show that A. fumigatus and A. niger conidia adhere with similar efficiency to lung epithelial A549 cells but A. fumigatus conidia internalized 17% more efficiently. Conidia of both aspergilli were taken up in phagolysosomes 8 h after the challenge. These organelles only acidified in the case of A. niger, which is probably due to the type of melanin coating of the conidia. Viability of both types of conidia was not affected after uptake in the phagolysosomes. Germination of A. fumigatus and A. niger conidia in the presence of epithelial cells was delayed when compared to conidia in the medium. However, germination of A. niger conidia was still higher than that of A. fumigatus 10 h after exposure to A549 cells. Remarkably, A. fumigatus hyphae grew mainly parallel to the epithelium, while growth direction of A. niger hyphae was predominantly perpendicular to the plane of the cells. Neutrophils reduced germination and hyphal growth of A. niger, but not of A fumigatus, in presence of epithelial cells. Taken together, efficient internalization, delayed germination, and hyphal growth parallel to the epithelium gives a new insight into what could be the causes for the success of A. fumigatus compared to A. niger as an opportunistic pathogen in the lung. PMID:27092115

Germination of Spores of Astrobiologically Relevant Bacillus Species in High-Salinity Environments

NASA Astrophysics Data System (ADS)

Nagler, Katja; Julius, Christina; Moeller, Ralf

2016-07-01

In times of increasing space exploration and search for extraterrestrial life, new questions and challenges for planetary protection, aiming to avoid forward contamination of different planets or moons with terrestrial life, are emerging. Spore-forming bacteria such as Bacillus species have a high contamination potential due to their spores' extreme resistance, enabling them to withstand space conditions. Spores require liquid water for their conversion into a growing cell (i.e., spore germination and subsequent growth). If present, water on extraterrestrial planets or moons is likely to be closely associated with salts (e.g., in salty oceans or brines), thus constituting high-salinity environments. Spores of Bacillus subtilis can germinate despite very high salt concentrations, although salt stress does exert negative effects on this process. In this study, germination and metabolic reactivation ("outgrowth") of spores of five astrobiologically relevant Bacillus species (B. megaterium, B. pumilus SAFR-032, B. nealsonii, B. mojavensis, and B. vallismortis) in high salinity (≤3.6 M NaCl) were investigated. Spores of different species exhibited different germination and outgrowth capabilities in high salinity, which strongly depended on germination conditions, especially the exact composition of the medium. In this context, a new "universal" germination trigger for Bacillus spores, named KAGE (KCl, L-alanine, D-glucose, ectoine), was identified, which will be very useful for future comparative germination and outgrowth studies on different Bacillus species. Overall, this study yielded interesting new insights on salt stress effects on spore germination and points out the difficulty of predicting the potential of spores to contaminate salty environments on extraterrestrial celestial bodies.

Transcription factor AtTCP14 regulates embryonic growth potential during seed germination in Arabidopsis thaliana.

PubMed

Tatematsu, Kiyoshi; Nakabayashi, Kazumi; Kamiya, Yuji; Nambara, Eiji

2008-01-01

To understand the molecular mechanisms underlying regulation of seed germination, we searched enriched cis elements in the upstream regions of Arabidopsis genes whose transcript levels increased during seed germination. Using available published microarray data, we found that two cis elements, Up1 or Up2, which regulate outgrowth of Arabidopsis axillary shoots, were significantly over-represented. Classification of Up1- and Up2-containing genes by gene ontology revealed that protein synthesis-related genes, especially ribosomal protein genes, were highly over-represented. Expression analysis using a reporter gene driven by a synthetic promoter regulated by these elements showed that the Up1 is necessary and sufficient for germination-associated gene induction, whereas Up2 acts as an enhancer of Up1. Up1-mediated gene expression was suppressed by treatments that blocked germination. Up1 is almost identical to the site II motif, which is the predicted target of TCP transcription factors. Of 24 AtTCP genes, AtTCP14, which showed the highest transcript level just prior to germination, was functionally characterized to test its involvement in the regulation of seed germination. Transposon-tagged lines for AtTCP14 showed delayed germination. In addition, germination of attcp14 mutants exhibited hypersensitivity to exogenously applied abscisic acid and paclobutrazol, an inhibitor of gibberellin biosynthesis. AtTCP14 was predominantly expressed in the vascular tissues of the embryo, and affected gene expression in radicles in a non-cell-autonomous manner. Taken together, these results indicate that AtTCP14 regulates the activation of embryonic growth potential in Arabidopsis seeds.

A pollen-specific calmodulin-binding protein, NPG1, interacts with putative pectate lyases.

PubMed

Shin, Sung-Bong; Golovkin, Maxim; Reddy, Anireddy S N

2014-06-12

Previous genetic studies have revealed that a pollen-specific calmodulin-binding protein, No Pollen Germination 1 (NPG1), is required for pollen germination. However, its mode of action is unknown. Here we report direct interaction of NPG1 with pectate lyase-like proteins (PLLs). A truncated form of AtNPG1 lacking the N-terminal tetratricopeptide repeat 1 (TPR1) failed to interact with PLLs, suggesting that it is essential for NPG1 interaction with PLLs. Localization studies with AtNPG1 fused to a fluorescent reporter driven by its native promoter revealed its presence in the cytosol and cell wall of the pollen grain and the growing pollen tube of plasmolyzed pollen. Together, our data suggest that the function of NPG1 in regulating pollen germination is mediated through its interaction with PLLs, which may modify the pollen cell wall and regulate pollen tube emergence and growth.

Development of Photodynamic Antimicrobial Chemotherapy (PACT) for Clostridium difficile.

PubMed

De Sordi, Luisa; Butt, M Adil; Pye, Hayley; Kohoutova, Darina; Mosse, Charles A; Yahioglu, Gokhan; Stamati, Ioanna; Deonarain, Mahendra; Battah, Sinan; Ready, Derren; Allan, Elaine; Mullany, Peter; Lovat, Laurence B

2015-01-01

Clostridium difficile is the leading cause of antibiotic-associated diarrhoea and pseudo membranous colitis in the developed world. The aim of this study was to explore whether Photodynamic Antimicrobial Chemotherapy (PACT) could be used as a novel approach to treating C. difficile infections. PACT utilises the ability of light-activated photosensitisers (PS) to produce reactive oxygen species (ROS) such as free radical species and singlet oxygen, which are lethal to cells. We screened thirteen PS against C. difficile planktonic cells, biofilm and germinating spores in vitro, and cytotoxicity of effective compounds was tested on the colorectal adenocarcinoma cell-line HT-29. Three PS were able to kill 99.9% of bacteria in both aerobic and anaerobic conditions, both in the planktonic state and in a biofilm, after exposure to red laser light (0.2 J/cm2) without harming model colon cells. The applicability of PACT to eradicate C. difficile germinative spores indirectly was also shown, by first inducing germination with the bile salt taurocholate, followed by PACT. This innovative and simple approach offers the prospect of a new antimicrobial therapy using light to treat C. difficile infection of the colon.

Aborted germinal center reactions and B cell memory by follicular T cells specific for a B cell receptor V region peptide.

PubMed

Heiser, Ryan A; Snyder, Christopher M; St Clair, James; Wysocki, Lawrence J

2011-07-01

A fundamental problem in immunoregulation is how CD4(+) T cells react to immunogenic peptides derived from the V region of the BCR that are created by somatic mechanisms, presented in MHC II, and amplified to abundance by B cell clonal expansion during immunity. BCR neo Ags open a potentially dangerous avenue of T cell help in violation of the principle of linked Ag recognition. To analyze this issue, we developed a murine adoptive transfer model using paired donor B cells and CD4 T cells specific for a BCR-derived peptide. BCR peptide-specific T cells aborted ongoing germinal center reactions and impeded the secondary immune response. Instead, they induced the B cells to differentiate into short-lived extrafollicular plasmablasts that secreted modest quantities of Ig. These results uncover an immunoregulatory process that restricts the memory pathway to B cells that communicate with CD4 T cells via exogenous foreign Ag.

Activation of the Endoplasmic Reticulum Stress-Associated Transcription Factor X Box-Binding Protein-1 Occurs in a Subset of Normal Germinal-Center B Cells and in Aggressive B-Cell Lymphomas with Prognostic Implications

PubMed Central

Balague, Olga; Mozos, Ana; Martinez, Daniel; Hernandez, Luis; Colomo, Lluis; Mate, Jose Luis; Teruya-Feldstein, Julie; Lin, Oscar; Campo, Elias; Lopez-Guillermo, Armando; Martinez, Antonio

2009-01-01

X box-binding protein 1 (Xbp-1) is a transcription factor that is required for the terminal differentiation of B lymphocytes into plasma cells. The Xbp-1 gene is activated in response to endoplasmic reticulum stress signals, which generate a 50-kDa nuclear protein that acts as a potent transactivator and regulates the expression of genes related to the unfolded protein response. Activated Xbp-1 is essential for cell survival in plasma-cell tumors but its role in B-cell lymphomas is unknown. We analyzed the expression of activated Xbp-1 in reactive lymphoid tissues, 411 lymphomas and plasma-cell neoplasms, and 24 B-cell lines. In reactive tissues, Xbp-1 was only found in nuclear extracts. Nuclear expression of Xbp-1 was observed in occasional reactive plasma cells and in a subpopulation of Irf-4+/Bcl-6−/Pax-5− B cells in the light zones of reactive germinal centers, probably representing cells committed to plasma-cell differentiation. None of the low-grade lymphomas showed evidence of Xbp-1 activation; however, Xbp-1 activation was found in 28% of diffuse large B-cell lymphomas, independent of germinal or postgerminal center phenotype, as well as in 48% of plasmablastic lymphomas and 69% of plasma-cell neoplasms. Diffuse large B-cell lymphomas with nuclear Xbp-1 expression had a significantly worse response to therapy and shorter overall survival compared with negative tumors. These findings suggest that Xbp-1 activation may play a role in the pathogenesis of aggressive B-cell lymphomas. PMID:19389935

Precision medicine and lymphoma.

PubMed

Heward, James A; Kumar, Emil A; Korfi, Koorosh; Okosun, Jessica; Fitzgibbon, Jude

2018-05-05

The treatment of the germinal center lymphomas, diffuse large B cell (DLBCL) and follicular lymphoma, has changed little beyond the introduction of immunochemotherapies. However, there exists a substantial group of patients within both diseases for which improvements in care will involve appropriate tailoring of treatment. DLBCL consists of two major subtypes with striking differences in their clinical outcomes paralleling their underlying genetic heterogeneity. Recent studies have seen advances in the stratification of germinal center lymphomas, through comprehensive profiling of 1001 DLBCLs alongside refinements in the identification of high-risk follicular lymphoma patients using m7-FLIPI and 23G models. A new wave of novel therapeutic agents is now undergoing clinical trials for germinal center lymphomas, with BCR and EZH2 inhibitors demonstrating preferential benefit in subgroups of patients. The emergence of cell-free DNA has raised the possibility of dynamic disease monitoring to potentially mitigate the complexity of spatial and temporal heterogeneity, whilst predicting tumor evolution in real time. Altogether knowledge of the genomic landscape of germinal center lymphomas is offering welcome opportunities in patient risk stratification and therapeutics. The challenge ahead is to establish how best to combine upfront or dynamic prognostication with precision therapies, while retaining practicality in clinical trials and the real-world setting.

Bioherbicides: Current knowledge on weed control mechanism.

PubMed

Radhakrishnan, Ramalingam; Alqarawi, Abdulaziz A; Abd Allah, Elsayed Fathi

2018-04-17

Weed control is a challenging event during crop cultivation. Integrated management, including the application of bioherbicides, is an emerging method for weed control in sustainable agriculture. Plant extracts, allelochemicals and some microbes are utilized as bioherbicides to control weed populations. Bioherbicides based on plants and microbes inhibit the germination and growth of weeds; however,few studies conducted in weed physiology. This review ascribes the current knowledge of the physiological changes in weeds that occur during the exposure to bioherbicides. Plant extracts or metabolites are absorbed by weed seeds, which initiates damage to the cell membrane, DNA, mitosis, amylase activity and other biochemical processes and delays or inhibits seed germination. The growth of weeds is also retarded due to low rates of root-cell division, nutrient uptake, photosynthetic pigment synthesis, and plant growth hormone synthesis, while the productions of reactive oxygen species (ROS) and stress-mediated hormones increase, including irregular antioxidant activity. However, lytic enzymes and toxic substances secreted from microbes degrade the weed seed coat and utilize the endosperm for survival, which inhibits seed germination. The microbes grow through the intercellular spaces to reach the root core, and the deposition of toxins in the cells affects cell division and cellular functions. Some of the metabolites of deleterious microbes cause disease, necrosis and chlorosis,which inhibit the germination and growth of weed seeds by suppressing photosynthesis and gibberellin activities and enhancing ROS, abscisic acid and ethylene. This review explains the effects of bioherbicides (derived from plants and microbes) on weed-plant physiology to elucidate their modes of action. Copyright © 2018 Elsevier Inc. All rights reserved.

Exploring the Role of Cell Wall-Related Genes and Polysaccharides during Plant Development.

PubMed

Tucker, Matthew R; Lou, Haoyu; Aubert, Matthew K; Wilkinson, Laura G; Little, Alan; Houston, Kelly; Pinto, Sara C; Shirley, Neil J

2018-05-31

The majority of organs in plants are not established until after germination, when pluripotent stem cells in the growing apices give rise to daughter cells that proliferate and subsequently differentiate into new tissues and organ primordia. This remarkable capacity is not only restricted to the meristem, since maturing cells in many organs can also rapidly alter their identity depending on the cues they receive. One general feature of plant cell differentiation is a change in cell wall composition at the cell surface. Historically, this has been viewed as a downstream response to primary cues controlling differentiation, but a closer inspection of the wall suggests that it may play a much more active role. Specific polymers within the wall can act as substrates for modifications that impact receptor binding, signal mobility, and cell flexibility. Therefore, far from being a static barrier, the cell wall and its constituent polysaccharides can dictate signal transmission and perception, and directly contribute to a cell's capacity to differentiate. In this review, we re-visit the role of plant cell wall-related genes and polysaccharides during various stages of development, with a particular focus on how changes in cell wall machinery accompany the exit of cells from the stem cell niche.

Glycosylated chicken ZP2 accumulates in the egg coat of immature oocytes and remains localized to the germinal disc region of mature eggs.

PubMed

Nishio, Shunsuke; Kohno, Yoshinori; Iwata, Yuki; Arai, Mayumi; Okumura, Hiroki; Oshima, Kenzi; Nadano, Daita; Matsuda, Tsukasa

2014-11-01

Vertebrate eggs are surrounded by an egg coat, which is a specific extracellular egg matrix consisting of several glycoproteins with a conserved zona pellucida (ZP) domain. Two mammalian egg coat subunits, ZP2 and ZP3, have been suggested to act as sperm receptors. In bird eggs, however, ZP2 has never been identified in the egg coat of mature oocytes and ovulated eggs. Here we report that chicken ZP2 is expressed in immature small follicles and remains as an egg-coat component locally in the germinal disc region of mature eggs. RT-PCR analysis indicated marked expression of the ZP2 and ZP4 genes in the granulosa cells of immature white follicles, whereas the ZP3 and ZPD genes showed marked expression in the cells of maturing yellow follicles. ZP2 was identified in the egg coat isolated from immature follicles as a heavily N-glycosylated glycoprotein of ∼200 kDa, which was enzymatically converted to a 70-kDa deglycosylated form. Immunoblotting and immunohistological analyses showed that ZP2 was localized around the germinal disc region of mature follicles. ZP2 was accumulated in the egg coat of immature white follicles at the earlier stages of oocyte development and became a minor component in the egg coat of maturing yellow follicles, except for the germinal disc region. Localization of ZP2 in the germinal disc region of mature eggs, where sperm bind to the egg coat at high density, suggests some role for ZP2 in the preferential binding and penetration of sperm in the germinal disc region of bird eggs. © 2014 by the Society for the Study of Reproduction, Inc.

A 3D intestinal tissue model supports Clostridioides difficile germination, colonization, toxin production and epithelial damage.

PubMed

Shaban, Lamyaa; Chen, Ying; Fasciano, Alyssa C; Lin, Yinan; Kaplan, David L; Kumamoto, Carol A; Mecsas, Joan

2018-04-01

Endospore-forming Clostridioides difficile is a causative agent of antibiotic-induced diarrhea, a major nosocomial infection. Studies of its interactions with mammalian tissues have been hampered by the fact that C. difficile requires anaerobic conditions to survive after spore germination. We recently developed a bioengineered 3D human intestinal tissue model and found that low O 2 conditions are produced in the lumen of these tissues. Here, we compared the ability of C. difficile spores to germinate, produce toxin and cause tissue damage in our bioengineered 3D tissue model versus in a 2D transwell model in which human cells form a polarized monolayer. 3D tissue models or 2D polarized monolayers on transwell filters were challenged with the non-toxin producing C. difficile CCUG 37787 serotype X (ATCC 43603) and the toxin producing UK1 C. difficile spores in the presence of the germinant, taurocholate. Spores germinated in both the 3D tissue model as well as the 2D transwell system, however toxin activity was significantly higher in the 3D tissue models compared to the 2D transwells. Moreover, the epithelium damage in the 3D tissue model was significantly more severe than in 2D transwells and damage correlated significantly with the level of toxin activity detected but not with the amount of germinated spores. Combined, these results show that the bioengineered 3D tissue model provides a powerful system with which to study early events leading to toxin production and tissue damage of C. difficile with mammalian cells under anaerobic conditions. Furthermore, these systems may be useful for examining the effects of microbiota, novel drugs and other potential therapeutics directed towards C. difficile infections. Copyright © 2018 Elsevier Ltd. All rights reserved.

Effects of moist cold stratification on germination, plant growth regulators, metabolites and embryo ultrastructure in seeds of Acer morrisonense (Sapindaceae).

PubMed

Chen, Shun-Ying; Chou, Shih-Han; Tsai, Ching-Chu; Hsu, Wen-Yu; Baskin, Carol C; Baskin, Jerry M; Chien, Ching-Te; Kuo-Huang, Ling-Long

2015-09-01

Breaking of seed dormancy by moist cold stratification involves complex interactions in cells. To assess the effect of moist cold stratification on dormancy break in seeds of Acer morrisonense, we monitored percentages and rates of germination and changes in plant growth regulators, sugars, amino acids and embryo ultrastructure after various periods of cold stratification. Fresh seeds incubated at 25/15 °C for 24 weeks germinated to 61%, while those cold stratified at 5 °C for 12 weeks germinated to 87% in 1 week. Neither exogenous GA3 nor GA4 pretreatment significantly increased final seed germination percentage. Total ABA content of seeds cold stratified for 12 weeks was reduced about 3.3-fold, to a concentration similar to that in germinated seeds (radicle emergence). Endogenous GA3 and GA7 were detected in 8-week and 12-week cold stratified seeds but not in fresh seeds. Numerous protein and lipid bodies were present in the plumule, first true leaves and cotyledons of fresh seeds. Protein and lipid bodies decreased greatly during cold stratification, and concentrations of total soluble sugars and amino acids increased. The major non-polar sugars in fresh seeds were sucrose and fructose, but sucrose increased and fructose decreased significantly during cold stratification. The major free amino acids were proline and tryptophan in fresh seeds, and proline increased and tryptophan decreased during cold stratification. Thus, as dormancy break occurs during cold stratification seeds of A. morrisonense undergo changes in plant growth regulators, proteins, lipids, sugars, amino acids and cell ultrastructure. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Isolation of tissues and preservation of RNA from intact, germinated barley grain.

PubMed

Betts, Natalie S; Berkowitz, Oliver; Liu, Ruijie; Collins, Helen M; Skadhauge, Birgitte; Dockter, Christoph; Burton, Rachel A; Whelan, James; Fincher, Geoffrey B

2017-08-01

Isolated barley (Hordeum vulgare L.) aleurone layers have been widely used as a model system for studying gene expression and hormonal regulation in germinating cereal grains. A serious technological limitation of this approach has been the inability to confidently extrapolate conclusions obtained from isolated tissues back to the whole grain, where the co-location of several living and non-living tissues results in complex tissue-tissue interactions and regulatory pathways coordinated across the multiple tissues. Here we have developed methods for isolating fragments of aleurone, starchy endosperm, embryo, scutellum, pericarp-testa, husk and crushed cell layers from germinated grain. An important step in the procedure involves the rapid fixation of the intact grain to freeze the transcriptional activity of individual tissues while dissection is effected for subsequent transcriptomic analyses. The developmental profiles of 19 611 gene transcripts were precisely defined in the purified tissues and in whole grain during the first 24 h of germination by RNA sequencing. Spatial and temporal patterns of transcription were validated against well-defined data on enzyme activities in both whole grain and isolated tissues. Transcript profiles of genes involved in mitochondrial assembly and function were used to validate the very early stages of germination, while the profiles of genes involved in starch and cell wall mobilisation matched existing data on activities of corresponding enzymes. The data will be broadly applicable for the interrogation of co-expression and differential expression patterns and for the identification of transcription factors that are important in the early stages of grain and seed germination. © 2017 The Authors The Plant Journal © 2017 John Wiley & Sons Ltd.

MAPK1 is required for establishing the pattern of cell proliferation and for cell survival during lens development

PubMed Central

Upadhya, Dinesh; Ogata, Masato; Reneker, Lixing W.

2013-01-01

The mitogen-activated protein kinases (MAPKs; also known as ERKs) are key intracellular signaling molecules that are ubiquitously expressed in tissues and were assumed to be functionally equivalent. Here, we use the mouse lens as a model system to investigate whether MAPK1 plays a specific role during development. MAPK3 is known to be dispensable for lens development. We demonstrate that, although MAPK1 is uniformly expressed in the lens epithelium, its deletion significantly reduces cell proliferation in the peripheral region, an area referred to as the lens germinative zone in which most active cell division occurs during normal lens development. By contrast, cell proliferation in the central region is minimally affected by MAPK1 deletion. Cell cycle regulators, including cyclin D1 and survivin, are downregulated in the germinative zone of the MAPK1-deficient lens. Interestingly, loss of MAPK1 subsequently induces upregulation of phosphorylated MAPK3 (pMAPK3) levels in the lens epithelium; however, this increase in pMAPK3 is not sufficient to restore cell proliferation in the germinative zone. Additionally, MAPK1 plays an essential role in epithelial cell survival but is dispensable for fiber cell differentiation during lens development. Our data indicate that MAPK1/3 control cell proliferation in the lens epithelium in a spatially defined manner; MAPK1 plays a unique role in establishing the highly mitotic zone in the peripheral region, whereas the two MAPKs share a redundant role in controlling cell proliferation in the central region of the lens epithelium. PMID:23482492

Primary central nervous system diffuse large B-cell lymphoma in the immunocompetent: Immunophenotypic subtypes and Epstein-Barr virus association.

PubMed

Mahadevan, Anita; Rao, Clementina Rama; Shanmugham, M; Shankar, Susarla Krishna

2015-01-01

Primary central nervous system diffuse large B-cell lymphoma (PCNSL DLBCL) in the immunocompetent is an uncommon tumor that has an activated B-cell immunophenotype resembling germinal center exit B cells. They also differ from primary central nervous diffuse large B-cell lymphomas in the immunocompromised as they show no association with the Epstein-Barr virus. To determine if immunophenotypic subtyping of PCNS DLBCL from Asian subcontinent are also different similar to its systemic counterpart is unclear, as there are only limited studies from Asia, and none from India. The immunohistochemical profile of 24 South Indian patients with primary central nervous system diffuse large B-cell lymphoma was studied using germinal center markers - CD10 and Bcl-6, and activation markers - MUM1 and CD138, which are markers for late/post germinal centre B cells. Insitu hybridization for EBV genome and LMP1 by immunohistochemistry was carried out in all cases to determine association with EBV. Centroblastic morphology and uniform activated B-cell phenotype with positivity for MUM1 was seen in 91.6% of tumors. Co-expression of Bcl-6 and MUM1 was evident in 50%, which is more frequent than in systemic diffuse large B-cell lymphomas. All cases were negative for Epstein-Barr virus using EBER in-situ hybridization and LMP1 immunohistochemistry. Primary diffuse large B-cell lymphoma in the immunocompetent is a distinct clinicopathological entity with centroblastic morphology, a uniform activated B-cell immunophenotype that is not associated with the Epstein-Barr virus regardless of geographic origin.

Cytokinins and Expression of SWEET, SUT, CWINV and AAP Genes Increase as Pea Seeds Germinate

PubMed Central

Jameson, Paula E.; Dhandapani, Pragatheswari; Novak, Ondrej; Song, Jiancheng

2016-01-01

Transporter genes and cytokinins are key targets for crop improvement. These genes are active during the development of the seed and its establishment as a strong sink. However, during germination, the seed transitions to being a source for the developing root and shoot. To determine if the sucrose transporter (SUT), amino acid permease (AAP), Sugar Will Eventually be Exported Transporter (SWEET), cell wall invertase (CWINV), cytokinin biosynthesis (IPT), activation (LOG) and degradation (CKX) gene family members are involved in both the sink and source activities of seeds, we used RT-qPCR to determine the expression of multiple gene family members, and LC-MS/MS to ascertain endogenous cytokinin levels in germinating Pisum sativum L. We show that genes that are actively expressed when the seed is a strong sink during its development, are also expressed when the seed is in the reverse role of being an active source during germination and early seedling growth. Cytokinins were detected in the imbibing seeds and were actively biosynthesised during germination. We conclude that, when the above gene family members are targeted for seed yield improvement, a downstream effect on subsequent seed germination or seedling vigour must be taken into consideration. PMID:27916945

Interference of the Histone Deacetylase Inhibits Pollen Germination and Pollen Tube Growth in Picea wilsonii Mast

PubMed Central

Zhou, Junhui; Li, Xiaojuan

2015-01-01

Histone deacetylase (HDAC) is a crucial component in the regulation of gene expression in various cellular processes in animal and plant cells. HDAC has been reported to play a role in embryogenesis. However, the effect of HDAC on androgamete development remains unclear, especially in gymnosperms. In this study, we used the HDAC inhibitors trichostatin A (TSA) and sodium butyrate (NaB) to examine the role of HDAC in Picea wilsonii pollen germination and pollen tube elongation. Measurements of the tip-focused Ca2+ gradient revealed that TSA and NaB influenced this gradient. Immunofluorescence showed that actin filaments were disrupted into disorganized fragments. As a result, the vesicle trafficking was disturbed, as determined by FM4-64 labeling. Moreover, the distribution of pectins and callose in cell walls was significantly altered in response to TSA and NaB. Our results suggest that HDAC affects pollen germination and polarized pollen tube growth in Picea wilsonii by affecting the intracellular Ca2+ concentration gradient, actin organization patterns, vesicle trafficking, as well as the deposition and configuration of cell wall components. PMID:26710276

Evaluating novel synthetic compounds active against Bacillus subtilis and Bacillus cereus spores using Live imaging with SporeTrackerX.

PubMed

Omardien, Soraya; Ter Beek, Alexander; Vischer, Norbert; Montijn, Roy; Schuren, Frank; Brul, Stanley

2018-06-14

An empirical approach was taken to screen a novel synthetic compound library designed to be active against Gram-positive bacteria. We obtained five compounds that were active against spores from the model organism Bacillus subtilis and the food-borne pathogen Bacillus cereus during our population based experiments. Using single cell live imaging we were able to observe effects of the compounds on spore germination and outgrowth. Difference in sensitivity to the compounds could be observed between B. subtilis and B. cereus using live imaging, with minor difference in the minimal inhibitory and bactericidal concentrations of the compounds against the spores. The compounds all delayed the bursting time of germinated spores and affected the generation time of vegetative cells at sub-inhibitory concentrations. At inhibitory concentrations spore outgrowth was prevented. One compound showed an unexpected potential for preventing spore germination at inhibitory concentrations, which merits further investigation. Our study shows the valuable role single cell live imaging can play in the final selection process of antimicrobial compounds.

Evaluation of Chromosomal Instability in Diabetic Rats Treated with Naringin

PubMed Central

A. Bakheet, Saleh; M. Attia, Sabry

2011-01-01

We used the bone marrow DNA strand breaks, micronucleus formations, spermatocyte chromosomal aberrations, and sperm characteristic assays to investigate the chromosomal instability in somatic and germinal cells of diabetic rats treated with multiple doses of naringin. The obtained results revealed that naringin was neither cytotoxic nor genotoxic for the rats at all tested doses. Moreover, naringin significantly reduced the diabetes-induced chromosomal instability in somatic and germinal cells in a dose-dependent manner. In addition, diabetes induced marked biochemical alterations characteristic of oxidative stress including enhanced lipid peroxidation, accumulation of oxidized glutathione, reduction in reduced glutathione, and accumulation of intracellular reactive oxygen species. Treatment with naringin ameliorated these biochemical markers dose-dependently. In conclusion, naringin confers an appealing protective effect against diabetes-induced chromosomal instability towards rat somatic and germinal cells which might be explained partially via diminishing the de novo free radical generation induced by hyperglycemia. Thus, naringin might be a good candidate to reduce genotoxic risk associated with hyperglycemia and may provide decreases in the development of secondary malignancy and abnormal reproductive outcomes risks, which seems especially important for diabetic patients. PMID:21941606

γδ T cells affect IL-4 production and B-cell tolerance

PubMed Central

Huang, Yafei; Heiser, Ryan A.; Detanico, Thiago O.; Getahun, Andrew; Kirchenbaum, Greg A.; Casper, Tamara L.; Aydintug, M. Kemal; Carding, Simon R.; Ikuta, Koichi; Huang, Hua; Cambier, John C.; Wysocki, Lawrence J.; O’Brien, Rebecca L.; Born, Willi K.

2015-01-01

γδ T cells can influence specific antibody responses. Here, we report that mice deficient in individual γδ T-cell subsets have altered levels of serum antibodies, including all major subclasses, sometimes regardless of the presence of αβ T cells. One strain with a partial γδ deficiency that increases IgE antibodies also displayed increases in IL-4–producing T cells (both residual γδ T cells and αβ T cells) and in systemic IL-4 levels. Its B cells expressed IL-4–regulated inhibitory receptors (CD5, CD22, and CD32) at diminished levels, whereas IL-4–inducible IL-4 receptor α and MHCII were increased. They also showed signs of activation and spontaneously formed germinal centers. These mice displayed IgE-dependent features found in hyper-IgE syndrome and developed antichromatin, antinuclear, and anticytoplasmic autoantibodies. In contrast, mice deficient in all γδ T cells had nearly unchanged Ig levels and did not develop autoantibodies. Removing IL-4 abrogated the increases in IgE, antichromatin antibodies, and autoantibodies in the partially γδ-deficient mice. Our data suggest that γδ T cells, controlled by their own cross-talk, affect IL-4 production, B-cell activation, and B-cell tolerance. PMID:25535377

γδ T cells affect IL-4 production and B-cell tolerance.

PubMed

Huang, Yafei; Heiser, Ryan A; Detanico, Thiago O; Getahun, Andrew; Kirchenbaum, Greg A; Casper, Tamara L; Aydintug, M Kemal; Carding, Simon R; Ikuta, Koichi; Huang, Hua; Cambier, John C; Wysocki, Lawrence J; O'Brien, Rebecca L; Born, Willi K

2015-01-06

γδ T cells can influence specific antibody responses. Here, we report that mice deficient in individual γδ T-cell subsets have altered levels of serum antibodies, including all major subclasses, sometimes regardless of the presence of αβ T cells. One strain with a partial γδ deficiency that increases IgE antibodies also displayed increases in IL-4-producing T cells (both residual γδ T cells and αβ T cells) and in systemic IL-4 levels. Its B cells expressed IL-4-regulated inhibitory receptors (CD5, CD22, and CD32) at diminished levels, whereas IL-4-inducible IL-4 receptor α and MHCII were increased. They also showed signs of activation and spontaneously formed germinal centers. These mice displayed IgE-dependent features found in hyper-IgE syndrome and developed antichromatin, antinuclear, and anticytoplasmic autoantibodies. In contrast, mice deficient in all γδ T cells had nearly unchanged Ig levels and did not develop autoantibodies. Removing IL-4 abrogated the increases in IgE, antichromatin antibodies, and autoantibodies in the partially γδ-deficient mice. Our data suggest that γδ T cells, controlled by their own cross-talk, affect IL-4 production, B-cell activation, and B-cell tolerance.

Akinetes May Be Representative of Past Nostocalean Blooms: a Case Study of Their Benthic Spatiotemporal Distribution and Potential for Germination in a Eutrophic Lake

PubMed Central

Le Jeune, Anne-Hélène; Colombet, Jonathan; Thouvenot, Antoine; Latour, Delphine

2017-01-01

ABSTRACT Monitoring of water and surface sediment in a French eutrophic lake (Lake Aydat) was carried out over a 2-year period in order to determine whether akinetes in sediment could be representative of the most recent bloom and to estimate their germination potential. Sediment analysis revealed two akinete species, Dolichospermum macrosporum and Dolichospermum flos-aquae, present in the same proportions as observed for the pelagic populations. Moreover, similar spatial patterns observed for vegetative cells in the water column and akinete distributions in the sediment suggest that akinetes in the sediment may be representative of the previous bloom. However, the relationship between akinetes in the sediment and vegetative cells in the water column was not linear, and other factors may interfere. For example, our results highlighted horizontal transport of akinetes during the winter. The benthic overwinter phase did not seem to influence the percentages of intact akinetes, which remained stable at approximately 7% and 60% for D. macrosporum and D. flos-aquae, respectively. These percentages may thus be the result of processes that occurred in the water column. The intact overwintering akinetes showed germination rates of up to 90% after 72 h for D. flos-aquae or 144 h for D. macrosporum. The difference in akinete germination rates between these two species demonstrates different ecological strategies, which serve to expand the window for germination in time and space and thus optimize colonization of the water column by nostocalean cyanobacteria. IMPORTANCE Cyanobacteria have the ability to proliferate and to form blooms. These blooms can then affect the local ecology, health, and economy. The akinete, a resistant cell type that persists in sediment, is an important intermediate phase between previous and future blooms. We monitored the water column and the surface sediment of a French eutrophic lake (Lake Aydat) to investigate the relationship between vegetative cells in the water column and akinetes in the sediment. This study focused on the characterization of spatiotemporal akinete distributions, cellular integrity, and germination potential. Species-specific ecological strategies were highlighted and may partly explain the temporal succession of species in the water column. Akinetes may also be used to understand past nostocalean blooms and to predict future ones. PMID:28970224

Green synthesis of gold nanoparticles using a cheap Sphaeranthus indicus extract: Impact on plant cells and the aquatic crustacean Artemia nauplii.

PubMed

Balalakshmi, Chinnasamy; Gopinath, Kasi; Govindarajan, Marimuthu; Lokesh, Ravi; Arumugam, Ayyakannu; Alharbi, Naiyf S; Kadaikunnan, Shine; Khaled, Jamal M; Benelli, Giovanni

2017-08-01

The impact of green-fabricated gold nanoparticles on plant cells and non-target aquatic species is scarcely studied. In this research, we reported an environment friendly technique for the synthesis of gold nanoparticles (Au NPs) using the Sphaeranthus indicus leaf extract. The formation of the metal NPs was characterized by UV-Visible and FT-IR spectroscopy, XRD, SEM and TEM analyses. The UV-Visible spectra of Au NPs showed a surface plasmon resonance peak at 531nm. FT-IR analysis indicated functional bio-molecules associated with Au NPs formation. The crystalline nature of Au nanoparticles was confirmed by their XRD diffraction pattern. TEM revealed the spherical shape with a mean particle size of 25nm. Au NPs was tested at 0, 1, 3, 5, 7 and 10% doses in mitotic cell division assays, pollen germination experiments, and in vivo toxicity trials against the aquatic crustacean Artemia nauplii. Au NPs did not show any toxic effects on plant cells and aquatic invertebrates. Notably, Au NPs promoted mitotic cell division in Allium cepa root tip cells and germination of Gloriosa superba pollen grains. Au NPs showed no mortality on A. nauplii, all the tested animals showed 100% survivability. Therefore, these Au NPs have potential applications in the development of pollen germination media and plant tissue culture. Copyright © 2017 Elsevier B.V. All rights reserved.

Novel population of small tumour-initiating stem cells in the ovaries of women with borderline ovarian cancer

PubMed Central

Virant-Klun, Irma; Stimpfel, Martin

2016-01-01

Small stem cells with diameters of up to 5 μm previously isolated from adult human ovaries indicated pluripotency and germinal lineage, especially primordial germ cells, and developed into primitive oocyte-like cells in vitro. Here, we show that a comparable population of small stem cells can be found in the ovarian tissue of women with borderline ovarian cancer, which, in contrast to small stem cells in “healthy” ovaries, formed spontaneous tumour-like structures and expressed some markers related to pluripotency and germinal lineage. The gene expression profile of these small putative cancer stem cells differed from similar cells sorted from “healthy” ovaries by 132 upregulated and 97 downregulated genes, including some important forkhead box and homeobox genes related to transcription regulation, developmental processes, embryogenesis, and ovarian cancer. These putative cancer stem cells are suggested to be a novel population of ovarian tumour-initiating cells in humans. PMID:27703207

Germination and Outgrowth of Single Spores of Saccharomyces cerevisiae Viewed by Scanning Electron and Phase-Contrast Microscopy

PubMed Central

Rousseau, Paul; Halvorson, Harlyn O.; Bulla, Lee A.; Julian, Grant St.

1972-01-01

Single spores of Saccharomyces cerevisiae were examined during germination and outgrowth by scanning electron and phase-contrast microscopy. Also determined were changes in cell weight and light absorbance, trehalose utilization, and synthesis of protein and KOH-soluble carbohydrates. These studies reveal that development of the vegetative cell from a spore follows a definite sequence of events involving dramatic physical and chemical modifications. These changes are: initial rapid loss in cellular absorbance followed later by an abrupt gain in absorbance; reduction in cell weight and a subsequent progressive increase; modification of the spore surface with concomitant diminution in refractility; elongation of the cell and augmentation of surface irregularities; rapid decline in trehalose content of the cell accompanied by extensive formation of KOH-soluble carbohydrates; and bud formation. Images PMID:4551750

Early carbon mobilization and radicle protrusion in maize germination.

PubMed

Sánchez-Linares, Luis; Gavilanes-Ruíz, Marina; Díaz-Pontones, David; Guzmán-Chávez, Fernando; Calzada-Alejo, Viridiana; Zurita-Villegas, Viridiana; Luna-Loaiza, Viridiana; Moreno-Sánchez, Rafael; Bernal-Lugo, Irma; Sánchez-Nieto, Sobeida

2012-07-01

Considerable amounts of information is available on the complex carbohydrates that are mobilized and utilized by the seed to support early seedling development. These events occur after radicle has protruded from the seed. However, scarce information is available on the role of the endogenous soluble carbohydrates from the embryo in the first hours of germination. The present work analysed how the soluble carbohydrate reserves in isolated maize embryos are mobilized during 6-24 h of water imbibition, an interval that exclusively embraces the first two phases of the germination process. It was found that sucrose constitutes a very significant reserve in the scutellum and that it is efficiently consumed during the time in which the adjacent embryo axis is engaged in an active metabolism. Sucrose transporter was immunolocalized in the scutellum and in vascular elements. In parallel, a cell-wall invertase activity, which hydrolyses sucrose, developed in the embryo axis, which favoured higher glucose uptake. Sucrose and hexose transporters were active in the embryo tissues, together with the plasma membrane H(+)-ATPase, which was localized in all embryo regions involved in both nutrient transport and active cell elongation to support radicle extension. It is proposed that, during the initial maize germination phases, a net flow of sucrose takes place from the scutellum towards the embryo axis and regions that undergo elongation. During radicle extension, sucrose and hexose transporters, as well as H(+)-ATPase, become the fundamental proteins that orchestrate the transport of nutrients required for successful germination.

Embryo growth, testa permeability, and endosperm weakening are major targets for the environmentally regulated inhibition of Lepidium sativum seed germination by myrigalone A

PubMed Central

2012-01-01

Myrigalone A (MyA) is a rare flavonoid in fruit leachates of Myrica gale, a deciduous shrub adapted to flood-prone habitats. As a putative allelochemical it inhibits seed germination and seedling growth. Using Lepidium sativum as a model target species, experiments were conducted to investigate how environmental cues modulate MyA’s interference with key processes of seed germination. Time course analyses of L. sativum testa and endosperm rupture under different light conditions and water potentials were combined with quantifying testa permeability, endosperm weakening, tissue-specific gibberellin (GA) and abscisic acid (ABA) contents, as well as embryo growth and apoplastic superoxide production important for cell expansion growth. Lepidium sativum testa permeability and early water uptake by imbibition is enhanced by MyA. During late germination, MyA inhibits endosperm weakening and embryo growth, both processes required for endosperm rupture. Inhibition of embryo cell expansion by MyA depends on environmental cues, which is evident from the light-modulated severity of the MyA-mediated inhibition of apoplastic superoxide accumulation. Several important key weakening and growth processes during early and late germination are targets for MyA. These effects are modulated by light conditions and ambient water potential. It is speculated that MyA is a soil seed bank-destroying allelochemical that secures the persistence of M. gale in its flood-prone environment. PMID:22821938

Conidial germination in Scedosporium apiospermum, S. aurantiacum, S. minutisporum and Lomentospora prolificans: influence of growth conditions and antifungal susceptibility profiles.

PubMed

Mello, Thaís Pereira de; Aor, Ana Carolina; Oliveira, Simone Santiago Carvalho de; Branquinha, Marta Helena; Santos, André Luis Souza Dos

2016-06-27

In the present study, we have investigated some growth conditions capable of inducing the conidial germination in Scedosporium apiospermum, S. aurantiacum, S. minutisporum and Lomentospora prolificans. Germination in Sabouraud medium (pH 7.0, 37ºC, 5% CO2) showed to be a typically time-dependent event, reaching ~75% in S. minutisporum and > 90% in S. apiospermum, S. aurantiacum and L. prolificans after 4 h. Similar germination rate was observed when conidia were incubated under different media and pHs. Contrarily, temperature and CO2 tension modulated the germination. The isotropic conidial growth (swelling) and germ tube-like projection were evidenced by microscopy and cytometry. Morphometric parameters augmented in a time-dependent fashion, evidencing changes in size and granularity of fungal cells compared with dormant 0 h conidia. In parallel, a clear increase in the mitochondrial activity was measured during the transformation of conidia-into-germinated conidia. Susceptibility profiles to itraconazole, fluconazole, voriconazole, amphotericin B and caspofungin varied regarding each morphotype and each fungal species. Overall, the minimal inhibitory concentrations for hyphae were higher than conidia and germinated conidia, except for caspofungin. Collectively, our study add new data about the conidia-into-hyphae transformation in Scedosporium and Lomentospora species, which is a relevant biological process of these molds directly connected to their antifungal resistance and pathogenicity mechanisms.

Conidial germination in Scedosporium apiospermum, S. aurantiacum, S. minutisporum and Lomentospora prolificans: influence of growth conditions and antifungal susceptibility profiles

PubMed Central

de Mello, Thaís Pereira; Aor, Ana Carolina; de Oliveira, Simone Santiago Carvalho; Branquinha, Marta Helena; dos Santos, André Luis Souza

2016-01-01

In the present study, we have investigated some growth conditions capable of inducing the conidial germination in Scedosporium apiospermum, S. aurantiacum, S. minutisporum and Lomentospora prolificans. Germination in Sabouraud medium (pH 7.0, 37ºC, 5% CO2) showed to be a typically time-dependent event, reaching ~75% in S. minutisporum and > 90% in S. apiospermum, S. aurantiacum and L. prolificans after 4 h. Similar germination rate was observed when conidia were incubated under different media and pHs. Contrarily, temperature and CO2 tension modulated the germination. The isotropic conidial growth (swelling) and germ tube-like projection were evidenced by microscopy and cytometry. Morphometric parameters augmented in a time-dependent fashion, evidencing changes in size and granularity of fungal cells compared with dormant 0 h conidia. In parallel, a clear increase in the mitochondrial activity was measured during the transformation of conidia-into-germinated conidia. Susceptibility profiles to itraconazole, fluconazole, voriconazole, amphotericin B and caspofungin varied regarding each morphotype and each fungal species. Overall, the minimal inhibitory concentrations for hyphae were higher than conidia and germinated conidia, except for caspofungin. Collectively, our study add new data about the conidia-into-hyphae transformation in Scedosporium and Lomentospora species, which is a relevant biological process of these molds directly connected to their antifungal resistance and pathogenicity mechanisms. PMID:27355215

An increase in pectin methyl esterase activity accompanies dormancy breakage and germination of yellow cedar seeds.

PubMed

Ren, C; Kermode, A R

2000-09-01

Pectin methyl esterase (PME) (EC 3.1.1.11) catalyzes the hydrolysis of methylester groups of cell wall pectins. We investigated the role of this enzyme in dormancy termination and germination of yellow cedar (Chamaecyparis nootkatensis [D. Don] Spach) seeds. PME activity was not detected in dormant seeds of yellow cedar but was induced and gradually increased during moist chilling; high activity coincided with dormancy breakage and germination. PME activity was positively correlated to the degree of dormancy breakage of yellow cedar seeds. The enzyme produced in different seed parts and in seeds at different times during moist chilling, germination, and early post-germinative growth consisted of two isoforms, both basic with isoelectric points of 8.7 and 8.9 and the same molecular mass of 62 kD. The pH optimum for the enzyme was between 7.4 and 8.4. In intact yellow cedar seeds, activities of the two basic isoforms of PME that were induced in embryos and in megagametophytes following dormancy breakage were significantly suppressed by abscisic acid. Gibberellic acid had a stimulatory effect on the activities of these isoforms in embryos and megagametophytes of intact seeds at the germinative stage. We hypothesize that PME plays a role in weakening of the megagametophyte, allowing radicle emergence and the completion of germination.

Biological characterization of adult MYC-translocation-positive mature B-cell lymphomas other than molecular Burkitt lymphoma.

PubMed

Aukema, Sietse M; Kreuz, Markus; Kohler, Christian W; Rosolowski, Maciej; Hasenclever, Dirk; Hummel, Michael; Küppers, Ralf; Lenze, Dido; Ott, German; Pott, Christiane; Richter, Julia; Rosenwald, Andreas; Szczepanowski, Monika; Schwaenen, Carsten; Stein, Harald; Trautmann, Heiko; Wessendorf, Swen; Trümper, Lorenz; Loeffler, Markus; Spang, Rainer; Kluin, Philip M; Klapper, Wolfram; Siebert, Reiner

2014-04-01

Chromosomal translocations affecting the MYC oncogene are the biological hallmark of Burkitt lymphomas but also occur in a subset of other mature B-cell lymphomas. If accompanied by a chromosomal break targeting the BCL2 and/or BCL6 oncogene these MYC translocation-positive (MYC(+)) lymphomas are called double-hit lymphomas, otherwise the term single-hit lymphomas is applied. In order to characterize the biological features of these MYC(+) lymphomas other than Burkitt lymphoma we explored, after exclusion of molecular Burkitt lymphoma as defined by gene expression profiling, the molecular, pathological and clinical aspects of 80 MYC-translocation-positive lymphomas (31 single-hit, 46 double-hit and 3 MYC(+)-lymphomas with unknown BCL6 status). Comparison of single-hit and double-hit lymphomas revealed no difference in MYC partner (IG/non-IG), genomic complexity, MYC expression or gene expression profile. Double-hit lymphomas more frequently showed a germinal center B-cell-like gene expression profile and had higher IGH and MYC mutation frequencies. Gene expression profiling revealed 130 differentially expressed genes between BCL6(+)/MYC(+) and BCL2(+)/MYC(+) double-hit lymphomas. BCL2(+)/MYC(+) double-hit lymphomas more frequently showed a germinal center B-like gene expression profile. Analysis of all lymphomas according to MYC partner (IG/non-IG) revealed no substantial differences. In this series of lymphomas, in which immunochemotherapy was administered in only a minority of cases, single-hit and double-hit lymphomas had a similar poor outcome in contrast to the outcome of molecular Burkitt lymphoma and lymphomas without the MYC break. Our data suggest that, after excluding molecular Burkitt lymphoma and pediatric cases, MYC(+) lymphomas are biologically quite homogeneous with single-hit and double-hit lymphomas as well as IG-MYC and non-IG-MYC(+) lymphomas sharing various molecular characteristics.

Roles of germination-specific lytic enzymes CwlJ and SleB in Bacillus anthracis.

PubMed

Heffron, Jared D; Orsburn, Benjamin; Popham, David L

2009-04-01

The structural characteristics of a spore enable it to withstand stresses that typically kill a vegetative cell. Spores remain dormant until small molecule signals induce them to germinate into vegetative bacilli. Germination requires degradation of the thick cortical peptidoglycan by germination-specific lytic enzymes (GSLEs). Bacillus anthracis has four putative GSLEs, based upon sequence similarities with enzymes in other species: SleB, CwlJ1, CwlJ2, and SleL. In this study, the roles of SleB, CwlJ1, and CwlJ2 were examined. The expression levels of all three genes peak 3.5 h into sporulation. Genetic analysis revealed that, similar to other known GSLEs, none of these gene products are individually required for growth, sporulation, or triggering of germination. However, later germination events are affected in spores lacking CwlJ1 or SleB. Compared to the wild type, germinating spores without CwlJ1 suffer a delay in optical density loss and cortex peptidoglycan release. The absence of SleB also causes a delay in cortex fragment release. A double mutant lacking both SleB and CwlJ1 is completely blocked in cortex hydrolysis and progresses through outgrowth to produce colonies at a frequency 1,000-fold lower than that of the wild-type strain. A null mutation eliminating CwlJ2 has no effect on germination. High-performance liquid chromatography and mass spectroscopy analysis revealed that SleB is required for lytic transglycosylase activity. CwlJ1 also clearly participates in cortex hydrolysis, but its specific mode of action remains unclear. Understanding the lytic germination activities that naturally diminish spore resistance can lead to methods for prematurely inducing them, thus simplifying the process of treating contaminated sites.

Investigating small molecules to inhibit germinal center kinase-like kinase (GLK/MAP4K3) upstream of PKCθ phosphorylation: Potential therapy to modulate T cell dependent immunity.

PubMed

May-Dracka, Tricia L; Arduini, Robert; Bertolotti-Ciarlet, Andrea; Bhisetti, Govinda; Brickelmaier, Margot; Cahir-McFarland, Ellen; Enyedy, Istvan; Fontenot, Jason D; Hesson, Thomas; Little, Kevin; Lyssikatos, Joe; Marcotte, Douglas; McKee, Timothy; Murugan, Paramasivam; Patterson, Thomas; Peng, Hairuo; Rushe, Mia; Silvian, Laura; Spilker, Kerri; Wu, Ping; Xin, Zhili; Burkly, Linda C

2018-06-01

Germinal center kinase-like kinase (GLK, also known as MAP4K3) has been hypothesized to have an effect on key cellular activities, including inflammatory responses. GLK is required for activation of protein kinase C-θ (PKCθ) in T cells. Controlling the activity of T helper cell responses could be valuable for the treatment of autoimmune diseases. This approach circumvents previous unsuccessful approaches to target PKCθ directly. The use of structure based drug design, aided by the first crystal structure of GLK, led to the discovery of several inhibitors that demonstrate potent inhibition of GLK biochemically and in relevant cell lines. Copyright © 2018 Elsevier Ltd. All rights reserved.

Disruption of the Aspergillus fumigatus ECM33 homologue results in rapid conidial germination, antifungal resistance and hypervirulence.

PubMed

Romano, Jacob; Nimrod, Guy; Ben-Tal, Nir; Shadkchan, Yona; Baruch, Koti; Sharon, Haim; Osherov, Nir

2006-07-01

The ECM33/SPS2 family of glycosylphosphatidylinositol-anchored proteins plays an important role in maintaining fungal cell wall integrity and virulence. However, the precise molecular role of these proteins is unknown. In this work, AfuEcm33, the gene encoding the ECM33 homologue in the important pathogenic fungus Aspergillus fumigatus, has been cloned and its function analysed. It is shown that disruption of AfuEcm33 results in rapid conidial germination, increased cell-cell adhesion, resistance to the antifungal agent caspofungin and increased virulence in an immunocompromised mouse model for disseminated aspergillosis. These results suggest that the protein encoded by AfuEcm33 is involved in key aspects of cell wall morphogenesis and plays an important role in A. fumigatus virulence.

Functional genomic characterization of neoblast-like stem cells in larval Schistosoma mansoni

PubMed Central

Wang, Bo; Collins, James J; Newmark, Phillip A

2013-01-01

Schistosomes infect hundreds of millions of people in the developing world. Transmission of these parasites relies on a stem cell-driven, clonal expansion of larvae inside a molluscan intermediate host. How this novel asexual reproductive strategy relates to current models of stem cell maintenance and germline specification is unclear. Here, we demonstrate that this proliferative larval cell population (germinal cells) shares some molecular signatures with stem cells from diverse organisms, in particular neoblasts of planarians (free-living relatives of schistosomes). We identify two distinct germinal cell lineages that differ in their proliferation kinetics and expression of a nanos ortholog. We show that a vasa/PL10 homolog is required for proliferation and maintenance of both populations, whereas argonaute2 and a fibroblast growth factor receptor-encoding gene are required only for nanos-negative cells. Our results suggest that an ancient stem cell-based developmental program may have enabled the evolution of the complex life cycle of parasitic flatworms. DOI: http://dx.doi.org/10.7554/eLife.00768.001 PMID:23908765

Cutting edge: double-stranded DNA breaks in the IgV region gene were detected at lower frequency in affinity-maturation impeded GANP-/- mice.

PubMed

Kawatani, Yousuke; Igarashi, Hideya; Matsui, Takeshi; Kuwahara, Kazuhiko; Fujimura, Satoru; Okamoto, Nobukazu; Takagi, Katsumasa; Sakaguchi, Nobuo

2005-11-01

Double-stranded DNA breaks (DSBs) at the IgV region (IgV) genes might be involved in somatic hypermutation and affinity-maturation of the B cell receptor in response to T cell-dependent Ag. By ligation-mediated PCR, we studied IgV DSBs that occurred in mature germinal center B cells in response to nitrophenyl-chicken gamma-globulin in a RAG1-independent, Ag-dependent, and IgV-selective manner. We quantified their levels in GANP-deficient B cells that have impaired generation of high-affinity Ab. GANP-/- B cells showed a decreased level of DSBs with blunt ends than control B cells and, on the contrary, the ganp gene transgenic (GANPTg) B cells showed an increased level. These results suggested that the level of IgV DSBs in germinal center B cells is associated with GANP expression, which is presumably required for B cell receptor affinity maturation.

Transcriptional regulation of germinal center B and plasma cell fates by dynamical control of IRF4

PubMed Central

Ochiai, Kyoko; Maienschein-Cline, Mark; Simonetti, Giorgia; Chen, Jianjun; Rosenthal, Rebecca; Brink, Robert; Chong, Anita S.; Klein, Ulf; Dinner, Aaron R.; Singh, Harinder; Sciammas, Roger

2013-01-01

Summary The transcription factor IRF4 regulates immunoglobulin class switch recombination and plasma cell differentiation. Its differing concentrations appear to regulate mutually antagonistic programs of B and plasma cell gene expression. We show IRF4 to be also required for generation of germinal center (GC) B cells. Its transient expression in vivo induced the expression of key GC genes including Bcl6 and Aicda. In contrast, sustained and higher concentrations of IRF4 promoted the generation of plasma cells while antagonizing the GC fate. IRF4 co-bound with the transcription factors PU.1 or BATF to Ets or AP-1 composite motifs, associated with genes involved in B cell activation and the GC response. At higher concentrations IRF4 binding shifted to interferon sequence response motifs; these enriched for genes involved in plasma cell differentiation. Our results support a model of “kinetic control” in which signaling induced dynamics of IRF4 in activated B cells control their cell fate outcomes. PMID:23684984

A dynamic T cell–limited checkpoint regulates affinity-dependent B cell entry into the germinal center

PubMed Central

Schwickert, Tanja A.; Victora, Gabriel D.; Fooksman, David R.; Kamphorst, Alice O.; Mugnier, Monica R.; Gitlin, Alexander D.; Dustin, Michael L.

2011-01-01

The germinal center (GC) reaction is essential for the generation of the somatically hypermutated, high-affinity antibodies that mediate adaptive immunity. Entry into the GC is limited to a small number of B cell clones; however, the process by which this limited number of clones is selected is unclear. In this study, we demonstrate that low-affinity B cells intrinsically capable of seeding a GC reaction fail to expand and become activated in the presence of higher-affinity B cells even before GC coalescence. Live multiphoton imaging shows that selection is based on the amount of peptide–major histocompatibility complex (pMHC) presented to cognate T cells within clusters of responding B and T cells at the T–B border. We propose a model in which T cell help is restricted to the B cells with the highest amounts of pMHC, thus allowing for a dynamic affinity threshold to be imposed on antigen-binding B cells. PMID:21576382

Interferon-gamma and T-bet expression in a patient with toxoplasmic lymphadenopathy.

PubMed

Jöhrens, Korinna; Moos, Verena; Schneider, Thomas; Stein, Harald; Anagnostopoulos, Ioannis

2010-04-01

Infection with Toxoplasma gondii (TG) presents in some individuals as a self-limited disease with a predominant lymphadenopathy characterized by prominent B-cell activation. As this is in contrast to the in vitro based concept of a T(h)1-immune response against TG, we investigated native lymphoid tissue and peripheral blood of a patient with serologic evidence of toxoplasmosis to verify which cells show T(h)1-response features. High-level expression of T-bet in monocytoid B-cells, in germinal center B-cells, and in a lesser amount in T cells could be demonstrated by immunohistochemistry. In vitro stimulation of lymph node cells with either TG, staphylococcus enterotoxin B, or phorbol 12-myristate 13-acetate/ionomycin revealed an interferon-gamma expression in T-bet(+) B cells only in the patient and not in controls. Similar results were found for T-bet(+) T cells which were also present in controls. CD4(+) peripheral blood cells stimulated with TG antigens showed a TG-specific but attenuated T(h)1-reactivity in the patient associated with a reduced expression of IL-2 when compared with controls. We conclude that the pathogenesis and course of toxoplasmic lymphadenopathy is based on a T(h)1-cell defect, which becomes compensated by the B cells mounting a T(h)1-like immune response.

Systemic Lupus Erythematosus: Molecular Mimicry between Anti-dsDNA CDR3 Idiotype, Microbial and Self Peptides-As Antigens for Th Cells.

PubMed

Aas-Hanssen, Kristin; Thompson, Keith M; Bogen, Bjarne; Munthe, Ludvig A

2015-01-01

Systemic lupus erythematosus (SLE) is marked by a T helper (Th) cell-dependent B cell hyperresponsiveness, with frequent germinal center reactions, and gammaglobulinemia. A feature of SLE is the finding of IgG autoantibodies specific for dsDNA. The specificity of the Th cells that drive the expansion of anti-dsDNA B cells is unresolved. However, anti-microbial, anti-histone, and anti-idiotype Th cell responses have been hypothesized to play a role. It has been entirely unclear if these seemingly disparate Th cell responses and hypotheses could be related or unified. Here, we describe that H chain CDR3 idiotypes from IgG(+) B cells of lupus mice have sequence similarities with both microbial and self peptides. Matched sequences were more frequent within the mutated CDR3 repertoire and when sequences were derived from lupus mice with expanded anti-dsDNA B cells. Analyses of histone sequences showed that particular histone peptides were similar to VDJ junctions. Moreover, lupus mice had Th cell responses toward histone peptides similar to anti-dsDNA CDR3 sequences. The results suggest that Th cells in lupus may have multiple cross-reactive specificities linked to the IgVH CDR3 Id-peptide sequences as well as similar DNA-associated protein motifs.

Candida albicans triggers interleukin-8 secretion by oral epithelial cells.

PubMed

Dongari-Bagtzoglou, A; Kashleva, H

2003-04-01

Oropharyngeal candidiasis is a frequent opportunistic infection associated with immunocompromised hosts. Candida albicans is the principal species responsible for this infection. Production of interleukin-8 (IL-8), by oral epithelial cells can be expected to play a major role in the recruitment and activation of professional phagocytes at the infected site. The purpose of this study was to determine whether C. albicans triggers secretion of IL-8 by oral epithelial cells in vitro and investigate mechanisms of host cell-fungal interactions that trigger such responses. Oral epithelial cell lines (SCC4, SCC15, and OKF6/TERT-2) as well as primary gingival epithelial cells were used. Epithelial cells were cocultured with C. albicans, strains SC5314, ATCC28366 or ATCC32077, for 24-48 hr, and supernatants were analyzed for IL-8 content by ELISA. A germination-deficient mutant (efg1/efg1 cph1/cph1), otherwise isogenic to strain SC5314, was used to assess the requirement for germination in triggering IL-8 responses. In order to ascertain whether direct contact of yeast with host cells is required to trigger cytokine production, epithelial cells were separated from yeast using cell culture inserts. To test whether IL-8 secretion is dependent on IL-1alpha activity, epithelial cells were challenged with viable C. albicans in the presence or absence of neutralizing anti-IL-1alpha antibody or IL-1ra, and IL-8 secretion was measured in the supernatants. All cell lines and primary cultures responded to C. albicans with an increase in IL-8 secretion. IL-8 responses were contact-dependent, strain-specific, required yeast viability and germination into hyphae, and were in part autoregulated by IL-1alpha. Copyright 2003 Elsevier Science Ltd.

Adenosine Monophosphate-Based Detection of Bacterial Spores

NASA Technical Reports Server (NTRS)

Kern, Roger G.; Chen, Fei; Venkateswaran, Kasthuri; Hattori, Nori; Suzuki, Shigeya

2009-01-01

A method of rapid detection of bacterial spores is based on the discovery that a heat shock consisting of exposure to a temperature of 100 C for 10 minutes causes the complete release of adenosine monophosphate (AMP) from the spores. This method could be an alternative to the method described in the immediately preceding article. Unlike that method and related prior methods, the present method does not involve germination and cultivation; this feature is an important advantage because in cases in which the spores are those of pathogens, delays involved in germination and cultivation could increase risks of infection. Also, in comparison with other prior methods that do not involve germination, the present method affords greater sensitivity. At present, the method is embodied in a laboratory procedure, though it would be desirable to implement the method by means of a miniaturized apparatus in order to make it convenient and economical enough to encourage widespread use.

Starry Sky Pattern in Hematopoietic Neoplasms: A Review of Pathophysiology and Differential Diagnosis.

PubMed

Dy-Ledesma, Janelyn L; Khoury, Joseph D; Agbay, Rose Lou Marie C; Garcia, Mar; Miranda, Roberto N; Medeiros, L Jeffrey

2016-11-01

The starry sky pattern is a distinctive histologic feature wherein a rapidly proliferating hematolymphoid neoplasm contains scattered histiocytes with abundant pale cytoplasm in a background of monomorphic neoplastic cells. The cytoplasm of these histiocytes typically contains cellular remnants, also known as tingible bodies, incorporated through active phagocytosis. Although common and widely recognized, relatively little is known about the pathophysiological underpinnings of the starry sky pattern. Its resemblance to a similar pattern seen in the germinal centers of secondary follicles suggests a possible starting point for understanding the molecular basis of the starry sky pattern and potential routes for its exploitation for therapeutic purposes. In this review, we discuss the historical, pathophysiological, and clinical implications of the starry sky pattern.

[The dynamics of chromosomal instability of welsh onion (Allium fistulosum L.): the influence of seed storage temperature].

PubMed

Lazarenko, L M; Bezrukov, V F

2008-01-01

The age-related dynamics of chromosomal instability and germination capacity of welsh onion (Allium fistulosum L.) seeds have been studied under two different storage temperatures during six years after harvesting. Seeds that were kept at the room temperature (14-28 degrees C) during 6 years of storage have lost their germination capacity. The frequencies of aberrant anaphases grew from 2% on the first month of storage up to 80% on the 75th month of storage. The germination capacity of seeds kept at the lower temperature (4-9 degrees C) was 73-77% on the 6th year of storage and the frequency of aberrant anaphases remained within the limits of 2-4%. Thus, storage of welsh onion seeds during 6 years at the lower temperature allows to retain germination capacity and restrains the augmentation of chromosomal instability in root meristem cells of seedlings during this period.

Germinal center hypoxia potentiates immunoglobulin class switch recombination

PubMed Central

Abbott, Robert K.; Thayer, Molly; Labuda, Jasmine; Silva, Murillo; Philbrook, Phaethon; Cain, Derek W.; Kojima, Hidefumi; Hatfield, Stephen; Sethumadhavan, Shalini; Ohta, Akio; Reinherz, Ellis L.; Kelsoe, Garnett; Sitkovsky, Michail

2016-01-01

Germinal centers (GCs) are anatomic sites where B cells undergo secondary diversification to produce high affinity, class switched antibodies. We hypothesized that proliferating B cells in GCs create a hypoxic microenvironment that governs their further differentiation. Using molecular markers, we found GCs to be predominantly hypoxic. Compared to normoxia (21% O2), hypoxic culture conditions (1% O2) in vitro accelerated class switching and plasma cell formation and enhanced expression of GL-7 on B and CD4+ T cells. Reversal of GC hypoxia in vivo by breathing 60% O2 during immunization resulted in reduced frequencies of GC B cells, T follicular helper (TFH) cells and plasmacytes, as well as lower expression of ICOS on TFH. Importantly, this reversal of GC hypoxia decreased antigen-specific serum IgG1 and reduced the frequency of IgG1+ B cells within the antigen specific GC. Taken together, these observations reveal a critical role for hypoxia in GC B cell differentiation. PMID:27798169

Strategy to inactivate Clostridium perfringens spores in meat products.

PubMed

Akhtar, Saeed; Paredes-Sabja, Daniel; Torres, J Antonio; Sarker, Mahfuzur R

2009-05-01

The current study aimed to develop an inactivation strategy for Clostridium perfringens spores in meat through a combination of spore activation at low pressure (100-200 MPa, 7 min) and elevated temperature (80 degrees C, 10 min); spore germination at high temperatures (55, 60 or 65 degrees C); and inactivation of germinated spores with elevated temperatures (80 and 90 degrees C, 10 and 20 min) and high pressure (586 MPa, at 23 and 73 degrees C, 10 min). Low pressures (100-200 MPa) were insufficient to efficiently activate C. perfringens spores for germination. However, C. perfringens spores were efficiently activated with elevated temperature (80 degrees C, 10 min), and germinated at temperatures lethal for vegetative cells (>or= 55 degrees C) when incubated for 60 min with a mixture of L-asparagine and KCl (AK) in phosphate buffer (pH 7) and in poultry meat. Inactivation of spores (approximately 4 decimal reduction) in meat by elevated temperatures (80-90 degrees C for 20 min) required a long germination period (55 degrees C for 60 min). However, similar inactivation level was reached with shorter germination period (55 degrees C for 15 min) when spore contaminated-meat was treated with pressure-assisted thermal processing (568 MPa, 73 degrees C, 10 min). Therefore, the most efficient strategy to inactivate C. perfringens spores in poultry meat containing 50 mM AK consisted: (i) a primary heat treatment (80 degrees C, 10 min) to pasteurize and denature the meat proteins and to activate C. perfringens spores for germination; (ii) cooling of the product to 55 degrees C in about 20 min and further incubation at 55 degrees C for about 15 min for spore germination; and (iii) inactivation of germinated spores by pressure-assisted thermal processing (586 MPa at 73 degrees C for 10 min). Collectively, this study demonstrates the feasibility of an alternative and novel strategy to inactivate C. perfringens spores in meat products formulated with germinants specific for C. perfringens.

Quantification of healthy and atretic germ cells and follicles in the developing and post-natal ovary of the South American plains vizcacha, Lagostomus maximus: evidence of continuous rise of the germinal reserve.

PubMed

Inserra, P I F; Leopardo, N P; Willis, M A; Freysselinard, A L; Vitullo, A D

2014-02-01

The female germ line in mammals is subjected to massive cell death that eliminates 60-85% of the germinal reserve by birth and continues from birth to adulthood until the exhaustion of the germinal pool. Germ cell demise occurs mainly through apoptosis by means of a biased expression in favour of pro-apoptotic members of the BCL2 gene family. By contrast, the South American plains vizcacha, Lagostomus maximus, exhibits sustained expression of the anti-apoptotic BCL2 gene throughout gestation and a low incidence of germ cell apoptosis. This led to the proposal that, in the absence of death mechanisms other than apoptosis, the female germ line should increase continuously from foetal life until after birth. In this study, we quantified all healthy germ cells and follicles in the ovaries of L. maximus from early foetal life to day 60 after birth using unbiased stereological methods and detected apoptosis by labelling with TUNEL assay. The healthy germ cell population increased continuously from early-developing ovary reaching a 50 times higher population number by the end of gestation. TUNEL-positive germ cells were <0.5% of the germ cell number, except at mid-gestation (3.62%). Mitotic proliferation, entrance into prophase I stage and primordial follicle formation occurred as overlapping processes from early pregnancy to birth. Germ cell number remained constant in early post-natal life, but a remnant population of non-follicular VASA- and PCNA-positive germ cells still persisted at post-natal day 60. L. maximus is the first mammal so far described in which female germ line develops in the absence of constitutive massive germ cell elimination.

Studying molecular changes during gravity perception and response in a single cell.

PubMed

Cannon, Ashley E; Salmi, Mari L; Bushart, Thomas J; Roux, Stanley J

2015-01-01

Early studies revealed a highly predictable pattern of gravity-directed growth and development in Ceratopteris richardii spores. This makes the spores a valuable model system for the study of how a single cell senses and responds to the force of gravity. Gravity regulates both the direction and magnitude of a trans-cell calcium current in germinating spores, and the orientation of this current predicts the polarization of spore development. Molecular techniques have been developed to evaluate the transcriptomic and proteomic profiles of spores before and after gravity establishes the polarity of their development. Here we describe these techniques, along with protocols for sterilizing the spores, sowing them in a solid or liquid growth media, and evaluating germination.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Velletri, P.A.; Aquilano, D.R.; Bruckwick, E.

Hypophysectomy of prepubescent (3-week-old) rats prevented the pubertal development of testicular, but not pulmonary, angiotensin-converting enzyme (EC 3.4.15.1). Additionally, hypophysectomy resulted in a loss of testicular converting enzyme activity in 10-week-old rats that had achieved puberty and had developed enzyme activity. Hormone regimens consisting of FSH/LH (7.5 U/rat X day), hCG (10 U/rat X day), or testosterone (1 mg/rat X day) were employed to ascertain their ability to maintain activity in hypophysectomized rats. All three of the above hormone regimens, if initiated on the first day after hypophysectomy of 10-week-old rats, were capable of maintaining testicular converting enzyme activity. Centrifugalmore » elutriation of dispersed testicular cells indicated that the majority of enzyme activity in mature rats was associated with the germinal cells, a result consistent with the data accumulated from the hormonal studies. Lastly, (/sup 3/H)captopril bound specifically to cellular fractions enriched in germinal cells. The above studies suggest that the pituitary gland is required for the development and maintenance of testicular angiotensin-converting enzyme in the rat by stimulating steroidogenesis in the testes. Furthermore, the sensitivity of converting enzyme activity to androgen coupled with the centrifugal elutriation and (/sup 3/H) captopril binding studies strongly support the notion that testicular converting enzyme is associated with germinal cells.« less

The structural bases of long-term anabiosis in non-spore-forming bacteria

NASA Astrophysics Data System (ADS)

Suzina, Natalia E.; Mulyukin, Andrey L.; Dmitriev, Vladimir V.; Nikolaev, Yury A.; Shorokhova, Anna P.; Bobkova, Yulia S.; Barinova, Ekaterina S.; Plakunov, Vladimir K.; El-Registan, Galina I.; Duda, Vitalii I.

2006-01-01

Peculiarities of the structural organization in non-spore-forming bacteria associated with long-term anabiosis were revealed both in laboratory cultures and in natural populations isolated from 1 3-Myr-old Eastern Siberian permafrost and tundra soil. Different advanced methods were used, including (a) high-resolution electron microscopy; (b) simulation of in situ conditions in the laboratory by varying the composition of growth medium and cultivation conditions; (c) low-temperature fractionation to isolate and concentrate microbial cells from natural soils; (d) comparative morphological analysis of microbial cells in model cultures and natural soils (in situ). Under laboratory conditions, the intense formation of resting cells by representatives of various taxa of eubacteria and halophilic archaea occurred in 2 9-month-old cultures grown in carbon-, nitrogen-, or phosphorus-limited media, in starved cell suspensions in the presence of sodium silicate, or on soil agar. Among resting cells, we revealed cystlike forms having a complicated structure and common features. These included a thick capsule; a thickened and multiprofile cell wall; the presence of large intramembrane particles on PF- and EF-fracture surfaces; fine-grained or lumpy cytoplasm; and a condensed nucleoid. The general morphological properties, ultrastructural organization, physiological features of cystlike cells, and their ability to germinate under the appropriate conditions suggest the existence of constitutive dormancy in non-spore-forming bacteria. It was found that the majority of microorganisms in permafrost and tundra soil are cystlike cells, very similar to those in laboratory cultures. Anabiotic (resting) cystlike cells are responsible for the survival of non-spore-formers in extreme Earth habitats and may be regarded as possible analogs of extraterrestrial forms of microbial life.

Elevated levels of N-lauroylethanolamine, an endogenous constituent of desiccated seeds, disrupt normal root development in Arabidopsis thaliana seedlings

NASA Technical Reports Server (NTRS)

Blancaflor, Elison B.; Hou, Guichuan; Chapman, Kent D.

2003-01-01

N-Acylethanolamines (NAEs) are prevalent in desiccated seeds of various plant species, and their levels decline substantially during seed imbibition and germination. Here, seeds of Arabidopsis thaliana (L.) Heynh. were germinated in, and seedlings maintained on, micromolar concentrations of N-lauroylethanolamine (NAE 12:0). NAE 12:0 inhibited root elongation, increased radial swelling of root tips, and reduced root hair numbers in a highly selective and concentration-dependent manner. These effects were reversible when seedlings were transferred to NAE-free medium. Older seedlings (14 days old) acclimated to exogenous NAE by increased formation of lateral roots, and generally, these lateral roots did not exhibit the severe symptoms observed in primary roots. Cells of NAE-treated primary roots were swollen and irregular in shape, and in many cases showed evidence, at the light- and electron-microscope levels, of improper cell wall formation. Microtubule arrangement was disrupted in severely distorted cells close to the root tip, and endoplasmic reticulum (ER)-localized green fluorescent protein (mGFP5-ER) was more abundant, aggregated and distributed differently in NAE-treated root cells, suggesting disruption of proper cell division, endomembrane organization and vesicle trafficking. These results suggest that NAE 12:0 likely influences normal cell expansion in roots by interfering with intracellular membrane trafficking to and/or from the cell surface. The rapid metabolism of NAEs during seed imbibition/germination may be a mechanism to remove this endogenous class of lipid mediators to allow for synchronized membrane reorganization associated with cell expansion.

EZH2 is required for germinal center formation and somatic EZH2 mutations promote lymphoid transformation

PubMed Central

Béguelin, Wendy; Popovic, Relja; Teater, Matt; Jiang, Yanwen; Bunting, Karen L.; Rosen, Monica; Shen, Hao; Yang, Shao Ning; Wang, Ling; Ezponda, Teresa; Martinez-Garcia, Eva; Zhang, Haikuo; Zhang, Yupeng; Verma, Sharad K.; McCabe, Michael T.; Ott, Heidi M.; Van Aller, Glenn S.; Kruger, Ryan G.; Liu, Yan; McHugh, Charles F.; Scott, David W.; Chung, Young Rock; Kelleher, Neil; Shaknovich, Rita; Creasy, Caretha L.; Gascoyne, Randy D.; Wong, Kwok-Kin; Cerchietti, Leandro C.; Levine, Ross L.; Abdel-Wahab, Omar; Licht, Jonathan D.; Elemento, Olivier; Melnick, Ari M.

2013-01-01

The EZH2 histone methyltransferase is highly expressed in germinal center (GC) B-cells and targeted by somatic mutations in B-cell lymphomas. Here we find that EZH2 deletion or pharmacologic inhibition suppresses GC formation and functions in mice. EZH2 represses proliferation checkpoint genes and helps establish bivalent chromatin domains at key regulatory loci to transiently suppress GC B-cell differentiation. Somatic mutations reinforce these physiological effects through enhanced silencing of EZH2 targets in B-cells, and in human B-cell lymphomas. Conditional expression of mutant EZH2 in mice induces GC hyperplasia and accelerated lymphomagenesis in cooperation with BCL2. GCB-type DLBCLs are mostly addicted to EZH2, regardless of mutation status, but not the more differentiated ABC-type DLBCLs, thus clarifying the therapeutic scope of EZH2 targeting. PMID:23680150

The dead seed coat functions as a long-term storage for active hydrolytic enzymes

PubMed Central

Raviv, Buzi; Aghajanyan, Lusine; Granot, Gila; Makover, Vardit; Frenkel, Omer; Gutterman, Yitzchak

2017-01-01

Seed development culminates in programmed cell death (PCD) and hardening of organs enclosing the embryo (e.g., pericarp, seed coat) providing essentially a physical shield for protection during storage in the soil. We examined the proposal that dead organs enclosing embryos are unique entities that store and release upon hydration active proteins that might increase seed persistence in soil, germination and seedling establishment. Proteome analyses of dead seed coats of Brassicaceae species revealed hundreds of proteins being stored in the seed coat and released upon hydration, many are stress-associated proteins such as nucleases, proteases and chitinases. Functional analysis revealed that dead seed coats function as long-term storage for multiple active hydrolytic enzymes (e.g., nucleases) that can persist in active forms for decades. Substances released from the dead seed coat of the annual desert plant Anastatica hierochuntica displayed strong antimicrobial activity. Our data highlighted a previously unrecognized feature of dead organs enclosing embryos (e.g., seed coat) functioning not only as a physical shield for embryo protection but also as a long-term storage for active proteins and other substances that are released upon hydration to the “seedsphere” and could contribute to seed persistence in the soil, germination and seedling establishment. PMID:28700755

The dead seed coat functions as a long-term storage for active hydrolytic enzymes.

PubMed

Raviv, Buzi; Aghajanyan, Lusine; Granot, Gila; Makover, Vardit; Frenkel, Omer; Gutterman, Yitzchak; Grafi, Gideon

2017-01-01

Seed development culminates in programmed cell death (PCD) and hardening of organs enclosing the embryo (e.g., pericarp, seed coat) providing essentially a physical shield for protection during storage in the soil. We examined the proposal that dead organs enclosing embryos are unique entities that store and release upon hydration active proteins that might increase seed persistence in soil, germination and seedling establishment. Proteome analyses of dead seed coats of Brassicaceae species revealed hundreds of proteins being stored in the seed coat and released upon hydration, many are stress-associated proteins such as nucleases, proteases and chitinases. Functional analysis revealed that dead seed coats function as long-term storage for multiple active hydrolytic enzymes (e.g., nucleases) that can persist in active forms for decades. Substances released from the dead seed coat of the annual desert plant Anastatica hierochuntica displayed strong antimicrobial activity. Our data highlighted a previously unrecognized feature of dead organs enclosing embryos (e.g., seed coat) functioning not only as a physical shield for embryo protection but also as a long-term storage for active proteins and other substances that are released upon hydration to the "seedsphere" and could contribute to seed persistence in the soil, germination and seedling establishment.

Receptor revision in CD4 T cells is influenced by follicular helper T cell formation and germinal-center interactions.

PubMed

Higdon, Lauren E; Deets, Katherine A; Friesen, Travis J; Sze, Kai-Yin; Fink, Pamela J

2014-04-15

Peripheral CD4 T cells in Vβ5 transgenic (Tg) C57BL/6J mice undergo tolerance to an endogenous superantigen encoded by mouse mammary tumor virus 8 (Mtv-8) by either deletion or T-cell receptor (TCR) revision. Revision is a process by which surface expression of the Vβ5(+) TCR is down-regulated in response to Mtv-8 and recombination activating genes are expressed to drive rearrangement of the endogenous TCRβ locus, effecting cell rescue through the expression of a newly generated, non-self-reactive TCR. In an effort to identify the microenvironment in which revision takes place, we show here that the proportion of T follicular helper cells (Tfh) and production of high-affinity antibody during a primary response are increased in Vβ5 Tg mice in an Mtv-8-dependent manner. Revising T cells have a Tfh-like surface phenotype and transcription factor profile, with elevated expression of B-cell leukemia/lymphoma 6 (Bcl-6), CXC chemokine receptor 5, programmed death-1, and other Tfh-associated markers. Efficient revision requires Bcl-6 and is inhibited by B lymphocyte-induced maturation protein-1. Revision completes less efficiently in the absence of signaling lymphocytic activation molecule-associated protein although initiation proceeds normally. These data indicate that Tfh formation is required for the initiation of revision and germinal-center interactions for its completion. The germinal center is known to provide a confined space in which B-cell antigen receptors undergo selection. Our data extend the impact of this selective microenvironment into the arena of T cells, suggesting that this fluid structure also provides a regulatory environment in which TCR revision can safely take place.

Methods to isolate a large amount of generative cells, sperm cells and vegetative nuclei from tomato pollen for "omics" analysis.

PubMed

Lu, Yunlong; Wei, Liqin; Wang, Tai

2015-01-01

The development of sperm cells (SCs) from microspores involves a set of finely regulated molecular and cellular events and the coordination of these events. The mechanisms underlying these events and their interconnections remain a major challenge. Systems analysis of genome-wide molecular networks and functional modules with high-throughput "omics" approaches is crucial for understanding the mechanisms; however, this study is hindered because of the difficulty in isolating a large amount of cells of different types, especially generative cells (GCs), from the pollen. Here, we optimized the conditions of tomato pollen germination and pollen tube growth to allow for long-term growth of pollen tubes in vitro with SCs generated in the tube. Using this culture system, we developed methods for isolating GCs, SCs and vegetative cell nuclei (VN) from just-germinated tomato pollen grains and growing pollen tubes and their purification by Percoll density gradient centrifugation. The purity and viability of isolated GCs and SCs were confirmed by microscopy examination and fluorescein diacetate staining, respectively, and the integrity of VN was confirmed by propidium iodide staining. We could obtain about 1.5 million GCs and 2.0 million SCs each from 180 mg initiated pollen grains, and 10 million VN from 270 mg initiated pollen grains germinated in vitro in each experiment. These methods provide the necessary preconditions for systematic biology studies of SC development and differentiation in higher plants.

MYC/BCL2/BCL6 triple hit lymphoma: a study of 40 patients with a comparison to MYC/BCL2 and MYC/BCL6 double hit lymphomas.

PubMed

Huang, Wenting; Medeiros, L Jeffrey; Lin, Pei; Wang, Wei; Tang, Guilin; Khoury, Joseph; Konoplev, Sergej; Yin, C Cameron; Xu, Jie; Oki, Yasuhiro; Li, Shaoying

2018-05-21

High-grade B-cell lymphomas with MYC, BCL2, and BCL6 rearrangements (triple hit lymphoma) are uncommon. We studied the clinicopathologic features of 40 patients with triple hit lymphoma and compared them to 157 patients with MYC/BCL2 double hit lymphoma and 13 patients with MYC/BCL6 double hit lymphoma. The triple hit lymphoma group included 25 men and 15 women with a median age of 61 years (range, 34-85). Nine patients had a history of B-cell lymphoma. Histologically, 23 (58%) cases were diffuse large B-cell lymphoma and 17 cases had features of B-cell lymphoma, unclassifiable, with features intermediate between diffuse large B-cell lymphoma and Burkitt lymphoma. Most cases of triple hit lymphoma were positive for CD10 (100%), BCL2 (95%), BCL6 (82%), MYC (74%), and 71% with MYC and BCL2 coexpression. P53 was overexpressed in 29% of triple hit lymphoma cases. The clinicopathological features of triple hit lymphoma patients were similar to patients with MYC/BCL2 and MYC/BCL6 double hit lymphoma, except that triple hit lymphoma cases were more often CD10 positive compared with MYC/BCL6 double hit lymphoma (p < 0.05). Induction chemotherapy used was similar for patients with triple hit lymphoma and double hit lymphoma and overall survival in triple hit lymphoma patients was 17.6 months, similar to the overall survival of patients with double hit lymphoma (p = 0.67). Patients with triple hit lymphoma showing P53 overexpression had significantly worse overall survival compared with those without P53 overexpression (p = 0.04). On the other hand, double expressor status and prior history of B-cell lymphoma did not correlate with overall survival. In conclusion, most patients with triple hit lymphoma have an aggressive clinical course and poor prognosis and these tumors have a germinal center B-cell immunophenotype, similar to patients with double hit lymphomas. P53 expression is a poor prognostic factor in patients with triple hit lymphoma.

MyD88 signaling in T cells directs IgA-mediated control of the microbiota to promote health

PubMed Central

Kubinak, Jason L.; Petersen, Charisse; Stephens, W. Zac; Soto, Ray; Bake, Erin; O’Connell, Ryan M.; Round, June L.

2015-01-01

SUMMARY Altered commensal communities are associated with human disease. IgA mediates intestinal homeostasis and regulates microbiota composition. Intestinal IgA is produced at high levels as a result of T follicular helper cell (TFH) and B cell interactions in germinal centers. However, the pathways directing host IgA responses towards the microbiota remain unknown. Here, we report that signaling through the innate adaptor MyD88 in gut T cells coordinates germinal center responses, including TFH and IgA+ B cell development. TFH development is deficient in germfree mice and can be restored by feeding TLR2 agonists that activate T cell intrinsic MyD88 signaling. Loss of this pathway diminishes high affinity IgA targeting of the microbiota and fails to control the bacterial community, leading to worsened disease. Our findings identify that T cells converge innate and adaptive immune signals to coordinate IgA against the microbiota, constraining microbial community membership to promote symbiosis. PMID:25620548

Interleukin-10 from CD4+ follicular regulatory T cells promotes the germinal center response.

PubMed

Laidlaw, Brian J; Lu, Yisi; Amezquita, Robert A; Weinstein, Jason S; Vander Heiden, Jason A; Gupta, Namita T; Kleinstein, Steven H; Kaech, Susan M; Craft, Joe

2017-10-20

CD4 + follicular regulatory T (T fr ) cells suppress B cell responses through modulation of follicular helper T (T fh ) cells and germinal center (GC) development. We found that T fr cells can also promote the GC response through provision of interleukin-10 (IL-10) after acute infection with lymphocytic choriomeningitis virus (LCMV). Sensing of IL-10 by B cells was necessary for optimal development of the GC response. GC B cells formed in the absence of T reg cell-derived IL-10 displayed an altered dark zone state and decreased expression of the transcription factor Forkhead box protein 1 (FOXO1). IL-10 promoted nuclear translocation of FOXO1 in activated B cells. These data indicate that T fr cells play a multifaceted role in the fine-tuning of the GC response and identify IL-10 as an important mediator by which T fr cells support the GC reaction. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

Hodgkin's disease biology: recent advances.

PubMed

Jox, A; Wolf, J; Diehl, V

1997-11-01

The cellular origin of H-RS cells has been questioned for a long time. Recently, using single cell amplification of Ig genes evidence was obtained that H-RS cells clonally arise from B-cells. Sequence analysis of rearranged Ig genes demonstrated that H-RS cells develop within the germinal centre. H-RS cells in classical HD grow despite loss of function of their rearranged Ig genes. In contrast, the mutation pattern of rearranged Ig genes in L & H cells of lymphocyte-predominant HD frequently shows ongoing mutations indicating that these cell are still antigen selected. These molecular differences show that LP HD genetically differs from classical HD. H-RS cells escape from apoptosis within the germinal centre. However, the events leading to malignant transformation are still unknown. The association between EBV and HD has been repeatedly described, but the occurrence of EBV negative cases is hard to explain just by loss of EBV. The analysis of chromosomal aberrations in H-RS cells did not result in the description of a specific 'HD-gene'. Also the role of the T-lymphocytes surrounding the H-RS cells has remained an open question.

Use of Green Fluorescent Protein-Transgenic Strains to Study Pathogenic and Nonpathogenic Lifestyles in Colletotrichum acutatum.

PubMed

Horowitz, Sigal; Freeman, Stanley; Sharon, Amir

2002-07-01

ABSTRACT Colletotrichum acutatum, which causes anthracnose disease on strawberry, can also persist on several other plant species without causing disease symptoms. The genetic and molecular bases that determine pathogenic and nonpathogenic lifestyles in C. acutatum are unclear. We developed a transformation system for C. acutatum by electroporation of germinating conidia, and transgenic isolates that express the green fluorescent protein (GFP) were produced. Details of the pathogenic and nonpathogenic lifestyles of C. acutatum were determined by using GFP-transgenic isolates. Major differences between colonization-mediating processes of strawberry and of other plants were observed. On the main host, strawberry, the germinating conidia formed branched, thick hyphae, and large numbers of appressoria were produced that were essential for plant penetration. In strawberry, the fungus developed rapidly, filling the mesophyll with dense mycelium that invaded the cells and caused necrosis of the tissue. In nonpathogenic interactions on pepper, eggplant, and tomato, the conidia germinated, producing thin, straight germ tubes. Appressoria were produced but failed to germinate and penetrate leaf tissue, resulting in epiphytic growth without invasion of the plant. Penetration of the plant occurred only several days after inoculation and was restricted to the intercellular spaces of the first cell layers of infected tissue without causing any visible damage. Much of the new fungal biomass continued to develop on the surface of inoculated organs in the nonpathogenic interaction. The differences in fungal development on strawberry compared with the other plant species suggest that signal molecules, which may be present only in strawberry, trigger appressorial germination and penetration of the primary host.

Germinal-center development of memory B cells driven by IL-9 from follicular helper T cells.

PubMed

Wang, Yifeng; Shi, Jingwen; Yan, Jiacong; Xiao, Zhengtao; Hou, Xiaoxiao; Lu, Peiwen; Hou, Shiyue; Mao, Tianyang; Liu, Wanli; Ma, Yuanwu; Zhang, Lianfeng; Yang, Xuerui; Qi, Hai

2017-08-01

Germinal centers (GCs) support high-affinity, long-lived humoral immunity. How memory B cells develop in GCs is not clear. Through the use of a cell-cycle-reporting system, we identified GC-derived memory precursor cells (GC-MP cells) that had quit cycling and reached G0 phase while in the GC, exhibited memory-associated phenotypes with signs of affinity maturation and localized toward the GC border. After being transferred into adoptive hosts, GC-MP cells reconstituted a secondary response like genuine memory B cells. GC-MP cells expressed the interleukin 9 (IL-9) receptor and responded to IL-9. Acute treatment with IL-9 or antibody to IL-9 accelerated or retarded the positioning of GC-MP cells toward the GC edge and exit from the GC, and enhanced or inhibited the development of memory B cells, which required B cell-intrinsic responsiveness to IL-9. Follicular helper T cells (T FH cells) produced IL-9, and deletion of IL-9 from T cells or, more specifically, from GC T FH cells led to impaired memory formation of B cells. Therefore, the GC development of memory B cells is promoted by T FH cell-derived IL-9.

Somatic mutation of EZH2 (Y641) in follicular and diffuse large B-cell lymphomas of germinal center origin | Office of Cancer Genomics

Cancer.gov

Morin et al. describe recurrent somatic mutations in EZH2, a polycomb group oncogene. The mutation, found in the SET domain of this gene encoding a histone methyltransferase, is found only in a subset of lymphoma samples. Specifically, EZH2 mutations are found in about 12% of follicular lymphomas (FL) and almost 23% of diffuse large B-cell lymphomas (DLBCL) of germinal center origin. This paper goes on to demonstrate that altered EZH2 proteins, corresponding to the most frequent mutations found in human lymphomas, have reduced activity using in vitro histone methylation assays.

Saliva affects the antifungal activity of exogenously added histatin 3 towards Candida albicans.

PubMed

Yamagishi, Hisako; Fitzgerald, Deirdre H; Sein, Tin; Walsh, Thomas J; O'Connell, Brian C

2005-03-01

Antifungal activity of histatin 3 against two Candida albicans clinical isolates was determined in assays containing rabbit submandibular gland saliva. Histatin 3 inhibited the cell growth and germination of both isolates dose-dependently (10-100 microg ml(-1)) with maximum inhibition occurring after 60 min incubation. Adding fresh histatin 3 after 60 min caused further reduction in the viable cell count. Higher histatin 3 concentrations (50-100 microg ml(-1)) and prolonged exposure to peptide were required to inhibit germination. Histatin 3 was rapidly degraded in rabbit submandibular gland saliva and this may explain why fresh addition of histatin 3 increases candidacidal activity.

Pharmacogenomic identification of small molecules for lineage specific manipulation of subventricular zone germinal activity.

PubMed

Azim, Kasum; Angonin, Diane; Marcy, Guillaume; Pieropan, Francesca; Rivera, Andrea; Donega, Vanessa; Cantù, Claudio; Williams, Gareth; Berninger, Benedikt; Butt, Arthur M; Raineteau, Olivier

2017-03-01

Strategies for promoting neural regeneration are hindered by the difficulty of manipulating desired neural fates in the brain without complex genetic methods. The subventricular zone (SVZ) is the largest germinal zone of the forebrain and is responsible for the lifelong generation of interneuron subtypes and oligodendrocytes. Here, we have performed a bioinformatics analysis of the transcriptome of dorsal and lateral SVZ in early postnatal mice, including neural stem cells (NSCs) and their immediate progenies, which generate distinct neural lineages. We identified multiple signaling pathways that trigger distinct downstream transcriptional networks to regulate the diversity of neural cells originating from the SVZ. Next, we used a novel in silico genomic analysis, searchable platform-independent expression database/connectivity map (SPIED/CMAP), to generate a catalogue of small molecules that can be used to manipulate SVZ microdomain-specific lineages. Finally, we demonstrate that compounds identified in this analysis promote the generation of specific cell lineages from NSCs in vivo, during postnatal life and adulthood, as well as in regenerative contexts. This study unravels new strategies for using small bioactive molecules to direct germinal activity in the SVZ, which has therapeutic potential in neurodegenerative diseases.

Alexandrium fundyense cyst viability and germling survival in light vs. dark at a constant low temperature

NASA Astrophysics Data System (ADS)

Vahtera, Emil; Crespo, Bibiana G.; McGillicuddy, Dennis J.; Olli, Kalle; Anderson, Donald M.

2014-05-01

Both observations and models suggest that large-scale coastal blooms of Alexandrium fundyense in the Gulf of Maine are seeded by deep-bottom cyst accumulation zones (“seed beds”) where cysts germinate from the sediment surface or the overlying near-bottom nepheloid layers at water depths exceeding 100 m. The germling cells and their vegetative progeny are assumed to be subject to mortality while in complete darkness, as they swim to illuminated surface waters. To test the validity of this assumption we conducted laboratory investigations of cyst viability and the survival of the germling cells and their vegetative progeny during prolonged exposure to darkness at a temperature of 6 °C, simulating the conditions in deep Gulf of Maine waters. We isolated cysts from bottom sediments collected in the Gulf of Maine under low red light and incubated them in 96-well tissue culture-plates in culture medium under a 10:14 h light:dark cycle and under complete darkness. Cyst viability was high, with excystment frequency reaching 90% in the illuminated treatment after 30 days and in the dark treatment after 50 days. Average germination rates were 0.062 and 0.038 d-1 for light and dark treatments, respectively. The dark treatment showed an approximately 2-week time lag in maximum germination rates compared to the light treatment. Survival of germlings was considerably lower in the dark treatment. In the light treatments, 47% of germinated cysts produced germlings that were able to survive for 7 days and produce vegetative progeny, i.e., there were live cells in the well along with an empty cyst at least once during the experiment. In the dark treatments 12% of the cysts produced germlings that were able to survive for the same length of time. When dark treatments are scaled to take into account non-darkness related mortality, approximately 28% of the cysts produced germlings that were able to survive for at least 7 days. Even though cysts are able to germinate in darkness, the lack of illumination considerably reduces survival rate of germling cells. In addition to viability of cysts in surface sediments and the near-bottom nepheloid layer, survivability of germling cells and their vegetative progeny at aphotic depths is an important consideration in assessing the quantitative role of deep-coastal cyst seed beds in bloom formation.

CD22 is required for formation of memory B cell precursors within germinal centers.

PubMed

Chappell, Craig P; Draves, Kevin E; Clark, Edward A

2017-01-01

CD22 is a BCR co-receptor that regulates B cell signaling, proliferation and survival and is required for T cell-independent Ab responses. To investigate the role of CD22 during T cell-dependent (TD) Ab responses and memory B cell formation, we analyzed Ag-specific B cell responses generated by wild-type (WT) or CD22-/- B cells following immunization with a TD Ag. CD22-/- B cells mounted normal early Ab responses yet failed to generate either memory B cells or long-lived plasma cells, whereas WT B cells formed both populations. Surprisingly, B cell expansion and germinal center (GC) differentiation were comparable between WT and CD22-/- B cells. CD22-/- B cells, however, were significantly less capable of generating a population of CXCR4hiCD38hi GC B cells, which we propose represent memory B cell precursors within GCs. These results demonstrate a novel role for CD22 during TD humoral responses evident during primary GC formation and underscore that CD22 functions not only during B cell maturation but also during responses to both TD and T cell-independent antigens.

CD22 is required for formation of memory B cell precursors within germinal centers

PubMed Central

Chappell, Craig P.; Draves, Kevin E.

2017-01-01

CD22 is a BCR co-receptor that regulates B cell signaling, proliferation and survival and is required for T cell-independent Ab responses. To investigate the role of CD22 during T cell-dependent (TD) Ab responses and memory B cell formation, we analyzed Ag-specific B cell responses generated by wild-type (WT) or CD22-/- B cells following immunization with a TD Ag. CD22-/- B cells mounted normal early Ab responses yet failed to generate either memory B cells or long-lived plasma cells, whereas WT B cells formed both populations. Surprisingly, B cell expansion and germinal center (GC) differentiation were comparable between WT and CD22-/- B cells. CD22-/- B cells, however, were significantly less capable of generating a population of CXCR4hiCD38hi GC B cells, which we propose represent memory B cell precursors within GCs. These results demonstrate a novel role for CD22 during TD humoral responses evident during primary GC formation and underscore that CD22 functions not only during B cell maturation but also during responses to both TD and T cell-independent antigens. PMID:28346517

N‐methyl‐D‐aspartate receptor antibody production from germinal center reactions: Therapeutic implications

PubMed Central

Makuch, Mateusz; Wilson, Robert; Al‐Diwani, Adam; Varley, James; Kienzler, Anne‐Kathrin; Taylor, Jennifer; Berretta, Antonio; Fowler, Darren; Lennox, Belinda; Leite, M. Isabel; Waters, Patrick

2018-01-01

Introduction N‐methyl‐D‐aspartate receptor (NMDAR) antibody encephalitis is mediated by immunoglobulin G (IgG) autoantibodies directed against the NR1 subunit of the NMDAR. Around 20% of patients have an underlying ovarian teratoma, and the condition responds to early immunotherapies and ovarian teratoma removal. However, despite clear therapeutic relevance, mechanisms of NR1‐IgG production and the contribution of germinal center B cells to NR1‐IgG levels are unknown. Methods Clinical data and longitudinal paired serum NR1‐reactive IgM and IgG levels from 10 patients with NMDAR‐antibody encephalitis were determined. Peripheral blood mononuclear cells from these 10 patients, and two available ovarian teratomas, were stimulated with combinations of immune factors and tested for secretion of total IgG and NR1‐specific antibodies. Results In addition to disease‐defining NR1‐IgG, serum NR1‐IgM was found in 6 of 10 patients. NR1‐IgM levels were typically highest around disease onset and detected for several months into the disease course. Moreover, circulating patient B cells were differentiated into CD19+CD27++CD38++ antibody‐secreting cells in vitro and, from 90% of patients, secreted NR1‐IgM and NR1‐IgG. Secreted levels of NR1‐IgG correlated with serum NR1‐IgG (p < 0.0001), and this was observed across the varying disease durations, suggestive of an ongoing process. Furthermore, ovarian teratoma tissue contained infiltrating lymphocytes which produced NR1‐IgG in culture. Interpretation Serum NR1‐IgM and NR1‐IgG, alongside the consistent production of NR1‐IgG from circulating B cells and from ovarian teratomas suggest that ongoing germinal center reactions may account for the peripheral cell populations which secrete NR1‐IgG. Cells participating in germinal center reactions might be a therapeutic target for the treatment of NMDAR‐antibody encephalitis. Ann Neurol 2018;83:553–561 PMID:29406578

Dynamic distribution and the role of abscisic acid during seed development of a lady’s slipper orchid, Cypripedium formosanum

PubMed Central

Lee, Yung-I; Chung, Mei-Chu; Yeung, Edward C.; Lee, Nean

2015-01-01

Background and Aims Although abscisic acid (ABA) is commonly recognized as a primary cause of seed dormancy, there is a lack of information on the role of ABA during orchid seed development. In order to address this issue, the localization and quantification of ABA were determined in developing seeds of Cypripedium formosanum. Methods The endogenous ABA profile of seeds was measured by enzyme-linked immunosorbent assay (ELISA). Temporal and spatial distributions of ABA in developing seeds were visualized by immunohistochemical staining with monoclonal ABA antibodies. Fluoridone was applied to test the causal relationship between ABA content and seed germinability. Key Results ABA content was low at the proembryo stage, then increased rapidly from 120 to 150 days after pollination (DAP), accompanied by a progressive decrease in water content and seed germination. Immunofluorescence signals indicated an increase in fluorescence over time from the proembryo stage to seed maturation. From immunogold labelling, gold particles could be seen within the cytoplasm of embryo-proper cells during the early stages of seed development. As seeds approached maturity, increased localization of gold particles was observed in the periplasmic space, the plasmalemma between embryo-proper cells, the surface wall of the embryo proper, and the inner walls of inner seed-coat cells. At maturity, gold particles were found mainly in the apoplast, such as the surface wall of the embryo proper, and the shrivelled inner and outer seed coats. Injection of fluoridone into capsules resulted in enhanced germination of mature seeds. Conclusions The results indicate that ABA is the key inhibitor of germination in C. formosanum. The distinct accumulation pattern of ABA suggests that it is synthesized in the cytosol of embryo cells during the early stages of seed development, and then exported to the apoplastic region of the cells for subsequent regulatory processes as seeds approach maturity. PMID:26105185

Endoreduplication intensity as a marker of seed developmental stage in the Fabaceae.

PubMed

Rewers, Monika; Sliwinska, Elwira

2012-12-01

Flow cytometry (FCM) can be used to study cell cycle activity in developing, mature and germinating seeds. It provides information about a seed's physiological state and therefore can be used by seed growers for assessing optimal harvest times and presowing treatments. Because an augmented proportion of 4C nuclei usually is indicative of high mitotic activity, the 4C/2C ratio is commonly used to follow the progress of seed development and germination. However, its usefulness for polysomatic (i.e., containing cells with different DNA content) seeds is questioned. Changes in cell cycle/endoreduplication activity in developing seeds of five members of the Fabaceae were studied to determine a more suitable marker of seed developmental stages for polysomatic species based on FCM measurements. Seeds of Phaseolus vulgaris, Medicago sativa, Pisum sativum, Vicia sativa, and Vicia faba var. minor were collected 20, 30, 40, 50, and 60 days after flowering (DAF), embryos were isolated and the proportion of nuclei with different DNA contents in the embryo axis and cotyledon was established. The ratios 4C/2C and (Σ>2C)/2C were calculated. Dried seeds were subjected to laboratory germination tests following international seed testing association (ISTA) rules. Additionally, the absolute nuclear DNA content was estimated in the leaves of the studied species. During seed development nuclei with DNA contents from 2C to 128C were detected; the endopolyploidy pattern depended on the species, seed organ and developmental stage. The cell cycle/endoreduplication parameters correlated negatively with genome size. The (Σ>2C)/2C ratio in the cotyledons reflected the seed developmental stage and corresponded with seed germinability. Therefore, this ratio is recommended as a marker in polysomatic seed research and production instead of the 4C/2C ratio, which does not consider the occurrence of endopolyploid cells. Copyright © 2012 International Society for Advancement of Cytometry.

A novel monoclonal antibody, C41, reveals IL-13Ralpha1 expression by murine germinal center B cells and follicular dendritic cells.

PubMed

Poudrier, J; Graber, P; Herren, S; Berney, C; Gretener, D; Kosco-Vilbois, M H; Gauchat, J F

2000-11-01

Responsiveness to IL-13 involves at least two chains, IL-4Ralpha and IL-13Ralpha1. Although mouse B cells express IL-4Ralpha, little is known about their expression of IL-13Ralpha chains. To investigate this topic further, we have generated a monoclonal antibody (C41) specific for murine IL-13Ralpha1. Using C41, IL-13Ralpha1 expression was detected on germinal center (GC) B cells by flow cytometry and immunohistochemistry. In addition, IL-13Ralpha1 was observed on follicular dendritic cells, but not interdigitating dendritic cells in the T cell areas. Furthermore, resting B cells also expressed IL-13Ralpha1, and in the presence of IL-13 produced increased amounts of IgM in response to in vitro CD40 stimulation. However, C41 was unable to neutralize this bioactivity. The distribution of IL-13Ralpha1 on murine B cells and during GC reactions suggests a role for IL-13 during B cell differentiation.

Apoplastic interactions between plants and plant root intruders.

PubMed

Mitsumasu, Kanako; Seto, Yoshiya; Yoshida, Satoko

2015-01-01

Numerous pathogenic or parasitic organisms attack plant roots to obtain nutrients, and the apoplast including the plant cell wall is where the plant cell meets such organisms. Root parasitic angiosperms and nematodes are two distinct types of plant root parasites but share some common features in their strategies for breaking into plant roots. Striga and Orobanche are obligate root parasitic angiosperms that cause devastating agricultural problems worldwide. Parasitic plants form an invasion organ called a haustorium, where plant cell wall degrading enzymes (PCWDEs) are highly expressed. Plant-parasitic nematodes are another type of agriculturally important plant root parasite. These nematodes breach the plant cell walls by protruding a sclerotized stylet from which PCWDEs are secreted. Responding to such parasitic invasion, host plants activate their own defense responses against parasites. Endoparasitic nematodes secrete apoplastic effectors to modulate host immune responses and to facilitate the formation of a feeding site. Apoplastic communication between hosts and parasitic plants also contributes to their interaction. Parasitic plant germination stimulants, strigolactones, are recently identified apoplastic signals that are transmitted over long distances from biosynthetic sites to functioning sites. Here, we discuss recent advances in understanding the importance of apoplastic signals and cell walls for plant-parasite interactions.

Apoplastic interactions between plants and plant root intruders

PubMed Central

Mitsumasu, Kanako; Seto, Yoshiya; Yoshida, Satoko

2015-01-01

Numerous pathogenic or parasitic organisms attack plant roots to obtain nutrients, and the apoplast including the plant cell wall is where the plant cell meets such organisms. Root parasitic angiosperms and nematodes are two distinct types of plant root parasites but share some common features in their strategies for breaking into plant roots. Striga and Orobanche are obligate root parasitic angiosperms that cause devastating agricultural problems worldwide. Parasitic plants form an invasion organ called a haustorium, where plant cell wall degrading enzymes (PCWDEs) are highly expressed. Plant-parasitic nematodes are another type of agriculturally important plant root parasite. These nematodes breach the plant cell walls by protruding a sclerotized stylet from which PCWDEs are secreted. Responding to such parasitic invasion, host plants activate their own defense responses against parasites. Endoparasitic nematodes secrete apoplastic effectors to modulate host immune responses and to facilitate the formation of a feeding site. Apoplastic communication between hosts and parasitic plants also contributes to their interaction. Parasitic plant germination stimulants, strigolactones, are recently identified apoplastic signals that are transmitted over long distances from biosynthetic sites to functioning sites. Here, we discuss recent advances in understanding the importance of apoplastic signals and cell walls for plant–parasite interactions. PMID:26322059

Heparan sulfate niche for cell proliferation in the adult brain.

PubMed

Mercier, Frederic; Arikawa-Hirasawa, Eri

2012-02-29

In adulthood, new neurons and glial cells are generated from stem cells in restricted zones of the brain, namely the olfactory bulb (OB), rostral migratory stream (RMS), subventricular zone (SVZ) of the lateral ventricle, sub-callosum zone (SCZ) and sub-granular layer (SGL) of the dentate gyrus. What makes these zones germinal? We previously reported that N-sulfated heparan sulfates (N-sulfated HS) present in specialized extracellular matrix structures (fractones) and vascular basement membranes bind the neurogenic factor FGF-2 (fibroblast growth factor-2) next to stem cells in the anterior SVZ of the lateral ventricle, the most neurogenic zone in adulthood. To determine to which extent cell proliferation is associated with N-sulfated HS, we mapped N-sulfated HS and proliferating cells by immunohistochemistry throughout the adult mouse brain. We found that cell proliferation is associated with N-sulfated HS in the OB, RMS, the whole germinal SVZ, and the SCZ. Cell proliferation was weakly associated with N-sulfated HS in the SGL, but the SGL was directly connected to a sub-cortical N-sulfated HS+ extension of the meninges. The NS-sulfated HS+ structures were blood vessels in the OB, RMS and SCZ, and primarily fractones in the SVZ. N-sulfated HS+ fractones, blood vessels and meninges formed a continuum that coursed along the OB, SVZ, RMS, SCZ and SGL, challenging the view that these structures are independent germinal entities. These results support the possibility that a single anatomical system might be globally responsible for mitogenesis and ultimately the production of new neurons and glial cells in the adult brain. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Recent advances in Echinococcus genomics and stem cell research.

PubMed

Koziol, U; Brehm, K

2015-10-30

Alveolar and cystic echinococcosis, caused by the metacestode larval stages of the tapeworms Echinococcus multilocularis and Echinococcus granulosus, respectively, are life-threatening diseases and very difficult to treat. The introduction of benzimidazole-based chemotherapy, which targets parasite β-tubulin, has significantly improved the life-span and prognosis of echinococcosis patients. However, benzimidazoles show only parasitostatic activity, are associated with serious adverse side effects and have to be administered for very long time periods, underlining the need for new drugs. Very recently, the nuclear genomes of E. multilocularis and E. granulosus have been characterised, revealing a plethora of data for gaining a deeper understanding of host-parasite interaction, parasite development and parasite evolution. Combined with extensive transcriptome analyses of Echinococcus life cycle stages these investigations also yielded novel clues for targeted drug design. Recent years also witnessed significant advancements in the molecular and cellular characterisation of the Echinococcus 'germinative cell' population, which forms a unique stem cell system that differs from stem cells of other organisms in the expression of several genes associated with the maintenance of pluripotency. As the only parasite cell type capable of undergoing mitosis, the germinative cells are central to all developmental transitions of Echinococcus within the host and to parasite expansion via asexual proliferation. In the present article, we will briefly introduce and discuss recent advances in Echinococcus genomics and stem cell research in the context of drug design and development. Interestingly, it turns out that benzimidazoles seem to have very limited effects on Echinococcus germinative cells, which could explain the high recurrence rates observed after chemotherapeutic treatment of echinococcosis patients. This clearly indicates that future efforts into the development of parasitocidal drugs should also target the parasite's stem cell system. Copyright © 2015 Elsevier B.V. All rights reserved.

The crucial role of the Pls1 tetraspanin during ascospore germination in Podospora anserina provides an example of the convergent evolution of morphogenetic processes in fungal plant pathogens and saprobes.

PubMed

Lambou, Karine; Malagnac, Fabienne; Barbisan, Crystel; Tharreau, Didier; Lebrun, Marc-Henri; Silar, Philippe

2008-10-01

Pls1 tetraspanins were shown for some pathogenic fungi to be essential for appressorium-mediated penetration into their host plants. We show here that Podospora anserina, a saprobic fungus lacking appressorium, contains PaPls1, a gene orthologous to known PLS1 genes. Inactivation of PaPls1 demonstrates that this gene is specifically required for the germination of ascospores in P. anserina. These ascospores are heavily melanized cells that germinate under inducing conditions through a specific pore. On the contrary, MgPLS1, which fully complements a DeltaPaPls1 ascospore germination defect, has no role in the germination of Magnaporthe grisea nonmelanized ascospores but is required for the formation of the penetration peg at the pore of its melanized appressorium. P. anserina mutants with mutation of PaNox2, which encodes the NADPH oxidase of the NOX2 family, display the same ascospore-specific germination defect as the DeltaPaPls1 mutant. Both mutant phenotypes are suppressed by the inhibition of melanin biosynthesis, suggesting that they are involved in the same cellular process required for the germination of P. anserina melanized ascospores. The analysis of the distribution of PLS1 and NOX2 genes in fungal genomes shows that they are either both present or both absent. These results indicate that the germination of P. anserina ascospores and the formation of the M. grisea appressorium penetration peg use the same molecular machinery that includes Pls1 and Nox2. This machinery is specifically required for the emergence of polarized hyphae from reinforced structures such as appressoria and ascospores. Its recurrent recruitment during fungal evolution may account for some of the morphogenetic convergence observed in fungi.

Effect of Essential Oils on Germination and Growth of Some Pathogenic and Spoilage Spore-Forming Bacteria.

PubMed

Voundi, Stève Olugu; Nyegue, Maximilienne; Lazar, Iuliana; Raducanu, Dumitra; Ndoye, Florentine Foe; Marius, Stamate; Etoa, François-Xavier

2015-06-01

The use of essential oils as a food preservative has increased due to their capacity to inhibit vegetative growth of some bacteria. However, only limited data are available on their effect on bacterial spores. The aim of the present study was to evaluate the effect of some essential oils on the growth and germination of three Bacillus species and Geobacillus stearothermophilus. Essential oils were chemically analyzed using gas chromatography and gas chromatography coupled to mass spectrometry. The minimal inhibitory and bactericidal concentrations of vegetative growth and spore germination were assessed using the macrodilution method. Germination inhibitory effect of treated spores with essential oils was evaluated on solid medium, while kinetic growth was followed using spectrophotometry in the presence of essential oils. Essential oil from Drypetes gossweileri mainly composed of benzyl isothiocyanate (86.7%) was the most potent, with minimal inhibitory concentrations ranging from 0.0048 to 0.0097 mg/mL on vegetative cells and 0.001 to 0.002 mg/mL on spore germination. Furthermore, essential oil from D. gossweileri reduced 50% of spore germination after treatment at 1.25 mg/mL, and its combination with other oils improved both bacteriostatic and bactericidal activities with additive or synergistic effects. Concerning the other essential oils, the minimal inhibitory concentration ranged from 5 to 0.63 mg/mL on vegetative growth and from 0.75 to 0.09 mg/mL on the germination of spores. Spectrophotometric evaluation showed an inhibitory effect of essential oils on both germination and outgrowth. From these results, it is concluded that some of the essential oils tested might be a valuable tool for bacteriological control in food industries. Therefore, further research regarding their use as food preservatives should be carried out.

New insights into the post-translational modification of multiple phosphoenolpyruvate carboxylase isoenzymes by phosphorylation and monoubiquitination during sorghum seed development and germination.

PubMed

Ruiz-Ballesta, Isabel; Baena, Guillermo; Gandullo, Jacinto; Wang, Liqun; She, Yi-Min; Plaxton, William Charles; Echevarría, Cristina

2016-05-01

Phosphoenolpyruvate carboxylase (PEPC; E.C. 4.1.1.31) was characterized in developing and germinating sorghum seeds, focusing on the transcript and polypeptide abundance of multiple plant-type phosphoenolpyruvate carboxylase (PTPC) genes, and the post-translational modification of each isoenzyme by phosphorylation versus monoubiquitination during germination. We observed high levels of SbPPC4 (Sb07g014960) transcripts during early development (stage I), and extensive transcript abundance of SbPPC2 (Sb02g021090) and SbPPC3 (Sb04g008720) throughout the entire life cycle of the seed. Although tandem mass spectrometry (MS) analysis of immunopurified PTPC indicated that four different PTPC isoenzymes were expressed in the developing and germinating seeds, SbPPC3 was the most abundant isozyme of the developing seed, and of the embryo and the aleurone layer of germinating seeds. In vivo phosphorylation of the different PTPC isoenzymes at their conserved N-terminal seryl phosphorylation site during germination was also established by MS/MS analysis. Furthermore, three of the four isoenzymes were partially monoubiquitinated, with MS/MS pinpointing SbPPC2 and SbPPC3 monoubiquitination at the conserved Lys-630 and Lys-624 residues, respectively. Our results demonstrate that monoubiquitination and phosphorylation simultaneously occur in vivo with different PTPC isozymes during seed germination. In addition, we show that PTPC monoubiquitination in germinating sorghum seeds always increases at stage II (emergence of the radicle), is maintained during the aerobic period of rapid cell division and reserve mobilization, and remains relatively constant until stage IV-V when coleoptiles initiate the formation of the photosynthetic tissues. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Growth dynamics of Salmonella enterica strains on alfalfa sprouts and in waste seed irrigation water.

PubMed

Howard, Michael B; Hutcheson, Steven W

2003-01-01

Alfalfa sprouts and other seed sprouts have been implicated in numerous outbreaks of salmonellosis. The source of these epidemics appears to have been low-level contamination of seeds by Salmonella bacteria that developed into clinically significant populations during the seed germination process. To test the possibility that Salmonella enterica strains carry host range determinants that allow them to grow on alfalfa, strains isolated from alfalfa or other sources were surveyed for their ability to grow on germinating alfalfa seeds. An S. enterica serovar Cubana strain originally isolated from contaminated alfalfa sprouts multiplied most rapidly during the initial 24 h of the seed germination process. Germinating alfalfa seeds supported the multiplication of S. enterica cells prior to the emergence of the root radicle at 72 h. Thereafter, much lower rates of multiplication were apparent. The ability of S. enterica to grow on germinating alfalfa seeds was independent of the serovar, isolation source, or virulence of the strain. Isolates obtained from alfalfa attained population levels similar to those observed for strains isolated from contaminated meat products or stools. Each of the strains could be detected in the waste irrigation water, with populations being strongly correlated with those detected on the germinating alfalfa seeds. The S. enterica strains were capable of utilizing the waste irrigation water as a sole carbon and nitrogen source. S. enterica strains thus appear to grow saprophytically on soluble organics released from seeds during early phases of germination. The ability to detect S. enterica in the waste irrigation water early in the germination process indicates that this method may be used as a simple way to monitor the contamination of sprouts during commercial operations.

New aspects in acne inflammation.

PubMed

Toyoda, Masahiko; Morohashi, Masaaki

2003-01-01

There is ample clinical evidence suggesting that the nervous system such as emotional stress can influence the course of acne. We examined possible participation of cutaneous neurogenic factors including neuropeptides, neuropeptide-degrading enzymes and neurotrophic factors, in association with inflammation in the pathogenesis of acne. Immunohistochemical studies revealed that substance P (SP)-immunoreactive nerve fibers were in close apposition to the sebaceous glands, and that neutral endopeptidase (NEP) was expressed in the germinative cells of the sebaceous glands in the skin from acne patients. Nerve growth factor showed immunoreactivity only within the germinative cells. In addition, an increase in the number of mast cells and a strong expression of endothelial leukocyte adhesion molecule-1 on the postcapillary venules were observed in adjacent areas to the sebaceous glands. In vitro, the levels and the expression of stem cell factor by fibroblasts were upregulated by SP. When organ-cultured normal skin specimens were exposed to SP, we observed significant increases in the sizes of the sebaceous glands and in the number of sebum vacuoles in sebaceous cells. Furthermore, supplementation of SP to organ-cultured skin induced expression of NEP, and we demonstrated the subcellular localization of NEP in the endoplasmic reticulum and the Golgi apparatus within the sebaceous germinative cells using preembedding immunoelectron microscopy. These findings suggest that SP may stimulate lipogenesis of the sebaceous glands which may be followed by proliferation of Propionibacterium acnes, and may yield a potent influence on the sebaceous glands by provocation of inflammatory reactions via mast cells. Thus, cutaneous neurogenic factors should contribute to onset and/or exacerbation of acne inflammation. Copyright 2003 S. Karger AG, Basel

PD-1HIGH Follicular CD4 T Helper Cell Subsets Residing in Lymph Node Germinal Centers Correlate with B Cell Maturation and IgG Production in Rhesus Macaques

PubMed Central

Xu, Huanbin; Wang, Xiaolei; Lackner, Andrew A.; Veazey, Ronald S.

2014-01-01

CD4+ T follicular helper (TFH) cells guide development and maturation of B cells and are crucial for effective antibody responses. Here we found rhesus macaque TFH cells, defined as CXCR5+CD4 T cells, contain two major populations: PD-1INT and PD-1HIGH cells. Of these, PD-1HIGHCD4+ T cells highly co-express ICOS but little CCR7, and reside in lymph node germinal centers (GCs), but not in blood. These cells secrete IL-21 and express transcriptional factor Bcl-6 at higher levels than CXCR5+PD-1INTCD4+ T cells. In addition, the frequency of PD-1HIGHCD4+ T cells is low in lymph nodes of newborns, but increases with age. Levels of PD-1HIGHCD4+ T cells correlate with mature B cells in lymph nodes, and PD-1 blockade in PD-1HIGHCD4+ T and B cell co-cultures significantly inhibits IgG production. In summary, PD-1HIGHCD4+ T cells residing in GC represent a specific TFH subset that contributes to maturation of B cells and IgG production. PMID:24678309

The t(14;18) translocation is absent from endothelial and follicular dendritic cells of follicular lymphoma (FL) and shows heterogeneous presence in preserved FL mantle zones.

PubMed

Kosmidis, Perikles; Mankel, Barbara; Fend, Falko; Adam, Patrick

2018-05-02

The translocation t(14;18)(q32;q21) is the genetic hallmark of follicular lymphoma (FL) and can be observed in 85-90% of cases. Whether the translocation is restricted to cells with germinal center B-cell phenotype or can be observed in other cell types of the microenvironment remains debated. Of interest, cases of associated histiocytic and dendritic cell sarcomas arising in the background of FL have been shown to be clonally related and carry the t(14;18), suggesting a "transdifferentiation" of the malignant FL clone into a neoplasm of a different hematopoietic lineage. We analyzed the presence of the t(14;18)(q32;q21) as a surrogate marker of the malignant clone in cells of the FL microenvironment using combined fluorescence immunophenotyping and interphase cytogenetics targeting the BCL2 gene locus. In addition to non-lymphoid cells in FL, we analysed FL with preserved IgD+ mantle zones and cases of in situ follicular neoplasia (ISFN) to investigate whether cells of non-germinal center B-cell phenotype are part of the malignant clone. Six (40%) of 15 manifest FL cases with preserved IgD+ mantle zones did not harbour the t(14;18)(q32;q21) translocation. In all t(14;18) + FL cases, follicular dendritic cells and endothelial cells lacked the t(14;18) translocation. 2/9 FL revealed t(14;18)- IgD+ mantle zone B-cells. In the seven ISFN cases, the t(14;18) translocation was strictly confined to germinal center cells. The t(14;18) translocation in follicular lymphoma is limited to B-cells. The origin of IgD+ mantle cells is heterogeneous, in the majority of cases belonging to the neoplastic clone, whereas a minority of cases of manifest FL show nonneoplastic mantle zones, similar to ISFN.

Plasma cell output from germinal centers is regulated by signals from Tfh and stromal cells

PubMed Central

George, Laura A.; Acs, Andreas; Durrett, Russell E.

2018-01-01

Germinal centers (GCs) are the sites where B cells undergo affinity maturation. The regulation of cellular output from the GC is not well understood. Here, we show that from the earliest stages of the GC response, plasmablasts emerge at the GC–T zone interface (GTI). We define two main factors that regulate this process: Tfh-derived IL-21, which supports production of plasmablasts from the GC, and TNFSF13 (APRIL), which is produced by a population of podoplanin+ CD157high fibroblastic reticular cells located in the GTI that are also rich in message for IL-6 and chemokines CXCL12, CCL19, and CCL21. Plasmablasts in the GTI express the APRIL receptor TNFRSF13B (TACI), and blocking TACI interactions specifically reduces the numbers of plasmablasts appearing in the GTI. Plasma cells generated in the GTI may provide an early source of affinity-matured antibodies that may neutralize pathogens or provide feedback regulating GC B cell selection. PMID:29549115

Cells of the connective tissue differentiate and migrate into pollen sacs

NASA Astrophysics Data System (ADS)

Iqbal, M. C. M.; Wijesekara, Kolitha B.

2002-01-01

In angiosperms, archesporial cells in the anther primordium undergo meiosis to form haploid pollen, the sole occupants of anther sacs. Anther sacs are held together by a matrix of parenchyma cells, the connective tissue. Cells of the connective tissue are not known to differentiate. We report the differentiation of parenchyma cells in the connective tissue of two Gordonia species into pollen-like structures (described as pseudopollen), which migrate into the anther sacs before dehiscence. Pollen and pseudopollen were distinguishable by morphology and staining. Pollen were tricolpate to spherical while pseudopollen were less rigid and transparent with a ribbed surface. Both types were different in size, shape, staining and surface architecture. The ratio of the number of pseudopollen to pollen was 1:3. During ontogeny in the connective tissue, neither cell division nor tetrad formation was observed and hence pseudopollen were presumed to be diploid. Only normal pollen germinated on a germination medium. Fixed preparations in time seemed to indicate that pseudopollen migrate from the connective tissue into the anther sac.

Ethylene Glycol-Induced Alteration of Conidial Germination in Neurospora crassa

PubMed Central

Bates, W. K.; Wilson, J. F.

1974-01-01

In nutrient medium containing 3.22 M ethylene glycol or glycerol, conidia of Neurospora crassa grow as single cells, without forming the germ tubes characteristic of normal morphological germination. Ethylene glycol is more effective than glycerol in producing this response. After growth in ethylene glycol medium for a suitable time, the cells are easily disrupted by an abrupt decrease in osmotic pressure. Osmotic disruption yields intact nuclei and mitochondria, although mitochondrial fractions obtained in this way show significantly reduced concentrations of cytochromes c + c1, as compared to those observed for comparable fractions obtained from vegetative hyphae. Cell cultures gradually adapted to lower concentrations of the glycol show a much higher degree of synchrony in the formation of germ tubes than do untreated conidia. Images PMID:4359649

CCR6 Defines Memory B Cell Precursors in Mouse and Human Germinal Centers, Revealing Light-Zone Location and Predominant Low Antigen Affinity.

PubMed

Suan, Dan; Kräutler, Nike J; Maag, Jesper L V; Butt, Danyal; Bourne, Katherine; Hermes, Jana R; Avery, Danielle T; Young, Clara; Statham, Aaron; Elliott, Michael; Dinger, Marcel E; Basten, Antony; Tangye, Stuart G; Brink, Robert

2017-12-19

Memory B cells (MBCs) and plasma cells (PCs) constitute the two cellular outputs of germinal center (GC) responses that together facilitate long-term humoral immunity. Although expression of the transcription factor BLIMP-1 identifies cells undergoing PC differentiation, no such marker exists for cells committed to the MBC lineage. Here, we report that the chemokine receptor CCR6 uniquely marks MBC precursors in both mouse and human GCs. CCR6 + GC B cells were highly enriched within the GC light zone (LZ), were the most quiescent of all GC B cells, exhibited a cell-surface phenotype and gene expression signature indicative of an MBC transition, and possessed the augmented response characteristics of MBCs. MBC precursors within the GC LZ predominantly possessed a low affinity for antigen but also included cells from within the high-affinity pool. These data indicate a fundamental dichotomy between the processes that drive MBC and PC differentiation during GC responses. Copyright © 2017 Elsevier Inc. All rights reserved.

DISTAG/TBCCd1 Is Required for Basal Cell Fate Determination in Ectocarpus[OPEN

PubMed Central

Godfroy, Olivier; Uji, Toshiki; Nagasato, Chikako; Colin, Sebastien; Mignerot, Laure; Motomura, Taizo

2017-01-01

Brown algae are one of the most developmentally complex groups within the eukaryotes. As in many land plants and animals, their main body axis is established early in development, when the initial cell gives rise to two daughter cells that have apical and basal identities, equivalent to shoot and root identities in land plants, respectively. We show here that mutations in the Ectocarpus DISTAG (DIS) gene lead to loss of basal structures during both the gametophyte and the sporophyte generations. Several abnormalities were observed in the germinating initial cell in dis mutants, including increased cell size, disorganization of the Golgi apparatus, disruption of the microtubule network, and aberrant positioning of the nucleus. DIS encodes a TBCCd1 protein, which has a role in internal cell organization in animals, Chlamydomonas reinhardtii, and trypanosomes. Our study highlights the key role of subcellular events within the germinating initial cell in the determination of apical/basal cell identities in a brown alga and emphasizes the remarkable functional conservation of TBCCd1 in regulating internal cell organization across extremely distant eukaryotic groups. PMID:29208703

Basal Cell Carcinoma With Matrical Differentiation: Clinicopathologic, Immunohistochemical, and Molecular Biological Study of 22 Cases.

PubMed

Kyrpychova, Liubov; Carr, Richard A; Martinek, Petr; Vanecek, Tomas; Perret, Raul; Chottová-Dvořáková, Magdalena; Zamecnik, Michal; Hadravsky, Ladislav; Michal, Michal; Kazakov, Dmitry V

2017-06-01

Basal cell carcinoma (BCC) with matrical differentiation is a fairly rare neoplasm, with about 30 cases documented mainly as isolated case reports. We studied a series of this neoplasm, including cases with an atypical matrical component, a hitherto unreported feature. Lesions coded as BCC with matrical differentiation were reviewed; 22 cases were included. Immunohistochemical studies were performed using antibodies against BerEp4, β-catenin, and epithelial membrane antigen (EMA). Molecular genetic studies using Ion AmpliSeq Cancer Hotspot Panel v2 by massively parallel sequencing on Ion Torrent PGM were performed in 2 cases with an atypical matrical component (1 was previously subjected to microdissection to sample the matrical and BCC areas separately). There were 13 male and 9 female patients, ranging in age from 41 to 89 years. Microscopically, all lesions manifested at least 2 components, a BCC area (follicular germinative differentiation) and areas with matrical differentiation. A BCC component dominated in 14 cases, whereas a matrical component dominated in 4 cases. Matrical differentiation was recognized as matrical/supramatrical cells (n=21), shadow cells (n=21), bright red trichohyaline granules (n=18), and blue-gray corneocytes (n=18). In 2 cases, matrical areas manifested cytologic atypia, and a third case exhibited an infiltrative growth pattern, with the tumor metastasizing to a lymph node. BerEP4 labeled the follicular germinative cells, whereas it was markedly reduced or negative in matrical areas. The reverse pattern was seen with β-catenin. EMA was negative in BCC areas but stained a proportion of matrical/supramatrical cells. Genetic studies revealed mutations of the following genes: CTNNB1, KIT, CDKN2A, TP53, SMAD4, ERBB4, and PTCH1, with some differences between the matrical and BCC components. It is concluded that matrical differentiation in BCC in most cases occurs as multiple foci. Rare neoplasms manifest atypia in the matrical areas. Immunohistochemical analysis for BerEP4, EMA, and β-catenin can be helpful in limited biopsy specimens. From a molecular biological prospective, BCC and matrical components appear to share some of the gene mutations but have differences in others, but this observation must be validated in a large series.

Varying susceptibility of clinical and environmental Scedosporium isolates to chemical oxidative stress in conidial germination.

PubMed

Staerck, Cindy; Godon, Charlotte; Bouchara, Jean-Philippe; Fleury, Maxime J J

2018-04-01

Scedosporium species are opportunistic pathogens causing a great variety of infections in both immunocompetent and immunocompromised individuals. The Scedosporium genus ranks the second among the filamentous fungi colonizing the airways of patients with cystic fibrosis (CF), after Aspergillus fumigatus, and most species are capable to chronically colonize the respiratory tract of these patients. Nevertheless, few data are available regarding evasion of the inhaled conidia to the host immune response. Upon microbial infection, macrophages and neutrophils release reactive oxygen species (ROS). To colonize the respiratory tract, the conidia need to germinate despite the oxidative stress generated by phagocytic cells. Germination of spores from different clinical or environmental isolates of the major Scedosporium species was investigated in oxidative stress conditions. All tested species showed susceptibility to oxidative stress. However, when comparing clinical and environmental isolates, differences in germination capabilities under oxidative stress conditions were seen between species as well as within each species. Among environmental isolates, Scedosporium aurantiacum isolates were the most resistant to oxidative stress whereas Scedosporium dehoogii were the most susceptible. Overall, the differences observed between Scedosporium species in the capacity to germinate under oxidative stress conditions could explain their varying prevalence and pathogenicity.

Dynamic DNA methylation reconfiguration during seed development and germination.

PubMed

Kawakatsu, Taiji; Nery, Joseph R; Castanon, Rosa; Ecker, Joseph R

2017-09-15

Unlike animals, plants can pause their life cycle as dormant seeds. In both plants and animals, DNA methylation is involved in the regulation of gene expression and genome integrity. In animals, reprogramming erases and re-establishes DNA methylation during development. However, knowledge of reprogramming or reconfiguration in plants has been limited to pollen and the central cell. To better understand epigenetic reconfiguration in the embryo, which forms the plant body, we compared time-series methylomes of dry and germinating seeds to publicly available seed development methylomes. Time-series whole genome bisulfite sequencing reveals extensive gain of CHH methylation during seed development and drastic loss of CHH methylation during germination. These dynamic changes in methylation mainly occur within transposable elements. Active DNA methylation during seed development depends on both RNA-directed DNA methylation and heterochromatin formation pathways, whereas global demethylation during germination occurs in a passive manner. However, an active DNA demethylation pathway is initiated during late seed development. This study provides new insights into dynamic DNA methylation reprogramming events during seed development and germination and suggests possible mechanisms of regulation. The observed sequential methylation/demethylation cycle suggests an important role of DNA methylation in seed dormancy.

An in silico evaluation of treatment regimens for recurrent Clostridium difficile infection

PubMed Central

Blanco, Natalia; Foxman, Betsy; Malani, Anurag N.; Zhang, Min; Walk, Seth; Rickard, Alexander H.

2017-01-01

Background Clostridium difficile infection (CDI) is a significant nosocomial infection worldwide, that recurs in as many as 35% of infections. Risk of CDI recurrence varies by ribotype, which also vary in sporulation and germination rates. Whether sporulation/germination mediate risk of recurrence and effectiveness of treatment of recurring CDI remains unclear. We aim to assess the role of sporulation/germination patterns on risk of recurrence, and the relative effectiveness of the recommended tapered/pulsing regimens using an in silico model. Methods We created a compartmental in-host mathematical model of CDI, composed of vegetative cells, toxins, and spores, to explore whether sporulation and germination have an impact on recurrence rates. We also simulated the effectiveness of three tapered/pulsed vancomycin regimens by ribotype. Results Simulations underscored the importance of sporulation/germination patterns in determining pathogenicity and transmission. All recommended regimens for recurring CDI tested were effective in reducing risk of an additional recurrence. Most modified regimens were still effective even after reducing the duration or dosage of vancomycin. However, the effectiveness of treatment varied by ribotype. Conclusion Current CDI vancomycin regimen for treating recurrent cases should be studied further to better balance associated risks and benefits. PMID:28800598

Phytotoxicity and Cytotoxicity of Essential Oil from Leaves of Plectranthus amboinicus, Carvacrol, and Thymol in Plant Bioassays.

PubMed

Pinheiro, Patrícia Fontes; Costa, Adilson Vidal; Alves, Thammyres de Assis; Galter, Iasmini Nicoli; Pinheiro, Carlos Alexandre; Pereira, Alexandre Fontes; Oliveira, Carlos Magno Ramos; Fontes, Milene Miranda Praça

2015-10-21

The essential oil of Plectranthus amboinicus and its chemotypes, carvacrol and thymol, were evaluated on the germination and root and aerial growth of Lactuca sativa and Sorghum bicolor and in acting on the cell cycle of meristematic root cells of L. sativa. The main component found in the oil by analysis in gas chromatography-mass spectrometry and gas chromatography flame ionization detection was carvacrol (88.61% in area). At a concentration of 0.120% (w v(-1)), the oil and its chemotypes retarded or inhibited the germination and decreased root and aerial growth in monocot and dicot species used in the bioassays. In addition, all substances caused changes in the cell cycle of the meristematic cells of L. sativa, with chromosomal alterations occurring from the 0.015% (w v(-1)) concentration. The essential oil of P. amboinicus, carvacrol, and thymol have potential for use as bioherbicides.

Gene activity during germination of spores of the fern, Onoclea sensibilis. Cell-free translation analysis of mRNA of spores and the effect of alpha-amanitin on spore germination

NASA Technical Reports Server (NTRS)

Raghavan, V.

1992-01-01

Poly(A)-RNA fractions of dormant, dark-imbibed (non-germinating) and photoinduced (germinating) spores of Onoclea sensibilis were poor templates in the rabbit reticulocyte lysate protein synthesizing system, but the translational efficiency of poly(A)+RNA was considerably higher than that of unfractionated RNA. Poly(A)+RNA isolated from photoinduced spores had a consistently higher translational efficiency than poly(A)+RNA from dark-imbibed spores. Analysis of the translation products by one-dimensional polyacrylamide gel electrophoresis showed no qualitative differences in the mRNA populations of dormant, dark-imbibed, and photoinduced spores. However, poly(A)+RNA from dark-imbibed spores appeared to encode in vitro fewer detectable polypeptides at a reduced intensity than photoinduced spores. A DNA clone encoding the large subunit of maize ribulose bisphosphate carboxylase hybridized at strong to moderate intensity to RNA isolated from dark-imbibed spores, indicating the absence of mRNA degradation. Although alpha-amanitin did not inhibit the germination of spores, the drug prevented the elongation of the rhizoid and protonemal initial with a concomitant effect on the synthesis of poly(A)+RNA. These results are consistent with the view that some form of translational control involving stored mRNA operates during dark-imbibition and photoinduced germination of spores.

The role of thioredoxin h in protein metabolism during wheat (Triticum aestivum L.) seed germination.

PubMed

Guo, Hongxiang; Wang, Shaoxin; Xu, Fangfang; Li, Yongchun; Ren, Jiangping; Wang, Xiang; Niu, Hongbin; Yin, Jun

2013-06-01

Thioredoxin h can regulate the redox environment in the cell and play an important role in the germination of cereals. In the present study, the thioredoxin s antisense transgenic wheat with down-regulation of thioredoxin h was used to study the role of thioredoxin h in protein metabolism during germination of wheat seeds, and to explore the mechanism of the thioredoxin s antisense transgenic wheat seeds having high resistance to pre-harvest sprouting. The qRT-PCR results showed that the expression of protein disulfide isomerase in the thioredoxin s antisense transgenic wheat was up-regulated, which induced easily forming glutenin macropolymers and the resistance of storage proteins to degradation. The expression of serine protease inhibitor was also up-regulated in transgenic wheat, which might be responsible for the decreased activity of thiocalsin during the germination. The expression of WRKY6 in transgenic wheat was down-regulated, which was consistent with the decreased activity of glutamine oxoglutarate aminotransferase. In transgenic wheat, the activities of glutamate dehydrogenase, glutamic pyruvic transaminase and glutamic oxaloacetic transaminase were down-regulated, indicating that the metabolism of amino acid was lower than that in wild-type wheat during seed germination. A putative model for the role of thioredoxin h in protein metabolism during wheat seed germination was proposed and discussed. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

Early quantitative method for measuring germination in non-green spores of Dryopteris paleacea using an epifluorescence-microscope technique

NASA Technical Reports Server (NTRS)

Scheuerlein, R.; Wayne, R.; Roux, S. J.

1988-01-01

A method is described to determine germination by blue-light excited red fluorescence in the positively photoblastic spores of Dryopteris paleacea Sw. This fluorescence is due to chlorophyll as evidenced from 1) a fluorescence-emission spectrum in vivo, where a bright fluorescence around 675 nm is obtained only in red light (R)-irradiated spores and 2) in vitro measurements with acetone extracts prepared from homogenized spores. Significant amounts of chlorophyll can be found only in R-treated spores; this chlorophyll exhibits an emission band around 668 nm, when irradiated with 430 nm light at 21 degrees C. Compared to other criteria for germination, such as swelling of the cell, coat splitting, greening, and rhizoid formation, which require longer periods after induction for their expression, chlorophyll fluorescence can be used to quantify germination after two days. This result is confirmed by fluence-response curves for R-induced spore germination; the same relationship between applied R and germination is obtained by the evaluation with the epifluorescence method 2 days after the light treatment as compared with the evaluation with bright-field microscopy 5 days after the inducing R. Using this technique we show for the first time that Ca2+ contributes to the signal-transduction chain in phytochrome-mediated chlorophyll synthesis in spores of Dryopteris paleacea.

Regulation of normal B-cell differentiation and malignant B-cell survival by OCT2

PubMed Central

Hodson, Daniel J.; Shaffer, Arthur L.; Xiao, Wenming; Wright, George W.; Schmitz, Roland; Phelan, James D.; Yang, Yandan; Webster, Daniel E.; Rui, Lixin; Kohlhammer, Holger; Nakagawa, Masao; Waldmann, Thomas A.; Staudt, Louis M.

2016-01-01

The requirement for the B-cell transcription factor OCT2 (octamer-binding protein 2, encoded by Pou2f2) in germinal center B cells has proved controversial. Here, we report that germinal center B cells are formed normally after depletion of OCT2 in a conditional knockout mouse, but their proliferation is reduced and in vivo differentiation to antibody-secreting plasma cells is blocked. This finding led us to examine the role of OCT2 in germinal center-derived lymphomas. shRNA knockdown showed that almost all diffuse large B-cell lymphoma (DLBCL) cell lines are addicted to the expression of OCT2 and its coactivator OCA-B. Genome-wide chromatin immunoprecipitation (ChIP) analysis and gene-expression profiling revealed the broad transcriptional program regulated by OCT2 that includes the expression of STAT3, IL-10, ELL2, XBP1, MYC, TERT, and ADA. Importantly, genetic alteration of OCT2 is not a requirement for cellular addiction in DLBCL. However, we detected amplifications of the POU2F2 locus in DLBCL tumor biopsies and a recurrent mutation of threonine 223 in the DNA-binding domain of OCT2. This neomorphic mutation subtly alters the DNA-binding preference of OCT2, leading to the transactivation of noncanonical target genes including HIF1a and FCRL3. Finally, by introducing mutations designed to disrupt the OCT2–OCA-B interface, we reveal a requirement for this protein–protein interface that ultimately might be exploited therapeutically. Our findings, combined with the predominantly B-cell–restricted expression of OCT2 and the absence of a systemic phenotype in our knockout mice, suggest that an OCT2-targeted therapeutic strategy would be efficacious in both major subtypes of DLBCL while avoiding systemic toxicity. PMID:26993806

Increased growth and germination success in plants following hydrogen sulfide administration.

PubMed

Dooley, Frederick D; Nair, Suven P; Ward, Peter D

2013-01-01

This study presents a novel way of enhancing plant growth through the use of a non-petroleum based product. We report here that exposing either roots or seeds of multicellular plants to extremely low concentrations of dissolved hydrogen sulfide at any stage of life causes statistically significant increases in biomass including higher fruit yield. Individual cells in treated plants were smaller (~13%) than those of controls. Germination success and seedling size increased in, bean, corn, wheat, and pea seeds while time to germination decreases. These findings indicated an important role of H2S as a signaling molecule that can increase the growth rate of all species yet tested. The increased crop yields reported here has the potential to effect the world's agricultural output.

miR-199a and miR-497 Are Associated with Better Overall Survival due to Increased Chemosensitivity in Diffuse Large B-Cell Lymphoma Patients

PubMed Central

Troppan, Katharina; Wenzl, Kerstin; Pichler, Martin; Pursche, Beata; Schwarzenbacher, Daniela; Feichtinger, Julia; Thallinger, Gerhard G.; Beham-Schmid, Christine; Neumeister, Peter; Deutsch, Alexander

2015-01-01

Micro-RNAs (miRNAs) are short non-coding single-stranded RNA molecules regulating gene expression at the post-transcriptional level. miRNAs are involved in cell development, differentiation, apoptosis, and proliferation. miRNAs can either function as tumor suppressor genes or oncogenes in various important pathways. The expression of specific miRNAs has been identified to correlate with tumor prognosis. For miRNA expression analysis real-time PCR on 81 samples was performed, including 63 diffuse large B-cell lymphoma (DLBCL, 15 of germinal center B-cell like subtype, 17 non germinal center B-cell, 23 transformed, and eight unclassified) and 18 controls, including nine peripheral B-cells, 5 germinal-center B-cells, four lymphadenitis samples, and 4 lymphoma cell lines (RI-1, SUDHL4, Karpas, U2932). Expression levels of a panel of 11 miRNAs that have been previously involved in other types of cancer (miR-15b_2, miR-16_1*, miR-16_2, miR-16_2*, miR-27a, miR-27a*, miR-98-1, miR-103a, miR-185, miR-199a, and miR-497) were measured and correlated with clinical data. Furthermore, cell lines, lacking miR-199a and miR-497 expression, were electroporated with the two respective miRNAs and treated with standard immunochemotherapy routinely used in patients with DLBCL, followed by functional analyses including cell count and apoptosis assays. Seven miRNAs (miR-16_1*, miR-16_2*, miR-27a, miR-103, miR-185, miR-199, and miR-497) were statistically significantly up-regulated in DLBCL compared to normal germinal cells. However, high expression of miR-497 or miR-199a was associated with better overall survival (p = 0.042 and p = 0.007). Overexpression of miR-199a and miR-497 led to a statistically significant decrease in viable cells in a dose-dependent fashion after exposure to rituximab and various chemotherapeutics relevant in multi-agent lymphoma therapy. Our data indicate that elevated miR-199a and miR-497 levels are associated with improved survival in aggressive lymphoma patients most likely by modifying drug sensitivity to immunochemotherapy. This functional impairment may serve as a potential novel therapeutic target in future treatment of patients with DLBCL. PMID:26251897

Regulation and Characterization of a Newly Deduced Cell Wall Hydrolase Gene (cwlJ) Which Affects Germination of Bacillus subtilis Spores

PubMed Central

Ishikawa, Shu; Yamane, Kunio; Sekiguchi, Junichi

1998-01-01

The predicted amino acid sequence of Bacillus subtilis ycbQ (renamed cwlJ) exhibits high similarity to those of the deduced C-terminal catalytic domain of SleBs, the specific cortex-hydrolyzing enzyme of B. cereus and the deduced one of B. subtilis. We constructed a cwlJ::lacZ fusion in the B. subtilis chromosome. The β-galactosidase activity and results of Northern hybridization and primer extension analyses of the cwlJ gene indicated that it is transcribed by EςE RNA polymerase. cwlJ-deficient spores responded to both l-alanine and AGFK, the A580 values of spore suspensions decreased more slowly than in the case of the wild-type strain, and the mutant spores released less dipicolinic acid than did those of the wild-type strain during germination. However, the mutant spores released only slightly less hexosamine than did the wild-type spores. In contrast, B. subtilis sleB spores did not release hexosamine at a significant level. While cwlJ and sleB spores were able to germinate, CJSB (cwlJ sleB) spores could not germinate but exhibited initial germination reactions, e.g., partial decrease in A580 and slow release of dipicolinic acid. CJSB spores became slightly gray after 6 h in the germinant, but their refractility was much greater than that of sleB mutant spores. The roles of the sleB and cwlJ mutations in germination and spore maturation are also discussed. PMID:9515903

miR-28 regulates the germinal center reaction and blocks tumor growth in preclinical models of non-Hodgkin lymphoma

PubMed Central

Bartolomé-Izquierdo, Nahikari; Mur, Sonia M.

2017-01-01

Non-Hodgkin lymphoma comprises a variety of neoplasms, many of which arise from germinal center (GC)-experienced B cells. microRNA-28 (miR-28) is a GC-specific miRNA whose expression is lost in numerous mature B-cell neoplasms. Here we show that miR-28 regulates the GC reaction in primary B cells by impairing class switch recombination and memory B and plasma cell differentiation. Deep quantitative proteomics combined with transcriptome analysis identified miR-28 targets involved in cell-cycle and B-cell receptor signaling. Accordingly, we found that miR-28 expression diminished proliferation in primary and lymphoma cells in vitro. Importantly, miR-28 reexpression in human Burkitt (BL) and diffuse large B-cell lymphoma (DLBCL) xenografts blocked tumor growth, both when delivered in viral vectors or as synthetic, clinically amenable, molecules. Further, the antitumoral effect of miR-28 is conserved in a primary murine in vivo model of BL. Thus, miR-28 replacement is uncovered as a novel therapeutic strategy for DLBCL and BL treatment. PMID:28188132

B cell–derived IL-6 initiates spontaneous germinal center formation during systemic autoimmunity

PubMed Central

Arkatkar, Tanvi

2017-01-01

Recent studies have identified critical roles for B cells in triggering autoimmune germinal centers (GCs) in systemic lupus erythematosus (SLE) and other disorders. The mechanisms whereby B cells facilitate loss of T cell tolerance, however, remain incompletely defined. Activated B cells produce interleukin 6 (IL-6), a proinflammatory cytokine that promotes T follicular helper (TFH) cell differentiation. Although B cell IL-6 production correlates with disease severity in humoral autoimmunity, whether B cell–derived IL-6 is required to trigger autoimmune GCs has not, to our knowledge, been addressed. Here, we report the unexpected finding that a lack of B cell–derived IL-6 abrogates spontaneous GC formation in mouse SLE, resulting in loss of class-switched autoantibodies and protection from systemic autoimmunity. Mechanistically, B cell IL-6 production was enhanced by IFN-γ, consistent with the critical roles for B cell–intrinsic IFN-γ receptor signals in driving autoimmune GC formation. Together, these findings identify a key mechanism whereby B cells drive autoimmunity via local IL-6 production required for TFH differentiation and autoimmune GC formation. PMID:28899868

Nucleoside-modified mRNA vaccines induce potent T follicular helper and germinal center B cell responses.

PubMed

Pardi, Norbert; Hogan, Michael J; Naradikian, Martin S; Parkhouse, Kaela; Cain, Derek W; Jones, Letitia; Moody, M Anthony; Verkerke, Hans P; Myles, Arpita; Willis, Elinor; LaBranche, Celia C; Montefiori, David C; Lobby, Jenna L; Saunders, Kevin O; Liao, Hua-Xin; Korber, Bette T; Sutherland, Laura L; Scearce, Richard M; Hraber, Peter T; Tombácz, István; Muramatsu, Hiromi; Ni, Houping; Balikov, Daniel A; Li, Charles; Mui, Barbara L; Tam, Ying K; Krammer, Florian; Karikó, Katalin; Polacino, Patricia; Eisenlohr, Laurence C; Madden, Thomas D; Hope, Michael J; Lewis, Mark G; Lee, Kelly K; Hu, Shiu-Lok; Hensley, Scott E; Cancro, Michael P; Haynes, Barton F; Weissman, Drew

2018-06-04

T follicular helper (Tfh) cells are required to develop germinal center (GC) responses and drive immunoglobulin class switch, affinity maturation, and long-term B cell memory. In this study, we characterize a recently developed vaccine platform, nucleoside-modified, purified mRNA encapsulated in lipid nanoparticles (mRNA-LNPs), that induces high levels of Tfh and GC B cells. Intradermal vaccination with nucleoside-modified mRNA-LNPs encoding various viral surface antigens elicited polyfunctional, antigen-specific, CD4 + T cell responses and potent neutralizing antibody responses in mice and nonhuman primates. Importantly, the strong antigen-specific Tfh cell response and high numbers of GC B cells and plasma cells were associated with long-lived and high-affinity neutralizing antibodies and durable protection. Comparative studies demonstrated that nucleoside-modified mRNA-LNP vaccines outperformed adjuvanted protein and inactivated virus vaccines and pathogen infection. The incorporation of noninflammatory, modified nucleosides in the mRNA is required for the production of large amounts of antigen and for robust immune responses. © 2018 Pardi et al.

Prospective Isolation and Comparison of Human Germinal Matrix and Glioblastoma EGFR+ Populations with Stem Cell Properties.

PubMed

Tome-Garcia, Jessica; Tejero, Rut; Nudelman, German; Yong, Raymund L; Sebra, Robert; Wang, Huaien; Fowkes, Mary; Magid, Margret; Walsh, Martin; Silva-Vargas, Violeta; Zaslavsky, Elena; Friedel, Roland H; Doetsch, Fiona; Tsankova, Nadejda M

2017-05-09

Characterization of non-neoplastic and malignant human stem cell populations in their native state can provide new insights into gliomagenesis. Here we developed a purification strategy to directly isolate EGFR +/- populations from human germinal matrix (GM) and adult subventricular zone autopsy tissues, and from de novo glioblastoma (GBM) resections, enriching for cells capable of binding EGF ligand ( LB EGFR + ), and uniquely compared their functional and molecular properties. LB EGFR + populations in both GM and GBM encompassed all sphere-forming cells and displayed proliferative stem cell properties in vitro. In xenografts, LB EGFR + GBM cells showed robust tumor initiation and progression to high-grade, infiltrative gliomas. Whole-transcriptome sequencing analysis confirmed enrichment of proliferative pathways in both developing and neoplastic freshly isolated EGFR + populations, and identified both unique and shared sets of genes. The ability to prospectively isolate stem cell populations using native ligand-binding capacity opens new doors onto understanding both normal human development and tumor cell biology. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Analysis of Peroxisome Biogenesis in Pollen by Confocal Microscopy and Transmission Electron Microscopy.

PubMed

Jia, Peng-Fei; Li, Hong-Ju; Yang, Wei-Cai

2017-01-01

Peroxisome is an essential single-membrane bound organelle in most eukaryotic cells and functions in diverse cellular processes. De novo formation, division, and turnover of peroxisomes contribute to its biogenesis, morphology, and population regulation. In plants, peroxisome plays multiple roles, including metabolism, development, and stress response. Defective peroxisome biogenesis and development retard plant growth, adaption, and reproduction. Through tracing the subcellular localization of fluorescent reporter tagged matrix protein of peroxisome, fluorescence microscopy is a reliable and fast way to detect peroxisome biogenesis. Further fine-structural observation of peroxisome by TEM enables researchers to observe the detailed ultrastructure of its morphology and spatial contact with other organelles. Pollen grain is a specialized structure where two small sperm cells are enclosed in the cytoplasm of a large vegetative cell. Two features make pollen grain a good system to study peroxisome biogenesis: indispensable requirement of peroxisome for germination on the stigma and homogeneity. Here, we describe the methods of studying peroxisome biogenesis in Arabidopsis pollen grains by fluorescent live-imaging with confocal laser scanning microscopy (CLSM) and by DAB-staining based transmission electron microscopy (TEM).

AtNG1 encodes a protein that is required for seed germination.

PubMed

Yang, Libo; Peng, Xiongbo; Sun, Meng-xiang

2011-10-01

The pentatricopeptide repeat (PPR) family of eukaryotic proteins has numerous members in plants and is important for plant development. In the present study, we cloned a novel PPR gene, designated AtNG1, and characterized the ng1 Arabidopsis mutant. Morphological and structural observation of an ng1 mutant revealed that its sexual reproduction and seed formation processes are essentially normal. The mature embryonic root of ng1 is fully developed and has a well-differentiated structure; however, ng1 seeds cannot germinate, even when supplied with supplemental hormones and nutrition. Further investigation showed that embryo expansion and root cell elongation fails to occur after water imbibitions. Transient gene expression analysis indicated that AtNG1 localizes in mitochondrion. This implies that the deficiency of mitochondrion function might be the reason for the failed seed germination. Thus, our finding confirmed that AtNG1 plays a critical role in the early process of seed germination. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

Spore coat architecture of Clostridium novyi NT spores.

PubMed

Plomp, Marco; McCaffery, J Michael; Cheong, Ian; Huang, Xin; Bettegowda, Chetan; Kinzler, Kenneth W; Zhou, Shibin; Vogelstein, Bert; Malkin, Alexander J

2007-09-01

Spores of the anaerobic bacterium Clostridium novyi NT are able to germinate in and destroy hypoxic regions of tumors in experimental animals. Future progress in this area will benefit from a better understanding of the germination and outgrowth processes that are essential for the tumorilytic properties of these spores. Toward this end, we have used both transmission electron microscopy and atomic force microscopy to determine the structure of both dormant and germinating spores. We found that the spores are surrounded by an amorphous layer intertwined with honeycomb parasporal layers. Moreover, the spore coat layers had apparently self-assembled, and this assembly was likely to be governed by crystal growth principles. During germination and outgrowth, the honeycomb layers, as well as the underlying spore coat and undercoat layers, sequentially dissolved until the vegetative cell was released. In addition to their implications for understanding the biology of C. novyi NT, these studies document the presence of proteinaceous growth spirals in a biological organism.

Control of seed development in Arabidopsis thaliana by atmospheric oxygen

NASA Technical Reports Server (NTRS)

Kuang, A.; Crispi, M.; Musgrave, M. E.

1998-01-01

Seed development is known to be inhibited completely when plants are grown in oxygen concentrations below 5.1 kPa, but apart from reports of decreased seed weight little is known about embryogenesis at subambient oxygen concentrations above this critical level. Arabidopsis thaliana (L.) Heynh. plants were grown full term under continuous light in premixed atmospheres with oxygen partial pressures of 2.5, 5.1, 10.1, 16.2 and 21.3 kPa O2, 0.035 kPa CO2 and the balance nitrogen. Seeds were harvested for germination tests and microscopy when siliques had yellowed. Seed germination was depressed in O2 treatments below 16.2 kPa, and seeds from plants grown in 2.5 kPa O2 did not germinate at all. Fewer than 25% of the seeds from plants grown in 5.1 kPa oxygen germinated and most of the seedlings appeared abnormal. Light and scanning electron microscopic observation of non-germinated seeds showed that these embryos had stopped growing at different developmental stages depending upon the prevailing oxygen level. Embryos stopped growing at the heart-shaped to linear cotyledon stage in 5.1 kPa O2, at around the curled cotyledon stage in 10.1 kPa O2, and at the premature stage in 16.2 kPa O2. Globular and heart-shaped embryos were observed in sectioned seeds from plants grown in 2.5 kPa O2. Tissue degeneration caused by cell autolysis and changes in cell structure were observed in cotyledons and radicles. Transmission electron microscopy of mature seeds showed that storage substances, such as protein bodies, were reduced in subambient oxygen treatments. The results demonstrate control of embryo development by oxygen in Arabidopsis.

Seed priming with iron and zinc in bread wheat: effects in germination, mitosis and grain yield.

PubMed

Reis, Sara; Pavia, Ivo; Carvalho, Ana; Moutinho-Pereira, José; Correia, Carlos; Lima-Brito, José

2018-07-01

Currently, the biofortification of crops like wheat with micronutrients such as iron (Fe) and zinc (Zn) is extremely important due to the deficiencies of these micronutrients in the human diet and in soils. Agronomic biofortification with Fe and Zn can be done through different exogenous strategies such as soil application, foliar spraying, and seed priming. However, the excess of these micronutrients can be detrimental to the plants. Therefore, in the last decade, a high number of studies focused on the evaluation of their phytotoxic effects to define the best strategies for biofortification of bread wheat. In this study, we investigated the effects of seed priming with different dosages (1 mg L -1 to 8 mg L -1 ) of Fe and/or Zn in germination, mitosis and yield of bread wheat cv. 'Jordão' when compared with control. Overall, our results showed that: micronutrient dosages higher than 4 mg L -1 negatively affect the germination; Fe and/or Zn concentrations higher than 2 mg L -1 significantly decrease the mitotic index and increase the percentage of dividing cells with anomalies; treatments performed with 8 mg L -1 of Fe and/or 8 mg L -1 Zn caused negative effects in germination, mitosis and grain yield. Moreover, seed priming with 2 mg L -1 Fe + 2 mg L -1 Zn has been shown to be non-cytotoxic, ensuring a high rate of germination (80%) and normal dividing cells (90%) as well as improving tillering and grain yield. This work revealed that seed priming with Fe and Zn micronutrients constitutes a useful and alternative approach for the agronomic biofortification of bread wheat.

A Diffusible Signal from Arbuscular Mycorrhizal Fungi Elicits a Transient Cytosolic Calcium Elevation in Host Plant Cells1[W

PubMed Central

Navazio, Lorella; Moscatiello, Roberto; Genre, Andrea; Novero, Mara; Baldan, Barbara; Bonfante, Paola; Mariani, Paola

2007-01-01

The implication of calcium as intracellular messenger in the arbuscular mycorrhizal (AM) symbiosis has not yet been directly demonstrated, although often envisaged. We used soybean (Glycine max) cell cultures stably expressing the bioluminescent Ca2+ indicator aequorin to detect intracellular Ca2+ changes in response to the culture medium of spores of Gigaspora margarita germinating in the absence of the plant partner. Rapid and transient elevations in cytosolic free Ca2+ were recorded, indicating that diffusible molecules released by the mycorrhizal fungus are perceived by host plant cells through a Ca2+-mediated signaling. Similar responses were also triggered by two Glomus isolates. The fungal molecules active in generating the Ca2+ transient were constitutively released in the medium, and the induced Ca2+ signature was not modified by the coculture of germinating spores with plant cells. Even ungerminated spores were able to generate the signaling molecules, as proven when the germination was blocked by a low temperature. The fungal molecules were found to be stable to heat treatment, of small molecular mass (<3 kD), and, on the basis of extraction with an organic solvent, partially lipophilic. Evidence for the specificity of such an early fungal signal to the AM symbiosis is suggested by the lack of a Ca2+ response in cultured cells of the nonhost plant Arabidopsis (Arabidopsis thaliana) and by the up-regulation in soybean cells of genes related to Medicago truncatula DMI1, DMI2, and DMI3 and considered essential for the establishment of the AM symbiosis. PMID:17142489

The barley anion channel, HvALMT1, has multiple roles in guard cell physiology and grain metabolism.

PubMed

Xu, Muyun; Gruber, Benjamin D; Delhaize, Emmanuel; White, Rosemary G; James, Richard A; You, Jiangfeng; Yang, Zhenming; Ryan, Peter R

2015-01-01

The barley (Hordeum vulgare) gene HvALMT1 encodes an anion channel in guard cells and in certain root tissues indicating that it may perform multiple roles. The protein localizes to the plasma membrane and facilitates malate efflux from cells when constitutively expressed in barley plants and Xenopus oocytes. This study investigated the function of HvALMT1 further by identifying its tissue-specific expression and by generating and characterizing RNAi lines with reduced HvALMT1 expression. We show that transgenic plants with 18-30% of wild-type HvALMT1 expression had impaired guard cell function. They maintained higher stomatal conductance in low light intensity and lost water more rapidly from excised leaves than the null segregant control plants. Tissue-specific expression of HvALMT1 was investigated in developing grain and during germination using transgenic barley lines expressing the green fluorescent protein (GFP) with the HvALMT1 promoter. We found that HvALMT1 is expressed in the nucellar projection, the aleurone layer and the scutellum of developing barley grain. Malate release measured from isolated aleurone layers prepared from imbibed grain was significantly lower in the RNAi barley plants compared with control plants. These data provide molecular and physiological evidence that HvALMT1 functions in guard cells, in grain development and during germination. We propose that HvALMT1 releases malate and perhaps other anions from guard cells to promote stomatal closure. The likely roles of HvALMT1 during seed development and grain germination are also discussed. © 2014 Scandinavian Plant Physiology Society.

Morphogenetic circuitry regulating growth and development in the dimorphic pathogen Penicillium marneffei.

PubMed

Boyce, Kylie J; Andrianopoulos, Alex

2013-02-01

Penicillium marneffei is an emerging human-pathogenic fungus endemic to Southeast Asia. Like a number of other fungal pathogens, P. marneffei exhibits temperature-dependent dimorphic growth and grows in two distinct cellular morphologies, hyphae at 25°C and yeast cells at 37°C. Hyphae can differentiate to produce the infectious agents, asexual spores (conidia), which are inhaled into the host lung, where they are phagocytosed by pulmonary alveolar macrophages. Within macrophages, conidia germinate into unicellular yeast cells, which divide by fission. This minireview focuses on the current understanding of the genes required for the morphogenetic control of conidial germination, hyphal growth, asexual development, and yeast morphogenesis in P. marneffei.

[Malignant ovarian tumors and second look operations].

PubMed

Yasuda, M

1983-08-01

There are many problems that, the timing of S.L.O. the range of reduction, and side effects etc., we studied retrospectively for the S.L.O. The number of subjects were 302 of common epithelial and 251 of germinal cell tumors, totalling 553 cases. Among them, 41 cases were common epithelial and 49 cases germ cell origine respectively, totalling 90 cases of S.L.O. were carried out. Conclusion The 5 year survival rate of S.L.O. was observed to a significant extent. The mean survival time of S.L.O. group: were significantly prolonged. Complete surgery at the S.L.O. of the patient with epithelial and germinal affected the survival rate significantly.

Encystment of Azotobacter nigricans grown diazotrophically on kerosene as sole carbon source.

PubMed

García-Esquivel, Gabriela; Calva-Calva, Graciano; Ferrera-Cerrato, Ronald; Fernández-Linares, Luis Carlos; Vázquez, Refugio Rodríguez; Esparza-García, Fernando José

2009-03-01

Encystment of Azotobacter nigricans was induced by its diazotrophic cultivation on kerosene. Its growth and nitrogenase activity were affected by kerosene in comparison to cultures grown on sucrose. Electron microscopy of vegetative cells showed that when nitrogenase activity was higher and the poly-beta-hydroxybutyrate granules were not present to a significant extent, peripheral bodies were abundant. After 8 days of culture on kerosene, the presence of cysts with intracellular bunches of poly-beta-hydroxybutyrate granules was observed. Germination of cysts bears germinating multicelled yet unbroken capsule cysts with up to three cells inside. This is the first report of encystment induction of Azotobacter species grown on kerosene.

Mouse model for acute Epstein-Barr virus infection.

PubMed

Wirtz, Tristan; Weber, Timm; Kracker, Sven; Sommermann, Thomas; Rajewsky, Klaus; Yasuda, Tomoharu

2016-11-29

Epstein-Barr Virus (EBV) infects human B cells and drives them into continuous proliferation. Two key viral factors in this process are the latent membrane proteins LMP1 and LMP2A, which mimic constitutively activated CD40 receptor and B-cell receptor signaling, respectively. EBV-infected B cells elicit a powerful T-cell response that clears the infected B cells and leads to life-long immunity. Insufficient immune surveillance of EBV-infected B cells causes life-threatening lymphoproliferative disorders, including mostly germinal center (GC)-derived B-cell lymphomas. We have modeled acute EBV infection of naive and GC B cells in mice through timed expression of LMP1 and LMP2A. Although lethal when induced in all B cells, induction of LMP1 and LMP2A in just a small fraction of naive B cells initiated a phase of rapid B-cell expansion followed by a proliferative T-cell response, clearing the LMP-expressing B cells. Interfering with T-cell activity prevented clearance of LMP-expressing B cells. This was also true for perforin deficiency, which in the human causes a life-threatening EBV-related immunoproliferative syndrome. LMP expression in GC B cells impeded the GC reaction but, upon loss of T-cell surveillance, led to fatal B-cell expansion. Thus, timed expression of LMP1 together with LMP2A in subsets of mouse B cells allows one to study major clinically relevant features of human EBV infection in vivo, opening the way to new therapeutic approaches.

Fluorescence-based methods for the detection of pressure-induced spore germination and inactivation

NASA Astrophysics Data System (ADS)

Baier, Daniel; Reineke, Kai; Doehner, Isabel; Mathys, Alexander; Knorr, Dietrich

2011-03-01

The application of high pressure (HP) provides an opportunity for the non-thermal preservation of high-quality foods, whereas highly resistant bacterial endospores play an important role. It is known that the germination of spores can be initiated by the application of HP. Moreover, the resistance properties of spores are highly dependent on their physiological states, which are passed through during the germination. To distinguish between different physiological states and to detect the amount of germinated spores after HP treatments, two fluorescence-based methods were applied. A flow cytometric method using a double staining with SYTO 16 as an indicator for germination and propidium iodide as an indicator for membrane damage was used to detect different physiological states of the spores. During the first step of germination, the spore-specific dipicolinic acid (DPA) is released [P. Setlow, Spore germination, Curr. Opin. Microbiol. 6 (2003), pp. 550-556]. DPA reacts with added terbium to form a distinctive fluorescent complex. After measuring the fluorescence intensity at 270 nm excitation wavelength in a fluorescence spectrophotometer, the amount of germinated spores can be determined. Spores of Bacillus subtilis were treated at pressures from 150 to 600 MPa and temperatures from 37 °C to 60 °C in 0.05 M ACES buffer solution (pH 7) for dwell times of up to 2 h. During the HP treatments, inactivation up to 2log 10 cycles and thermal sensitive populations up to 4log 10 cycles could be detected by plate counts. With an increasing number of thermal sensitive spores, an increased proportion of spores in germinated states was detected by flow cytometry. Also the released amount of DPA increased during the dwell times. Moreover, a clear pressure-temperature-time-dependency was shown by screening different conditions. The fluorescence-based measurement of the released DPA can provide the opportunity of an online monitoring of the germination of spores under HP inside the HP vessel. Implementation can be done using diamond anvil cells, units with inspection glasses or by inserting an optical fiber into the HP vessel. The analytical methods used can help to understand the complex mechanism of germination and inactivation of bacterial spores. Due to its universal, process-independent character, the application of these methods is feasible for established and emerging technologies.

Autoradiography and the Cell Cycle.

ERIC Educational Resources Information Center

Jones, C. Weldon

1992-01-01

Outlines the stages of a cell biology "pulse-chase" experiment in which the students apply autoradiography techniques to learn about the concept of the cell cycle. Includes (1) seed germination and plant growth; (2) radioactive labeling and fixation of root tips; (3) feulgen staining of root tips; (4) preparation of autoradiograms; and…

B cells in tertiary lymphoid structures are associated with favorable prognosis in gastric cancer.

PubMed

Sakimura, Chie; Tanaka, Hiroaki; Okuno, Takahiro; Hiramatsu, Soichiro; Muguruma, Kazuya; Hirakawa, Kosei; Wanibuchi, Hideki; Ohira, Masaichi

2017-07-01

The role of tumor-infiltrating B cells in the tumor microenvironment is still unclear. Recent studies have reported that B cells and tertiary lymphoid structures (TLSs) that contain B cell follicles correlate with the favorable prognosis of cancer patients. The aim of this study was to investigate the association between tumor-infiltrating B cells and clinicopathological features in gastric cancer. Tumor blocks were obtained from 226 patients with stage Ib to stage IV gastric cancer. The density of CD20 + B cells within the tumor and in the invasive margin area was assessed using immunohistochemistry. We also evaluated CD3 + T cells, CD21 + follicular dendritic cells, Bcl6 + germinal center B cells, and PNAd + high endothelial venules to show the presence of TLSs. Tumor-infiltrating B cells were mostly organized as clusters that were surrounded by CD3 + T cells. The B cell area contained follicular dendritic cells and some clusters contained Bcl6 + B cells. High endothelial venules were present around follicles. We identified these follicles as TLSs. A high number of CD20 + B cells were associated with significantly better overall survival, and multivariate analysis also showed that CD20 high was one of the independent predictors of prognosis. In addition, there was a significant correlation between CD20 + B cell and CD8 + T cell infiltration. B cells mostly infiltrated tumors as TLSs and were associated with better prognosis in patients with gastric cancer. Copyright © 2017 Elsevier Inc. All rights reserved.

Pediatric nodal marginal zone lymphoma may develop in the adult population.

PubMed

Gitelson, Elena; Al-Saleem, Tahseen; Robu, Valentin; Millenson, Michael M; Smith, Mitchell R

2010-01-01

Pediatric nodal marginal zone lymphoma (NMZL) is described as a separate variant of NMZL in the most recent WHO classification of tumors of hematologic and lymphoid tissues. It has distinctive morphology and clinical presentation and stands out as an indolent disease with remarkably better overall prognosis compared to classic NMZL. Here we report two adult patients with NMZL with clinical and morphologic features consistent with pediatric NMZL (pNMZL) and review available literature describing the clinical and histologic presentation of pNMZL. Two men, ages 44 and 18 years, each presented with localized cervical lymphadenopathy, both demonstrated florid proliferation of the marginal zone and disruption of reactive germinal centers, progressive transformation of germinal centers-like morphologic features typical for pNMZL and clonal disease with immunophenotype consistent with NMZL. This is the first report of pNMZL in a middle-aged person. Distinct histologic features and characteristic benign clinical course will help to distinguish this rare variant from other NMZL in the adults. Clinically, recognition is important to understand the true incidence of this rare form in the adult population and to avoid unnecessary overtreatment of this indolent form.

Taka-amylase A in the conidia of Aspergillus oryzae RIB40.

PubMed

Nguyen, Cong Ha; Tsurumizu, Ryoji; Sato, Tsutomu; Takeuchi, Michio

2005-11-01

A study of Taka-amylase A of conidia from Aspergillus oryzae RIB40 was done. During the research, proteins from conidia and germinated conidia were analyzed using SDS-PAGE, 2-D gel electrophoresis, Western blot analysis, MALDI-TOF Mass spectrometry, and native-PAGE combined with activity staining of TAA. The results showed that TAA exists not only in germinated conidia but also in conidia. Some bands representing degraded products of TAA were detected. Conidia, which formed on starch (SCYA), glucose (DCYA), and glycerol (GCYA) plates, contained mature TAA. Only one active band of TAA was detected after native-PAGE activity staining. In addition, TAA activity was detected in cell extracts of conidia using 0.5 M acetate buffer, pH 5.2, as extraction buffer, but was not detected in whole conidia or cell debris. The results indicate that TAA exists in conidia in active form even when starch, glucose, or glycerol is used as carbon source. TAA might belong to a set of basal proteins inside conidia, which helps in imbibition and germination of conidia.

Utility of LRF/Pokemon and NOTCH1 Protein Expression in the Distinction of Nodular Lymphocyte-Predominant Hodgkin Lymphoma and Classical Hodgkin Lymphoma

PubMed Central

Bohn, Olga; Maeda, Takahiro; Filatov, Alexander; Lunardi, Andrea; Pandolfi, Pier Paolo; Teruya-Feldstein, Julie

2014-01-01

Classical Hodgkin lymphoma (CHL) and nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL) are considered separate entities with different prognosis and treatment. However, morphologic features can be similar and immunohistochemical studies are essential in the distinction; thus, determination of additional biomarkers is of utmost importance. LRF/Pokemon is a protooncogene, an interacting partner co-expressed with BCL6 in germinal centers and highly expressed in diffuse large B-cell lymphoma and follicular lymphoma. Conversely, loss of the LRF gene in mouse hematopoietic stem cells results in complete block of early B cell development with concomitant Notch derepression, indicating its critical role in B versus T cell fate decision at the hematopoietic stem cell stage. For the first time, we show that LRF/Pokemon is predominantly expressed in NLPHL cases as is BCL6 with low to absent NOTCH1 protein expression; while Hodgkin Reed-Sternberg (HRS) cells in CHL show low to absent BCL6 and LRF/Pokemon expression with higher NOTCH1 expression. We illustrate a potential functional interaction between LRF and BCL6 in NLPHL pathogenesis, and differential expression of LRF/Pokemon and NOTCH1 proteins in CHL thus showing differential expression, making for an additional diagnostic marker and therapeutic target. PMID:24326827

Utility of LRF/Pokemon and NOTCH1 protein expression in the distinction between nodular lymphocyte-predominant Hodgkin lymphoma and classical Hodgkin lymphoma.

PubMed

Bohn, Olga; Maeda, Takahiro; Filatov, Alexander; Lunardi, Andrea; Pandolfi, Pier Paolo; Teruya-Feldstein, Julie

2014-02-01

Classical Hodgkin lymphoma (CHL) and nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL) are considered separate entities with different prognosis and treatment. However, morphologic features can be similar and immunohistochemical studies are essential in the distinction; thus, determination of additional biomarkers is of utmost importance. LRF/Pokemon is a proto-oncogene, an interacting partner co-expressed with BCL6 in germinal centers and highly expressed in diffuse large B-cell lymphoma and follicular lymphoma. Conversely, loss of the LRF gene in mouse hematopoietic stem cells results in complete block of early B cell development with concomitant Notch de-repression, indicating its critical role in B versus T cell fate decision at the hematopoietic stem cell stage. For the first time, we show that LRF/Pokemon is predominantly expressed in NLPHL cases as is BCL6 with low to absent NOTCH1 protein expression; while Hodgkin Reed-Sternberg (HRS) cells in CHL show low to absent BCL6 and LRF/Pokemon expression with higher NOTCH1 expression. We illustrate a potential functional interaction between LRF and BCL6 in NLPHL pathogenesis, and differential expression of LRF/Pokemon and NOTCH1 proteins in CHL thus showing differential expression, making for an additional diagnostic marker and therapeutic target.

Analysis of immunoglobulin heavy and light chain variable genes in post-transplant lymphoproliferative disorders.

PubMed

Capello, Daniela; Cerri, Michaela; Muti, Giuliana; Lucioni, Marco; Oreste, Pierluigi; Gloghini, Annunziata; Berra, Eva; Deambrogi, Clara; Franceschetti, Silvia; Rossi, Davide; Alabiso, Oscar; Morra, Enrica; Rambaldi, Alessandro; Carbone, Antonino; Paulli, Marco; Gaidano, Gianluca

2006-12-01

Post-transplant lymphoproliferative disorders (PTLD) derive from antigen-experienced B-cells and represent a major complication of solid organ transplantation. We characterized usage, mutation frequency and mutation pattern of immunoglobulin variable (IGV) gene rearrangements in 50 PTLD (polymorphic PTLD, n=10; diffuse large B-cell lymphoma, n=35; and Burkitt/Burkitt-like lymphoma, n=5). Among PTLD yielding clonal IGV amplimers, a functional IGV heavy chain (IGHV) rearrangement was found in 40/50 (80.0%) cases, whereas a potentially functional IGV light chain rearrangement was identified in 36/46 (78.3%) PTLD. By combining IGHV and IGV light chain rearrangements, 10/50 (20.0%) PTLD carried crippling mutations, precluding expression of a functional B-cell receptor (BCR). Immunohistochemistry showed detectable expression of IG light chains in only 18/43 (41.9%) PTLD. Failure to detect a functional IGV rearrangement associated with lack of IGV expression. Our data suggest that a large fraction of PTLD arise from germinal centre (GC)-experienced B-cells that display impaired BCR. Since a functional BCR is required for normal B-cell survival during GC transit, PTLD development may implicate rescue from apoptosis and expansion of B-cells that have failed the GC reaction. The high frequency of IGV loci inactivation appears to be a peculiar feature of PTLD among immunodeficiency-associated lymphoproliferations.

The Achene Mucilage Hydrated in Desert Dew Assists Seed Cells in Maintaining DNA Integrity: Adaptive Strategy of Desert Plant Artemisia sphaerocephala

PubMed Central

Yang, Xuejun; Zhang, Wenhao; Dong, Ming; Boubriak, Ivan; Huang, Zhenying

2011-01-01

Despite proposed ecological importance of mucilage in seed dispersal, germination and seedling establishment, little is known about the role of mucilage in seed pre-germination processes. Here we investigated the role of mucilage in assisting achene cells to repair DNA damage during dew deposition in the desert. Artemisia sphaerocephala achenes were first treated γ-irradiation to induce DNA damage, and then they were repaired in situ in the desert dew. Dew deposition duration can be as long as 421 min in early mornings. Intact achenes absorbed more water than demucilaged achenes during dew deposition and also carried water for longer time following sunrise. After 4-d dew treatment, DNA damage of irradiated intact and demucilaged achenes was reduced to 24.38% and 46.84%, respectively. The irradiated intact achenes exhibited much higher DNA repair ratio than irradiated demucilaged achenes. Irradiated intact achenes showed an improved germination and decreased nonviable achenes after dew treatment, and significant differences in viability between the two types of achenes were detected after 1020 min of dew treatment. Achene mucilage presumably plays an ecologically important role in the life cycle of A. sphaerocephala by aiding DNA repair of achene cells in genomic-stressful habitats. PMID:21912689

Lineage-specific functions of Bcl-6 in immunity and inflammation are mediated through distinct biochemical mechanisms

PubMed Central

Huang, Chuanxin; Hatzi, Katerina; Melnick, Ari

2013-01-01

The transcription factor Bcl-6 orchestrates the germinal center reaction through its actions in B and T cells, and regulates inflammatory signaling in macrophages. We report that genetic replacement by mutant Bcl-6, which cannot bind corepressors to its BTB domain, disrupted germinal center formation and immunoglobulin affinity maturation, due to a defect in B cell proliferation and survival. In contrast, BTB loss of function had no effect on T follicular helper cell differentiation and function, nor other T helper subsets. Bcl6 null mice displayed a lethal inflammatory phenotype, whereas BTB mutant mice experienced normal healthy lives with no inflammation. Bcl-6 repression of inflammatory responses in macrophages was accordingly independent of the BTB domain repressor function. Bcl-6 thus mediates its actions through lineage-specific biochemical functions. PMID:23455674

Coexistence of age-related EBV-associated follicular hyperplasia and large B-cell EBV+ lymphoma of the elderly. Two distinct features of the same T-cell dysfunction related to senescence?

PubMed

Gibier, Jean-Baptiste; Bouchindhomme, Brigitte; Dubois, Romain; Hivert, Benedicte; Grardel, Nathalie; Copin, Marie-Christine

2017-03-01

Age-related EBV-associated lymphoproliferative disorders form a new clinicopathological group. Until recently, this group was associated with diffuse large B-cell lymphoma (DLBCL), characterised by an aggressive clinical presentation and a poor prognosis. Recent findings in Western Caucasian patients, however, suggest that this entity covers a wide spectrum of diseases, ranging from reactive follicular hyperplasia (HR) to DLBCL. We report the case of an 85-year-old Caucasian man showing lymphadenopathy and numerous hypodense lesions of the liver. Examination of a lymph node revealed follicular hyperplasia with EBV expression confined to germinal centres. The patient was treated with Rituximab and subsequently, the lesions of the liver were explored. They showed extensive necrosis and a polymorphic large B-cell population with strong EBV expression. This is the first report to describe age-related EBV-associated follicular hyperplasia at one site coexisting with DLBCL at another. This case warrants undertaking further investigations each time a diagnosis of age-related EBV-HR is associated with extranodal lesions. Copyright © 2016 Elsevier GmbH. All rights reserved.

Tonsillar CD56brightNKG2A+ NK cells restrict primary Epstein-Barr virus infection in B cells via IFN-γ.

PubMed

Jud, Aurelia; Kotur, Monika; Berger, Christoph; Gysin, Claudine; Nadal, David; Lünemann, Anna

2017-01-24

Natural killer (NK) cells constitute the first line of defense against viruses and cancers cells. Epstein-Barr virus (EBV) was the first human virus to be directly implicated in carcinogenesis, and EBV infection is associated with a broad spectrum of B cell lymphomas. How NK cells restrict EBV-associated oncogenesis is not understood. Here, we investigated the efficacies and mechanisms of distinct NK cell subsets from tonsils, the portal of entry of EBV, in limiting EBV infection in naïve, germinal center-associated and memory B cells. We found that CD56bright and NKG2A expression sufficiently characterizes the potent anti-EBV capacity of tonsillar NK cells. We observed restriction of EBV infection in B cells as early as 18 hours after infection. The restriction was most efficient in naïve B cells and germinal center-associated B cells, the B cell subsets that exhibited highest susceptibility to EBV infection in vitro. IFN-γ release by and partially NKp44 engagement of CD56bright and NKG2A positive NK cells mediated the restriction that eventually inhibited B-cell transformation. Thus, harnessing CD56brightNKG2A+ NK cell function might be promising to improve treatment strategies that target EBV-associated B cell lymphomas.

The dual function of the splenic marginal zone: essential for initiation of anti-TI-2 responses but also vital in the general first-line defense against blood-borne antigens

PubMed Central

ZANDVOORT, A; TIMENS, W

2002-01-01

The splenic marginal zone (S-MZ) is especially well equipped for rapid humoral responses and is unique in its ability to initiate an immune response to encapsulated bacteria (T-cell independent type 2 (TI-2) antigens). Because of the rapid spreading through the blood, infections with blood-borne bacteria form a major health risk. To cope with blood-borne antigens, a system is needed that can respond rapidly to a great diversity of organisms. Because of a number of unique features, S-MZ B cells can respond rapid and efficient to all sorts of blood-borne antigens. These unique features include a low blood flow microenvironment, low threshold for activation, high expression of complement receptor 2 (CR2, CD21) and multireactivity. Because of the unique high expression of CD21 in a low flow compartment, S-MZ B cells can bind and respond to TI-2 antigens even with relatively low-avid B cell receptors. Although TI-2 antigens are in general poorly opsonized by classic opsonins, a particular characteristic of these antigens is their ability to bind very rapidly to complement fragment C3d without the necessity of previous immunoglobulin binding. TI-2 primed S-MZ B cells, already by first passage through the germinal centre, will meet antigen-C3d complexes bound to follicular dendritic cells, allowing unique immediate isotype switching. This explains that the primary humoral response to TI-2 antigens is unique in its characterization by a rapid increase in IgM concurrent with IgG antibody levels. PMID:12296846

Mouse model of Epstein-Barr virus LMP1- and LMP2A-driven germinal center B-cell lymphoproliferative disease.

PubMed

Minamitani, Takeharu; Ma, Yijie; Zhou, Hufeng; Kida, Hiroshi; Tsai, Chao-Yuan; Obana, Masanori; Okuzaki, Daisuke; Fujio, Yasushi; Kumanogoh, Atsushi; Zhao, Bo; Kikutani, Hitoshi; Kieff, Elliott; Gewurz, Benjamin E; Yasui, Teruhito

2017-05-02

Epstein-Barr virus (EBV) is a major cause of immunosuppression-related B-cell lymphomas and Hodgkin lymphoma (HL). In these malignancies, EBV latent membrane protein 1 (LMP1) and LMP2A provide infected B cells with surrogate CD40 and B-cell receptor growth and survival signals. To gain insights into their synergistic in vivo roles in germinal center (GC) B cells, from which most EBV-driven lymphomas arise, we generated a mouse model with conditional GC B-cell LMP1 and LMP2A coexpression. LMP1 and LMP2A had limited effects in immunocompetent mice. However, upon T- and NK-cell depletion, LMP1/2A caused massive plasmablast outgrowth, organ damage, and death. RNA-sequencing analyses identified EBV oncoprotein effects on GC B-cell target genes, including up-regulation of multiple proinflammatory chemokines and master regulators of plasma cell differentiation. LMP1/2A coexpression also up-regulated key HL markers, including CD30 and mixed hematopoietic lineage markers. Collectively, our results highlight synergistic EBV membrane oncoprotein effects on GC B cells and provide a model for studies of their roles in immunosuppression-related lymphoproliferative diseases.

Splenic TFH expansion participates in B-cell differentiation and antiplatelet-antibody production during immune thrombocytopenia.

PubMed

Audia, Sylvain; Rossato, Marzia; Santegoets, Kim; Spijkers, Sanne; Wichers, Catharina; Bekker, Cornelis; Bloem, Andries; Boon, Louis; Flinsenberg, Thijs; Compeer, Ewoud; van den Broek, Theo; Facy, Olivier; Ortega-Deballon, Pablo; Berthier, Sabine; Leguy-Seguin, Vanessa; Martin, Laurent; Ciudad, Marion; Samson, Maxime; Trad, Malika; Lorcerie, Bernard; Janikashvili, Nona; Saas, Philippe; Bonnotte, Bernard; Radstake, Timothy R D J

2014-10-30

Antiplatelet-antibody-producing B cells play a key role in immune thrombocytopenia (ITP) pathogenesis; however, little is known about T-cell dysregulations that support B-cell differentiation. During the past decade, T follicular helper cells (TFHs) have been characterized as the main T-cell subset within secondary lymphoid organs that promotes B-cell differentiation leading to antibody class-switch recombination and secretion. Herein, we characterized TFHs within the spleen of 8 controls and 13 ITP patients. We show that human splenic TFHs are the main producers of interleukin (IL)-21, express CD40 ligand (CD154), and are located within the germinal center of secondary follicles. Compared with controls, splenic TFH frequency is higher in ITP patients and correlates with germinal center and plasma cell percentages that are also increased. In vitro, IL-21 stimulation combined with an anti-CD40 agonist antibody led to the differentiation of splenic B cells into plasma cells and to the secretion of antiplatelet antibodies in ITP patients. Overall, these results point out the involvement of TFH in ITP pathophysiology and the potential interest of IL-21 and CD40 as therapeutic targets in ITP. © 2014 by The American Society of Hematology.

Plants that attack plants: molecular elucidation of plant parasitism.

PubMed

Yoshida, Satoko; Shirasu, Ken

2012-12-01

Obligate parasitic plants in the family Orobanchaceae, such as Striga and Orobanche (including Phelipanche) spp., parasitize important crops and cause severe agricultural damage. Recent molecular studies have begun to reveal how these parasites have adapted to hosts in a parasitic lifecycle. The parasites detect nearby host roots and germinate by a mechanism that seems to have evolved from a conserved germination system found in non-parasites. The development of a specialized infecting organ called a haustorium is a unique feature of plant parasites and is triggered by host compounds and redox signals. Newly developed genomic and genetic resources will facilitate more rapid progress toward a molecular understanding of plant parasitism. Copyright © 2012 Elsevier Ltd. All rights reserved.

PD-1(HIGH) Follicular CD4 T Helper Cell Subsets Residing in Lymph Node Germinal Centers Correlate with B Cell Maturation and IgG Production in Rhesus Macaques.

PubMed

Xu, Huanbin; Wang, Xiaolei; Lackner, Andrew A; Veazey, Ronald S

2014-01-01

CD4+ T follicular helper (TFH) cells guide development and maturation of B cells and are crucial for effective antibody responses. Here we found rhesus macaque TFH cells, defined as CXCR5+CD4 T cells, contain two major populations: PD-1(INT) and PD-1(HIGH) cells. Of these, PD-1(HIGH)CD4+ T cells highly co-express ICOS but little CCR7, and reside in lymph node germinal centers (GCs), but not in blood. These cells secrete IL-21 and express transcriptional factor Bcl-6 at higher levels than CXCR5+PD-1(INT)CD4+ T cells. In addition, the frequency of PD-1(HIGH)CD4+ T cells is low in lymph nodes of newborns, but increases with age. Levels of PD-1(HIGH)CD4+ T cells correlate with mature B cells in lymph nodes, and PD-1 blockade in PD-1(HIGH)CD4+ T and B cell co-cultures significantly inhibits IgG production. In summary, PD-1(HIGH)CD4+ T cells residing in GC represent a specific TFH subset that contributes to maturation of B cells and IgG production.

[Effects of exogenous salicylic acid on seed germination and physiological characteristics of Coronilla varia under drought stress.

PubMed

Ma, Le Yuan; Chen, Nian Lai; Han, Guo Jun; Li, Liang

2017-10-01

This research investigated the effects of different concentrations (0, 0.5, 1.0, 2.0 mmol·L -1 ) of salicylic acid on the seed germination and physiological characteristics of legume forage Coronilla varia (cultivar 'Lvbaoshi') under PEG-6000 (concentration 8% and 12%) simulated drought stress. The results showed that under drought stress, 0.5-1.0 mmol·L -1 salicylic acid significantly increased germination percentage, germination vigour, germination index, vitality index and bud length of C. varia. Under the stress of 12% PEG, the dry mass of C. varia seedlings processed by 1.0 mmol·L -1 salicylic acid was significantly higher than that under drought stress. 0.5-1.0 mmol·L -1 salicylic acid processing significantly increased proline, soluble protein content, the activities of catalase, peroxidase and superoxide dismutase of C. varia seedlings under drought stress, but cell electrolyte permeability, H2O2 content and O2 - · production rate of seedlings were significantly decreased. 1.0 mmol·L -1 salicylic acid produced the best results. When the concentration of salicylic acid was beyond 2.0 mmol·L -1 , no mitigation effect was observed on the seed germination and growth of seedlings under drought stress. It was concluded that salicylic acid at appropriate concentrations could effectively improve osmotic regulation, antioxidation and mitigate the damage of drought stress so as to promote the growth of C. varia seedlings.

Comparative seed germination and seedling development of the ghost orchid, Dendrophylax lindenii (Orchidaceae), and molecular identification of its mycorrhizal fungus from South Florida

PubMed Central

Hoang, Nguyen H.; Kane, Michael E.; Radcliffe, Ellen N.; Zettler, Lawrence W.; Richardson, Larry W.

2017-01-01

Background and Aims The endangered leafless ghost orchid, Dendrophylax lindenii, one of the most renowned orchids in the world, is difficult to grow under artificial conditions. Published information on asymbiotic and symbiotic (co-culture with a mycobiont) seed germination, seedling anatomy and developmental morphology of this leafless orchid is completely lacking. This information is critical for the development of efficient procedures for ghost orchid production for successful reintroduction. Methods Ghost orchid seedling early development stages were morphologically and anatomically defined to compare germination, embryo and protocorm maturation and seedling development during asymbiotic and symbiotic culture with one of two mycorrhizal strains (Dlin-379 and Dlin-394) isolated from ghost orchid roots in situ. Key Results Seeds symbiotically germinated at higher rates when cultured with fungal strain Dlin-394 than with strain Dlin-379 or asymbiotically on P723 medium during a 10-week culture period. Fungal pelotons were observed in protocorm cells co-cultured with strain Dlin-394 but not Dlin-379. Some 2-year-old seedlings produced multinode inflorescences in vitro. Production of keikis from inflorescence nodes indicated the capacity for clonal production in the ghost orchid. Conclusions Ghost orchid embryo and seedling development were characterized into seven stages. Fungal strain Dlin-394 was confirmed as a possible ghost orchid germination mycobiont, which significantly promoted seed germination and seedling development. Internal transcribed spacer sequencing data confirmed that Dlin-394 belongs within the genus Ceratobasidium. These results offer the opportunity to examine the benefits of using a mycobiont to enhance in vitro germination and possibly ex vitro acclimatization and sustainability following outplanting. PMID:28025292

The Arabidopsis KINβγ Subunit of the SnRK1 Complex Regulates Pollen Hydration on the Stigma by Mediating the Level of Reactive Oxygen Species in Pollen

PubMed Central

Zhao, Ting Ting; Li, Fei; Jia, Xiao Na; Zhao, Xin-Ying; Zhang, Xian Sheng

2016-01-01

Pollen–stigma interactions are essential for pollen germination. The highly regulated process of pollen germination includes pollen adhesion, hydration, and germination on the stigma. However, the internal signaling of pollen that regulates pollen–stigma interactions is poorly understood. KINβγ is a plant-specific subunit of the SNF1-related protein kinase 1 complex which plays important roles in the regulation of plant development. Here, we showed that KINβγ was a cytoplasm- and nucleus-localized protein in the vegetative cells of pollen grains in Arabidopsis. The pollen of the Arabidopsis kinβγ mutant could not germinate on stigma, although it germinated normally in vitro. Further analysis revealed the hydration of kinβγ mutant pollen on the stigma was compromised. However, adding water to the stigma promoted the germination of the mutant pollen in vivo, suggesting that the compromised hydration of the mutant pollen led to its defective germination. In kinβγ mutant pollen, the structure of the mitochondria and peroxisomes was destroyed, and their numbers were significantly reduced compared with those in the wild type. Furthermore, we found that the kinβγ mutant exhibited reduced levels of reactive oxygen species (ROS) in pollen. The addition of H2O2 in vitro partially compensated for the reduced water absorption of the mutant pollen, and reducing ROS levels in pollen by overexpressing Arabidopsis CATALASE 3 resulted in compromised hydration of pollen on the stigma. These results indicate that Arabidopsis KINβγ is critical for the regulation of ROS levels by mediating the biogenesis of mitochondria and peroxisomes in pollen, which is required for pollen–stigma interactions during pollination. PMID:27472382

Comparative seed germination and seedling development of the ghost orchid, Dendrophylax lindenii (Orchidaceae), and molecular identification of its mycorrhizal fungus from South Florida.

PubMed

Hoang, Nguyen H; Kane, Michael E; Radcliffe, Ellen N; Zettler, Lawrence W; Richardson, Larry W

2017-02-01

The endangered leafless ghost orchid, Dendrophylax lindenii, one of the most renowned orchids in the world, is difficult to grow under artificial conditions. Published information on asymbiotic and symbiotic (co-culture with a mycobiont) seed germination, seedling anatomy and developmental morphology of this leafless orchid is completely lacking. This information is critical for the development of efficient procedures for ghost orchid production for successful reintroduction. Ghost orchid seedling early development stages were morphologically and anatomically defined to compare germination, embryo and protocorm maturation and seedling development during asymbiotic and symbiotic culture with one of two mycorrhizal strains (Dlin-379 and Dlin-394) isolated from ghost orchid roots in situ KEY RESULTS: Seeds symbiotically germinated at higher rates when cultured with fungal strain Dlin-394 than with strain Dlin-379 or asymbiotically on P723 medium during a 10-week culture period. Fungal pelotons were observed in protocorm cells co-cultured with strain Dlin-394 but not Dlin-379. Some 2-year-old seedlings produced multinode inflorescences in vitro Production of keikis from inflorescence nodes indicated the capacity for clonal production in the ghost orchid. Ghost orchid embryo and seedling development were characterized into seven stages. Fungal strain Dlin-394 was confirmed as a possible ghost orchid germination mycobiont, which significantly promoted seed germination and seedling development. Internal transcribed spacer sequencing data confirmed that Dlin-394 belongs within the genus Ceratobasidium These results offer the opportunity to examine the benefits of using a mycobiont to enhance in vitro germination and possibly ex vitro acclimatization and sustainability following outplanting. © The Author 2016. Published by Oxford University Press on behalf of the Annals of Botany Company.

Superoxide and its metabolism during germination and axis growth of Vigna radiata (L.) Wilczek seeds

PubMed Central

Singh, Khangembam Lenin; Chaudhuri, Abira; Kar, Rup Kumar

2014-01-01

Involvement of reactive oxygen species in regulation of plant growth and development is recently being demonstrated with various results depending on the experimental system and plant species. Role of superoxide and its metabolism in germination and axis growth was investigated in case of Vigna radiata seeds, a non-endospermous leguminous species having epigeal germination, by studying the effect of different reactive oxygen species (ROS) inhibitors, distribution of O2•ˉ and H2O2 and ROS enzyme profile in axes. Germination percentage and axis growth were determined under treatment with ROS inhibitors and scavengers. Localization of O2•ˉ and H2O2 was done using nitroblue tetrazolium (NBT) and 3,3′,5,5′-tetramethyl benzidine dihydrochloride hydrate (TMB), respectively. Apoplastic level of O2•ˉ was monitored by spectrophotometric analysis of bathing medium of axes. Profiles of NADPH oxidase and superoxide dismutase (SOD) were studied by in-gel assay. Germination was retarded by treatments affecting ROS level except H2O2 scavengers, while axis growth was retarded by all. Superoxide synthesis inhibitor and scavenger prevented H2O2 accumulation in axes in later phase as revealed from TMB staining. Activity of Cu/Zn SOD1 was initially high and declined thereafter. Superoxide being produced in apoplast possibly by NADPH oxidase activity is further metabolized to •OH via H2O2. Germination process depends possibly on •OH production in the axes. Post-germinative axis growth requires O2•ˉ while the differentiating zone of axis (radicle) requires H2O2 for cell wall stiffening. PMID:25763616

RNA and ribosomal protein patterns during aerial spore germination in Streptomyces granaticolor.

PubMed

Mikulík, K; Janda, I; Weiser, J; Stastná, J; Jiránová, A

1984-12-03

Disruption of the external sheath of Streptomyces granaticolor aerial spores and subsequent cultivation in a rich medium result in a synchronous germination. This method was used to analyze RNA and protein patterns during the germination. The germination process took place through a sequence of time-ordered events. RNA and protein synthesis started during the first 5 min and net DNA synthesis at 60-70 min of germination. Within the first 10 min of germination, synthesis of RNA was not sensitive to the inhibitory effect of rifamycin. During this period rRNA and other species including 4-5-S RNA were synthesized. Dormant spores contained populations of ribosomes or ribosomal precursors that were structurally and functionally defective. The ribosomal particles bound a sporulation pigment(s) of the melanine type. The ribosomal proteins complexed to the pigments formed insoluble aggregates which were easily removed from the ribosomes by one wash with 1 M NH4Cl. During the first 10 min of germination, pigment(s) were liberated from the complexes with the ribosomes and protein extracts of the washed ribosomes had essentially the same pattern as the extracts of ribosomes of vegetative cells. These structural alterations were accompanied by enhancement of the ribosome activities in polypeptide synthesis in vivo and in vitro. When the spores were incubated with a 14C-labelled amino acid mixture in the presence of rifamycin, only three proteins (GS1, GL1 and GS9) were identified to be radiolabelled in the extracts from the washed ribosomes. These experiments indicate that liberation of the sporulation pigment(s) from the complexes with ribosomal proteins and assembly of de novo synthesized proteins and proteins from a preexisting pool in the spore are involved in the reactivation of the ribosomes of dormant spores of S. granaticolor.

PATHWAYS OF GLUCOSE CATABOLISM IN BACILLUS CEREUS1

PubMed Central

Goldman, Manuel; Blumenthal, Harold J.

1964-01-01

Goldman, Manuel (The University of Michigan, Ann Arbor), and Harold J. Blumenthal. Pathways of glucose catabolism in Bacillus cereus. J. Bacteriol. 87:377–386. 1964.—Estimates by a radiorespirometric method of the pathways of glucose catabolism of resting-cell suspensions of Bacillus cereus strain terminalis indicate that the Embden-Meyerhof pathway predominates at every stage of development, including the sporogenic and germinative phases. At the filamentous, granular, forespore, and transitional stages, 98% of the glucose was catabolized by the Embden-Meyerhof pathway, and the remainder by the hexose monophosphate oxidative pathway. Estimates of the pathways in resting spore-suspensions arrested at defined stages of development indicate that 20% of the glucose was catabolized through the hexose monophosphate pathway in germinated spores, and 10% in the swollen and elongated stages of postgermination. In cells which had completed the first cell division, the figure fell to about 2%, a level similar to that found for vegetative cells at later stages of development. The key Embden-Meyerhof enzymes, hexokinase, phosphohexoisomerase, phosphofructokinase, and aldolase, as well as several other enzymes, were present at all stages of germination and postgerminative development, supporting the radioisotopic data obtained with whole cells. As indicated by the release of C14O2 from glucose-6-C14, terminal respiration of resting-cell suspensions operates maximally in vegetative cells at the granular, fore-spore, and transitional stages. There was marked inhibition of terminal respiration during the development of spores into vegetative cells. Only slight activity occurred in the earliest vegetative stages, and maximal operation developed after about ten cell divisions. Fumarase was absent in spores until sometime late in the elongation stage. At this point, a weak but definite activity appeared which increased during later stages of development so that, by the end of about the sixth cell division, fumarase had a specific activity about 80 times that observed at elongation. PMID:14151060

Aurelia aurita (Cnidaria) Oocytes' Contact Plate Structure and Development

PubMed Central

Adonin, Leonid S.; Shaposhnikova, Tatyana G.; Podgornaya, Olga

2012-01-01

One of the A. aurita medusa main mesoglea polypeptides, mesoglein, has been described previously. Mesoglein belongs to ZP-domain protein family and therefore we focused on A.aurita oogenesis. Antibodies against mesoglein (AB RA47) stain the plate in the place where germinal epithelium contacts oocyte on the paraffin sections. According to its position, we named the structure found the “contact plate”. Our main instrument was AB against mesoglein. ZP-domain occupies about half of the whole amino acid sequence of the mesoglein. Immunoblot after SDS-PAGE and AU-PAGE reveals two charged and high Mr bands among the female gonad germinal epithelium polypeptides. One of the gonads' polypeptides Mr corresponds to that of mesogleal cells, the other ones' Mr is higher. The morphological description of contact plate formation is the subject of the current work. Two types of AB RA47 positive granules were observed during progressive oogenesis stages. Granules form the contact plate in mature oocyte. Contact plate of A.aurita oocyte marks its animal pole and resembles Zona Pellucida by the following features: (1) it attracts spermatozoids; (2) the material of the contact plate is synthesized by oocyte and stored in granules; (3) these granules and the contact plate itself contain ZP domain protein(s); (4) contact plate is an extracellular structure made up of fiber bundles similar to those of conventional Zona Pellucida. PMID:23185235

Compartment-specific pH monitoring in Bacillus subtilis using fluorescent sensor proteins: a tool to analyze the antibacterial effect of weak organic acids.

PubMed

van Beilen, Johan W A; Brul, Stanley

2013-01-01

The internal pH (pHi) of a living cell is one of its most important physiological parameters. To monitor the pH inside Bacillus subtilis during various stages of its life cycle, we constructed an improved version (IpHluorin) of the ratiometric, pH-sensitive fluorescent protein pHluorin by extending it at the 5' end with the first 24 bp of comGA. The new version, which showed an approximate 40% increase in fluorescence intensity, was expressed from developmental phase-specific, native promoters of B. subtilis that are specifically active during vegetative growth on glucose (PptsG) or during sporulation (PspoIIA, PspoIIID, and PsspE). Our results show strong, compartment-specific expression of IpHluorin that allowed accurate pHi measurements of live cultures during exponential growth, early and late sporulation, spore germination, and during subsequent spore outgrowth. Dormant spores were characterized by an pHi of 6.0 ± 0.3. Upon full germination the pHi rose dependent on the medium to 7.0-7.4. The presence of sorbic acid in the germination medium inhibited a rise in the intracellular pH of germinating spores and inhibited germination. Such effects were absent when acetic was added at identical concentrations.

Response of soybean seed germination to cadmium and acid rain.

PubMed

Liu, Ting Ting; Wu, Peng; Wang, Li Hong; Zhou, Qing

2011-12-01

Cadmium (Cd) pollution and acid rain are the main environmental issues, and they often occur in the same agricultural region. Nevertheless, up to now, little information on the combined pollution of Cd(2+) and acid rain action on crops were presented. Here, we investigated the combined effect of Cd(2+) and acid rain on the seed germination of soybean. The results indicated that the single treatment with the low level of Cd(2+) (0.18, 1.0, 3.0 mg L(-1)) or acid rain (pH ≥3.0) could not affect the seed germination of soybean, which was resulted in the increased activities of peroxidase and catalase. The single treatment with the high concentration of Cd(2+) (>6 mg L(-1)) or acid rain at pH 2.5 decreased the activities of peroxidase and catalase, damaged the cell membrane and then decreased the seed germination of soybean. Meanwhile, the same toxic effect was observed in the combined treatment with Cd(2+) and acid rain, and the combined treatment had more toxic effect than the single treatment with Cd(2+) or acid rain. Thus, the combined pollution of Cd(2+) and acid rain had more potential threat to the seed germination of soybean than the single pollution of Cd(2+) or acid rain.

A role for antimicrobial peptides in intestinal microsporidiosis

PubMed Central

Leitch, Gordon J.; Ceballos, Carolina

2009-01-01

SUMMARY Clinical isolates from three microsporidia species, Encephalitozoon intestinalis and Encephalitozoon hellem, and the insect parasite Anncaliia (Brachiola, Nosema) algerae, were used in spore germination and enterocyte-like (C2Bbe1) cell infection assays to determine the effect of a panel of antimicrobial peptides. Spores were incubated with lactoferrin (Lf), lysozyme (Lz), and human beta defensin 2 (HBD2), human alpha defensin 5 (HD5), and human alpha defensin 1 (HNP1), alone and in combination with Lz, prior to germination. Of the Encephalitozoon species only E. hellem spore germination was inhibited by HNP1, while A. algerae spore germination was inhibited by Lf, HBD2, HD5 and HNP1, although HBD2 and HD5 inhibition required the presence of Lz. The effects of HBD2 and HD5 on microsporidia enterocyte infection paralleled their effects on spore germination. Lysozyme alone only inhibited infection with A. algerae, while Lf inhibited infection by E. intestinalis and A. algerae. HNP1 significantly reduced enterocyte infection by all three parasite species and a combination of Lf, Lz and HNP1 caused a further reduced infection with A. algerae. These data suggest that intestinal antimicrobial peptides contribute to the defense of the intestine against infection by luminal microsporidia spores and may partially determine which parasite species infects the intestine. PMID:19079820

The Crucial Role of the Pls1 Tetraspanin during Ascospore Germination in Podospora anserina Provides an Example of the Convergent Evolution of Morphogenetic Processes in Fungal Plant Pathogens and Saprobes▿ †

PubMed Central

Lambou, Karine; Malagnac, Fabienne; Barbisan, Crystel; Tharreau, Didier; Lebrun, Marc-Henri; Silar, Philippe

2008-01-01

Pls1 tetraspanins were shown for some pathogenic fungi to be essential for appressorium-mediated penetration into their host plants. We show here that Podospora anserina, a saprobic fungus lacking appressorium, contains PaPls1, a gene orthologous to known PLS1 genes. Inactivation of PaPls1 demonstrates that this gene is specifically required for the germination of ascospores in P. anserina. These ascospores are heavily melanized cells that germinate under inducing conditions through a specific pore. On the contrary, MgPLS1, which fully complements a ΔPaPls1 ascospore germination defect, has no role in the germination of Magnaporthe grisea nonmelanized ascospores but is required for the formation of the penetration peg at the pore of its melanized appressorium. P. anserina mutants with mutation of PaNox2, which encodes the NADPH oxidase of the NOX2 family, display the same ascospore-specific germination defect as the ΔPaPls1 mutant. Both mutant phenotypes are suppressed by the inhibition of melanin biosynthesis, suggesting that they are involved in the same cellular process required for the germination of P. anserina melanized ascospores. The analysis of the distribution of PLS1 and NOX2 genes in fungal genomes shows that they are either both present or both absent. These results indicate that the germination of P. anserina ascospores and the formation of the M. grisea appressorium penetration peg use the same molecular machinery that includes Pls1 and Nox2. This machinery is specifically required for the emergence of polarized hyphae from reinforced structures such as appressoria and ascospores. Its recurrent recruitment during fungal evolution may account for some of the morphogenetic convergence observed in fungi. PMID:18757568

29 CFR 1990.103 - Definitions.

Code of Federal Regulations, 2012 CFR

2012-07-01

... Health and Human Services, or designee. Director of NCI means the Director of the National Cancer... means the induction of heritable changes in the genetic material of either somatic or germinal cells..., Neurospora or Drosophila melanogaster; (3) Mutagenesis in mammalian somatic cells; (4) Mutagenesis in...

29 CFR 1990.103 - Definitions.

Code of Federal Regulations, 2013 CFR

2013-07-01

... Health and Human Services, or designee. Director of NCI means the Director of the National Cancer... means the induction of heritable changes in the genetic material of either somatic or germinal cells..., Neurospora or Drosophila melanogaster; (3) Mutagenesis in mammalian somatic cells; (4) Mutagenesis in...

29 CFR 1990.103 - Definitions.

Code of Federal Regulations, 2011 CFR

2011-07-01

... Health and Human Services, or designee. Director of NCI means the Director of the National Cancer... means the induction of heritable changes in the genetic material of either somatic or germinal cells..., Neurospora or Drosophila melanogaster; (3) Mutagenesis in mammalian somatic cells; (4) Mutagenesis in...

29 CFR 1990.103 - Definitions.

Code of Federal Regulations, 2014 CFR

2014-07-01

... Health and Human Services, or designee. Director of NCI means the Director of the National Cancer... means the induction of heritable changes in the genetic material of either somatic or germinal cells..., Neurospora or Drosophila melanogaster; (3) Mutagenesis in mammalian somatic cells; (4) Mutagenesis in...

Severe Malaria Infections Impair Germinal Center Responses by Inhibiting T Follicular Helper Cell Differentiation.

PubMed

Ryg-Cornejo, Victoria; Ioannidis, Lisa Julia; Ly, Ann; Chiu, Chris Yu; Tellier, Julie; Hill, Danika Lea; Preston, Simon Peter; Pellegrini, Marc; Yu, Di; Nutt, Stephen Laurence; Kallies, Axel; Hansen, Diana Silvia

2016-01-05

Naturally acquired immunity to malaria develops only after years of repeated exposure to Plasmodium parasites. Despite the key role antibodies play in protection, the cellular processes underlying the slow acquisition of immunity remain unknown. Using mouse models, we show that severe malaria infection inhibits the establishment of germinal centers (GCs) in the spleen. We demonstrate that infection induces high frequencies of T follicular helper (Tfh) cell precursors but results in impaired Tfh cell differentiation. Despite high expression of Bcl-6 and IL-21, precursor Tfh cells induced during infection displayed low levels of PD-1 and CXCR5 and co-expressed Th1-associated molecules such as T-bet and CXCR3. Blockade of the inflammatory cytokines TNF and IFN-γ or T-bet deletion restored Tfh cell differentiation and GC responses to infection. Thus, this study demonstrates that the same pro-inflammatory mediators that drive severe malaria pathology have detrimental effects on the induction of protective B cell responses. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Pulmonary Embolism as the Initial Presentation of Testicular Carcinoma

PubMed Central

Berber, Ilhami; Erkurt, Mehmet Ali; Ulutas, Ozkan; Ediz, Caner; Nizam, Ilknur; Kırıcı Berber, Nurcan; Unlu, Serkan; Koroglu, Reyhan; Koroglu, Mustafa; Akpolat, Nusret

2013-01-01

Objective. The risk of pulmonary embolism is well recognized as showing an increase in oncological patients. We report a case presenting with pulmonary embolism initially, which was then diagnosed with testicular cancer. Clinical Presentation and Intervention. A 25-year-old man was admitted to the emergency department with a complaint of dyspnoea. Thoracic tomography, lung ventilation/perfusion scintigraphy, and an increased D-dimer level revealed pulmonary embolism. For the aetiology of pulmonary embolism, a left orchiectomy was performed and the patient was diagnosed with a germinal cell tumour of the testicle. Conclusion. In this paper, we present a patient for whom pulmonary embolism was the initial presentation, and a germinal cell tumour was diagnosed later during the search for the aetiology. PMID:24383024

Viral Superantigen Drives Extrafollicular and Follicular B Cell Differentiation Leading to Virus-specific Antibody Production

PubMed Central

Luther, Sanjiv A.; Gulbranson-Judge, Adam; Acha-Orbea, Hans; MacLennan, Ian C.M.

1997-01-01

Mouse mammary tumor virus (MMTV[SW]) encodes a superantigen expressed by infected B cells. It evokes an antibody response specific for viral envelope protein, indicating selective activation of antigen-specific B cells. The response to MMTV(SW) in draining lymph nodes was compared with the response to haptenated chicken gamma globulin (NP-CGG) using flow cytometry and immunohistology. T cell priming occurs in both responses, with T cells proliferating in association with interdigitating dendritic cells in the T zone. T cell proliferation continues in the presence of B cells in the outer T zone, and B blasts then undergo exponential growth and differentiation into plasma cells in the medullary cords. Germinal centers develop in both responses, but those induced by MMTV(SW) appear later and are smaller. Most T cells activated in the T zone and germinal centers in the MMTV(SW) response are superantigen specific and these persist for weeks in lymph nodes draining the site MMTV(SW) injection; this contrasts with the selective loss of superantigen-specific T cells from other secondary lymphoid tissues. The results indicate that this viral superantigen, when expressed by professional antigen-presenting cells, drives extrafollicular and follicular B cell differentiation leading to virus-specific antibody production. PMID:9053455

Transcriptional Profiling of Antigen-Dependent Murine B Cell Differentiation and Memory Formation1

PubMed Central

Bhattacharya, Deepta; Cheah, Ming T.; Franco, Christopher B.; Hosen, Naoki; Pin, Christopher L.; Sha, William C.; Weissman, Irving L.

2015-01-01

Humoral immunity is characterized by the generation of Ab-secreting plasma cells and memory B cells that can more rapidly generate specific Abs upon Ag exposure than their naive counterparts. To determine the intrinsic differences that distinguish naive and memory B cells and to identify pathways that allow germinal center B cells to differentiate into memory B cells, we compared the transcriptional profiles of highly purified populations of these three cell types along with plasma cells isolated from mice immunized with a T-dependent Ag. The transcriptional profile of memory B cells is similar to that of naive B cells, yet displays several important differences, including increased expression of activation-induced deaminase and several antiapoptotic genes, chemotactic receptors, and costimulatory molecules. Retroviral expression of either Klf2 or Ski, two transcriptional regulators specifically enriched in memory B cells relative to their germinal center precursors, imparted a competitive advantage to Ag receptor and CD40-engaged B cells in vitro. These data suggest that humoral recall responses are more rapid than primary responses due to the expression of a unique transcriptional program by memory B cells that allows them to both be maintained at high frequencies and to detect and rapidly respond to antigenic re-exposure. PMID:17982071

Phytotoxicity and cytogenotoxicity of hydroalcoholic extracts from Solanum muricatum Ait. and Solanum betaceum Cav. (Solanaceae) in the plant model Lactuca sativa.

PubMed

Dos Santos, Fabio Eduardo; Carvalho, Marcos Schleiden Sousa; Silveira, Graciele Lurdes; Correa, Felipe Folgaroli; Cardoso, Maria das Graças; Andrade-Vieira, Larissa Fonseca; Vilela, Luciane Resende

2018-03-05

Plants are rich in biologically active compounds. They can be explored for the production of bioherbicides. In this context, the present work aimed to evaluate the allelopathic effect of hydroalcoholic extracts from two Solanaceae species: Solanum muricatum Ait. and Solanum betaceum Cav. For this end, we conducted phytochemical screening and biological assays, determining the effects of the extracts on germination, early development, cell cycle, and DNA fragmentation in plantlets and meristematic cells of the plant model Lactuca sativa L. (lettuce). The percentage of seeds germinated under effect of S. muricatum extract did not differ from the control, but plantlet growth was reduced at the highest concentrations. For S. betaceum extract, dose dependence was observed for both germination and plantlet development, with the highest concentrations inhibiting germination. The growth curves revealed the concentrations of 2.06 and 1.93 g/L for S. muricatum and S. betaceum extracts, respectively, as those reducing 50% of root growth (RG). At these concentrations, both extracts presented mitodepressive effect, besides inducing significant increase in the frequency of condensed nuclei, associated to DNA fragmentation and cytoplasmic shrinkage. The frequency of chromosome alterations was not significant. We further discuss the mechanisms of action related to the chemical composition of the extracts, which presented organic acids, reducing sugars, proteins, amino acids, and tannins, besides catechins and flavonoids, only found in the extract of S. betaceum.

Overexpression of a novel salt stress-induced glycine-rich protein gene from alfalfa causes salt and ABA sensitivity in Arabidopsis.

PubMed

Long, Ruicai; Yang, Qingchuan; Kang, Junmei; Zhang, Tiejun; Wang, Huimin; Li, Mingna; Zhang, Ze

2013-08-01

We cloned a novel salt stress-induced glycine-rich protein gene ( MsGRP ) from alfalfa. Its overexpression retards seed germination and seedling growth of transgenic Arabidopsis after salt and ABA treatments. Since soil salinity is one of the most significant abiotic stresses, salt tolerance is required to overcome salinity-induced reductions in crop productivity. Many glycine-rich proteins (GRPs) have been implicated in plant responses to environmental stresses, but the function and importance of some GRPs in stress responses remain largely unknown. Here, we report on a novel salt stress-induced GRP gene (MsGRP) that we isolated from alfalfa. Compared with some glycine-rich RNA-binding proteins, MsGRP contains no RNA recognition motifs and localizes in the cell membrane or cell wall according to the subcellular localization result. MsGRP mRNA is induced by salt, abscisic acid (ABA), and drought stresses in alfalfa seedlings, and its overexpression driven by a constitutive cauliflower mosaic virus-35S promoter in Arabidopsis plants confers salinity and ABA sensitivity compared with WT plants. MsGRP retards seed germination and seedling growth of transgenic Arabidopsis plants after salt and ABA treatments, which implies that MsGRP may affect germination and growth through an ABA-dependent regulation pathway. These results provide indirect evidence that MsGRP plays important roles in seed germination and seedling growth of alfalfa under some abiotic stress conditions.

Intranasal Live Influenza Vaccine Priming Elicits Localized B Cell Responses in Mediastinal Lymph Nodes.

PubMed

Jegaskanda, Sinthujan; Mason, Rosemarie D; Andrews, Sarah F; Wheatley, Adam K; Zhang, Ruijun; Reynoso, Glennys V; Ambrozak, David R; Santos, Celia P; Luke, Catherine J; Matsuoka, Yumiko; Brenchley, Jason M; Hickman, Heather D; Talaat, Kawsar R; Permar, Sallie R; Liao, Hua-Xin; Yewdell, Jonathan W; Koup, Richard A; Roederer, Mario; McDermott, Adrian B; Subbarao, Kanta

2018-05-01

Pandemic live attenuated influenza vaccines (pLAIV) prime subjects for a robust neutralizing antibody response upon subsequent administration of a pandemic inactivated subunit vaccine (pISV). However, a difference was not detected in H5-specific memory B cells in the peripheral blood between pLAIV-primed and unprimed subjects prior to pISV boost. To investigate the mechanism underlying pLAIV priming, we vaccinated groups of 12 African green monkeys (AGMs) with H5N1 pISV or pLAIV alone or H5N1 pLAIV followed by pISV and examined immunity systemically and in local draining lymph nodes (LN). The AGM model recapitulated the serologic observations from clinical studies. Interestingly, H5N1 pLAIV induced robust germinal center B cell responses in the mediastinal LN (MLN). Subsequent boosting with H5N1 pISV drove increases in H5-specific B cells in the axillary LN, spleen, and circulation in H5N1 pLAIV-primed animals. Thus, H5N1 pLAIV primes localized B cell responses in the MLN that are recalled systemically following pISV boost. These data provide mechanistic insights for the generation of robust humoral responses via prime-boost vaccination. IMPORTANCE We have previously shown that pandemic live attenuated influenza vaccines (pLAIV) prime for a rapid and robust antibody response on subsequent administration of inactivated subunit vaccine (pISV). This is observed even in individuals who had undetectable antibody (Ab) responses following the initial vaccination. To define the mechanistic basis of pLAIV priming, we turned to a nonhuman primate model and performed a detailed analysis of B cell responses in systemic and local lymphoid tissues following prime-boost vaccination with pLAIV and pISV. We show that the nonhuman primate model recapitulates the serologic observations from clinical studies. Further, we found that pLAIVs induced robust germinal center B cell responses in the mediastinal lymph node. Subsequent boosting with pISV in pLAIV-primed animals resulted in detection of B cells in the axillary lymph nodes, spleen, and peripheral blood. We demonstrate that intranasally administered pLAIV elicits a highly localized germinal center B cell response in the mediastinal lymph node that is rapidly recalled following pISV boost into germinal center reactions at numerous distant immune sites. Copyright © 2018 American Society for Microbiology.

Antifungal Attributes of Lactobacillus plantarum MYS6 against Fumonisin Producing Fusarium proliferatum Associated with Poultry Feeds

PubMed Central

Deepthi, B. V.; Poornachandra Rao, K.; Chennapa, G.; Naik, M. K.; Chandrashekara, K. T.; Sreenivasa, M. Y.

2016-01-01

Fumonisins, being common in occurrence in maize-based feeds, pose a great threat to animal and human health. The present study is aimed at determining the antifungal activity of Lactobacillus plantarum MYS6 against a fumonisin producing fungus, Fusarium proliferatum MYS9. The isolate was subjected to standard tests for determining its probiotic attributes and antifungal properties. L. plantarum MYS6 thrived well at pH 3.0 and 6.0, and exhibited strong resistance up to 3% bile. The isolate showed a high degree of cell surface hydrophobicity corresponding to its strong adhesion to chicken crop epithelial cells. Co-inoculation with the fungus on modified de Man Rogosa Sharpe medium revealed the inhibitory effect of L. plantarum MYS6 on fungal growth and biomass. Observation using scanning electron microscopy showed distortion of hyphal structures, swollen tips and disrupted conidia. Conidia germination inhibition assay restrained germination and showed deformed hyphae. The bioprotective feature of the isolate was evident by the inhibition of fungal development in maize-kernel treated with the cell free supernatant of L. plantarum MYS6. Both the isolate and its extracellular metabolites lowered fumonisin content in feed model up to 0.505 mg/Kg of feed and 0.3125 mg/Kg of feed respectively when compared to the level of 0.870 mg/Kg of feed in control. The major antifungal compounds produced by the isolate were 10-Octadecenoic acid, methyl ester; palmitic acid, methyl ester; heptadecanoic acid, 16-methyl ester; stearic acid and lauric acid. L. plantarum MYS6 reduced 61.7% of fumonisin possibly by a binding mechanism. These findings suggest the application of L. plantarum MYS6 as an efficient probiotic additive and biocontrol agent in feed used in poultry industry. Additionally, the antifungal metabolites pose a conspicuous inhibition of Fusarium growth and fumonisin production. PMID:27285317

Antifungal Attributes of Lactobacillus plantarum MYS6 against Fumonisin Producing Fusarium proliferatum Associated with Poultry Feeds.

PubMed

Deepthi, B V; Poornachandra Rao, K; Chennapa, G; Naik, M K; Chandrashekara, K T; Sreenivasa, M Y

2016-01-01

Fumonisins, being common in occurrence in maize-based feeds, pose a great threat to animal and human health. The present study is aimed at determining the antifungal activity of Lactobacillus plantarum MYS6 against a fumonisin producing fungus, Fusarium proliferatum MYS9. The isolate was subjected to standard tests for determining its probiotic attributes and antifungal properties. L. plantarum MYS6 thrived well at pH 3.0 and 6.0, and exhibited strong resistance up to 3% bile. The isolate showed a high degree of cell surface hydrophobicity corresponding to its strong adhesion to chicken crop epithelial cells. Co-inoculation with the fungus on modified de Man Rogosa Sharpe medium revealed the inhibitory effect of L. plantarum MYS6 on fungal growth and biomass. Observation using scanning electron microscopy showed distortion of hyphal structures, swollen tips and disrupted conidia. Conidia germination inhibition assay restrained germination and showed deformed hyphae. The bioprotective feature of the isolate was evident by the inhibition of fungal development in maize-kernel treated with the cell free supernatant of L. plantarum MYS6. Both the isolate and its extracellular metabolites lowered fumonisin content in feed model up to 0.505 mg/Kg of feed and 0.3125 mg/Kg of feed respectively when compared to the level of 0.870 mg/Kg of feed in control. The major antifungal compounds produced by the isolate were 10-Octadecenoic acid, methyl ester; palmitic acid, methyl ester; heptadecanoic acid, 16-methyl ester; stearic acid and lauric acid. L. plantarum MYS6 reduced 61.7% of fumonisin possibly by a binding mechanism. These findings suggest the application of L. plantarum MYS6 as an efficient probiotic additive and biocontrol agent in feed used in poultry industry. Additionally, the antifungal metabolites pose a conspicuous inhibition of Fusarium growth and fumonisin production.

Involvement of Pinus taeda MYB1 and MYB8 in phenylpropanoid metabolism and secondary cell wall biogenesis: a comparative in planta analysis

PubMed Central

Bomal, Claude; Bedon, Frank; Caron, Sébastien; Mansfield, Shawn D.; Levasseur, Caroline; Cooke, Janice E. K.; Blais, Sylvie; Tremblay, Laurence; Morency, Marie-Josée; Pavy, Nathalie; Grima-Pettenati, Jacqueline; Séguin, Armand; MacKay, John

2008-01-01

The involvement of two R2R3-MYB genes from Pinus taeda L., PtMYB1 and PtMYB8, in phenylpropanoid metabolism and secondary cell wall biogenesis was investigated in planta. These pine MYBs were constitutively overexpressed (OE) in Picea glauca (Moench) Voss, used as a heterologous conifer expression system. Morphological, histological, chemical (lignin and soluble phenols), and transcriptional analyses, i.e. microarray and reverse transcription quantitative PCR (RT-qPCR) were used for extensive phenotyping of MYB-overexpressing spruce plantlets. Upon germination of somatic embryos, root growth was reduced in both transgenics. Enhanced lignin deposition was also a common feature but ectopic secondary cell wall deposition was more strongly associated with PtMYB8-OE. Microarray and RT-qPCR data showed that overexpression of each MYB led to an overlapping up-regulation of many genes encoding phenylpropanoid enzymes involved in lignin monomer synthesis, while misregulation of several cell wall-related genes and other MYB transcription factors was specifically associated with PtMYB8-OE. Together, the results suggest that MYB1 and MYB8 may be part of a conserved transcriptional network involved in secondary cell wall deposition in conifers. PMID:18805909

Germinal center reentries of BCL2-overexpressing B cells drive follicular lymphoma progression

PubMed Central

Sungalee, Stéphanie; Mamessier, Emilie; Morgado, Ester; Grégoire, Emilie; Brohawn, Philip Z.; Morehouse, Christopher A.; Jouve, Nathalie; Monvoisin, Céline; Menard, Cédric; Debroas, Guilhaume; Faroudi, Mustapha; Mechin, Violaine; Navarro, Jean-Marc; Drevet, Charlotte; Eberle, Franziska C.; Chasson, Lionel; Baudimont, Fannie; Mancini, Stéphane J.; Tellier, Julie; Picquenot, Jean-Michel; Kelly, Rachel; Vineis, Paolo; Ruminy, Philippe; Chetaille, Bruno; Jaffe, Elaine S.; Schiff, Claudine; Hardwigsen, Jean; Tice, David A.; Higgs, Brandon W.; Tarte, Karin; Nadel, Bertrand; Roulland, Sandrine

2014-01-01

It has recently been demonstrated that memory B cells can reenter and reengage germinal center (GC) reactions, opening the possibility that multi-hit lymphomagenesis gradually occurs throughout life during successive immunological challenges. Here, we investigated this scenario in follicular lymphoma (FL), an indolent GC-derived malignancy. We developed a mouse model that recapitulates the FL hallmark t(14;18) translocation, which results in constitutive activation of antiapoptotic protein B cell lymphoma 2 (BCL2) in a subset of B cells, and applied a combination of molecular and immunofluorescence approaches to track normal and t(14;18)+ memory B cells in human and BCL2-overexpressing B cells in murine lymphoid tissues. BCL2-overexpressing B cells required multiple GC transits before acquiring FL-associated developmental arrest and presenting as GC B cells with constitutive activation–induced cytidine deaminase (AID) mutator activity. Moreover, multiple reentries into the GC were necessary for the progression to advanced precursor stages of FL. Together, our results demonstrate that protracted subversion of immune dynamics contributes to early dissemination and progression of t(14;18)+ precursors and shapes the systemic presentation of FL patients. PMID:25384217

GANP regulates recruitment of AID to immunoglobulin variable regions by modulating transcription and nucleosome occupancy

PubMed Central

Singh, Shailendra Kumar; Maeda, Kazuhiko; Eid, Mohammed Mansour Abbas; Almofty, Sarah Ameen; Ono, Masaya; Pham, Phuong; Goodman, Myron F.; Sakaguchi, Nobuo

2013-01-01

Somatic hypermutation in B cells is initiated by activation-induced cytidine deaminase-catalyzed C→U deamination at immunoglobulin variable regions. Here we investigate the role of the germinal centre-associated nuclear protein (GANP) in enhancing the access of activation-induced cytidine deaminase (AID) to immunoglobulin variable regions. We show that the nuclear export factor GANP is involved in chromatin modification at rearranged immunoglobulin variable loci, and its activity requires a histone acetyltransferase domain. GANP interacts with the transcription stalling protein Spt5 and facilitates RNA Pol-II recruitment to immunoglobulin variable regions. Germinal centre B cells from ganp-transgenic mice showed a higher AID occupancy at the immunoglobulin variable region, whereas B cells from conditional ganp-knockout mice exhibit a lower AID accessibility. These findings suggest that GANP-mediated chromatin modification promotes transcription complex recruitment and positioning at immunoglobulin variable loci to favour AID targeting. PMID:23652018

Spatio-Temporal Accumulation and Activity of Calcium-Dependent Protein Kinases during Embryogenesis, Seed Development, and Germination in Sandalwood1

PubMed Central

Anil, Veena S.; Harmon, Alice C.; Rao, K. Sankara

2000-01-01

Western-blot analysis and protein kinase assays identified two Ca2+-dependent protein kinases (CDPKs) of 55 to 60 kD in soluble protein extracts of embryogenic cultures of sandalwood (Santalum album L.). However, these sandalwood CDPKs (swCDPKs) were absent in plantlets regenerated from somatic embryos. swCDPKs exhibited differential expression (monitored at the level of the protein) and activity in different developmental stages. Zygotic embryos, seedlings, and endosperm showed high accumulation of swCDPK, but the enzyme was not detected in the soluble proteins of shoots and flowers. swCDPK exhibited a temporal pattern of expression in endosperm, showing high accumulation and activity in mature fruit and germinating stages; the enzyme was localized strongly in the storage bodies of the endosperm cells. The study also reports for the first time to our knowledge a post-translational inhibition/inactivation of swCDPK in zygotic embryos during seed dormancy and early stages of germination. The temporal expression of swCDPK during somatic/zygotic embryogenesis, seed maturation, and germination suggests involvement of the enzyme in these developmental processes. PMID:10759499

Spatio-temporal accumulation and activity of calcium-dependent protein kinases during embryogenesis, seed development, and germination in sandalwood.

PubMed

Anil, V S; Harmon, A C; Rao, K S

2000-04-01

Western-blot analysis and protein kinase assays identified two Ca(2+)-dependent protein kinases (CDPKs) of 55 to 60 kD in soluble protein extracts of embryogenic cultures of sandalwood (Santalum album L.). However, these sandalwood CDPKs (swCDPKs) were absent in plantlets regenerated from somatic embryos. swCDPKs exhibited differential expression (monitored at the level of the protein) and activity in different developmental stages. Zygotic embryos, seedlings, and endosperm showed high accumulation of swCDPK, but the enzyme was not detected in the soluble proteins of shoots and flowers. swCDPK exhibited a temporal pattern of expression in endosperm, showing high accumulation and activity in mature fruit and germinating stages; the enzyme was localized strongly in the storage bodies of the endosperm cells. The study also reports for the first time to our knowledge a post-translational inhibition/inactivation of swCDPK in zygotic embryos during seed dormancy and early stages of germination. The temporal expression of swCDPK during somatic/zygotic embryogenesis, seed maturation, and germination suggests involvement of the enzyme in these developmental processes.

Fat Metabolism in Higher Plants XXVI. Biosynthesis of Fatty Acids in Tissues of Developing Seeds and Germinating Seedlings of Safflower (Carthamus tinctorius L.) 1

PubMed Central

McMahon, Vern; Stumpf, P. K.

1966-01-01

The capacity of both developing seeds and germinating seedlings of safflower for the incorporation of acetate-C14 into long-chain fatty acids is examined. Intact tissue of the developing seed shows a low rate of acetate incorporation into fatty acid initially but between the tenth and twenty-fifth day after flowering the tissue has a high rate of synthesis, in particular with respect to the unsaturated fatty acids. Centrifuged fractionation of homogenates of this developmental tissue yielded several active fractions, the most active being the PI fraction consisting mostly of plastids. Cofactor requirements and pH effects are examined. Germinating tissue shows a more uniform capacity for synthesis of fatty acids since there is no marked change in synthetic capacity. The newly synthesized fatty acids are consistently palmitic, stearic, and oleic acid. No linoleic synthesis could be detected. The most active fraction of cell-free preparation of germinating tissue is the plastid fraction (PI), similar to what was formed with developing tissue. PMID:5904587

Determination of carbohydrate profile in sugarbeet (Beta vulgaris) cell walls

USDA-ARS?s Scientific Manuscript database

Sugarbeet germplasms USH20, C869, EL55, EL54 were used, and different tissues at different developmental stages were sampled, including dry seeds, germinating seedlings, developing leaves, mature leaves, petioles, hypocotyls, mature roots, flowering stems and inflorescences. Cell Wall Composition An...

The effect of cyanide compounds, fluorides, aluminum, and inorganic oxides present in spent pot liner on germination and root tip cells of Lactuca sativa.

PubMed

Andrade, L F; Davide, L C; Gedraite, L S

2010-05-01

SPL (spent pot liner) is a solid waste produced by the aluminum industry. This waste has a highly variable composition, consisting of cyanides, fluorides, organics, and metals. The aim of this work was to study the effect of SPL on root tips of Lactuca sativa using current plant bioassays. We observed a decrease in the germination rate with increasing concentrations of SPL. In addition, SPL was found to reduce root growth, which is correlated with a decrease in the mitotic index. Nevertheless, we noticed a significant enhancement in the percentage of stickiness, c-metaphase, anaphase bridges, and laggard chromosomes in dividing cells and also an increase in the number of cells with condensed nuclei. Moreover, SPL was found to alter the root tip surface, resulting in a reduction in the amount of root hair. These results demonstrate that SPL is a toxic agent that leads to cell damage and disturbance. Copyright 2009 Elsevier Inc. All rights reserved.

Sensitivity of Aspergillus nidulans to the Cellulose Synthase Inhibitor Dichlobenil: Insights from Wall-Related Genes’ Expression and Ultrastructural Hyphal Morphologies

PubMed Central

Obersriebnig, Michael; Salerno, Marco; Pum, Dietmar; Strauss, Joseph

2013-01-01

The fungal cell wall constitutes an important target for the development of antifungal drugs, because of its central role in morphogenesis, development and determination of fungal-specific molecular features. Fungal walls are characterized by a network of interconnected glycoproteins and polysaccharides, namely α-, β-glucans and chitin. Cell walls promptly and dynamically respond to environmental stimuli by a signaling mechanism, which triggers, among other responses, modulations in wall biosynthetic genes’ expression. Despite the absence of cellulose in the wall of the model filamentous fungus Aspergillus nidulans, we found in this study that fungal growth, spore germination and morphology are affected by the addition of the cellulose synthase inhibitor dichlobenil. Expression analysis of selected genes putatively involved in cell wall biosynthesis, carried out at different time points of drug exposure (i.e. 0, 1, 3, 6 and 24 h), revealed increased expression for the putative mixed linkage β-1,3;1,4 glucan synthase celA together with the β-1,3-glucan synthase fksA and the Rho-related GTPase rhoA. We also compared these data with the response to Congo Red, a known plant/fungal drug affecting both chitin and cellulose biosynthesis. The two drugs exerted different effects at the cell wall level, as shown by gene expression analysis and the ultrastructural features observed through atomic force microscopy and scanning electron microscopy. Although the concentration of dichlobenil required to affect growth of A. nidulans is approximately 10-fold higher than that required to inhibit plant cellulose biosynthesis, our work for the first time demonstrates that a cellulose biosynthesis inhibitor affects fungal growth, changes fungal morphology and expression of genes connected to fungal cell wall biosynthesis. PMID:24312197

IRF8 Governs Expression of Genes Involved in Innate and Adaptive Immunity in Human and Mouse Germinal Center B Cells

PubMed Central

Morse, Herbert C.

2011-01-01

IRF8 (Interferon Regulatory Factor 8) is a transcription factor expressed throughout B cell differentiation except for mature plasma cells. Previous studies showed it is part of the transcriptional network governing B cell specification and commitment in the bone marrow, regulates the distribution of mature B cells into the splenic follicular and marginal zone compartments, and is expressed at highest levels in germinal center (GC) B cells. Here, we investigated the transcriptional programs and signaling pathways affected by IRF8 in human and mouse GC B cells as defined by ChIP-chip analyses and transcriptional profiling. We show that IRF8 binds a large number of genes by targeting two distinct motifs, half of which are also targeted by PU.1. Over 70% of the binding sites localized to proximal and distal promoter regions with ∼25% being intragenic. There was significant enrichment among targeted genes for those involved in innate and adaptive immunity with over 30% previously defined as interferon stimulated genes. We also showed that IRF8 target genes contributes to multiple aspects of the biology of mature B cells including critical components of the molecular crosstalk among GC B cells, T follicular helper cells, and follicular dendritic cells. PMID:22096565

Co-stimulatory function in primary germinal center responses: CD40 and B7 are required on distinct antigen-presenting cells.

PubMed

Watanabe, Masashi; Fujihara, Chiharu; Radtke, Andrea J; Chiang, Y Jeffrey; Bhatia, Sumeena; Germain, Ronald N; Hodes, Richard J

2017-09-04

T cell-dependent germinal center (GC) responses require coordinated interactions of T cells with two antigen-presenting cell (APC) populations, B cells and dendritic cells (DCs), in the presence of B7- and CD40-dependent co-stimulatory pathways. Contrary to the prevailing paradigm, we found unique cellular requirements for B7 and CD40 expression in primary GC responses to vaccine immunization with protein antigen and adjuvant: B7 was required on DCs but was not required on B cells, whereas CD40 was required on B cells but not on DCs in the generation of antigen-specific follicular helper T cells, antigen-specific GC B cells, and high-affinity class-switched antibody production. There was, in fact, no requirement for coexpression of B7 and CD40 on the same cell in these responses. Our findings support a substantially revised model for co-stimulatory function in the primary GC response, with crucial and distinct contributions of B7- and CD40-dependent pathways expressed by different APC populations and with important implications for understanding how to optimize vaccine responses or limit autoimmunity. This is a work of the U.S. Government and is not subject to copyright protection in the United States. Foreign copyrights may apply.

Persistent Simian Immunodeficiency Virus Infection Causes Ultimate Depletion of Follicular Th Cells in AIDS.

PubMed

Xu, Huanbin; Wang, Xiaolei; Malam, Naomi; Lackner, Andrew A; Veazey, Ronald S

2015-11-01

CD4(+) T follicular helper (Tfh) cells are critical for the generation of humoral immune responses to pathogenic infections, providing help for B cell development, survival, and affinity maturation of Abs. Although CD4(+) Tfh cells are reported to accumulate in HIV or SIV infection, we found that germinal center Tfh cells, defined in this study as CXCR5(+)PD-1(HIGH)CD4(+) T cells, did not consistently accumulate in chronically SIV-infected rhesus macaques compared with those infected with less pathogenic simian HIV, vaccinated and SIVmac-challenged, or SIVmac-infected Mamu-A*01(+) macaques, all of which are associated with some control of virus replication and slower disease progression. Interestingly, CXCR5(+)PD-1(HIGH) Tfh cells in lymphoid tissues were eventually depleted in macaques with AIDS compared with the other cohorts. Chronic activation and proliferation of CXCR5(+)PD-1(HIGH) Tfh were increased, but PD-L2 expression was downregulated on B cells, possibly resulting in germinal center Tfh cell apoptosis. Together, these findings suggest that changes in CXCR5(+)PD-1(HIGH) Tfh cells in lymph nodes correlate with immune control during infection, and their loss or dysregulation contribute to impairment of B cell responses and progression to AIDS. Copyright © 2015 by The American Association of Immunologists, Inc.

Molecular Characteristics of Mantle Cell Lymphoma Presenting with Clonal Plasma Cell Component

PubMed Central

Visco, Carlo; Hoeller, Sylvia; Malik, Jeffrey T.; Xu-Monette, Zijun Y.; Wiggins, Michele L.; Liu, Jessica; Sanger, Warren G.; Liu, Zhongfeng; Chang, Julie; Ranheim, Erik A.; Gradowski, Joel F.; Serrrano, Sergio; Wang, Huan-You; Liu, Qingquan; Dave, Sandeep; Olsen, Brian; Gascoyne, Randy D.; Campo, Elias; Swerdlow, Steven H.; Chan, Wing C.; Tzankov, Alexander; Young, Ken H.

2011-01-01

The normal counterparts of mantle cell lymphoma (MCL) are naïve quiescent B-cells that have not been processed through the germinal center (GC). For this reason, while lymphomas arising from GC or post-GC B-cells often exhibit plasmacytic differentiation, MCL rarely presents with plasmacytic features. Seven cases of MCL with a monotypic plasma cell (PC) population were collected from six centers and studied by immunohistochemistry, FICTION (Fluorescence immunophenotyping and Interphase Cytogenetics as a Tool for the Investigation of Neoplasms), capillary gel electrophoresis, and restriction fragment length polymorphism of immunoglobulin heavy chain analysis (RFLP/IgH) of microdissections of each of the MCL and PC populations to assess their clonal relationship. Clinical presentation was rather unusual compared to typical MCL, with two cases arising from extranodal soft-tissues of the head. All MCL cases were morphologically and immunohistochemically typical, bearing the t(11;14)(q13;q32). In all cases PC populations were clonal. In 5 of the 7 cases, the MCL and PC clones showed identical restriction fragments, indicating a common clonal origin of the neoplastic populations. The two cases with clonal diversity denoted the coexistence of two different tumors in a composite lymphoma/plasma cell neoplasm. Our findings suggest that MCL can present with a PC component that is often clonally related to the lymphoma, representing a rare but unique biological variant of this tumor. PMID:21263238

Metagenomic Analyses of the Viruses Detected in Mycorrhizal Fungi and Their Host Orchid.

PubMed

Shimura, Hanako; Masuta, Chikara; Koda, Yasunori

2018-01-01

In nature, mycorrhizal association with soilborne fungi is indispensable for orchid families. Fungal structures from compatible endo-mycorrhizal fungi in orchid cells are digested in cells to be supplied to orchids as nutrition. Because orchid seeds lack the reserves for germination, they keep receiving nutrition through mycorrhizal formation from seed germination until shoots develop (leaves) and become photoautotrophic. Seeds of all orchid species surely geminate with the help of their own fungal partners, and this specific partnership has been acquired for a long evolutional history between orchids and fungi.We have studied the interactions between orchids and mycorrhizal fungi and recently conducted transcriptome analyses (RNAseq) by a next-generation sequencing (NGS) approach. It is possible that orchid RNA isolated form naturally grown plants is contaminated with RNAs derived from mycorrhizal fungi in the orchid cells. To avoid such contamination, we here prepared aseptically germinated orchid plants (i.e., fungus-free plants) together with a pure-cultured fungal isolate and field-growing orchid samples. In the cDNA library prepared from orchid and fungal tissues, we found that partitivirus-like sequences were common in an orchid and its mycorrhizal fungus. These partitivirus-like sequences were closely related by a phylogenetic analysis, suggesting that transmission of an ancestor virus between the two organisms occurred through the specific relation of the orchid and its associated fungus.

[On the similarity in the effects of different cryopreservation conditions on the growth and development of plants].

PubMed

Chetverikova, E P; Shabaeva, E V; Iashina, S G

2008-01-01

The morphological characteristics of 35 wild plant species were studied after freezing of seeds under the conditions of deep, fast, and programmed freezing (-196 degrees C) and non-deep freezing (-10 degrees C). The seeds were stored frozen for a month. The seeds of all the species were characterized by a low humidity. The field and laboratory seed germination capacity, leaf growth, the quantity and length of shoots, the quantity of generative organs, and the variability of these characteristics were studied. It was shown that the direction of changes under different cooling conditions was the same except for the laboratory germination capacity of some species. The direction was determined by the species features rather than cooling conditions.

Ecology and thermal inactivation of microbes in and on interplanetary space vehicle components. [heat sensitivity of bacterial spores

NASA Technical Reports Server (NTRS)

Campbell, J. E.; Reyes, A. L.; Wehby, A. J.; Crawford, R. G.; Wimsatt, J. C.; Peeler, J. T.

1973-01-01

The mechanism for thermal inactivation of bacterial spores under moist or dry heat was studied. Experimental conditions were established relating to spore loss of heat resistance and loss of optical density as a measure of the rate and extent of germination in spore suspensions. Events occurring during germination were correlated with phase darkening (refractility and non-refractility of spores), stainability characteristics of heat and non-heat treated spores, morphological characteristics, and studies on swelling of spores by an increase in packed cell volume.

Effect of prior refrigeration on botulinal outgrowth in perishable canned cured meat when temperature abused.

PubMed Central

Tompkin, R B; Christiansen, L N; Shaparis, A B

1978-01-01

Perishable canned cured meat inoculated with Clostridium botulinum spores was placed at 4.4 or 10 degrees C after manufacture. Spore germination occurred at 10 degrees C. The germinated cell count remained stable over a period of 16 to 18 weeks. During that time period the inhibitory system and residual nitrite descreased. These factors combine to make perishable canned cured meats more prone to spoilage and potential hazard if they are temperature abused after a period of refrigerated storage. PMID:350155

Very-long-chain aldehydes induce appressorium formation in ascospores of the wheat powdery mildew fungus Blumeria graminis.

PubMed

Zhu, Mo; Riederer, Markus; Hildebrandt, Ulrich

2017-08-01

Asexually produced conidia of the wheat powdery mildew fungus Blumeria graminis f. sp. tritici (Bgt) are known to perceive cuticular very-long-chain aldehydes as signal substances strongly stimulating germination and differentiation of infection structures in a concentration- and chain-length-dependent manner. Conidial germination and appressorium formation are widely prevented by the presence of free water on the host surface. However, sexually produced ascospores can differentiate immersed in water. Applying a Formvar ® -based in vitro-system showed that ascospore appressorium formation was strongly induced by the presence of wheat leaf cuticular wax. Similar to conidia, ascospore appressorium formation is triggered by the presence of very-long-chain aldehydes in a chain-length-dependent manner with n-octacosanal as the most inducing aldehyde. Surface hydrophobicity positively affected ascospore germination but not appressorium formation. Ascospores required significantly more time to complete the differentiation of appressoria and exhibited a more distinct dependence on the availability of free water than their conidial counterparts. Unlike conidia, ascospores showed a more variable germination and differentiation pattern even with a single germ tube differentiating an appressorium. Despite these differences our results demonstrate that a host surface recognition principle based on cuticular very-long-chain aldehydes is a common feature of B. graminis f. sp. tritici ascospores and conidia. Copyright © 2017 British Mycological Society. Published by Elsevier Ltd. All rights reserved.

Epstein-Barr Virus Latent Membrane Protein 2A (LMP2A)-Mediated changes in Fas Expression and Fas-Dependent Apoptosis: Role of Lyn/Syk activation

PubMed Central

Incrocci, Ryan; Hussain, Samira; Stone, Amanda; Bieging, Kathryn; Alt, Lauren A.C.; Fay, Michael J.; Swanson-Mungerson, Michelle

2015-01-01

Epstein-Barr virus Latent Membrane Protein 2A (LMP2A) is expressed in EBV-infected B cells in the germinal center, a site of significant apoptosis induced by engagement of Fas on activated B cells. Signals from the B cell receptor (BCR) protect germinal center B cells from Fas-mediated apoptosis, and since LMP2A is a BCR mimic, we hypothesized that LMP2A would also protect B cells from Fas-mediated apoptosis. Surprisingly, latently-infected human and murine B cell lines expressing LMP2A were more sensitive to Fas-mediated apoptosis, as determined by increases in Annexin-V staining, and cleavage of caspase-8, −3 and PARP. Additional studies show that LMP2A-expressing B cell lines demonstrate a Lyn- and Syk-dependent increase in sensitivity to Fas-mediated apoptosis, due to an LMP2A-dependent enhancement in Fas expression. These findings demonstrate the ability for LMP2A to directly increase a pro-apoptotic molecule and have implications for EBV latency as well as the treatment of EBV-associated malignancies. PMID:26255694

BIIDXI, the At4g32460 DUF642 gene, is involved in pectin methyl esterase regulation during Arabidopsis thaliana seed germination and plant development.

PubMed

Zúñiga-Sánchez, Esther; Soriano, Diana; Martínez-Barajas, Eleazar; Orozco-Segovia, Alma; Gamboa-deBuen, Alicia

2014-12-02

DUF642 proteins constitute a highly conserved family of proteins that are associated with the cell wall and are specific to spermatophytes. Transcriptome studies have suggested that members of this family are involved in seed development and germination processes. Previous in vitro studies have revealed that At4g32460- and At5g11420-encoded proteins interact with the catalytic domain of pectin methyl esterase 3 (AtPME3, which is encoded by At3g14310). PMEs play an important role in plant development, including seed germination. The aim of this study was to evaluate the function of the DUF642 gene At4g32460 during seed germination and plant development and to determine its relation to PME activity regulation. Our results indicated that the DUF642 proteins encoded by At4g32460 and At5g11420 could be positive regulators of PME activity during several developmental processes. Transgenic lines overexpressing these proteins showed increased PME activity during seed germination, and improved seed germination performance. In plants expressing At4g32460 antisense RNA, PME activity was decreased in the leaves, and the siliques were very short and contained no seeds. This phenotype was also present in the SALK_142260 and SALK_054867 lines for At4g32460. Our results suggested that the DUF642 family contributes to the complexity of the methylesterification process by participating in the fine regulation of pectin status during plant development.

Identification of a new gene regulatory circuit involving B cell receptor activated signaling using a combined analysis of experimental, clinical and global gene expression data

PubMed Central

Schrader, Alexandra; Meyer, Katharina; Walther, Neele; Stolz, Ailine; Feist, Maren; Hand, Elisabeth; von Bonin, Frederike; Evers, Maurits; Kohler, Christian; Shirneshan, Katayoon; Vockerodt, Martina; Klapper, Wolfram; Szczepanowski, Monika; Murray, Paul G.; Bastians, Holger; Trümper, Lorenz; Spang, Rainer; Kube, Dieter

2016-01-01

To discover new regulatory pathways in B lymphoma cells, we performed a combined analysis of experimental, clinical and global gene expression data. We identified a specific cluster of genes that was coherently expressed in primary lymphoma samples and suppressed by activation of the B cell receptor (BCR) through αIgM treatment of lymphoma cells in vitro. This gene cluster, which we called BCR.1, includes numerous cell cycle regulators. A reduced expression of BCR.1 genes after BCR activation was observed in different cell lines and also in CD10+ germinal center B cells. We found that BCR activation led to a delayed entry to and progression of mitosis and defects in metaphase. Cytogenetic changes were detected upon long-term αIgM treatment. Furthermore, an inverse correlation of BCR.1 genes with c-Myc co-regulated genes in distinct groups of lymphoma patients was observed. Finally, we showed that the BCR.1 index discriminates activated B cell-like and germinal centre B cell-like diffuse large B cell lymphoma supporting the functional relevance of this new regulatory circuit and the power of guided clustering for biomarker discovery. PMID:27166259

Activation of B Cells by a Dendritic Cell-Targeted Oral Vaccine

PubMed Central

Sahay, Bikash; Owen, Jennifer L.; Yang, Tao; Zadeh, Mojgan; Lightfoot, Yaíma L.; Ge, Jun-Wei; Mohamadzadeh, Mansour

2015-01-01

Production of long-lived, high affinity humoral immunity is an essential characteristic of successful vaccination and requires cognate interactions between T and B cells in germinal centers. Within germinal centers, specialized T follicular helper cells assist B cells and regulate the antibody response by mediating the differentiation of B cells into memory or plasma cells after exposure to T cell-dependent antigens. It is now appreciated that local immune responses are also essential for protection against infectious diseases that gain entry to the host by the mucosal route; therefore, targeting the mucosal compartments is the optimum strategy to induce protective immunity. However, because the gastrointestinal mucosae are exposed to large amounts of environmental and dietary antigens on a daily basis, immune regulatory mechanisms exist to favor tolerance and discourage autoimmunity at these sites. Thus, mucosal vaccination strategies must ensure that the immunogen is efficiently taken up by the antigen presenting cells, and that the vaccine is capable of activating humoral and cellular immunity, while avoiding the induction of tolerance. Despite significant progress in mucosal vaccination, this potent platform for immunotherapy and disease prevention must be further explored and refined. Here we discuss recent progress in the understanding of the role of different phenotypes of B cells in the development of an efficacious mucosal vaccine against infectious disease. PMID:24372255

RELATIONSHIP OF GERMINAL CENTERS IN LYMPHOID TISSUE TO IMMUNOLOGICAL MEMORY

PubMed Central

Wakefield, J. D.; Thorbecke, G. J.

1968-01-01

The fate, proliferation, and developmental potentialities of cell suspensions made from white pulp containing large germinal centers have been studied in the mouse by transfer of cells labeled with thymidine-3H to lethally irradiated, syngeneic recipients. Radioautographic analyses were made using both smears and sections of a variety of tissues. Thymidine-3H-labeling patterns of white pulp showed that, initially, labeling occurred in a majority of blast and "intermediate cells" but in very few or no small lymphocytes. After intravenous transfer, most of the labeled cells localized in the lymphoid tissues of spleen, lymph nodes, and Peyer's patches. Few cells migrated to the thymus, lung, liver, and intestinal mucosa. Both after intravenous and after intraperitoneal transfer there was a rapid increase in the incidence of labeled small lymphocytes and a decrease of labeled blasts and intermediate cells. This was accompanied by an increase in the grain count of the small lymphocytes and a progressive decrease in the grain counts of the blast cells. Exposure of nonlabeled donor cells to thymidine-3H at various time intervals after transfer indicated that dividing cells were present early after transfer but that their incidence progressively decreased. Between 24 and 48 hr, very little cell division was detectable. PMID:5662013

Germinal and Somatic Activity of the Maize Element Activator (Ac) in Arabidopsis

PubMed Central

Keller, J.; Lim, E.; James-Jr., D. W.; Dooner, H. K.

1992-01-01

We have investigated the germinal and somatic activity of the maize Activator (Ac) element in Arabidopsis with the objective of developing an efficient transposon-based system for gene isolation in that plant. Transposition activity was assayed with a chimeric marker that consists of the cauliflower mosaic virus 35S promoter and a bacterial streptomycin phosphotransferase gene (SPT). Somatic activity was detected in seedlings germinated on plates containing streptomycin as green-resistant sectors against a background of white-sensitive cells. Germinal excisions resulted in fully green seedlings. The transposition frequency was extremely low when a single copy of the transposon was present, but appeared to increase with an increase in Ac copy number. Plants that were selected as variegated produced an increased number of green progeny. The methylation state of the Ac elements in lines with either low or high levels of excision was assessed by restriction analysis. No difference was found between these lines, indicating that the degree of methylation did not contribute to the level of Ac activity. Germinal excision events were analyzed molecularly and shown to carry reinserted transposons in about 50% of the cases. In several instances, streptomycin-resistant siblings carried the same transposed Ac element, indicating that excision had occurred prior to meiosis in the parent. We discuss parameters that need to be considered to optimize the use of Ac as a transposon tag in Arabidopsis. PMID:1322854

Photodynamic treatment with phenothiazinium photosensitizers kills both ungerminated and germinated microconidia of the pathogenic fungi Fusarium oxysporum, Fusarium moniliforme and Fusarium solani.

PubMed

de Menezes, Henrique Dantas; Tonani, Ludmilla; Bachmann, Luciano; Wainwright, Mark; Braga, Gilberto Úbida Leite; von Zeska Kress, Marcia Regina

2016-11-01

The search for alternatives to control microorganisms is necessary both in clinical and agricultural areas. Antimicrobial photodynamic treatment (APDT) is a promising light-based approach that can be used to control both human and plant pathogenic fungi. In the present study, we evaluated the effects of photodynamic treatment with red light and four phenothiazinium photosensitizers (PS): methylene blue (MB), toluidine blue O (TBO), new methylene blue N (NMBN) and the phenothiazinium derivative S137 on ungerminated and germinated microconidia of Fusarium oxysporum, F. moniliforme, and F. solani. APDT with each PS killed efficiently both the quiescent ungerminated microconidia and metabolically active germinated microconidia of the three Fusarium species. Washing away the unbound PS from the microconidia (both ungerminated and germinated) before red light exposure reduced but did not prevent the effect of APDT. Subcelullar localization of PS in ungerminated and germinated microconidia and the effects of photodynamic treatment on cell membranes were also evaluated in the three Fusarium species. APDT with MB, TBO, NMBN or S137 increased the membrane permeability in microconidia and APDT with NMBN or S137 increased the lipids peroxidation in microconidia of the three Fusarium species. These findings expand the understanding of photodynamic inactivation of filamentous fungi with phenothiazinium PS. Copyright © 2016 Elsevier B.V. All rights reserved.

Compartment-specific pH monitoring in Bacillus subtilis using fluorescent sensor proteins: a tool to analyze the antibacterial effect of weak organic acids

PubMed Central

van Beilen, Johan W. A.; Brul, Stanley

2013-01-01

The internal pH (pHi) of a living cell is one of its most important physiological parameters. To monitor the pH inside Bacillus subtilis during various stages of its life cycle, we constructed an improved version (IpHluorin) of the ratiometric, pH-sensitive fluorescent protein pHluorin by extending it at the 5′ end with the first 24 bp of comGA. The new version, which showed an approximate 40% increase in fluorescence intensity, was expressed from developmental phase-specific, native promoters of B. subtilis that are specifically active during vegetative growth on glucose (PptsG) or during sporulation (PspoIIA, PspoIIID, and PsspE). Our results show strong, compartment-specific expression of IpHluorin that allowed accurate pHi measurements of live cultures during exponential growth, early and late sporulation, spore germination, and during subsequent spore outgrowth. Dormant spores were characterized by an pHi of 6.0 ± 0.3. Upon full germination the pHi rose dependent on the medium to 7.0–7.4. The presence of sorbic acid in the germination medium inhibited a rise in the intracellular pH of germinating spores and inhibited germination. Such effects were absent when acetic was added at identical concentrations. PMID:23785365

B cell-intrinsic TLR7 signaling is required for optimal B cell responses during chronic viral infection

PubMed Central

Clingan, Jonathan M.; Matloubian, Mehrdad

2013-01-01

The importance for activation of innate immunity by pattern recognition receptors in forming an effective adaptive immune response is well known. Toll-like receptors (TLRs) have been demonstrated to be critical for antibody responses to a variety of immunizations. In particular, recent evidence suggests that B cell-intrinsic TLR signaling is required for optimal responses to virus-like antigens, but mechanisms by which TLR signaling impacts antibody responses during infection in vivo is unclear. In the present study, we demonstrate that deficiency of TLR7 in B cells alone is sufficient to significantly impact antibody responses in mice during chronic viral infection. This effect was independent of T follicular helper cells, and resulted in a loss of plasma cells generated later, but not early, in the response. The defect in plasma cell formation appeared to be secondary to a qualitative effect of TLR signaling on the germinal center (GC) B cell response. GC B cells in TLR7-deficient mice proliferated to a lesser extent and had a greater proportion of cells with phenotypic characteristics of light zone, relative to dark zone GC B cells. These results suggest that B cell-intrinsic TLR signaling in vivo likely affects plasma cell output by altered selection of antigen-specific B cells in the germinal center. PMID:23761632

Norway spruce embryogenesis: changes in carbohydrate profile, structural development and response to polyethylene glycol

PubMed Central

Hudec, Lukáš; Konrádová, Hana; Hašková, Anna; Lipavská, Helena

2016-01-01

Two unrelated, geographically distinct, highly embryogenic lines of Norway spruce (Picea abies (L.) Karst.) were analysed to identify metabolic traits characteristic for lines with good yields of high-quality embryos. The results were compared with corresponding characteristics of a poorly productive line (low embryo yield, scarce high-quality embryos). The following carbohydrate profiles and spectra during maturation, desiccation and germination were identified as promising characteristics for line evaluation: a gradual decrease in total soluble carbohydrates with an increasing sucrose : hexose ratio during maturation; accumulation of raffinose family oligosaccharides resulting from desiccation and their rapid degradation at the start of germination; and a decrease in sucrose, increase in hexoses and the appearance of pinitol with proceeding germination. We propose that any deviation from this profile in an embryonic line is a symptom of inferior somatic embryo development. We further propose that a fatty acid spectrum dominated by linoleic acid (18 : 2) was a common feature of healthy spruce somatic embryos, although it was quite different from zygotic embryos mainly containing oleic acid (18 : 1). The responses of the lines to osmotic stress were evaluated based on comparison of control (without osmoticum) and polyethylene glycol (PEG)-exposed (PEG 4000) variants. Although genetically distinct, both highly embryogenic lines responded in a very similar manner, with the only difference being sensitivity to high concentrations of PEG. At an optimum PEG concentration (3.75 and 5%), which was line specific, negative effects of PEG on embryo germination were compensated for by a higher maturation efficiency so that the application of PEG at an appropriate concentration improved the yield of healthy germinants per gram of initial embryonal mass and accelerated the process. Polyethylene glycol application, however, resulted in no improvement of the poorly productive line. PMID:27052433

Genome-wide analyses of late pollen-preferred genes conserved in various rice cultivars and functional identification of a gene involved in the key processes of late pollen development.

PubMed

Moon, Sunok; Oo, Moe Moe; Kim, Backki; Koh, Hee-Jong; Oh, Sung Aeong; Yi, Gihwan; An, Gynheung; Park, Soon Ki; Jung, Ki-Hong

2018-04-23

Understanding late pollen development, including the maturation and pollination process, is a key component in maintaining crop yields. Transcriptome data obtained through microarray or RNA-seq technologies can provide useful insight into those developmental processes. Six series of microarray data from a public transcriptome database, the Gene Expression Omnibus of the National Center for Biotechnology Information, are related to anther and pollen development. We performed a systematic and functional study across the rice genome of genes that are preferentially expressed in the late stages of pollen development, including maturation and germination. By comparing the transcriptomes of sporophytes and male gametes over time, we identified 627 late pollen-preferred genes that are conserved among japonica and indica rice cultivars. Functional classification analysis with a MapMan tool kit revealed a significant association between cell wall organization/metabolism and mature pollen grains. Comparative analysis of rice and Arabidopsis demonstrated that genes involved in cell wall modifications and the metabolism of major carbohydrates are unique to rice. We used the GUS reporter system to monitor the expression of eight of those genes. In addition, we evaluated the significance of our candidate genes, using T-DNA insertional mutant population and the CRISPR/Cas9 system. Mutants from T-DNA insertion and CRISPR/Cas9 systems of a rice gene encoding glycerophosphoryl diester phosphodiesterase are defective in their male gamete transfer. Through the global analyses of the late pollen-preferred genes from rice, we found several biological features of these genes. First, biological process related to cell wall organization and modification is over-represented in these genes to support rapid tube growth. Second, comparative analysis of late pollen preferred genes between rice and Arabidopsis provide a significant insight on the evolutional disparateness in cell wall biogenesis and storage reserves of pollen. In addition, these candidates might be useful targets for future examinations of late pollen development, and will be a valuable resource for accelerating the understanding of molecular mechanisms for pollen maturation and germination processes in rice.

Toxicity of selected acaricides in a glass-vial bioassay to two-spotted spider mite (Acari: Tetranychidae)

USDA-ARS?s Scientific Manuscript database

Two-spotted spider mite (TSSM), Tetranychus urticae Koch, feeds on epidermal cells of cotton foliage, destroys photosynthetic cells, and reduces yields, fiber quality and seed germination. With a short life cycle, prolific fecundity, an arrhenotokous reproduction, and an ability to expeditiously dig...

Influence of microgravity on root-cap regeneration and the structure of columella cells in Zea mays

NASA Technical Reports Server (NTRS)

Moore, R.; McClelen, C. E.; Fondren, W. M.; Wang, C. L.

1987-01-01

We launched imbibed seeds and seedlings of Zea mays into outer space aboard the space shuttle Columbia to determine the influence of microgravity on 1) root-cap regeneration, and 2) the distribution of amyloplasts and endoplasmic reticulum (ER) in the putative statocytes (i.e., columella cells) of roots. Decapped roots grown on Earth completely regenerated their caps within 4.8 days after decapping, while those grown in microgravity did not regenerate caps. In Earth-grown seedlings, the ER was localized primarily along the periphery of columella cells, and amyloplasts sedimented in response to gravity to the lower sides of the cells. Seeds germinated on Earth and subsequently launched into outer space had a distribution of ER in columella cells similar to that of Earth-grown controls, but amyloplasts were distributed throughout the cells. Seeds germinated in outer space were characterized by the presence of spherical and ellipsoidal masses of ER and randomly distributed amyloplasts in their columella cells. These results indicate that 1) gravity is necessary for regeneration of the root cap, 2) columella cells can maintain their characteristic distribution of ER in microgravity only if they are exposed previously to gravity, and 3) gravity is necessary to distribute the ER in columella cells of this cultivar of Z. mays.

Radiosensitivity of antibody responses and radioresistant secondary tetanus antitoxin responses

DOE Office of Scientific and Technical Information (OSTI.GOV)

Stoner, R.; Terres, G.; Cottier, H.

1976-01-01

Primary tetanus antitoxin responses were increasingly repressed in mice when gamma radiation doses of 100 to 400 rads were delivered by whole-body exposure prior to immunization with fluid tetanus toxoid (FTT). Nearly normal secondary antitoxin responses were obtained in mice exposed to 600 rads of gamma radiation 4 days after secondary antigenic stimulation with FTT. A rapid transition from radiosensitivity of the antibody-forming system on days 1 to 3 was followed by relative radioresistance on day 4 after the booster injection of toxoid. Studies on lymphoid cellular kinetics in popliteal lymph nodes after injection of $sup 3$H--thymidine ($sup 3$H--TdR) andmore » incorporation of $sup 3$H--L- histidine into circulating antitoxin were carried out. Analysis of tritium radioactivity in antigen--antibody precipitates of serums 2 hr after injection of the labeled amino acid revealed maximum incorporation into antibody around day 7 after the booster in nonirradiated controls and about day 12, i.e., 8 days after irradiation, in experimental mice. The shift from radiosensitivity to relative radioresistance was attributed to a marked peak of plasma-cell proliferation in the medulla of lymph nodes on day 3. Many medullary plasma cells survived and continued to proliferate after exposure to radiation. Germinal centers were destroyed by radiation within 1 day. Since antibody formation continued after exposure to radiation and after the loss of germinal centers, this supports the view that germinal-center cells were involved more in the generation of memory cells than in antibody synthesis. (auth)« less

Oogenesis: From Oogonia to Ovulation in the Flagfish, Jordanella floridae Goode and Bean, 1879 (Teleostei: Cyprinodontidae).

PubMed

Uribe, Mari Carmen; Grier, Harry J; García-Alarcón, Adriana; Parenti, Lynne R

2016-10-01

We provide histological details of the development of oocytes in the cyprinodontid flagfish, Jordanella floridae. There are six stages of oogenesis: Oogonial proliferation, chromatin nucleolus, primary growth (previtellogenesis [PG]), secondary growth (vitellogenesis), oocyte maturation and ovulation. The ovarian lamellae are lined by a germinal epithelium composed of epithelial cells and scattered oogonia. During primary growth, the development of cortical alveoli and oil droplets, are initiated simultaneously. During secondary growth, yolk globules coalesce into a fluid mass. The full-grown oocyte contains a large globule of fluid yolk. The germinal vesicle is at the animal pole, and the cortical alveoli and oil droplets are located at the periphery. The disposition of oil droplets at the vegetal pole of the germinal vesicle during late secondary growth stage is a unique characteristic. The follicular cell layer is composed initially of a single layer of squamous cells during early PG which become columnar during early vitellogenesis. During primary and secondary growth stages, filaments develop among the follicular cells and also around the micropyle. The filaments are seen extending from the zona pellucida after ovulation. During ovulation, a space is evident between the oocyte and the zona pellucida. Asynchronous spawning activity is confirmed by the observation that, after ovulation, the ovarian lamellae contain follicles in both primary and secondary growth stages; in contrast, when the seasonal activity of oogenesis and spawning ends, after ovulation, the ovarian lamellae contain only follicles in the primary growth stage. J. Morphol. 277:1339-1354, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Identification of novel tumor antigens with patient-derived immune-selected antibodies

PubMed Central

Rodriguez-Pinto, Daniel; Sparkowski, Jason; Keough, Martin P.; Phoenix, Kathryn N.; Vumbaca, Frank; Han, David K.; Gundelfinger, Eckart D.; Beesley, Philip

2010-01-01

The identification of tumor antigens capable of eliciting an immune response in vivo may be an effective method to identify therapeutic cancer targets. We have developed a method to identify such antigens using frozen tumor-draining lymph node samples from breast cancer patients. Immune responses in tumor-draining lymph nodes were identified by immunostaining lymph node sections for B-cell markers (CD20&CD23) and Ki67 which revealed cell proliferation in germinal center zones. Antigen-dependent somatic hypermutation (SH) and clonal expansion (CE) were present in heavy chain variable (VH) domain cDNA clones obtained from these germinal centers, but not from Ki67 negative germinal centers. Recombinant VH single-domain antibodies were used to screen tumor proteins and affinity select potential tumor antigens. Neuroplastin (NPTN) was identified as a candidate breast tumor antigen using proteomic identification of affinity selected tumor proteins with a recombinant VH single chain antibody. NPTN was found to be highly expressed in approximately 20% of invasive breast carcinomas and 50% of breast carcinomas with distal metastasis using a breast cancer tissue array. Additionally, NPTN over-expression in a breast cancer cell line resulted in a significant increase in tumor growth and angiogenesis in vivo which was related to increased VEGF production in the transfected cells. These results validate NPTN as a tumor-associated antigen which could promote breast tumor growth and metastasis if aberrantly expressed. These studies also demonstrate that humoral immune responses in tumor-draining lymph nodes can provide antibody reagents useful in identifying tumor antigens with applications for biomarker screening, diagnostics and therapeutic interventions. PMID:18568347

Histology and immunohistochemistry of the gut-associated lymphoid tissue of the eastern grey kangaroo, Macropus giganteus.

PubMed

Old, J M; Deane, E M

2001-12-01

Mesenteric lymph nodes and gut-associated lymphoid tissue (GALT) from juvenile eastern grey kangaroos were investigated. The mesenteric nodes had a similar structure to that described for eutherian mammals. They contained distinct regions of medulla and cortex, with prominent follicles and germinal centres. Gut associated lymphoid tissue consisted of areas of submucosal follicles. These varied from areas of densely packed lymphocytes with darkly staining, prominent coronas to areas with no defined follicles. The distribution of T cells in these tissues was documented by use of species-crossreactive antibodies to the surface markers CD3 and CD5; B cells were identified by antibodies to CD79b. Within the lymph nodes T cells were located mainly in the paracortex and cortex, with limited numbers observed in the follicles; B cells were located on the marginal zone of the follicles. In GALT, T cells were located in the peripheral regions of the germinal centres of secondary follicles, while B cells were abundant in primary follicles. These observations are consistent with those made in a range of other marsupials (metatherian) and eutherian mammals and are indicative of the capacity to respond to antigens entering via the mouth.

Inhibition of Orobanche crenata seed germination and radicle growth by allelochemicals identified in cereals.

PubMed

Fernández-Aparicio, Mónica; Cimmino, Alessio; Evidente, Antonio; Rubiales, Diego

2013-10-16

Orobanche crenata is a parasitic weed that causes severe yield losses in important grain and forage legume crops. Cereals have been reported to inhibit O. crenata parasitism when grown intercropped with susceptible legumes, but the responsible metabolites have not been identified. A number of metabolites have been reported in cereals that have allelopathic properties against weeds, pests, and pathogens. We tested the effect of several allelochemicals identified in cereals on O. crenata seed germination and radicle development. We found that 2-benzoxazolinone, its derivative 6-chloroacetyl-2-benzoxazolinone, and scopoletin significantly inhibited O. crenata seed germination. Benzoxazolinones, l-tryptophan, and coumalic acid caused the stronger inhibition of radicle growth. Also, other metabolites reduced radicle length, this inhibition being dose-dependent. Only scopoletin caused cell necrotic-like darkening in the young radicles. Prospects for their application to parasitic weed management are discussed.

Responses of indigenous microorganisms to soil incubation as viewed by transmission electron microscopy of cell thin sections.

NASA Technical Reports Server (NTRS)

Bae, h. C.; Casida, L. E., Jr.

1973-01-01

Indigenous soil microorganisms were cultivated in their soil habitat with 50% moisture capacity at 30 C for two weeks. Changes in microorganism cells were studied by electron microscopy during incubation, with particular attention to the dormant cell growth and to the ability of cystlike cells to germinate and reencyst. The responses of various cell species to incubation conditions are described and illustrated by photomicrographs.

Nucleoli from two-cell embryos support the development of enucleolated germinal vesicle oocytes in the pig.

PubMed

Kyogoku, Hirohisa; Ogushi, Sugako; Miyano, Takashi

2012-11-01

Recent research has shown that nucleoli of oocytes at the germinal vesicle (GV) stage (GV nucleoli) are not necessary for oocyte maturation but are essential for early embryonic development. Nucleoli of 2-cell embryos (2-cell nucleoli) have morphology similar to that of nucleoli in oocytes at the GV stage. In this study, we examined the ability of 2-cell nucleoli to substitute for GV nucleoli in terms of supporting early embryonic development by nucleolus aspiration (enucleolation) and transfer into metaphase II (MII) oocytes or 2-cell embryos that were derived from enucleolated oocytes at the GV stage in the pig. When 2-cell embryos were centrifuged to move the lipid droplets to one side of the blastomere, multiple nucleoli in the nucleus fused into a single nucleolus. The nucleoli were then aspirated from the 2-cell embryos by micromanipulation. The injection of 2-cell nucleoli to GV enucleolated oocytes at the MII stage rescued the embryos from the early embryonic arrest, and the resulting oocytes developed to blastocysts. However, the injection of 2-cell and GV nucleoli to 2-cell embryos derived from GV enucleolated oocytes rarely restored the development to blastocysts. These results indicate that 2-cell nucleoli support early embryonic development as GV nucleoli and that the presence of nucleoli is essential for pig embryos before the 2-cell stage.

B cell IFN-γ receptor signaling promotes autoimmune germinal centers via cell-intrinsic induction of BCL-6

PubMed Central

Jackson, Shaun W.; Jacobs, Holly M.; Arkatkar, Tanvi; Dam, Elizabeth M.; Scharping, Nicole E.; Kolhatkar, Nikita S.; Hou, Baidong; Buckner, Jane H.

2016-01-01

Dysregulated germinal center (GC) responses are implicated in the pathogenesis of human autoimmune diseases, including systemic lupus erythematosus (SLE). Although both type 1 and type 2 interferons (IFNs) are involved in lupus pathogenesis, their respective impacts on the establishment of autoimmune GCs has not been addressed. In this study, using a chimeric model of B cell-driven autoimmunity, we demonstrate that B cell type 1 IFN receptor signals accelerate, but are not required for, lupus development. In contrast, B cells functioning as antigen-presenting cells initiate CD4+ T cell activation and IFN-γ production, and strikingly, B cell–intrinsic deletion of the IFN-γ receptor (IFN-γR) abrogates autoimmune GCs, class-switched autoantibodies (auto-Abs), and systemic autoimmunity. Mechanistically, although IFN-γR signals increase B cell T-bet expression, B cell–intrinsic deletion of T-bet exerts an isolated impact on class-switch recombination to pathogenic auto-Ab subclasses without impacting GC development. Rather, in both mouse and human B cells, IFN-γ synergized with B cell receptor, toll-like receptor, and/or CD40 activation signals to promote cell-intrinsic expression of the GC master transcription factor, B cell lymphoma 6 protein. Our combined findings identify a novel B cell–intrinsic mechanism whereby IFN signals promote lupus pathogenesis, implicating this pathway as a potential therapeutic target in SLE. PMID:27069113

Isolation of Small SSEA-4-Positive Putative Stem Cells from the Ovarian Surface Epithelium of Adult Human Ovaries by Two Different Methods

PubMed Central

Virant-Klun, Irma; Skutella, Thomas; Hren, Matjaz; Gruden, Kristina; Cvjeticanin, Branko; Vogler, Andrej; Sinkovec, Jasna

2013-01-01

The adult ovarian surface epithelium has already been proposed as a source of stem cells and germinal cells in the literature, therefore it has been termed the “germinal epithelium”. At present more studies have confirmed the presence of stem cells expressing markers of pluripotency in adult mammalian ovaries, including humans. The aim of this study was to isolate a population of stem cells, based on the expression of pluripotency-related stage-specific embryonic antigen-4 (SSEA-4) from adult human ovarian surface epithelium by two different methods: magnetic-activated cell sorting and fluorescence-activated cell sorting. Both methods made it possible to isolate a similar, relatively homogenous population of small, SSEA-4-positive cells with diameters of up to 4 μm from the suspension of cells retrieved by brushing of the ovarian cortex biopsies in reproductive-age and postmenopausal women and in women with premature ovarian failure. The immunocytochemistry and genetic analyses revealed that these small cells—putative stem cells—expressed some primordial germ cell and pluripotency-related markers and might be related to the in vitro development of oocyte-like cells expressing some oocyte-specific transcription factors in the presence of donated follicular fluid with substances important for oocyte growth and development. The stemness of these cells needs to be further researched. PMID:23509763

Analysis of the Spore Membrane Proteome in Clostridium perfringens Implicates Cyanophycin in Spore Assembly.

PubMed

Liu, Hualan; Ray, W Keith; Helm, Richard F; Popham, David L; Melville, Stephen B

2016-06-15

Heat-resistant endospore formation plays an important role in Clostridium perfringens-associated foodborne illnesses. The spores allow the bacterium to survive heating during normal cooking processes, followed by germination and outgrowth of the bacterium in contaminated foods. To identify proteins associated with germination and other spore functions, a comparative spore membrane proteome analysis of dormant and germinated spores of C. perfringens strain SM101 was performed by using gel-based protein separation and liquid chromatography coupled with matrix-assisted laser desorption ionization-tandem time of flight (MALDI-TOF/TOF) mass spectrometry. A total of 494 proteins were identified, and 117 of them were predicted to be integral membrane or membrane-associated proteins. Among these membrane proteins, 16 and 26 were detected only in dormant and germinated spores, respectively. One protein that was detected only in germinated spore membranes was the enzyme cyanophycinase, a protease that cleaves the polymer cyanophycin, which is composed of l-arginine-poly(l-aspartic acid), to β-Asp-Arg. Genes encoding cyanophycinase and cyanophycin synthetase have been observed in many species of Clostridium, but their role has not been defined. To determine the function of cyanophycin in C. perfringens, a mutation was introduced into the cphA gene, encoding cyanophycin synthetase. In comparison to parent strain SM101, the spores of the mutant strain retained wild-type levels of heat resistance, but fewer spores were made, and they were smaller, suggesting that cyanophycin synthesis plays a role in spore assembly. Although cyanophycin could not be extracted from sporulating C. perfringens cells, an Escherichia coli strain expressing the cphA gene made copious amounts of cyanophycin, confirming that cphA encodes a cyanophycin synthetase. Clostridium perfringens is a common cause of food poisoning, and germination of spores after cooking is thought to play a significant role in the disease. How C. perfringens controls the germination process is still not completely understood. We characterized the proteome of the membranes from dormant and germinated spores and discovered that large-scale changes occur after germination is initiated. One of the proteins that was detected after germination was the enzyme cyanophycinase, which degrades the storage compound cyanophycin, which is found in cyanobacteria and other prokaryotes. A cyanophycin synthetase mutant was constructed and found to make spores with altered morphology but normal heat resistance, suggesting that cyanophycin plays a different role in C. perfringens than it does in cyanobacteria. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Dispersed phase volume fraction, weak acids and Tween 80 in a model emulsion: Effect on the germination and growth of Bacillus weihenstephanensis KBAB4 spores.

PubMed

Léonard-Akkari, Lucie; Guégan, Stéphanie; Courand, Fabienne; Couvert, Olivier; Lepage, Jean-François; Rondeau-Mouro, Corinne; Desriac, Noémie; Mathot, Anne-Gabrielle; Leguérinel, Ivan; Coroller, Louis; Decourcelle, Nicolas

2018-07-01

In foodstuffs, physico-chemical interactions and/or physical constraints between spores, inhibitors and food components may exist. Thus, the objective of this study was to investigate such interactions using a model emulsion as a microbial medium in order to improve bacterial spore control with better knowledge of the interactions in the formulation. Emulsions were prepared with hexadecane mixed with nutrient broth using sonication and were stabilized by Tween 80 and Span 80. The hexadecane ratio was either 35% (v/v) or 50% (v/v) and each emulsion was studied in the presence of organic acid (acetic, lactic or hexanoic) at two pH levels (5.5 and 6). Self-diffusion coefficients of emulsion components and the organic acids were measured by Pulsed Field Gradient-Nuclear Magnetic Resonance (PFG-NMR). The inhibition effect on the spore germination and cell growth of Bacillus weihenstephanensis KBAB4 was characterized by the measure of the probability of growth using the most probable number methodology, and the measure of the time taken for the cells to germinate and grow using a single cell Bioscreen® method and using flow cytometry. The inhibition of spore germination and growth in the model emulsion depended on the dispersed phase volume fraction and the pH value. The effect of the dispersed phase volume fraction was due to a combination of (i) the lipophilicity of the biocide, hexanoic acid, that may have had an impact on the distribution of organic acid between hexadecane and the aqueous phases and (ii) the antimicrobial activity of the emulsifier Tween 80 detected at the acidic pH value. The interface phenomena seemed to have a major influence. Future work will focus on the exploration of these phenomena at the interface. Copyright © 2018 Elsevier Ltd. All rights reserved.

Members of the gibberellin receptor gene family GID1 (GIBBERELLIN INSENSITIVE DWARF1) play distinct roles during Lepidium sativum and Arabidopsis thaliana seed germination

PubMed Central

Voegele, Antje; Linkies, Ada; Müller, Kerstin; Leubner-Metzger, Gerhard

2011-01-01

Germination of endospermic seeds is partly regulated by the micropylar endosperm, which acts as constraint to radicle protrusion. Gibberellin (GA) signalling pathways control coat-dormancy release, endosperm weakening, and organ expansion during seed germination. Three GIBBERELLIN INSENSITIVE DWARF1 (GID1) GA receptors are known in Arabidopsis thaliana: GID1a, GID1b, and GID1c. Molecular phylogenetic analysis of angiosperm GID1s reveals that they cluster into two eudicot (GID1ac, GID1b) groups and one monocot group. Eudicots have at least one gene from each of the two groups, indicating that the different GID1 receptors fulfil distinct roles during plant development. A comparative Brassicaceae approach was used, in which gid1 mutant and whole-seed transcript analyses in Arabidopsis were combined with seed-tissue-specific analyses of its close relative Lepidium sativum (garden cress), for which three GID1 orthologues were cloned. GA signalling via the GID1ac receptors is required for Arabidopsis seed germination, GID1b cannot compensate for the impaired germination of the gid1agid1c mutant. Transcript expression patterns differed temporarily, spatially, and hormonally, with GID1b being distinct from GID1ac in both species. Endosperm weakening is mediated, at least in part, through GA-induced genes encoding cell-wall-modifying proteins. A suppression subtraction hybridization (SSH) cDNA library enriched for sequences that are highly expressed during early germination in the micropylar endosperm contained expansins and xyloglucan endo-transglycosylases/hydrolases (XTHs). Their transcript expression patterns in both species strongly suggest that they are regulated by distinct GID1-mediated GA signalling pathways. The GID1ac and GID1b pathways seem to fulfil distinct regulatory roles during Brassicaceae seed germination and seem to control their downstream targets distinctly. PMID:21778177

Circulating CXCR5+CD4+ T cells participate in the IgE accumulation in allergic asthma.

PubMed

Gong, Fang; Zhu, Hua-Yan; Zhu, Jie; Dong, Qiao-Jing; Huang, Xuan; Jiang, Dong-Jin

2018-05-01

The pathogenesis of allergic asthma is primarily characterized by abnormality in immunoglobin(Ig)E pathway, suggesting a possible role for follicular helper T cells (Tfh) in the genesis of excessive IgE accumulation. The blood chemokine (C-X-C motif) receptor 5 (CXCR)5 + CD4 + T cells, known as "circulating" Tfh, share common functional characteristics with Tfh cells from germinal centers. The aim of this study was to determine the phenotypes and functions of circulating CXCR5 + CD4 + T cells in allergic asthmatics. Here we found the frequency of the circulating CXCR5 + CD4 + T cells was raised in allergic asthma compared with healthy control (HC). Phenotypic assays showed that activated circulating CXCR5 + CD4 + T cells display the key features of Tfh cells, including invariably coexpressed programmed cell death (PD)-1 and inducible costimulator (ICOS). The frequency of interleukin IL-4 + -, IL-21 + -producing CXCR5 + CD4 + T cells was increased in allergic asthma patients compared with HC. Furthermore, sorted circulating CXCR5 + CD4 + T cells from allergic asthma patients boosted IgE production in coculture assay which could be inhibited by IL-4 or IL-21 blockage. Interestingly, IL-4 + -, IL-21 + -CXCR5 + CD4 + T cells positively correlated with total IgE in the blood. Our data indicated that circulating CXCR5 + CD4 + T cells may have a significant role in facilitating IgE production in allergic asthma patients. Copyright © 2018 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.

Isolation and structure elucidation of avocado seed (Persea americana) lipid derivatives that inhibit Clostridium sporogenes endospore germination.

PubMed

Rodríguez-Sánchez, Dariana Graciela; Pacheco, Adriana; García-Cruz, María Isabel; Gutiérrez-Uribe, Janet Alejandra; Benavides-Lozano, Jorge Alejandro; Hernández-Brenes, Carmen

2013-07-31

Avocado fruit extracts are known to exhibit antimicrobial properties. However, the effects on bacterial endospores and the identity of antimicrobial compounds have not been fully elucidated. In this study, avocado seed extracts were tested against Clostridium sporogenes vegetative cells and active endospores. Bioassay-guided purification of a crude extract based on inhibitory properties linked antimicrobial action to six lipid derivatives from the family of acetogenin compounds. Two new structures and four compounds known to exist in nature were identified as responsible for the activity. Structurally, most potent molecules shared features of an acetyl moiety and a trans-enone group. All extracts produced inhibition zones on vegetative cells and active endospores. Minimum inhibitory concentrations (MIC) of isolated molecules ranged from 7.8 to 15.6 μg/mL, and bactericidal effects were observed for an enriched fraction at 19.5 μg/mL. Identified molecules showed potential as natural alternatives to additives and antibiotics used by the food and pharmaceutical industries to inhibit Gram-positive spore-forming bacteria.

Phytochrome-mediated germination and early development in spores of Dryopteris filix-mas L.: phase-specific and non phase-specific inhibition by staurosporine

NASA Technical Reports Server (NTRS)

Haas, C. J.; Scheuerlein, R.; Roux, S. J.

1991-01-01

The alkaloid staurosporine, currently known as the most potent inhibitor of protein kinase C, PKC, was tested for its ability to inhibit phytochrome-mediated spore germination in Dryopteris filix-mas L., evaluated by the induction of chlorophyll synthesis. Approximately half-maximal inhibition was obtained at a concentration of 10(-5) M. This effect of staurosporine was phase-specific and was found during the same period in which the presence of extracellular calcium is necessary for realization of the light signal. Furthermore, the ability of staurosporine to prevent progression of a germinated spore into early gametophyte development, evaluated by the accumulation of chlorophyll, was examined. Again, staurosporine (10(-5) M) significantly diminished chlorophyll accumulation, determined quantitatively in vivo by single-cell measurements, in a non-phase specific way. The fact that the phase-specific inhibitory effect of staurosporine in preventing germination was coincident with the phase-specific requirement of Ca2+ suggests that both Ca2+ and staurosporine affect the same step in the signal-transduction chain. A phosphorylation event catalysed by PKC or any Ca2+ -dependent protein kinase is proposed as the target of staurosporine and Ca2+.

Effect of stationary magnetic field strengths of 150 and 200 mT on reactive oxygen species production in soybean.

PubMed

Shine, M B; Guruprasad, K N; Anand, Anjali

2012-07-01

Our previous investigation reported the beneficial effect of pre-sowing magnetic treatment for improving germination parameters and biomass accumulation in soybean. In this study, soybean seeds treated with static magnetic fields of 150 and 200 mT for 1 h were evaluated for reactive oxygen species (ROS) and activity of antioxidant enzymes. Superoxide and hydroxyl radicals were measured in embryos and hypocotyls of germinating seeds by electron paramagnetic resonance spectroscopy and kinetics of superoxide production; hydrogen peroxide and antioxidant activities were estimated spectrophotometrically. Magnetic field treatment resulted in enhanced production of ROS mediated by cell wall peroxidase while ascorbic acid content, superoxide dismutase and ascorbate peroxidase activity decreased in the hypocotyl of germinating seeds. An increase in the cytosolic peroxidase activity indicated that this antioxidant enzyme had a vital role in scavenging the increased H(2)O(2) produced in seedlings from the magnetically treated seeds. Hence, these studies contribute to our first report on the biochemical basis of enhanced germination and seedling growth in magnetically treated seeds of soybean in relation to increased production of ROS. Copyright © 2012 Wiley Periodicals, Inc.

Action of the Metalloproteinases in Gonadal Remodeling during Sex Reversal in the Sequential Hermaphroditism of the Teleostei Fish Synbranchus marmoratus (Synbranchiformes: Synbranchidae)

PubMed Central

Mazzoni, Talita Sarah; Lo Nostro, Fabiana Laura; Antoneli, Fernanda Natália; Quagio-Grassiotto, Irani

2018-01-01

Teleostei present great plasticity regarding sex change. During sex reversal, the whole gonad including the germinal epithelium undergoes significant changes, remodeling, and neoformation. However, there is no information on the changes that occur within the interstitial compartment. Considering the lack of information, especially on the role played by metalloproteinases (MMPs) in fish gonadal remodeling, the aim of this study was to evaluate the action of MMPs on gonads of sex reversed females of Synbranchus marmoratus, a fresh water protogynic diandric fish. Gonads were processed for light microscopy and blood samples were used for the determination of plasma sex steroid levels. During sex reversal, degeneration of the ovaries occurred and were gradually replaced by the germinal tissue of the male. The action of the MMPs induces significant changes in the interstitial compartment, allowing the reorganization of germinal epithelium. Leydig cells also showed an important role in female to male reversion. The gonadal transition coincides with changes in circulating sex steroid levels throughout sex reversion. The action of the MMPs, in the gonadal remodeling, especially on the basement membrane, is essential for the establishment of a new functional germinal epithelium. PMID:29695033

Bacillus sphaericus LMG 22257 is physiologically suitable for self-healing concrete.

PubMed

Wang, Jianyun; Jonkers, Henk M; Boon, Nico; De Belie, Nele

2017-06-01

The suitability of using a spore-forming ureolytic strain, Bacillus sphaericus, was evaluated for self-healing of concrete cracks. The main focus was on alkaline tolerance, calcium tolerance, oxygen dependence, and low-temperature adaptability. Experimental results show that B. sphaericus had a good tolerance. It can grow and germinate in a broad range of alkaline pH. The optimal pH range is 7 ∼ 9. High alkaline conditions (pH 10 ∼ 11) slow down but not stop the growth and germination. Oxygen was strictly needed during bacterial growth and germination, but not an essential factor during bacterial urea decomposition. B. sphaericus also had a good Ca tolerance, especially at a high bacterial concentration of 10 8  cells/mL; no significant influence was observed on bacterial ureolytic activity of the presence of 0.9M Ca 2+ . Furthermore, at a low temperature (10 °C), bacterial spores germinated and revived ureolytic activity with some retardation. However, this retardation can be counteracted by using a higher bacterial concentration and by supplementing yeast extract. It can be concluded that B. sphaericus is a suitable bacterium for application in bacteria-based self-healing concrete.

Richter transformation driven by Epstein-Barr virus reactivation during therapy-related immunosuppression in chronic lymphocytic leukaemia.

PubMed

García-Barchino, Maria J; Sarasquete, Maria E; Panizo, Carlos; Morscio, Julie; Martinez, Antonio; Alcoceba, Miguel; Fresquet, Vicente; Gonzalez-Farre, Blanca; Paiva, Bruno; Young, Ken H; Robles, Eloy F; Roa, Sergio; Celay, Jon; Larrayoz, Marta; Rossi, Davide; Gaidano, Gianluca; Montes-Moreno, Santiago; Piris, Miguel A; Balanzategui, Ana; Jimenez, Cristina; Rodriguez, Idoia; Calasanz, Maria J; Larrayoz, Maria J; Segura, Victor; Garcia-Muñoz, Ricardo; Rabasa, Maria P; Yi, Shuhua; Li, Jianyong; Zhang, Mingzhi; Xu-Monette, Zijun Y; Puig-Moron, Noemi; Orfao, Alberto; Böttcher, Sebastian; Hernandez-Rivas, Jesus M; Miguel, Jesus San; Prosper, Felipe; Tousseyn, Thomas; Sagaert, Xavier; Gonzalez, Marcos; Martinez-Climent, Jose A

2018-05-01

The increased risk of Richter transformation (RT) in patients with chronic lymphocytic leukaemia (CLL) due to Epstein-Barr virus (EBV) reactivation during immunosuppressive therapy with fludarabine other targeted agents remains controversial. Among 31 RT cases classified as diffuse large B-cell lymphoma (DLBCL), seven (23%) showed EBV expression. In contrast to EBV - tumours, EBV + DLBCLs derived predominantly from IGVH-hypermutated CLL, and they also showed CLL-unrelated IGVH sequences more frequently. Intriguingly, despite having different cellular origins, clonally related and unrelated EBV + DLBCLs shared a previous history of immunosuppressive chemo-immunotherapy, a non-germinal centre DLBCL phenotype, EBV latency programme type II or III, and very short survival. These data suggested that EBV reactivation during therapy-related immunosuppression can transform either CLL cells or non-tumoural B lymphocytes into EBV + DLBCL. To investigate this hypothesis, xenogeneic transplantation of blood cells from 31 patients with CLL and monoclonal B-cell lymphocytosis (MBL) was performed in Rag2 -/- IL2γc -/- mice. Remarkably, the recipients' impaired immunosurveillance favoured the spontaneous outgrowth of EBV + B-cell clones from 95% of CLL and 64% of MBL patients samples, but not from healthy donors. Eventually, these cells generated monoclonal tumours (mostly CLL-unrelated but also CLL-related), recapitulating the principal features of EBV + DLBCL in patients. Accordingly, clonally related and unrelated EBV + DLBCL xenografts showed indistinguishable cellular, virological and molecular features, and synergistically responded to combined inhibition of EBV replication with ganciclovir and B-cell receptor signalling with ibrutinib in vivo. Our study underscores the risk of RT driven by EBV in CLL patients receiving immunosuppressive therapies, and provides the scientific rationale for testing ganciclovir and ibrutinib in EBV + DLBCL. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Colonization strategy of the endophytic plant growth-promoting strains of Pseudomonas fluorescens and Klebsiella oxytoca on the seeds, seedlings and roots of the epiphytic orchid, Dendrobium nobile Lindl.

PubMed

Pavlova, A S; Leontieva, M R; Smirnova, T A; Kolomeitseva, G L; Netrusov, A I; Tsavkelova, E A

2017-04-29

Orchids form strong mycorrhizal associations, but their interactions with bacteria are poorly understood. We aimed to investigate the distribution of plant growth promoting rhizobacteria (PGPR) at different stages of orchid development and to study if there is any selective specificity in choosing PGPR partners. Colonization patterns of gfp-tagged Pseudomonas fluorescens and Klebsiella oxytoca were studied on roots, seeds, and seedlings of Dendrobium nobile. Endophytic rhizobacteria rapidly colonized velamen and core parenchyma entering through exodermis and the passage cells, whereas at the early stages, they stayed restricted to the surface and the outer layers of the protocorms and rhizoids. The highest amounts of auxin (indole-3-acetic acid) were produced by K. oxytoca and P. fluorescens in the nitrogen-limiting and NO 3 -containing media respectively. Bacterization of D. nobile seeds resulted in promotion of their in vitro germination. The plant showed no selective specificity to the tested strains. Klebsiella oxytoca demonstrated more intense colonization activity and more efficient growth promoting impact under tryptophan supplementation, while P. fluorescens revealed its growth-promoting capacity without tryptophan. Both strategies are regarded as complementary, improving adaptive potentials of the orchid when different microbial populations colonize the plant. This study enlarges our knowledge on orchid-microbial interactions, and provides new features on application of the nonorchid PGPR in orchid seed germination and conservation. © 2017 The Society for Applied Microbiology.

Computational investigations of the band structure, and thermodynamic and optical features of thorium-based oxide ThGeO4 using the full-potential linearized augmented plane-wave plus local orbital approach

NASA Astrophysics Data System (ADS)

Chiker, F.; Khachai, H.; Mathieu, C.; Bin-Omran, S.; Kada, Belkacem; Sun, Xiao-Wei; Sandeep; Rai, D. P.; Khenata, R.

2018-05-01

In this study, first-principles investigations were performed using the full-potential linearized augmented plane-wave method of the structural and optoelectronic properties of thorium germinate (ThGeO4), a high-K dielectric material. Under ambient conditions, the structural properties calculated for ThGeO4 in the zircon phase were in excellent agreement with the available experimental data. Furthermore, using the modified Becke -Johnson correction method, the calculated band gaps and optical constants accurately described this compound. Finally, the thermal properties were predicted over a temperature range of 0-700 K and pressures up to 11 GPa using the quasi-harmonic Debye model, where the variations in the heat capacity, primitive cell volume, and thermal expansion coefficients were determined successfully.

Ethylene Receptors Signal via a Noncanonical Pathway to Regulate Abscisic Acid Responses1[OPEN

PubMed Central

Bakshi, Arkadipta; Fernandez, Jessica C.

2018-01-01

Ethylene is a gaseous plant hormone perceived by a family of receptors in Arabidopsis (Arabidopsis thaliana) including ETHYLENE RESPONSE1 (ETR1) and ETR2. Previously we showed that etr1-6 loss-of-function plants germinate better and etr2-3 loss-of-function plants germinate worse than wild-type under NaCl stress and in response to abscisic acid (ABA). In this study, we expanded these results by showing that ETR1 and ETR2 have contrasting roles in the control of germination under a variety of inhibitory conditions for seed germination such as treatment with KCl, CuSO4, ZnSO4, and ethanol. Pharmacological and molecular biology results support a model where ETR1 and ETR2 are indirectly affecting the expression of genes encoding ABA signaling proteins to affect ABA sensitivity. The receiver domain of ETR1 is involved in this function in germination under these conditions and controlling the expression of genes encoding ABA signaling proteins. Epistasis analysis demonstrated that these contrasting roles of ETR1 and ETR2 do not require the canonical ethylene signaling pathway. To explore the importance of receptor-protein interactions, we conducted yeast two-hybrid screens using the cytosolic domains of ETR1 and ETR2 as bait. Unique interacting partners with either ETR1 or ETR2 were identified. We focused on three of these proteins and confirmed the interactions with receptors. Loss of these proteins led to faster germination in response to ABA, showing that they are involved in ABA responses. Thus, ETR1 and ETR2 have both ethylene-dependent and -independent roles in plant cells that affect responses to ABA. PMID:29158332

Proteomic Analysis Reveals Different Involvement of Embryo and Endosperm Proteins during Aging of Yliangyou 2 Hybrid Rice Seeds.

PubMed

Zhang, Ying-Xue; Xu, Heng-Heng; Liu, Shu-Jun; Li, Ni; Wang, Wei-Qing; Møller, Ian M; Song, Song-Quan

2016-01-01

Seed aging is a process that results in a delayed germination, a decreased germination percentage, and finally a total loss of seed viability. However, the mechanism of seed aging is poorly understood. In the present study, Yliangyou 2 hybrid rice ( Oryza sativa L.) seeds were artificially aged at 100% relative humidity and 40°C, and the effect of artificial aging on germination, germination time course and the change in protein profiles of embryo and endosperm was studied to understand the molecular mechanism behind seed aging. With an increasing duration of artificial aging, the germination percentage and germination rate of hybrid rice seeds decreased. By comparing the protein profiles from the seeds aged for 0, 10 and 25 days, a total of 91 and 100 protein spots were found to show a significant change of more than 2-fold ( P < 0.05) in abundance, and 71 and 79 protein spots were identified, in embryos and endosperms, respectively. The great majority of these proteins increased in abundance in embryos (95%) and decreased in abundance in endosperms (99%). In embryos, most of the identified proteins were associated with energy (30%), with cell defense and rescue (28%), and with storage protein (18%). In endosperms, most of the identified proteins were involved in metabolism (37%), in energy (27%), and in protein synthesis and destination (11%). The most marked change was the increased abundance of many glycolytic enzymes together with the two fermentation enzymes pyruvate decarboxylase and alcohol dehydrogenase in the embryos during aging. We hypothesize that the decreased viability of hybrid rice seeds during artificial aging is caused by the development of hypoxic conditions in the embryos followed by ethanol accumulation.

Endotrophic Calcium, Strontium, and Barium Spores of Bacillus megaterium and Bacillus cereus1

PubMed Central

Foerster, Harold F.; Foster, J. W.

1966-01-01

Foerster, Harold F. (The University of Texas, Austin), and J. W. Foster. Endotrophic calcium, strontium, and barium spores of Bacillus megaterium and Bacillus cereus. J. Bacteriol. 91:1333–1345. 1966.—Spores were produced by washed vegetative cells suspended in deionized water supplemented with CaCl2, SrCl2, or BaCl2. Normal, refractile spores were produced in each case; a portion of the barium spores lost refractility and darkened. Thin-section electron micrographs revealed no apparent anatomical differences among the three types of spores. Analyses revealed that the different spore types were enriched specifically in the metal to which they were exposed during sporogenesis. The calcium content of the strontium and the barium spores was very small. From binary equimolar mixtures of the metal salts, endotrophic spores accumulated both metals to nearly the same extent. Viability of the barium spores was considerably less than that of the other two types. Strontium and barium spores were heat-resistant; however, calcium was essential for maximal heat resistance. Significant differences existed in the rates of germination; calcium spores germinated fastest, strontium spores were slower, and barium spores were slowest. Calcium-barium and calcium-strontium spores germinated readily. Endotrophic calcium and strontium spores germinated without the prior heat activation essential for growth spores. Chemical germination of the different metal-type spores with n-dodecylamine took place at the same relative rates as physiological germination. Heat-induced release of dipicolinic acid occurred much faster with barium and strontium spores than with calcium spores. The washed “coat fraction” from disrupted spores contained little of the spore calcium but most of the spore barium. The metal in this fraction was released by dilute acid. The demineralized coats reabsorbed calcium and barium at neutral pH. Images PMID:4956334

Effect of Nanoencapsulated Vitamin B1 Derivative on Inhibition of Both Mycelial Growth and Spore Germination of Fusarium oxysporum f. sp. raphani

PubMed Central

Cho, Jeong Sub; Seo, Yong Chang; Yim, Tae Bin; Lee, Hyeon Yong

2013-01-01

Nanoencapsulation of thiamine dilauryl sulfate (TDS), a vitamin B1 derivative, was proved to effectively inhibit the spore germination of Fusarium oxysporum f. sp. raphani (F. oxysporum), as well as mycelial growth. The average diameter of nanoparticles was measured as 136 nm by being encapsulated with an edible encapsulant, lecithin, whose encapsulation efficiency was about 55% in containing 200 ppm of TDS concentration: the 100 ppm TDS nanoparticle solution showed a mycelial growth inhibition rate of 59%. These results were about similar or even better than the cases of treating 100 ppm of dazomet, a positive antifungal control (64%). Moreover, kinetic analysis of inhibiting spore germination were estimated as 6.6% reduction of spore germination rates after 24 h treatment, which were 3.3% similar to the case of treating 100 ppm of a positive control (dazomet) for the same treatment time. It was also found that TDS itself could work as an antifungal agent by inhibiting both mycelial growth and spore germination, even though its efficacy was lower than those of nanoparticles. Nanoparticles especially played a more efficient role in limiting the spore germination, due to their easy penetration into hard cell membranes and long resident time on the surface of the spore shell walls. In this work, it was first demonstrated that the nanoparticle of TDS not a harmful chemical can control the growth of F. oxysporum by using a lower dosage than commercial herbicides, as well as the inhibiting mechanism of the TDS. However, field trials of the TDS nanoparticles encapsulated with lecithin should be further studied to be effectively used for field applications. PMID:23429270

Proteomic Analysis Reveals Different Involvement of Embryo and Endosperm Proteins during Aging of Yliangyou 2 Hybrid Rice Seeds

PubMed Central

Zhang, Ying-Xue; Xu, Heng-Heng; Liu, Shu-Jun; Li, Ni; Wang, Wei-Qing; Møller, Ian M.; Song, Song-Quan

2016-01-01

Seed aging is a process that results in a delayed germination, a decreased germination percentage, and finally a total loss of seed viability. However, the mechanism of seed aging is poorly understood. In the present study, Yliangyou 2 hybrid rice (Oryza sativa L.) seeds were artificially aged at 100% relative humidity and 40°C, and the effect of artificial aging on germination, germination time course and the change in protein profiles of embryo and endosperm was studied to understand the molecular mechanism behind seed aging. With an increasing duration of artificial aging, the germination percentage and germination rate of hybrid rice seeds decreased. By comparing the protein profiles from the seeds aged for 0, 10 and 25 days, a total of 91 and 100 protein spots were found to show a significant change of more than 2-fold (P < 0.05) in abundance, and 71 and 79 protein spots were identified, in embryos and endosperms, respectively. The great majority of these proteins increased in abundance in embryos (95%) and decreased in abundance in endosperms (99%). In embryos, most of the identified proteins were associated with energy (30%), with cell defense and rescue (28%), and with storage protein (18%). In endosperms, most of the identified proteins were involved in metabolism (37%), in energy (27%), and in protein synthesis and destination (11%). The most marked change was the increased abundance of many glycolytic enzymes together with the two fermentation enzymes pyruvate decarboxylase and alcohol dehydrogenase in the embryos during aging. We hypothesize that the decreased viability of hybrid rice seeds during artificial aging is caused by the development of hypoxic conditions in the embryos followed by ethanol accumulation. PMID:27708655

OsLOL1, a C2C2-type zinc finger protein, interacts with OsbZIP58 to promote seed germination through the modulation of gibberellin biosynthesis in Oryza sativa.

PubMed

Wu, Jiahe; Zhu, Chuanfeng; Pang, Jinhuan; Zhang, Xiangrong; Yang, Chunlin; Xia, Guixian; Tian, Yingchuan; He, Chaozu

2014-12-01

Seed germination is a key developmental process in the plant life cycle that is influenced by various environmental cues and phytohormones through gene expression and a series of metabolism pathways. In the present study, we investigated a C2C2-type finger protein, OsLOL1, which promotes gibberellin (GA) biosynthesis and affects seed germination in Oryza sativa (rice). We used OsLOL1 antisense and sense transgenic lines to explore OsLOL1 functions. Seed germination timing in antisense plants was restored to wild type when exogenous GA3 was applied. The reduced expression of the GA biosynthesis gene OsKO2 and the accumulation of ent-kaurene were observed during germination in antisense plants. Based on yeast two-hybrid and firefly luciferase complementation analyses, OsLOL1 interacted with the basic leucine zipper protein OsbZIP58. The results from electrophoretic mobility shift and dual-luciferase reporter assays showed that OsbZIP58 binds the G-box cis-element of the OsKO2 promoter and activates LUC reporter gene expression, and that interaction between OsLOL1 and OsbZIP58 activates OsKO2 gene expression. In addition, OsLOL1 decreased SOD1 gene expression and accelerated programmed cell death (PCD) in the aleurone layer of rice grains. These findings demonstrate that the interaction between OsLOL1 and OsbZIP58 influences GA biosynthesis through the activation of OsKO2 via OsbZIP58, thereby stimulating aleurone PCD and seed germination. © 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.

Role of YpeB in Cortex Hydrolysis during Germination of Bacillus anthracis Spores

PubMed Central

Bernhards, Casey B.

2014-01-01

The infectious agent of the disease anthrax is the spore of Bacillus anthracis. Bacterial spores are extremely resistant to environmental stresses, which greatly hinders spore decontamination efforts. The spore cortex, a thick layer of modified peptidoglycan, contributes to spore dormancy and resistance by maintaining the low water content of the spore core. The cortex is degraded by germination-specific lytic enzymes (GSLEs) during spore germination, rendering the cells vulnerable to common disinfection techniques. This study investigates the relationship between SleB, a GSLE in B. anthracis, and YpeB, a protein necessary for SleB stability and function. The results indicate that ΔsleB and ΔypeB spores exhibit similar germination phenotypes and that the two proteins have a strict codependency for their incorporation into the dormant spore. In the absence of its partner protein, SleB or YpeB is proteolytically degraded soon after expression during sporulation, rather than escaping the developing spore. The three PepSY domains of YpeB were examined for their roles in the interaction with SleB. YpeB truncation mutants illustrate the necessity of a region beyond the first PepSY domain for SleB stability. Furthermore, site-directed mutagenesis of highly conserved residues within the PepSY domains resulted in germination defects corresponding to reduced levels of both SleB and YpeB in the mutant spores. These results identify residues involved in the stability of both proteins and reiterate their codependent relationship. It is hoped that the study of GSLEs and interacting proteins will lead to the use of GSLEs as targets for efficient activation of spore germination and facilitation of spore cleanup. PMID:25022853

Evasion of affinity-based selection in germinal centers by Epstein-Barr virus LMP2A.

PubMed

Minamitani, Takeharu; Yasui, Teruhito; Ma, Yijie; Zhou, Hufeng; Okuzaki, Daisuke; Tsai, Chiau-Yuang; Sakakibara, Shuhei; Gewurz, Benjamin E; Kieff, Elliott; Kikutani, Hitoshi

2015-09-15

Epstein-Barr virus (EBV) infects germinal center (GC) B cells and establishes persistent infection in memory B cells. EBV-infected B cells can cause B-cell malignancies in humans with T- or natural killer-cell deficiency. We now find that EBV-encoded latent membrane protein 2A (LMP2A) mimics B-cell antigen receptor (BCR) signaling in murine GC B cells, causing altered humoral immune responses and autoimmune diseases. Investigation of the impact of LMP2A on B-cell differentiation in mice that conditionally express LMP2A in GC B cells or all B-lineage cells found LMP2A expression enhanced not only BCR signals but also plasma cell differentiation in vitro and in vivo. Conditional LMP2A expression in GC B cells resulted in preferential selection of low-affinity antibody-producing B cells despite apparently normal GC formation. GC B-cell-specific LMP2A expression led to systemic lupus erythematosus-like autoimmune phenotypes in an age-dependent manner. Epigenetic profiling of LMP2A B cells found increased H3K27ac and H3K4me1 signals at the zinc finger and bric-a-brac, tramtrack domain-containing protein 20 locus. We conclude that LMP2A reduces the stringency of GC B-cell selection and may contribute to persistent EBV infection and pathogenesis by providing GC B cells with excessive prosurvival effects.

CD45RO enriches for activated, highly mutated human germinal center B cells

PubMed Central

Jackson, Stephen M.; Harp, Natessa; Patel, Darshna; Zhang, Jeffrey; Willson, Savannah; Kim, Yoon J.; Clanton, Christian

2007-01-01

To date, there is no consensus regarding the influence of different CD45 isoforms during peripheral B-cell development. Examining correlations between surface CD45RO expression and various physiologic processes ongoing during the germinal center (GC) reaction, we hypothesized that GC B cells, like T cells, that up-regulate surface RO should progressively acquire phenotypes commonly associated with activated, differentiating lymphocytes. GC B cells (IgD−CD38+) were subdivided into 3 surface CD45RO fractions: RO−, RO+/−, and RO+. We show here that the average number of mutations per IgVH transcript increased in direct correlation with surface RO levels. Conjunctional use of RO and CD69 further delineated low/moderately and highly mutated fractions. Activation-induced cytidine deaminase (AID) mRNA was slightly reduced among RO+ GC B cells, suggesting that higher mutation averages are unlikely due to elevated somatic mutation activity. Instead, RO+ GC B cells were negative for Annexin V, comprised mostly (93%) of CD77− centrocytes, and were enriched for CD69+ cells. Collectively, RO+ GC B cells occupy what seems to be a specialized niche comprised mostly of centrocytes that may be in transition between activation states. These findings are among the first to sort GC B cells into populations enriched for live mutated cells solely using a single extracellular marker. PMID:17644737

The genomic landscape of mantle cell lymphoma is related to the epigenetically determined chromatin state of normal B cells

PubMed Central

Zhang, Jenny; Jima, Dereje; Moffitt, Andrea B.; Liu, Qingquan; Czader, Magdalena; Hsi, Eric D.; Fedoriw, Yuri; Dunphy, Cherie H.; Richards, Kristy L.; Gill, Javed I.; Sun, Zhen; Love, Cassandra; Scotland, Paula; Lock, Eric; Levy, Shawn; Hsu, David S.; Dunson, David; Dave, Sandeep S.

2014-01-01

In this study, we define the genetic landscape of mantle cell lymphoma (MCL) through exome sequencing of 56 cases of MCL. We identified recurrent mutations in ATM, CCND1, MLL2, and TP53. We further identified a number of novel genes recurrently mutated in patients with MCL including RB1, WHSC1, POT1, and SMARCA4. We noted that MCLs have a distinct mutational profile compared with lymphomas from other B-cell stages. The ENCODE project has defined the chromatin structure of many cell types. However, a similar characterization of primary human mature B cells has been lacking. We defined, for the first time, the chromatin structure of primary human naïve, germinal center, and memory B cells through chromatin immunoprecipitation and sequencing for H3K4me1, H3K4me3, H3Ac, H3K36me3, H3K27me3, and PolII. We found that somatic mutations that occur more frequently in either MCLs or Burkitt lymphomas were associated with open chromatin in their respective B cells of origin, naïve B cells, and germinal center B cells. Our work thus elucidates the landscape of gene-coding mutations in MCL and the critical interplay between epigenetic alterations associated with B-cell differentiation and the acquisition of somatic mutations in cancer. PMID:24682267

Oral lymphoepithelial cyst: A clinicopathological study of 26 cases and review of the literature.

PubMed

Sykara, Maria; Ntovas, Panagiotis; Kalogirou, Eleni-Marina; Tosios, Konstantinos I; Sklavounou, Alexandra

2017-08-01

Τo describe the clinicopathological features of 26 oral lymphoepithelial cysts (LECs) and review the literature. Twenty-six cases of oral LECs diagnosed during a 37-year period were retrospectively collected. The patients' gender and age, as well as the main clinical features of the cysts were retrieved from the requisition forms. The main microscopic features were recorded after reevaluation of all cases. Pubmed and Google Scholar electronic databases were searched with the key word "oral LEC". Inclusion criteria were the microscopic confirmation of LEC diagnosis and the report at least two of three main clinical features (gender, age and cyst's location). The 26 oral LECs represented 0.08% of 31,564 biopsies accessioned during the study period. They affected 25 patients, 14 females and 11 males with a mean age of 33.04±9.81 years. They appeared as smooth (92%) nodules, with soft (24%) or firm (76%) consistency and normal (28%), yellow to normal (20%), yellow (32%) or white (20%) hue, in the tongue (69.23%) or the floor of mouth (30.77%). They were covered by parakeratinized squamous (92.31%) or non-keratinized (7.69%) epithelium and contained desquamated epithelial cells, amorphous eosinophilic material and/or inflammatory cells (100%). The lymphoid tissue surrounded the cystic cavity partially (34.62%) or completely (65.38%), often in a follicular pattern with prominent germinal centers (53.85%). Literature review yielded 316 cases of oral LECs derived from 25 case reports, 3 case studies/retrospective studies with detailed information for each case and 7 studies with summarized data. Oral LEC is a pathologic entity with discrete clinical presentation that is, however, commonly misdiagnosed in clinical practice as other, mostly benign, entities. Its pathogenesis remains obscure, as its clinicopathologic features are consistent with both theories suggested up to date. Key words: Oral lymphoepithelial cyst; developmental cyst; non odontogenic cyst; lymphoid tissue; oral tonsil.

Oral lymphoepithelial cyst: A clinicopathological study of 26 cases and review of the literature

PubMed Central

Sykara, Maria; Ntovas, Panagiotis; Tosios, Konstantinos I.; Sklavounou, Alexandra

2017-01-01

Introduction Τo describe the clinicopathological features of 26 oral lymphoepithelial cysts (LECs) and review the literature. Material and Methods Twenty-six cases of oral LECs diagnosed during a 37-year period were retrospectively collected. The patients’ gender and age, as well as the main clinical features of the cysts were retrieved from the requisition forms. The main microscopic features were recorded after reevaluation of all cases. Pubmed and Google Scholar electronic databases were searched with the key word “oral LEC”. Inclusion criteria were the microscopic confirmation of LEC diagnosis and the report at least two of three main clinical features (gender, age and cyst’s location). Results The 26 oral LECs represented 0.08% of 31,564 biopsies accessioned during the study period. They affected 25 patients, 14 females and 11 males with a mean age of 33.04±9.81 years. They appeared as smooth (92%) nodules, with soft (24%) or firm (76%) consistency and normal (28%), yellow to normal (20%), yellow (32%) or white (20%) hue, in the tongue (69.23%) or the floor of mouth (30.77%). They were covered by parakeratinized squamous (92.31%) or non-keratinized (7.69%) epithelium and contained desquamated epithelial cells, amorphous eosinophilic material and/or inflammatory cells (100%). The lymphoid tissue surrounded the cystic cavity partially (34.62%) or completely (65.38%), often in a follicular pattern with prominent germinal centers (53.85%). Literature review yielded 316 cases of oral LECs derived from 25 case reports, 3 case studies/retrospective studies with detailed information for each case and 7 studies with summarized data. Conclusions Oral LEC is a pathologic entity with discrete clinical presentation that is, however, commonly misdiagnosed in clinical practice as other, mostly benign, entities. Its pathogenesis remains obscure, as its clinicopathologic features are consistent with both theories suggested up to date. Key words:Oral lymphoepithelial cyst; developmental cyst; non odontogenic cyst; lymphoid tissue; oral tonsil. PMID:28936296

The cell wall pectic polymer rhamnogalacturonan-II is required for proper pollen tube elongation: implications of a putative sialyltransferase-like protein.

PubMed

Dumont, Marie; Lehner, Arnaud; Bouton, Sophie; Kiefer-Meyer, Marie Christine; Voxeur, Aline; Pelloux, Jérôme; Lerouge, Patrice; Mollet, Jean-Claude

2014-10-01

Rhamnogalacturonan-II (RG-II) is one of the pectin motifs found in the cell wall of all land plants. It contains sugars such as 2-keto-3-deoxy-d-lyxo-heptulosaric acid (Dha) and 2-keto-3-deoxy-d-manno-octulosonic acid (Kdo), and within the wall RG-II is mostly found as a dimer via a borate diester cross-link. To date, little is known regarding the biosynthesis of this motif. Here, after a brief review of our current knowledge on RG-II structure, biosynthesis and function in plants, this study explores the implications of the presence of a Golgi-localized sialyltransferase-like 2 (SIA2) protein that is possibly involved in the transfer of Dha or Kdo in the RG-II of Arabidopsis thaliana pollen tubes, a fast-growing cell type used as a model for the study of cell elongation. Two heterozygous mutant lines of arabidopsis (sia2-1+/- and qrt1 × sia2-2+/-) were investigated. sia2-2+/- was in a quartet1 background and the inserted T-DNA contained the reporter gene β-glucuronidase (GUS) under the pollen-specific promoter LAT52. Pollen germination and pollen tube phenotype and growth were analysed both in vitro and in vivo by microscopy. Self-pollination of heterozygous lines produced no homozygous plants in the progeny, which may suggest that the mutation could be lethal. Heterozygous mutants displayed a much lower germination rate overall and exhibited a substantial delay in germination (20 h of delay to reach 30 % of pollen grain germination compared with the wild type). In both lines, mutant pollen grains that were able to produce a tube had tubes that were either bursting, abnormal (swollen or dichotomous branching tip) or much shorter compared with wild-type pollen tubes. In vivo, mutant pollen tubes were restricted to the style, whereas the wild-type pollen tubes were detected at the base of the ovary. This study highlights that the mutation in arabidopsis SIA2 encoding a sialyltransferase-like protein that may transfer Dha or Kdo on the RG-II motif has a dramatic effect on the stability of the pollen tube cell wall. © The Author 2014. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

The new reports on life cycle of Heterosigma akashiwo (Raphidophyceae)

NASA Astrophysics Data System (ADS)

Kim, J. H.

2016-02-01

Heterosigma akashiwo (Hada) Hada (Raphidophyceae) is a noxious bloom-forming algal species that has damaged many fish farms in coastal waters during recent decades. Consequently, many studies focused on the population dynamics of H. akashiwo, while its life cycle was not well studied. In this study, we investigated veiled life cycle of H. akashiwo through culture based method. We cultured eight H. akashiwostrains originated from Korea, Japan, USA under various conditions (water temp., light intensity, salinity, pH). Morphological diversity of cells were observed via light microscopy and scanning electron microscopy. To observe nucleus of living cell, cells were stained with Hoechst and changes of cells in culture were observed through time-lapse. For observation of cysts and their germination process, cysts were isolated from sediment. Different from the previous knowledge, H. akashiwo has only vegetative cell stage and cyst stage, we discovered that H. akashiwo has extra small cell stage and large cell stage. Large cells are much bigger (20-45 µm) than vegetative cells. Large cell formation was resulted from fusion of vegetative cells. Small cells were very small (6.88 ± 0.85 µm), these cell divided from large cell or formed in germination process of cysts rarely. Small cells have lower motility than vegetative cells. These results improved the study of life stages of H. akashiwo and this fundamental investigation provide important new information and improve our understanding of the life cycle of H. akashiwo.

Two NADPH oxidase isoforms are required for sexual reproduction and ascospore germination in the filamentous fungus Podospora anserina.

PubMed

Malagnac, Fabienne; Lalucque, Hervé; Lepère, Gersende; Silar, Philippe

2004-11-01

NADPH oxidases are enzymes that produce reactive oxygen species (ROS) using electrons derived from intracellular NADPH. In plants and mammals, ROS have been proposed to be second messengers that signal defence responses or cell proliferation. By inactivating PaNox1 and PaNox2, two genes encoding NADPH oxidases, we demonstrate the crucial role of these enzymes in the control of two key steps of the filamentous fungus Podospora anserina life cycle. PaNox1 mutants are impaired in the differentiation of fruiting bodies from their progenitor cells, and the deletion of the PaNox2 gene specifically blocks ascospore germination. Furthermore, we show that PaNox1 likely acts upstream of PaASK1, a MAPKKK previously implicated in stationary phase differentiation and cell degeneration. Using nitro blue tetrazolium (NBT) and diaminobenzidine (DAB) assays, we detect a regulated secretion of both superoxide and peroxide during P. anserina vegetative growth. In addition, two oxidative bursts are shown to occur during fruiting body development and ascospore germination. Analysis of mutants establishes that PaNox1, PaNox2, and PaASK1, as well as a still unknown additional source of ROS, modulate these secretions. Altogether, our data point toward a role for NADPH oxidases in signalling fungal developmental transitions with respect to nutrient availability. These enzymes are conserved in other multicellular eukaryotes, suggesting that early eukaryotes were endowed with a redox network used for signalling purposes.

Nuclear dynamics during germination, conidiation, and hyphal fusion of Fusarium oxysporum.

PubMed

Ruiz-Roldán, M Carmen; Köhli, Michael; Roncero, M Isabel G; Philippsen, Peter; Di Pietro, Antonio; Espeso, Eduardo A

2010-08-01

In many fungal pathogens, infection is initiated by conidial germination. Subsequent stages involve germ tube elongation, conidiation, and vegetative hyphal fusion (anastomosis). Here, we used live-cell fluorescence to study the dynamics of green fluorescent protein (GFP)- and cherry fluorescent protein (ChFP)-labeled nuclei in the plant pathogen Fusarium oxysporum. Hyphae of F. oxysporum have uninucleated cells and exhibit an acropetal nuclear pedigree, where only the nucleus in the apical compartment is mitotically active. In contrast, conidiation follows a basopetal pattern, whereby mononucleated microconidia are generated by repeated mitotic cycles of the subapical nucleus in the phialide, followed by septation and cell abscission. Vegetative hyphal fusion is preceded by directed growth of the fusion hypha toward the receptor hypha and followed by a series of postfusion nuclear events, including mitosis of the apical nucleus of the fusion hypha, migration of a daughter nucleus into the receptor hypha, and degradation of the resident nucleus. These previously unreported patterns of nuclear dynamics in F. oxysporum could be intimately related to its pathogenic lifestyle.

Differentiation of Dictyostelium discoideum vegetative cells into spores during earth orbit in space

NASA Astrophysics Data System (ADS)

Takahashi, A.; Ohnishi, K.; Takahashi, S.; Masukawa, M.; Sekikawa, K.; Amano, T.; Nakano, T.; Nagaoka, S.; Ohnishi, T.

2001-01-01

We reported previously that emerged amoebae of Dictyosterium ( D.) discoideum grew, aggregated and differentiated to fruiting bodies with normal morphology in space. Here, we investigated the effects of space radiation and/or microgravity on the number, viability, kinetics of germination, growth rate and mutation frequency of spores formed in space in a radiation-sensitive strain, γs13, and the parental strain, NC4. In γs13, there were hardly spores in the fruiting bodies formed in space. In NC4, we found a decrease in the number of spores, a delay in germination of the spores and delayed start of cell growth of the spores formed in space when compared to the ground control. However, the mutation frequency of the NC4 spores formed in space was similar to that of the ground control. We conclude that the depression of spore formation might be induced by microgravity and/or space radiation through the depression of some stage(s) of DNA repair during cell differentiation in the slime mold.

Increased T follicular helper cells and germinal center B cells are required for cGVHD and bronchiolitis obliterans

PubMed Central

Flynn, Ryan; Du, Jing; Veenstra, Rachelle G.; Reichenbach, Dawn K.; Panoskaltsis-Mortari, Angela; Taylor, Patricia A.; Freeman, Gordon J.; Serody, Jonathan S.; Murphy, William J.; Munn, David H.; Sarantopoulos, Stefanie; Luznik, Leo; Maillard, Ivan; Koreth, John; Cutler, Corey; Soiffer, Robert J.; Antin, Joseph H.; Ritz, Jerome; Dubovsky, Jason A.; Byrd, John C.; MacDonald, Kelli P.; Hill, Geoff R.; Blazar, Bruce R.

2014-01-01

Chronic graft-versus-host disease (cGVHD) is a leading cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation. Having shown that germinal center (GC) formation and immunoglobulin deposition are required for multiorgan system cGVHD and associated bronchiolitis obliterans syndrome (BOS) in a murine model, we hypothesized that T follicular helper (Tfh) cells are necessary for cGVHD by supporting GC formation and maintenance. We show that increased frequency of Tfh cells correlated with increased GC B cells, cGVHD, and BOS. Although administering a highly depletionary anti-CD20 monoclonal antibody (mAb) to mice with established cGVHD resulted in peripheral B-cell depletion, B cells remained in the lung, and BOS was not reversed. BOS could be treated by eliminating production of interleukin-21 (IL-21) by donor T cells or IL-21 receptor (IL-21R) signaling of donor B cells. Development of BOS was dependent upon T cells expressing the chemokine receptor CXCR5 to facilitate T-cell trafficking to secondary lymphoid organ follicles. Blocking mAbs for IL-21/IL-21R, inducible T-cell costimulator (ICOS)/ICOS ligand, and CD40L/CD40 hindered GC formation and cGVHD. These data provide novel insights into cGVHD pathogenesis, indicate a role for Tfh cells in these processes, and suggest a new line of therapy using mAbs targeting Tfh cells to reverse cGVHD. PMID:24820310

Mutant IDH1 Disrupts the Mouse Subventricular Zone and Alters Brain Tumor Progression

PubMed Central

Pirozzi, Christopher J.; Carpenter, Austin B.; Waitkus, Matthew S.; Wang, Catherine Y.; Zhu, Huishan; Hansen, Landon J.; Chen, Lee H.; Greer, Paula K.; Feng, Jie; Wang, Yu; Bock, Cheryl B.; Fan, Ping; Spasojevic, Ivan; McLendon, Roger E.; Bigner, Darell D.; He, Yiping; Yan, Hai

2017-01-01

IDH1 mutations occur in the majority of low-grade gliomas and lead to the production of the oncometabolite, D-2-hydroxyglutarate (D-2HG). To understand the effects of tumor-associated mutant IDH1 (IDH1-R132H) on both the neural stem cell (NSC) population and brain tumorigenesis, genetically faithful cell lines and mouse model systems were generated. Here, it is reported that mouse NSCs expressing Idh1-R132H displayed reduced proliferation due to p53-mediated cell cycle arrest as well as a decreased ability to undergo neuronal differentiation. In vivo, Idh1-R132H expression reduced proliferation of cells within the germinal zone of the subventricular zone (SVZ). The NSCs within this area were dispersed and disorganized in mutant animals, suggesting that Idh1-R132H perturbed the NSCs and the microenvironment from which gliomas arise. Additionally, tumor-bearing animals expressing mutant Idh1 displayed a prolonged survival and also overexpressed Olig2, features consistent with IDH1-mutated human gliomas. These data indicate that mutant Idh1 disrupts the NSC microenvironment and the candidate cell of origin for glioma; thus, altering the progression of tumorigenesis. Additionally, this study provides a mutant Idh1 brain tumor model that genetically recapitulates human disease, laying the foundation for future investigations on mutant IDH1-mediated brain tumorigenesis and targeted therapy. PMID:28148827

Respiratory metabolism in the embryonic axis of germinating pea seed exposed to cadmium.

PubMed

Smiri, Moêz; Chaoui, Abdelilah; El Ferjani, Ezzedine

2009-02-15

Seeds of pea (Pisum sativum L.) were germinated for 5d by soaking in distilled water or 5mM cadmium nitrate. The relationships among cadmium stress, germination rate, changes in respiratory enzyme activities and carbohydrates mobilization were studied. Two cell fractions were obtained from embryonic axis: (1) mitochondria, used to determine enzyme activities of citric acid cycle and electron transport chain, and (2) soluble, to measure some enzyme activities involved in fermentation and pentose phosphate pathway. Activities of malate- and succinate-dehydrogenases (MDH, SDH) and NADH- and succinate-cytochrome c reductases (NCCR, SCCR) were rapidly inhibited, while cytochrome c oxidase (CCO) was unaltered by cadmium treatment. However, this stimulated the NADPH-generating enzyme activities of the pentose phosphate pathway, glucose-6-phosphate- and 6-phosphogluconate-dehydrogenases (G6PDH, 6PGDH), as well as enzyme activity of fermentation, alcohol dehydrogenase (ADH), with concomitant inhibition in the capacity of enzyme inactivator (INADH). Moreover, Cd restricted carbohydrate mobilization in the embryonic axis. Almost no glucose and less than 7% of control fructose and total soluble sugars were available in the embryo tissues after 5d of exposure to cadmium. Cotyledonary invertase isoenzyme activity was also inhibited by Cd. The results indicate that cadmium induces disorder in the resumption of respiration in germinating pea seeds. The contribution of Cd-stimulated alternative metabolic pathways to compensate for the failure in mitochondrial respiration is discussed in relation to the delay in seed germination and embryonic axis growth.

Content of Adenosine Phosphates and Adenylate Energy Charge in Germinating Ponderosa Pine Seeds

PubMed Central

Ching, Te May; Ching, Kim K.

1972-01-01

An average of 540 picomoles of total adenosine phosphates was found in the embryo of mature seeds of ponderosa pine (Pinus ponderosa Laws.) and 1140 picomoles in the gametophyte. Adenylate energy charges were 0.44 and 0.26, respectively. After stratification, total adenosine phosphates increased 7-fold and 6-fold in embryo and gametophyte, respectively, and energy charges rose to 0.85 and 0.75. During germination, total adenosine phosphates increased to a 20-fold peak on the 9th day in gametophytic tissue, parallel with the peak of reserve regradation and organellar synthesis, and then decreased. In embryo and seedling, total adenosine phosphates elevated 80-fold with two distinct oscillating increases of AMP and ADP. The oscillating increases occurred before the emergence of radicle and cotyledons during which the highest mitotic index prevailed in all tissues. Energy charges fluctuated between 0.65 at the rapid cell dividing stage to 0.85 at the fully differentiated stage of the seedling, while energy charges remained around 0.75 in the gametophyte. These data indicated that the content of adenosine phosphates of germinating seeds reflects growth, organogenesis, and morphogenesis, and that a compartmentalized energy metabolism must exist in dividing and growing plant cells. PMID:16658212

Cell hydration as a biomarker for estimation of biological effects of nonionizing radiation on cells and organisms.

PubMed

Ayrapetyan, Sinerik; De, Jaysankar

2014-01-01

"Changes in cell hydration" have been hypothesized as an input signal for intracellular metabolic cascade responsible for biological effects of nonionizing radiation (NIR). To test this hypothesis a comparative study on the impacts of different temperature and NIR (infrasound frequency mechanical vibration (MV), static magnetic field (SMF), extremely low frequency electromagnetic field (ELF EMF), and microwave (MW)) pretreated water on the hydration of barley seeds in its dormant and germination periods was performed. In dormant state temperature sensitivity (Q 10) of seed hydration in distilled water (DW) was less than 2, and it was nonsensitive to NIR treated DW, whereas during the germination period (48-72 hours) seeds hydration exhibited temperature sensitivity Q 10 > 2 and higher sensitivity to NIR treated DW. Obtained data allow us to suggest that the metabolic driving of intracellular water dynamics accompanied by hydrogen bonding and breaking is more sensitive to NIR-induced water structure changes in seed bathing aqua medium than the simple thermodynamic processes such as osmotic gradient driven water absorption by seeds in dormant state. Therefore, cell hydration is suggested to be a universal and extrasensitive biomarker for detection of biological effects of NIR on cells and organisms.

Distinct Differentiation Programs Triggered by IL-6 and LPS in Teleost IgM(+) B Cells in The Absence of Germinal Centers.

PubMed

Abós, Beatriz; Wang, Tiehui; Castro, Rosario; Granja, Aitor G; Leal, Esther; Havixbeck, Jeffrey; Luque, Alfonso; Barreda, Daniel R; Secombes, Chris J; Tafalla, Carolina

2016-08-02

Although originally identified as a B cell differentiation factor, it is now known that mammalian interleukin-6 (IL-6) only regulates B cells committed to plasma cells in response to T-dependent (TD) antigens within germinal centers (GCs). Even though adaptive immunity is present in teleost fish, these species lack lymph nodes and GCs. Thus, the aim of the present study was to establish the role of trout IL-6 on B cells, comparing its effects to those induced by bacterial lipopolysaccharide (LPS). We demonstrate that the effects of teleost IL-6 on naïve spleen B cells include proliferation, activation of NF-κB, increased IgM secretion, up-regulation of Blimp1 transcription and decreased MHC-II surface expression that point to trout IL-6 as a differentiation factor for IgM antibody-secreting cells (ASCs). However, LPS induced the secretion of IgM without up-regulating Blimp1, driving the cells towards an intermediate activation state in which antigen presenting mechanisms are elicited together with antibody secretion and expression of pro-inflammatory genes. Our results reveal that, in trout, IL-6 is a differentiation factor for B cells, stimulating IgM responses in the absence of follicular structures, and suggest that it was after follicular structures appeared that this cytokine evolved to modulate TD responses within the GC.

HIV and T follicular helper cells: a dangerous relationship

PubMed Central

Vinuesa, Carola G.

2012-01-01

HIV infection leads to progressive destruction of infected CD4 T cells, hypergammaglobulinemia, and loss of memory B cells. Germinal centers, which are key to memory B cell formation and protective antibody responses, are major HIV reservoirs in which the virus replicates within T follicular helper (TFH) cells. In this issue of the JCI, the Koup and Streeck groups report that chronic SIV/HIV infection promotes TFH cell accumulation, which may drive B cell dysregulation. Their discoveries suggest that HIV harnesses TFH cells to evade the antibody response. PMID:22922252

In vitro and in vivo analyses of the Bacillus anthracis spore cortex lytic protein SleL

PubMed Central

Lambert, Emily A.; Sherry, Nora

2012-01-01

The bacterial endospore is the most resilient biological structure known. Multiple protective integument layers shield the spore core and promote spore dehydration and dormancy. Dormancy is broken when a spore germinates and becomes a metabolically active vegetative cell. Germination requires the breakdown of a modified layer of peptidoglycan (PG) known as the spore cortex. This study reports in vitro and in vivo analyses of the Bacillus anthracis SleL protein. SleL is a spore cortex lytic enzyme composed of three conserved domains: two N-terminal LysM domains and a C-terminal glycosyl hydrolase family 18 domain. Derivatives of SleL containing both, one or no LysM domains were purified and characterized. SleL is incapable of digesting intact cortical PG of either decoated spores or purified spore sacculi. However, SleL derivatives can hydrolyse fragmented PG substrates containing muramic-δ-lactam recognition determinants. The muropeptides that result from SleL hydrolysis are the products of N-acetylglucosaminidase activity. These muropeptide products are small and readily released from the cortex matrix. Loss of the LysM domain(s) decreases both PG binding and hydrolysis activity but these domains do not appear to determine specificity for muramic-δ-lactam. When the SleL derivatives are expressed in vivo, those proteins lacking one or both LysM domains do not associate with the spore. Instead, these proteins remain in the mother cell and are apparently degraded. SleL with both LysM domains localizes to the coat or cortex of the endospore. The information revealed by elucidating the role of SleL and its domains in B. anthracis sporulation and germination is important in designing new spore decontamination methods. By exploiting germination-specific lytic enzymes, eradication techniques may be greatly simplified. PMID:22343356

The Bacillus subtilis yaaH Gene Is Transcribed by SigE RNA Polymerase during Sporulation, and Its Product Is Involved in Germination of Spores

PubMed Central

Kodama, Takeko; Takamatsu, Hiromu; Asai, Kei; Kobayashi, Kazuo; Ogasawara, Naotake; Watabe, Kazuhito

1999-01-01

The expression of 21 novel genes located in the region from dnaA to abrB of the Bacillus subtilis chromosome was analyzed. One of the genes, yaaH, had a predicted promoter sequence conserved among SigE-dependent genes. Northern blot analysis revealed that yaaH mRNA was first detected from 2 h after the cessation of logarithmic growth (T2) of sporulation in wild-type cells and in spoIIIG (SigG−) and spoIVCB (SigK−) mutants but not in spoIIAC (SigF−) and spoIIGAB (SigE−) mutants. The transcription start point was determined by primer extension analysis; the −10 and −35 regions are very similar to the consensus sequences recognized by SigE-containing RNA polymerase. A YaaH-His tag fusion encoded by a plasmid with a predicted promoter for the yaaH gene was produced from T2 of sporulation in a B. subtilis transformant and extracted from mature spores, indicating that the yaaH gene product is a spore protein. Inactivation of the yaaH gene by insertion of an erythromycin resistance gene did not affect vegetative growth or spore resistance to heat, chloroform, and lysozyme. The germination of yaaH mutant spores in a mixture of l-asparagine, d-glucose, d-fructose, and potassium chloride was almost the same as that of wild-type spores, but the mutant spores were defective in l-alanine-stimulated germination. These results suggest that yaaH is a novel gene encoding a spore protein produced in the mother cell compartment from T2 of sporulation and that it is required for the l-alanine-stimulated germination pathway. PMID:10419957

The potential for dispersal of microalgal resting cysts by migratory birds.

PubMed

Tesson, Sylvie Vm; Weißbach, Astrid; Kremp, Anke; Lindström, Åke; Rengefors, Karin

2018-06-11

Most microalgal species are geographically widespread, but little is known about how they are dispersed. One potential mechanism for long-distance dispersal is through birds, which may transport cells internally (endozoochory) and deposit them during, or in-between, their migratory stopovers. We hypothesize that dinoflagellates, in particular resting stages, can tolerate bird digestion; that bird temperature, acidity, and retention time negatively affect dinoflagellate viability; and that recovered cysts can germinate after passage through the birds' gut, contributing to species-specific dispersal of the dinoflagellates across scales. Tolerance of two dinoflagellate species (Peridiniopsis borgei, a warm-water species and Apocalathium malmogiense, a cold-water species) to Mallard gut passage was investigated using in vitro experiments simulating the gizzard and caeca conditions. The effect of in vitro digestion and retention time on cell integrity, cell viability and germination capacity of the dinoflagellate species was examined targeting both their vegetative and resting stages. Resting stages (cysts) of both species were able to survive simulated bird gut passage, even if their survival rate and germination were negatively affected by exposure to acidic condition and bird internal temperature. Cysts of A. malmogiense were more sensitive than P. borgei to treatments and to the presence of digestive enzymes. Vegetative cells did not survive conditions of bird internal temperature and formed pellicle cysts when exposed to gizzard-like acid conditions. We show that dinoflagellate resting cysts serve as dispersal propagules through migratory birds. Assuming a retention time of viable cysts of 2-12 h to duck stomach conditions, cysts could be dispersed 150-800 km and beyond. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Monitoring of Commitment, Blocking, and Continuation of Nutrient Germination of Individual Bacillus subtilis Spores

PubMed Central

Zhang, Pengfei; Liang, Jintao; Yi, Xuan; Setlow, Peter

2014-01-01

Short exposures of Bacillus spores to nutrient germinants can commit spores to germinate when germinants are removed or their binding to the spores' nutrient germinant receptors (GRs) is inhibited. Bacillus subtilis spores were exposed to germinants for various periods, followed by germinant removal to prevent further commitment. Release of spore dipicolinic acid (DPA) was then measured by differential interference contrast microscopy to monitor germination of multiple individual spores, and spores did not release DPA after 1 to 2 min of germinant exposure until ∼7 min after germinant removal. With longer germinant exposures, percentages of committed spores with times for completion of DPA release (Trelease) greater than the time of germinant removal (Tb) increased, while the time Tlag − Tb, where Tlag represents the time when rapid DPA release began, was decreased but rapid DPA release times (ΔTrelease = Trelease − Tlag) were increased; Factors affecting average Trelease values and the percentages of committed spores were germinant exposure time, germinant concentration, sporulation conditions, and spore heat activation, as previously shown for commitment of spore populations. Surprisingly, germination of spores given a 2nd short germinant exposure 30 to 45 min after a 1st exposure of the same duration was significantly higher than after the 1st exposure, but the number of spores that germinated in the 2nd germinant exposure decreased as the interval between germinant exposures increased up to 12 h. The latter results indicate that spores have some memory, albeit transient, of their previous exposure to nutrient germinants. PMID:24769693

Tolerance of cultivated and wild plants of different taxonomy to soil contamination by kerosene.

PubMed

Sharonova, Natalia; Breus, Irina

2012-05-01

In laboratory experiments on leached chernozem contaminated by kerosene (1-15 wt.%), germination of 50 plants from 21 families (cultivated and wild, annual and perennial, mono- and dicotyledonous) as affected by kerosene type and concentration and plant features was determined. Tested plants formed three groups: more tolerant, less tolerant, and intolerant, in which relative germination was more than 70%, 30-70% and less than 30%, respectively. As parameters of soil phytotoxicity, effective kerosene concentrations (EC) causing germination depression of 10%, 25% and 50% were determined. EC values depended on the plant species and varied in a wide range of kerosene concentrations: 0.02-7.3% (EC(10)), 0.05-8.1% (EC(25)), and 0.2-12.7% (EC(50)). The reported data on germination in soils contaminated by oil and petrochemicals were generalized. The comparison showed that at very high contamination levels (10 and 15%) kerosene was 1.3-1.6 times more phytotoxic than diesel fuel and 1.3-1.4 times more toxic than crude oil, and at low (1 and 2%) and medium (3 and 5%) levels the toxicity of these contaminants was close differing by a factor of 1.1-1.2. Tolerance of plants to soil contamination had a species-specific nature and, on the average, decreased in the following range of families: Fabaceae (germination decrease of 10-60% as compared to an uncontaminated control)>Brassicaceae (5-70%)>Asteraceae (25-95%)>Poaceae (10-100%). The monocotyledonous species tested were characterized as medium- and low-stable to contamination, whereas representatives of dicotyledonous plants were met in all groups of tolerance. Tested wild plants, contrary to reference data on oil toxicity, were more sensitive to kerosene than cultivated. No correlation was observed between degree of plant tolerance to kerosene and mass of seeds. The evidence indicates factors as structure and properties of testa, structure of germ, type of storage compounds, and type of seed germination (underground or aboveground) are more important. Copyright © 2012 Elsevier B.V. All rights reserved.

Similar prognosis of transformed and de novo diffuse large B-cell lymphomas in patients treated with immunochemotherapy.

PubMed

Sorigue, Marc; Garcia, Olga; Baptista, Maria Joao; Sancho, Juan-Manuel; Tapia, Gustavo; Mate, José Luis; Feliu, Evarist; Navarro, José-Tomás; Ribera, Josep-Maria

2017-03-22

The prognosis of diffuse large B-cell lymphomas (DLBCL) transformed from indolent lymphoma (TL) has been considered poorer than that of de novo DLBCL. However, it seems to have improved since the introduction of rituximab. We compared the characteristics (including the cell-of-origin), and the prognosis of 29 patients with TL and 101 with de novo DLBCL treated with immunochemotherapy. Patients with TL and de novo DLBCL had similar characteristics. All TL cases evolving from follicular lymphoma were germinal-center B-cell-like, while those TL from marginal zone lymphoma or chronic lymphocytic leukemia were non-germinal-center B-cell-like. The complete response rate was similar in TL and de novo DLBCL (62 vs. 66%, P=.825). The 5-year overall and progression-free survival probabilities (95% CI) were 59% (40-78) and 41% (22-60) for TL and 63% (53-73) and 60% (50-70) for de novo DLBCL, respectively (P=.732 for overall survival and P=.169 for progression-free survival). In this study, the prognosis of TL and de novo DLBCL treated with immunochemotherapy was similar. The role of intensification with stem cell transplantation in the management of TL may be questionable in the rituximab era. Copyright © 2016 Elsevier España, S.L.U. All rights reserved.

The Role of B Cells and Humoral Immunity in Mycobacterium tuberculosis Infection

PubMed Central

Kozakiewicz, Lee; Phuah, Jiayao; Flynn, JoAnne

2014-01-01

Tuberculosis (TB) remains a serious threat to public health, causing 2 million deaths annually world-wide. The control of TB has been hindered by the requirement of long duration of treatment involving multiple chemotherapeutic agents, the increased susceptibility to Mycobacterium tuberculosis infection in the HIV-infected population, and the development of multi-drug resistant and extensively resistant strains of tubercle bacilli. An efficacious and cost-efficient way to control TB is the development of effective anti-TB vaccines. This measure requires thorough understanding of the immune response to M. tuberculosis. While the role of cell-mediated immunity in the development of protective immune response to the tubercle bacillus has been well established, the role of B cells in this process is not clearly understood. Emerging evidence suggests that B cells and humoral immunity can modulate the immune response to various intracellular pathogens, including M. tuberculosis. These lymphocytes form conspicuous aggregates in the lungs of tuberculous humans, non-human primates, and mice, which display features of germinal center B cells. In murine TB, it has been shown that B cells can regulate the level of granulomatous reaction, cytokine production, and the T cell response. This chapter discusses the potential mechanisms by which specific functions of B cells and humoral immunity can shape the immune response to intracellular pathogens in general, and to M. tuberculosis in particular. Knowledge of the B cell-mediated immune response to M. tuberculosis may lead to the design of novel strategies, including the development of effective vaccines, to better control TB. PMID:23468112

Highly Resolved Intravital Striped-illumination Microscopy of Germinal Centers

PubMed Central

Andresen, Volker; Sporbert, Anje

2014-01-01

Monitoring cellular communication by intravital deep-tissue multi-photon microscopy is the key for understanding the fate of immune cells within thick tissue samples and organs in health and disease. By controlling the scanning pattern in multi-photon microscopy and applying appropriate numerical algorithms, we developed a striped-illumination approach, which enabled us to achieve 3-fold better axial resolution and improved signal-to-noise ratio, i.e. contrast, in more than 100 µm tissue depth within highly scattering tissue of lymphoid organs as compared to standard multi-photon microscopy. The acquisition speed as well as photobleaching and photodamage effects were similar to standard photo-multiplier-based technique, whereas the imaging depth was slightly lower due to the use of field detectors. By using the striped-illumination approach, we are able to observe the dynamics of immune complex deposits on secondary follicular dendritic cells – on the level of a few protein molecules in germinal centers. PMID:24748007

Porphyra yezoensis bei Helgoland — eine entwicklungsgeschichtliche Studie

NASA Astrophysics Data System (ADS)

Kornmann, P.

1986-09-01

A Porphyropsis-like epiphytic specimen found in the harbour of Helgoland was grown in culture and proved to be identical with the Japanese Porphyra yezoensis. Life history studies on this economically important alga resulted in some interesting and hitherto unknown details. The variability of the adult frond is fundamentally determined by the pattern of spore germination. Settled on Chaetomorpha filaments, monospores elongate within 20 minutes; the epiphytic germlings are attached to the substrate by a typical basal cell and give rise exclusively to elongated fronds provided with a cuneate base. Unattached spores, however, germinate into buds with rhizoids; they develop into elongated elliptical to oval fronds provided with round or cordate bases. Only plants with male areas were observed in the cultures, but Conchocelis was abundantly produced from cells of aged thalli. Grown in mussel-shells, the filamentous phase liberated conchospores for a long time.

Ripple/Carcinoid pattern sebaceoma with apocrine differentiation.

PubMed

Misago, Noriyuki; Narisawa, Yutaka

2011-02-01

Sebaceoma is a benign sebaceous neoplasm, which has been reported to show characteristic growth patterns, such as, ripple, labyrinthine/sinusoidal, and carcinoid-like patterns. Another recent finding regarding in sebaceoma is the observation of apocrine differentiation within the sebaceoma lesion. This report describes a case of carcinoid (a partial ripple and labyrinthine) pattern sebaceoma with apocrine differentiation with a literature review and immunohistochemical studies. The various characteristic growth patterns in sebaceoma were suggested to simply be variations of the same growth pattern arranged in cords, namely, a unified term "ripple/carcinoid pattern." The primitive sebaceous germinative cells in sebaceoma may still have the ability to undergo apocrine differentiation. Most of the reports so far on sebaceoma with apocrine differentiation, including the present case, describe a ripple/carcinoid pattern, thus suggesting that ripple/carcinoid pattern sebaceoma is composed of more primitive sebaceous germinative cells than conventional sebaceoma.

The Molecular Timeline of a Reviving Bacterial Spore

PubMed Central

Sinai, Lior; Rosenberg, Alex; Smith, Yoav; Segev, Einat; Ben-Yehuda, Sigal

2015-01-01

Summary The bacterial spore can rapidly convert from a dormant to a fully active cell. Here we study this remarkable cellular transition in Bacillus subtilis and reveal the identity of the newly synthesized proteins throughout spore revival. Our analysis uncovers a highly ordered developmental program that correlates with the spore morphological changes and reveals the spatial and temporal molecular events fundamental to reconstruct a cell. As opposed to current knowledge, we found that translation takes place during the earliest revival event, termed germination, a process hitherto considered to occur without the need for any macromolecule synthesis. Furthermore, we demonstrate that translation is required for execution of germination and relies on the bona fide translational factors RpmE and Tig. Our study sheds light on the spore revival process and on the vital building blocks underlying cellular awakening, thereby paving the way for designing new antimicrobial agents to eradicate spore-forming pathogens. PMID:25661487

DC-SIGN–expressing macrophages trigger activation of mannosylated IgM B-cell receptor in follicular lymphoma

PubMed Central

Amin, Rada; Mourcin, Frédéric; Uhel, Fabrice; Pangault, Céline; Ruminy, Philippe; Dupré, Loic; Guirriec, Marion; Marchand, Tony; Fest, Thierry; Lamy, Thierry

2015-01-01

Follicular lymphoma (FL) results from the accumulation of malignant germinal center (GC) B cells leading to the development of an indolent and largely incurable disease. FL cells remain highly dependent on B-cell receptor (BCR) signaling and on a specific cell microenvironment, including T cells, macrophages, and stromal cells. Importantly, FL BCR is characterized by a selective pressure to retain surface immunoglobulin M (IgM) BCR despite an active class-switch recombination process, and by the introduction, in BCR variable regions, of N-glycosylation acceptor sites harboring unusual high-mannose oligosaccharides. However, the relevance of these 2 FL BCR features for lymphomagenesis remains unclear. In this study, we demonstrated that IgM+ FL B cells activated a stronger BCR signaling network than IgG+ FL B cells and normal GC B cells. BCR expression level and phosphatase activity could both contribute to such heterogeneity. Moreover, we underlined that a subset of IgM+ FL samples, displaying highly mannosylated BCR, efficiently bound dendritic cell–specific intercellular adhesion molecule-3–grabbing nonintegrin (DC-SIGN), which could in turn trigger delayed but long-lasting BCR aggregation and activation. Interestingly, DC-SIGN was found within the FL cell niche in situ. Finally, M2 macrophages induced a DC-SIGN–dependent adhesion of highly mannosylated IgM+ FL B cells and triggered BCR-associated kinase activation. Interestingly, pharmacologic BCR inhibitors abolished such crosstalk between macrophages and FL B cells. Altogether, our data support an important role for DC-SIGN–expressing infiltrating cells in the biology of FL and suggest that they could represent interesting therapeutic targets. PMID:26272216

Spatial distribution of epigenetic modifications in Brachypodium distachyon embryos during seed maturation and germination.

PubMed

Wolny, Elzbieta; Braszewska-Zalewska, Agnieszka; Hasterok, Robert

2014-01-01

Seed development involves a plethora of spatially and temporally synchronised genetic and epigenetic processes. Although it has been shown that epigenetic mechanisms, such as DNA methylation and chromatin remodelling, act on a large number of genes during seed development and germination, to date the global levels of histone modifications have not been studied in a tissue-specific manner in plant embryos. In this study we analysed the distribution of three epigenetic markers, i.e. H4K5ac, H3K4me2 and H3K4me1 in 'matured', 'dry' and 'germinating' embryos of a model grass, Brachypodium distachyon (Brachypodium). Our results indicate that the abundance of these modifications differs considerably in various organs and tissues of the three types of Brachypodium embryos. Embryos from matured seeds were characterised by the highest level of H4K5ac in RAM and epithelial cells of the scutellum, whereas this modification was not observed in the coleorhiza. In this type of embryos H3K4me2 was most evident in epithelial cells of the scutellum. In 'dry' embryos H4K5ac was highest in the coleorhiza but was not present in the nuclei of the scutellum. H3K4me1 was the most elevated in the coleoptile but absent from the coleorhiza, whereas H3K4me2 was the most prominent in leaf primordia and RAM. In embryos from germinating seeds H4K5ac was the most evident in the scutellum but not present in the coleoptile, similarly H3K4me1 was the highest in the scutellum and very low in the coleoptile, while the highest level of H3K4me2 was observed in the coleoptile and the lowest in the coleorhiza. The distinct patterns of epigenetic modifications that were observed may be involved in the switch of the gene expression profiles in specific organs of the developing embryo and may be linked with the physiological changes that accompany seed desiccation, imbibition and germination.

[Study on method and its optimization of improving seed germination of Astragalus membranaceus as gansu traditional medicinal herb].

PubMed

Shi, Li-Ping; Ou, Qiao-Ming; Cui, Wen-Juan; Chen, Yu-Liang

2014-04-01

To break the hard testa and improve seed germination situation of Astragalus membranaceus var. mongholicus, in order to solve the problems of low success rate of seed germination and seedling. Longxi Astragalus membranaceus var. mongholicus seed was treated by soaking seed with 75% alcohol and concentrated sulfuric acid, warm-water incubating, grinding and comprehensive treating with warm-water incubating, grinding and sand culture. Its seed germination situation was evaluated by germination potential, germination rate and germination index. Different processing methods significantly improved seed germination with different effect. Comprehensive treatment with warm-water incubating, grinding and sand culture was the best one on Astragalus membranaceus var. mongholicus seed germination. Its germination potential, germination rate and germination index was 66.04%, 87.70% and 1.34,respectively. Comprehensive treatment with warm-water incubating, grinding and sand culture is an economic and effective processing method, which is suitable for actual production.

Membrane permeability and the loss of germination factor from Neurospora crassa at low water activities

NASA Technical Reports Server (NTRS)

Charlang, G.; Horowitz, N. H.

1974-01-01

Neurospora crassa conidia incubating in buffer at low water activities release a germination-essential component as well as 260-nm absorbing and ninhydrin-positive materials, regardless of whether an electrolyte or nonelectrolyte is used to reduce water activity. Chloroform and antibiotics known to increase cell-membrane permeability have a similar effect. This suggests that membrane damage occurs in media of low water activity and that an increase in permeability is responsible for the release of cellular components. The damage caused in media of low water activity is nonlethal in most cases, and the conidia recover when transferred to nutrient medium.

Invariant natural killer T cells ameliorate murine chronic GVHD by expanding donor regulatory T cells.

PubMed

Du, Jing; Paz, Katelyn; Thangavelu, Govindarajan; Schneidawind, Dominik; Baker, Jeanette; Flynn, Ryan; Duramad, Omar; Feser, Colby; Panoskaltsis-Mortari, Angela; Negrin, Robert S; Blazar, Bruce R

2017-06-08

Chronic graft-versus-host-disease (cGVHD) can cause multiorgan system disease, typically with autoimmune-like features, resulting in high mortality and morbidity caused by treatment limitations. Invariant natural killer T cells (iNKTs), a small population characterized by expression of a semi-invariant T-cell receptor, rapidly produce copious amounts of diverse cytokines on activation that exert potent immune regulatory function. Here, we show that iNKTs are significantly reduced in a cGVHD murine model that recapitulates several aspects of autoimmunity and organ fibrosis observed in patients with cGVHD. Low iNKT infused doses effectively prevented and, importantly, reversed established cGVHD, as did third-party iNKTs. iNKTs suppressed the autoimmune response by reducing the germinal center (GC) reaction, which was associated with an increase in total Tregs and follicular Tregs (Tfr) that control the GC reaction, along with pathogenic antibody production. Treg depletion during iNKT infusions completely abolished iNKT efficacy in treating cGVHD. iNKT cell interleukin 4 production and GC migration were critical to cGVHD reversal. In vivo stimulation of iNKT cells by α-galactosyl-ceramide was effective in both preventing and treating cGVHD. Together, this study demonstrates iNKT deficiency in cGVHD mice and highlights the key role of iNKTs in regulating cGVHD pathogenesis and as a potentially novel prophylactic and therapeutic option for patients with cGVHD. © 2017 by The American Society of Hematology.

Chitosan inhibits enterotoxigenic Clostridium perfringens type A in growth medium and chicken meat.

PubMed

Alnoman, Maryam; Udompijitkul, Pathima; Sarker, Mahfuzur R

2017-06-01

Clostridium perfringens is a spore-forming bacterium and a major cause of bacterial food-borne illness. In this study, we evaluated the inhibitory effects of chitosan against spore germination, spore outgrowth and vegetative growth of C. perfringens food poisoning (FP) isolates. Chitosan of differing molecular weights inhibited germination of spores of all tested FP isolates in a KCl germinant solution containing 0.1 mg/ml chitosan at pH 4.5. However, higher level (0.25 mg/ml) of chitosan was required to effectively arrest outgrowth of the germinated C. perfringens spores in Tripticase-yeast extract-glucose (TGY) medium. Furthermore, chitosan (1.0 mg/ml) was bacteriostatic against vegetative cells of C. perfringens in TGY medium. Although chitosan showed strong inhibitory activities against C. perfringens in laboratory medium, higher levels (2.0 mg/g) were required to achieve similar inhibition of spores inoculated into chicken meat. In summary, the inhibitory effects of chitosan against C. perfringens FP isolates was concentration dependent, and no major difference was observed when using different molecule weight chitosan as an inhibitor. Our results contribute to a better understanding on the potential application of chitosan in cooked meat products to control C. perfringens-associated disease. Copyright © 2016 Elsevier Ltd. All rights reserved.

Candida albicans adherence to resin-composite restorative dental material: influence of whole human saliva.

PubMed

Maza, José Luis; Elguezabal, Natalia; Prado, Carlota; Ellacuría, Joseba; Soler, Iñaki; Pontón, José

2002-11-01

Attachment of Candida albicans to oral surfaces is believed to be a critical event in the colonization of the oral cavity and in the development of oral diseases such as Candida-associated denture stomatitis. Although there is considerable information about the adhesion of C albicans to buccal epithelial cells and prosthetic materials, there is very little information about the adhesion of C albicans to composite restorative materials. The purpose of this study was to investigate the degree of adhesion of C albicans to a resin-composite restorative material (Herculite). The adhesion of 2 strains of C albicans, a germinative and a germ tube-deficient mutant, was studied by a visual method after incubating the fungus and the resin with and without human whole saliva. In absence of saliva, the adhesion of the C albicans germinative isolate to the resin showed an increase in parallel with the germination, reaching a maximum at the end of the experiment (120 minutes). However, no significant differences were observed in the adhesion of the agerminative mutant during the period of time studied. In the presence of saliva, the adhesion of both isolates to the resin was significantly lowered. Germination and the presence of human whole saliva are important factors in the adhesion of C albicans to the resin-composite restorative material Herculite.

Magnetic-time model at off-season germination

NASA Astrophysics Data System (ADS)

Mahajan, Tarlochan Singh; Pandey, Om Prakash

2014-03-01

Effect of static magnetic field on germination of mung beans is described. Seeds of mung beans, were exposed in batches to static magnetic fields of 87 to 226 mT intensity for 100 min. Magnetic time constant - 60.743 Th (Tesla hour) was determined experimentally. High value of magnetic time constant signifies lower effect of magnetic field on germination rate as this germination was carried out at off-season (13°C). Using decay function, germination magnetic constant was calculated. There was a linear increase in germination magnetic constant with increasing intensity of magnetic field. Calculated values of mean germination time, mean germination rate, germination rate coefficient, germination magnetic constant, transition time, water uptake, indicate that the impact of applied static magnetic field improves the germination of mung beans seeds even in off-season

A germination bioassay as a toxicological screening system for studying the effects of potential prodrugs of naproxen.

PubMed

Gonzalez-de la Parra, M; Ramos-Mundo, C; Jimenez-Estrada, M; Ponce-de Leon, C; Castillo, R; Tejeda, V; Cuevas, K G; Enriquez, R G

1998-01-01

A germination bioassay with radish (Raphanus sativus L.) seeds was developed as a toxicological screening system for assessing the effects of new potential prodrugs of naproxen, as an alternative to animals and animal cell toxicity screens. Both enantiomers of naproxen (6-methoxy-α-methyl-2-naphthaleneacetic acid) and naproxol (6-methoxy-β-2-naphthaleneethanol), and their racemic mixtures, inhibited the radicle growth of R. sativus at a concentration of 1mM, while only (R)-(+ )-naproxol and racemic naproxol inhibited the hypocotyl growth of R. sativus at the same concentration. Four novel combinatorial esters, naproxen naproxyl esters (6-methoxy-β-methyl-2-naphthaleneethyl 6-methoxy-α-methyl-2-naphthaleneacetate), resulting from the combinatorial chemistry of the esterification reaction between naproxen and naproxol, were synthesised and then tested in the germination bioassay, at a concentration of 0.5mM. It was found that they did not inhibit either the radicle or the hypocotyl growth of R. sativus. 1998 FRAME.

Infectious mononucleosis mimicking lymphoma: distinguishing morphological and immunophenotypic features.

PubMed

Louissaint, Abner; Ferry, Judith A; Soupir, Chad P; Hasserjian, Robert P; Harris, Nancy L; Zukerberg, Lawrence R

2012-08-01

The diagnosis of infectious mononucleosis (acute Epstein-Barr virus (EBV) infection) is usually made on the basis of clinical and laboratory findings. However, an atypical clinical presentation occasionally results in a lymph node or tonsillar biopsy. The morphological features of EBV-infected lymphoid tissue can easily mimic lymphoma. Furthermore, the immunophenotype of the immunoblasts has not been well characterized. To assess the morphological spectrum of acute EBV infection and the utility of immunohistochemistry in diagnosing difficult cases that resemble lymphoma, we reviewed 18 cases of acute EBV infection submitted in consultation to our institution with an initial diagnosis of/or suspicion for lymphoma. Patients included nine male and nine female individuals with a median age of 18 years (range 9-69). Biopsies were obtained from lymph nodes (3/18) or Waldeyer's ring (15/18). Infectious mononucleosis was confirmed by monospot or serological assays in 72% of cases (13/18). All cases featured architectural distortion by a polymorphous infiltrate with an immunoblastic proliferation, sometimes forming sheets. Reed-Sternberg-like cells were present in 8/18 (44%) of the cases. Infiltrates were often accompanied by necrosis (10/18) and mucosal ulceration (6/15). The majority of immunoblasts in all cases were CD20+ B cells with a post-germinal center immunophenotype (strongly positive for MUM1/IRF4 (18/18), CD10- (18/18 negative) and BCL-6- (16/18 negative; 2/18 faint BCL-6 expression in <10% of immunoblasts)). Immunoblasts showed variable weak expression of BCL-2 and polyclonal expression of κ and λ immunoglobulin light chains in 81% cases. Reed-Sternberg-like cells in 8/8 cases were CD30+, CD15-, BOB.1+ and OCT-2+. In conclusion, an atypical lymphoid infiltrate with numerous MUM1+, CD10-, BCL-6- immunoblasts should raise the suspicion of a reactive process, such as infectious mononucleosis, and warrants additional consideration before a diagnosis of lymphoma is made.

[The research of Valeriana amurensis seed germination characteristics].

PubMed

Liu, Juan; Yang, Chun-Rong; Jiang, Bo; Fang, Min; Du, Juan

2011-10-01

To study the effect of different treatments on the Valeriana amurensis seed germination rate. Used different chemical reagents and seed soakings on the routine germination test and the orthogonal test of the Valeriana amurensis seed, calculated the germination rate under different germination condition. Valeriana amurensis treated with different chemical reagends had different germination rate. The suitable immersion time could enhance Valeriana amurensis seed germination rate. Different treatment time, different disposal temperature, different germination temperature would have an impact on the Valeriana amurensis seed germination rate. In order to raise the Valeriana amurensis seed germination rate, use appropriate treatment on the seed before plant seeds; The seed growing must under suitable time and temperature.

iTRAQ and RNA-Seq Analyses Provide New Insights into Regulation Mechanism of Symbiotic Germination of Dendrobium officinale Seeds (Orchidaceae).

PubMed

Chen, Juan; Liu, Si Si; Kohler, Annegret; Yan, Bo; Luo, Hong Mei; Chen, Xiao Mei; Guo, Shun Xing

2017-06-02

Mycorrhizal fungi colonize orchid seeds and induce germination. This so-called symbiotic germination is a critical developmental process in the lifecycle of all orchid species. However, the molecular changes that occur during orchid seed symbiotic germination remain largely unknown. To better understand the molecular mechanism of orchid seed germination, we performed a comparative transcriptomic and proteomic analysis of the Chinese traditional medicinal orchid Dendrobium officinale to explore the change in protein expression at the different developmental stages during asymbiotic and symbiotic germination and identify the key proteins that regulate the symbiotic germination of orchid seeds. Among 2256 identified plant proteins, 308 were differentially expressed across three developmental stages during asymbiotic and symbiotic germination, and 229 were differentially expressed during symbiotic germination compared to asymbiotic development. Of these, 32 proteins were coup-regulated at both the proteomic and transcriptomic levels during symbiotic germination compared to asymbiotic germination. Our results suggest that symbiotic germination of D. officinale seeds shares a common signaling pathway with asymbiotic germination during the early germination stage. However, compared to asymbiotic germination, fungal colonization of orchid seeds appears to induce higher and earlier expression of some key proteins involved in lipid and carbohydrate metabolism and thus improves the efficiency of utilization of stored substances present in the embryo. This study provides new insight into the molecular basis of orchid seed germination.

Effect of day length on germination of seeds collected in Alaska

USGS Publications Warehouse

Densmore, R.V.

1997-01-01

Day length control can effectively limit seed germination to favorable seasons, but this phenomenon has been studied in relatively few wild plants. I tested species from interior Alaska for day length control of germination under controlled conditions, and I also monitored germination phenology in natural habitats. Unstratified and cold-stratified seeds were germinated on short (13 h) and long (22 h) day length and in the dark at constant and alternating temperatures. On long day length, unstratified Ledum decumbens and Saxifraga tricuspidata seeds germinated from 5??C to 20??C, but on short day length few or no seeds germinated at 5??C and 10??C and germination was reduced at higher temperatures. Unstratified seeds of Diapensia lapponica and Chamaedaphne calyculata germinated only at 15??C and 20??C on long day length, and short day length completely inhibited germination. Cold stratification widened the temperature range for germination on both long and short day lengths, but germination was still lower on short than long day length. Germination phenology in natural habitats was consistent with germination in controlled conditions. In these species, short day length and low temperatures interact to inhibit germination in the fall. After overwintering, seeds germinate in the spring at low temperatures and on long day lengths. The inhibitory effect of short day length is not important in the spring because day length is already long at snowmelt.

Rebamipide attenuates autoimmune arthritis severity in SKG mice via regulation of B cell and antibody production

PubMed Central

Byun, J-K; Moon, S-J; Jhun, J-Y; Kim, E-K; Park, J-S; Youn, J; Min, J-K; Park, S-H; Kim, H-Y; Cho, M-L

2014-01-01

Oxidative stress is involved in the pathophysiology of rheumatoid arthritis (RA). We investigated the therapeutic potential of rebamipide, a gastroprotective agent with a property of reactive oxygen species scavenger, on the development of inflammatory polyarthritis and the pathophysiological mechanisms by which rebamipide might confer anti-arthritic effects in SKG mice, an animal model of RA. Intraperitoneal (i.p.) injection of rebamipide attenuated the severity of clinical and histological arthritis. Rebampide treatment reduced the number of T helper type 1 (Th1), Th2, Th17, inducible T cell co-stimulator (ICOS)+ follicular helper T (Tfh) transitional type (T2) and mature B cells in the spleen, but increased the number of regulatory T (Treg), CD19+ CD1dhigh CD5high, CD19+ CD25high forkhead box protein 3 (FoxP3)+ regulatory B (Breg) cells, memory B cells, and transitional type 1 (T1) B cells. In addition, flow cytometric analysis revealed significantly decreased populations of FAS+GL-7+ germinal centre B cells and B220− CD138+ plasma cells in the spleens of rebamipide-treated SKG mice compared to controls. Rebamipide decreased germinal centre B cells and reciprocally induced Breg cells in a dose-dependent manner in vitro. Rebamipide-induced Breg cells had more suppressive capacity in relation to T cell proliferation and also inhibited Th17 differentiation from murine CD4+ T cells. Together, these data show that i.p. administration of rebamipide suppresses arthritis severity by inducing Breg and Treg cells and suppressing Tfh and Th17 cells in a murine model of RA. PMID:24749771

Phytochrome and Seed Germination. V. Changes of Phytochrome Content During the Germination of Cucumber Seeds 1

PubMed Central

Mancinelli, Alberto L.; Tolkowsky, Abby

1968-01-01

Cucumber seeds are light-sensitive, dark-germinating seeds. Inhibition of germination can be induced by prolonged exposure to continuous or intermittent FR. The dark germination process and the response to FR are phytochrome controlled. Phytochrome can be detected in these seeds by differential spectrophotometry in vivo. Spectrophotometrically measurable phytochrome increases during dark germination. The rate of increase is temperature dependent. Light treatments which are inhibitory for germination result in phytochrome contents lower than those of the seeds germinating in darkness. Treatments which restore germination also restore phytochrome formation. PMID:16656797

Fluorescence immunophenotyping and interphase cytogenetics (FICTION) detects BCL6 abnormalities, including gene amplification, in most cases of nodular lymphocyte-predominant Hodgkin lymphoma.

PubMed

Bakhirev, Alexei G; Vasef, Mohammad A; Zhang, Qian-Yun; Reichard, Kaaren K; Czuchlewski, David R

2014-04-01

BCL6 translocations are a frequent finding in B-cell lymphomas of diverse subtypes, including some cases of nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL). However, reliable analysis of BCL6 rearrangements using fluorescence in situ hybridization is difficult in NLPHL because of the relative paucity of neoplastic cells. Combined immunofluorescence microscopy and fluorescence in situ hybridization, or fluorescence immunophenotyping and interphase cytogenetics as a tool for the investigation of neoplasms (FICTION), permits targeted analysis of neoplastic cells. To better define the spectrum of BCL6 abnormalities in NLPHL using FICTION analysis. We performed an optimized FICTION analysis of 24 lymph nodes, including 11 NLPHL, 5 follicular hyperplasia with prominent progressive transformation of germinal centers, and 8 follicular hyperplasia without progressive transformation of germinal centers. BCL6 rearrangement was identified in 5 of 11 cases of NLPHL (46%). In addition, BCL6 gene amplification, with large clusters of BCL6 signals in the absence of chromosome 3 aneuploidy, was detected in 3 of 11 cases of NLPHL (27%). One NLPHL showed extra copies of BCL6 present in conjunction with multiple copies of chromosome 3. Altogether, we detected BCL6 abnormalities in 9 of 11 cases of NLPHL (82%). None of the progressive transformation of germinal centers or follicular hyperplasia cases showed BCL6 abnormalities by FICTION. To our knowledge, this is the first report of BCL6 gene amplification in NLPHL. Our optimized protocol for FICTION permits detection of cytogenetic abnormalities in most NLPHL cases and may represent a useful ancillary diagnostic technique.

Evaluation of the toxic potential of coffee wastewater on seeds, roots and meristematic cells of Lactuca sativa L.

PubMed

Aguiar, Luara Louzada; Andrade-Vieira, Larissa Fonseca; de Oliveira David, José Augusto

2016-11-01

Coffee wastewater (CWW) is an effluent produced through wet processing of coffee containing high concentration of organic matter, nutrients, salts and also agrochemicals. It is released directly into the argillaceous soil or into decantation tanks for later disposal into soils, by fertigation, subsurface infiltration or superficial draining. However, this practice is not followed by the monitoring the toxicity potential of this effluent. In this sense, the present work aimed to evaluate the phytotoxic, cytogenotoxic and mutagenic potential of CWW on seed germination, root elongation and cell cycle alterations in the plant model Lactuca sativa L. The effluent (CWW) collected was diluted in distilled water into six concentrations solutions (1.25%, 1.66%, 2.5%, 5.0%, 10%, 20%). A solution of raw CWW (100%) was also applied. Distilled water was used as negative control), and the DNA alkylating agent, metilmetano sulfonate (4×10(-4)M) as positive control. Physico-chemical parameters of the CWW was accessed and it was found that the effluent contained total phenols and inorganic matter in amounts within the limits established by the National Environment Council (CONAMA). Nevertheless, the biologicals assays performed demonstrated the phytotoxicity and cytogenotoxicty of CWW. Seed germination was totally inhibited after exposure of raw CWW. In addition, a decrease in seed germination speed as well as in root growth dose-dependently manner was noticed. Moreover, nuclear and chromosomal alterations were observed in the cell cycle, mostly arising from aneugenic action. Copyright © 2016 Elsevier Inc. All rights reserved.

Early root cap development and graviresponse in white clover (Trifolium repens) grown in space and on a two-axis clinostat

NASA Technical Reports Server (NTRS)

Smith, J. D.; Staehelin, L. A.; Todd, P.

1999-01-01

White clover (Trifolium repens) was germinated and grown in microgravity aboard the Space Shuttle (STS-60, 1994; STS-63, 1995), on Earth in stationary racks and in a slow-rotating two-axis clinostat. The objective of this study was to determine if normal root cap development and early plant gravity responses were dependent on gravitational cues. Seedlings were germinated in space and chemically fixed in orbit after 21, 40, and 72 h. Seedlings 96 h old were returned viable to earth. Germination and total seedling length were not dependent on gravity treatment. In space-flown seedlings, the number of cell stories in the root cap and the geometry of central columella cells did not differ from those of the Earth-grown seedlings. The root cap structure of clinorotated plants appeared similar to that of seedlings from microgravity, with the exception of three-day rotated plants, which displayed significant cellular damage in the columella region. Nuclear polarity did not depend on gravity; however, the positions of amyloplasts in the central columella cells were dependent on both the gravity treatment and the age of the seedlings. Seedlings from space, returned viable to earth, responded to horizontal stimulation as did 1 g controls, but seedlings rotated on the clinostat for the same duration had a reduced curvature response. This study demonstrates that initial root cap development is insensitive to either chronic clinorotation or microgravity. Soon after differentiation, however, clinorotation leads to loss of normal root cap structure and plant graviresponse while microgravity does not.

[An ultrastructural study of oogenesis in the planarian Schmidtea mediterranea (Platyhelminthe, Paludicola)].

PubMed

Harrath, Abdul Halim; Alwasal, Saleh H; Alhazza, Ibrahim; Zghal, Fathia; Tekaya, Saida

2011-07-01

The ovary of the freshwater planarian Schmidtea mediterranea has been studied for the first time using both light and electron microscopy methods. The ultrastructure of the ovary revealed two types of cells: accessory cells and germinal cells at various stages of differentiation, distributed along a maturation axis. Initially, oogonia underwent cytoplasm growth due to the development of organelles, such as endoplasmic reticulum, Golgi complex, and mitochondria, which are all involved in the production of cytoplasmic inclusions or yolk globules. It is shown that the chromatoid body and fibrogranular aggregates may participate in the synthesis of vitelline inclusions. When completely mature, the oocytes have become larger, due to the accumulation of nutritive inclusions, which are round in shape and have a paracrystalline structure. These inclusions are interpreted as being yolk globules and may represent a kind of nutritive material for the developing embryo. These ultrastructural features of the ovary agree with the available phylogenetic tree, based on morphological and karyological characters that considers Schmidtea group as a genus and not a subgenus. The presence of sperm between the oocytes suggests that fertilization may occur within the ovary, representing an uncommon condition within the Triclads, in which fertilization usually takes places outside of the ovaries. Copyright © 2011 Académie des sciences. Published by Elsevier SAS. All rights reserved.

An Antimicrobial Peptidomimetic Induces Mucorales Cell Death through Mitochondria-Mediated Apoptosis

PubMed Central

Barbu, E. Magda; Shirazi, Fazal; McGrath, Danielle M.; Albert, Nathaniel; Sidman, Richard L.; Pasqualini, Renata; Arap, Wadih; Kontoyiannis, Dimitrios P.

2013-01-01

The incidence of mucormycosis has dramatically increased in immunocompromised patients. Moreover, the array of cellular targets whose inhibition results in fungal cell death is rather limited. Mitochondria have been mechanistically identified as central regulators of detoxification and virulence in fungi. Our group has previously designed and developed a proteolytically-resistant peptidomimetic motif D(KLAKLAK)2 with pleiotropic action ranging from targeted (i.e., ligand-directed) activity against cancer and obesity to non-targeted activity against antibiotic resistant gram-negative rods. Here we evaluated whether this non-targeted peptidomimetic motif is active against Mucorales. We show that D(KLAKLAK)2 has marked fungicidal action, inhibits germination, and reduces hyphal viability. We have also observed cellular changes characteristic of apoptosis in D(KLAKLAK)2-treated Mucorales cells. Moreover, the fungicidal activity was directly correlated with vacuolar injury, mitochondrial swelling and mitochondrial membrane depolarization, intracellular reactive oxygen species accumulation (ROS), and increased caspase-like enzymatic activity. Finally, these apoptotic features were prevented by the addition of the ROS scavenger N-acetyl-cysteine indicating mechanistic pathway specificity. Together, these findings indicate that D(KLAKLAK)2 makes Mucorales exquisitely susceptible via mitochondrial injury-induced apoptosis. This prototype may serve as a candidate drug for the development of translational applications against mucormycosis and perhaps other fungal infections. PMID:24098573

An antimicrobial peptidomimetic induces Mucorales cell death through mitochondria-mediated apoptosis.

PubMed

Barbu, E Magda; Shirazi, Fazal; McGrath, Danielle M; Albert, Nathaniel; Sidman, Richard L; Pasqualini, Renata; Arap, Wadih; Kontoyiannis, Dimitrios P

2013-01-01

The incidence of mucormycosis has dramatically increased in immunocompromised patients. Moreover, the array of cellular targets whose inhibition results in fungal cell death is rather limited. Mitochondria have been mechanistically identified as central regulators of detoxification and virulence in fungi. Our group has previously designed and developed a proteolytically-resistant peptidomimetic motif D(KLAKLAK)2 with pleiotropic action ranging from targeted (i.e., ligand-directed) activity against cancer and obesity to non-targeted activity against antibiotic resistant gram-negative rods. Here we evaluated whether this non-targeted peptidomimetic motif is active against Mucorales. We show that D(KLAKLAK)2 has marked fungicidal action, inhibits germination, and reduces hyphal viability. We have also observed cellular changes characteristic of apoptosis in D(KLAKLAK)2-treated Mucorales cells. Moreover, the fungicidal activity was directly correlated with vacuolar injury, mitochondrial swelling and mitochondrial membrane depolarization, intracellular reactive oxygen species accumulation (ROS), and increased caspase-like enzymatic activity. Finally, these apoptotic features were prevented by the addition of the ROS scavenger N-acetyl-cysteine indicating mechanistic pathway specificity. Together, these findings indicate that D(KLAKLAK)2 makes Mucorales exquisitely susceptible via mitochondrial injury-induced apoptosis. This prototype may serve as a candidate drug for the development of translational applications against mucormycosis and perhaps other fungal infections.

Bcl-2 and BLIMP-1 expression predict worse prognosis in gastric diffuse large B cell lymphoma (DLCBL) while other markers for nodal DLBCL are not useful.

PubMed

Martin-Arruti, Maialen; Vaquero, Manuel; Díaz de Otazu, Ramón; Zabalza, Iñaki; Ballesteros, Javier; Roncador, Giovanna; García-Orad, Africa

2012-04-01

Previous studies have identified clinicopathological and immunohistochemical differences among diffuse large B cell lymphomas (DLBCL) as a function of disease location. Nevertheless, there is a continuing tendency to generalize the prognostic value of various identified markers without taking into account tumour site. Accordingly, we analysed the prognostic value of several of the immunohistochemical markers that have been proposed for nodal DLBCL in a group of patients with gastric DLBCL. Using histochemical methods, CD10, Bcl-6, Gcet1, MUM-1, Bcl-2 and BLIMP-1 expression was investigated in 43 cases of gastric DBLCL. As in nodal DLBCLs, expression of BLIMP-1, and of Bcl-2 in non-germinal centre B cell-like (non-GCB) patients, was associated with a worse prognosis. However, unlike nodal DBLCL, there was no significant association of prognosis with expression of CD10, Bcl-6, Gcet1 or MUM-1, or with categorization according to Hans or Muris algorithms. Although most markers of prognosis in nodal DLBCL are not useful indicators for gastric DLBCL, Bcl-2 or BLIMP-1 expression does correlate with worse prognosis. These data support the notion that clinicopathological features in DLBCL vary according to the disease location. © 2012 Blackwell Publishing Ltd.

Regulation of Seed Germination in the Close Arabidopsis Relative Lepidium sativum: A Global Tissue-Specific Transcript Analysis1[C][W][OA

PubMed Central

Morris, Karl; Linkies, Ada; Müller, Kerstin; Oracz, Krystyna; Wang, Xiaofeng; Lynn, James R.; Leubner-Metzger, Gerhard; Finch-Savage, William E.

2011-01-01

The completion of germination in Lepidium sativum and other endospermic seeds (e.g. Arabidopsis [Arabidopsis thaliana]) is regulated by two opposing forces, the growth potential of the radicle (RAD) and the resistance to this growth from the micropylar endosperm cap (CAP) surrounding it. We show by puncture force measurement that the CAP progressively weakens during germination, and we have conducted a time-course transcript analysis of RAD and CAP tissues throughout this process. We have also used specific inhibitors to investigate the importance of transcription, translation, and posttranslation levels of regulation of endosperm weakening in isolated CAPs. Although the impact of inhibiting translation is greater, both transcription and translation are required for the completion of endosperm weakening in the whole seed population. The majority of genes expressed during this process occur in both tissues, but where they are uniquely expressed, or significantly differentially expressed between tissues, this relates to the functions of the RAD as growing tissue and the CAP as a regulator of germination through weakening. More detailed analysis showed that putative orthologs of cell wall-remodeling genes are expressed in a complex manner during CAP weakening, suggesting distinct roles in the RAD and CAP. Expression patterns are also consistent with the CAP being a receptor for environmental signals influencing germination. Inhibitors of the aspartic, serine, and cysteine proteases reduced the number of isolated CAPs in which weakening developed, and inhibition of the 26S proteasome resulted in its complete cessation. This indicates that targeted protein degradation is a major control point for endosperm weakening. PMID:21321254

Involvement of reactive oxygen species in endosperm cap weakening and embryo elongation growth during lettuce seed germination

PubMed Central

Zhang, Yu; Chen, Bingxian; Xu, Zhenjiang; Shi, Zhaowan; Chen, Shanli; Huang, Xi; Chen, Jianxun; Wang, Xiaofeng

2014-01-01

Endosperm cap (CAP) weakening and embryo elongation growth are prerequisites for the completion of lettuce seed germination. Although it has been proposed that the cell wall loosening underlying these processes results from an enzymatic mechanism, it is still unclear which enzymes are involved. Here it is shown that reactive oxygen species (ROS), which are non-enzymatic factors, may be involved in the two processes. In Guasihong lettuce seeds imbibed in water, O2·– and H2O2 accumulated and peroxidase activity increased in the CAP, whereas its puncture force decreased. In addition, in the radicle, the increase in embryo growth potential was accompanied by accumulation of O2·– and an increase in peroxidase activity. Imbibing seeds in 0.3% sodium dichloroisocyanurate (SDIC) reduced endosperm viability and the levels of O2·–, H2O2, and peroxidase activity in the CAP, whereas the decrease in its puncture force was inhibited. However, in the embryo, SDIC did not affect the accumulation of O2·–, peroxidase activity, and the embryo growth potential. As a result, SDIC caused atypical germination, in which the endosperm ruptured at the boundary between the CAP and lateral endosperm. ROS scavengers and ROS generation inhibitors inhibited the CAP weakening and also decreased the embryo growth potential, thus decreasing the percentage of seed germination. Exogenous ROS and ROS generation inducers increased the percentage of CAP rupture to some extent, and the addition of H2O2 to 0.3% SDIC enabled some seeds to undergo typical germination. PMID:24744430