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Sample records for promote trichome formation

  1. Gibberellins Promote Trichome Formation by Up-Regulating GLABROUS1 in Arabidopsis1

    PubMed Central

    Perazza, Daniel; Vachon, Gilles; Herzog, Michel

    1998-01-01

    Trichome development is dependent on gibberellin (GA) signaling in Arabidopsis thaliana. Using the GA-deficient mutant ga1–3, the GA-response mutant spy-5, and uniconazol (a GA-biosynthesis inhibitor), we show that the GA level response correlates positively with both trichome number and trichome branch number. Two genes, GL1 and TTG, are required for trichome initiation. In ga1–3, coexpression of GL1 and R, the maize TTG functional homolog, under control of the constitutive 35S promoter, restored trichome development, whereas overexpression of neither GL1 nor R alone was sufficient to significantly suppress the glabrous phenotype. We next focused on GL1 regulation by GAs. In the double mutant the gl1–1 glabrous phenotype is epistatic to the spy-5 phenotype, suggesting that GL1 acts downstream of the GA signal transduction pathway. The activity of a β-glucuronidase reporter gene driven by the GL1 promoter was decreased in the wild type grown on uniconazol and showed a clear GA-dependent activation in ga1–3. Finally, quantification of GL1 transcript levels by reverse transcriptase-polymerase chain reaction demonstrated that relative to wild type, ga1–3 plants contained less transcript. These data support the hypothesis that GAs induce trichome development through up-regulation of GL1 and possibly TTG genes. PMID:9625690

  2. Auxin regulates first leaf development and promotes the formation of protocorm trichomes and rhizome-like structures in developing seedlings of Spathoglottis plicata (Orchidaceae)

    PubMed Central

    Novak, Stacey D.; Whitehouse, Grace A.

    2013-01-01

    Auxin flows in a polar manner to target tissues and exert its morphogenic effect. Preventing auxin movement, with polar auxin transport (PAT) inhibitors, or increasing auxin levels in tissues through exogenous application can provide a means for assessing the importance of appropriate tissue distribution and concentration of this hormone during development. The formulation of culture media for micropropagation has been the primary focus of most orchid tissue culture research, a goal that unveils seedling hormone responses at a single point in development. This study was unique because it evaluated the auxin response of orchids during three stages of seedling development. Seedlings were grown on standard culture media for 10, 35 and 85 days. Each group was sub-cultured onto auxin- and/or PAT inhibitor-containing media for an additional 10, 30 and 60 days, respectively. Data were collected on first leaf initiation, trichome formation and the appearance of propagative structures. In the 20-day seedlings, auxins and PAT inhibitors promoted precocious formation and random placement of protocorm hairs rather than in tufts, as seen in older, control seedlings. The 65-day seedlings formed protocorm-like bodies, rhizome-like growths from the stem, and fleshy leaves with trichomes. Seedlings cultured for 145 days developed microshoots or callus growth in the axils of older leaves and exhibited necrosis of original seedling roots and leaves. In general, exogenously applied auxin promoted the reversion of differentiated Spathoglottis plicata seedling tissue to a morphology that had propagative properties. Additionally, auxins commonly induced hair formation, which suggests that protocorm hairs may be root hair-like in nature. This work characterized three auxin growth responses in S. plicata seedlings that have not been reported in orchids: (i) the inhibition of first leaf initiation and abnormal first leaf morphology; (ii) the promotion of trichome formation; and (iii) the

  3. Arabidopsis GLASSY HAIR genes promote trichome papillae development

    PubMed Central

    Kirik, Viktor

    2013-01-01

    Specialized plant cells form cell walls with distinct composition and properties pertinent to their function. Leaf trichomes in Arabidopsis form thick cell walls that support the upright growth of these large cells and, curiously, have strong light-reflective properties. To understand the process of trichome cell-wall maturation and the molecular origins of this optical property, mutants affected in trichome light reflection were isolated and characterized. It was found that GLASSY HAIR (GLH) genes are required for the formation of surface papillae structures at late stages of trichome development. Trichomes in these mutants appeared transparent due to unobstructed light transmission. Genetic analysis of the isolated mutants revealed seven different gene loci. Two—TRICHOME BIREFRINGENCE (TBR) and NOK (Noeck)—have been reported previously to have the glassy trichome mutant phenotype. The other five glh mutants were analysed for cell-wall-related phenotypes. A significant reduction was found in cellulose content in glh2 and glh4 mutant trichomes. In addition to the glassy trichome phenotype, the glh6 mutants showed defects in leaf cuticular wax, and glh6 was found to represent a new allele of the eceriferum 10 (cer10) mutation. Trichomes of the glh1 and glh3 mutants did not show any other phenotypes beside reduced papillae formation. These data suggest that the GLH1 and GLH3 genes may have specific functions in trichome papillae formation, whereas GLH2, GLH4, and GLH6 genes are also involved in deposition of other cell-wall components. PMID:24014871

  4. TEMPRANILLO Reveals the Mesophyll as Crucial for Epidermal Trichome Formation1[OPEN

    PubMed Central

    Aguilar-Jaramillo, Andrea E.; Osnato, Michela; Shani, Eilon

    2016-01-01

    Plant trichomes are defensive specialized epidermal cells. In all accepted models, the epidermis is the layer involved in trichome formation, a process controlled by gibberellins (GAs) in Arabidopsis rosette leaves. Indeed, GA activates a genetic cascade in the epidermis for trichome initiation. Here we report that TEMPRANILLO (TEM) genes negatively control trichome initiation not only from the epidermis but also from the leaf layer underneath the epidermis, the mesophyll. Plants over-expressing or reducing TEM specifically in the mesophyll, display lower or higher trichome numbers, respectively. We surprisingly found that fluorescently labeled GA3 accumulates exclusively in the mesophyll of leaves, but not in the epidermis, and that TEM reduces its accumulation and the expression of several newly identified GA transporters. This strongly suggests that TEM plays an essential role, not only in GA biosynthesis, but also in regulating GA distribution in the mesophyll, which in turn directs epidermal trichome formation. Moreover, we show that TEM also acts as a link between GA and cytokinin signaling in the epidermis by negatively regulating downstream genes of both trichome formation pathways. Overall, these results call for a re-evaluation of the present theories of trichome formation as they reveal mesophyll essential during epidermal trichome initiation. PMID:26802039

  5. Genetic basis for glandular trichome formation in cotton

    PubMed Central

    Ma, Dan; Hu, Yan; Yang, Changqing; Liu, Bingliang; Fang, Lei; Wan, Qun; Liang, Wenhua; Mei, Gaofu; Wang, Lingjian; Wang, Haiping; Ding, Linyun; Dong, Chenguang; Pan, Mengqiao; Chen, Jiedan; Wang, Sen; Chen, Shuqi; Cai, Caiping; Zhu, Xiefei; Guan, Xueying; Zhou, Baoliang; Zhu, Shuijin; Wang, Jiawei; Guo, Wangzhen; Chen, Xiaoya; Zhang, Tianzhen

    2016-01-01

    Trichomes originate from epidermal cells and can be classified as either glandular or non-glandular. Gossypium species are characterized by the presence of small and darkly pigmented lysigenous glands that contain large amounts of gossypol. Here, using a dominant glandless mutant, we characterize GoPGF, which encodes a basic helix-loop-helix domain-containing transcription factor, that we propose is a positive regulator of gland formation. Silencing GoPGF leads to a completely glandless phenotype. A single nucleotide insertion in GoPGF, introducing a premature stop codon is found in the duplicate recessive glandless mutant (gl2gl3). The characterization of GoPGF helps to unravel the regulatory network of glandular structure biogenesis, and has implications for understanding the production of secondary metabolites in glands. It also provides a potential molecular basis to generate glandless seed and glanded cotton to not only supply fibre and oil but also provide a source of protein for human consumption. PMID:26795254

  6. Fine-mapping of the woolly gene controlling multicellular trichome formation and embryonic development in tomato.

    PubMed

    Yang, Changxian; Li, Hanxia; Zhang, Junhong; Wang, Taotao; Ye, Zhibiao

    2011-08-01

    Trichomes are small hairs that originate from the epidermal cells of nearly all land plants, and they exist in unicellular and multicellular forms. The regulatory pathway of unicellular trichomes in Arabidopsis is well characterized. However, little is known about the multicellular trichome formation in tomato (Solanum lycopersicum). The woolly (Wo) gene controls multicellular trichome initiation and leads to embryonic lethality when homozygous in tomato. To clone and characterize Wo, the gene was fine-mapped to a DNA fragment of ~200 kb using the map-based cloning strategy. A series of sequence-based molecular markers, including simple sequence repeat, sequence characterized amplified region, and cleaved amplified polymorphic sequence were utilized in this study. Analysis of the sequence indicated that this region carries 19 putative open reading frames. These results will provide not only the important information for the isolation and characterization of Wo but also the starting point for studying the regulatory pathway responsible for trichome formation and embryonic lethality in tomato.

  7. Promoters of AaGL2 and AaMIXTA-Like1 genes of Artemisia annua direct reporter gene expression in glandular and non-glandular trichomes

    PubMed Central

    Jindal, Sunita; Longchar, Bendangchuchang; Singh, Alka; Gupta, Vikrant

    2015-01-01

    Herein, we report cloning and analysis of promoters of GLABRA2 (AaGL2) homolog and a MIXTA-Like (AaMIXTA-Like1) gene from Artemisia annua. The upstream regulatory regions of AaGL2 and AaMIXTA-Like1 showed the presence of several crucial cis-acting elements. Arabidopsis and A. annua seedlings were transiently transfected with the promoter-GUS constructs using a robust agro-infiltration method. Both AaGL2 and AaMIXTA-Like1 promoters showed GUS expression preferentially in Arabidopsis single-celled trichomes and glandular as well as T-shaped trichomes of A. annua. Transgenic Arabidopsis harboring constructs in which AaGL2 or AaMIXTA-Like1 promoters would control GFP expression, showed fluorescence emanating specifically from trichome cells. Our study provides a fast and efficient method to study trichome-specific expression, and 2 promoters that have potential for targeted metabolic engineering in plants. PMID:26340695

  8. Two IIIf Clade-bHLHs from Freesia hybrida Play Divergent Roles in Flavonoid Biosynthesis and Trichome Formation when Ectopically Expressed in Arabidopsis

    PubMed Central

    Li, Yueqing; Shan, Xiaotong; Gao, Ruifang; Yang, Song; Wang, Shucai; Gao, Xiang; Wang, Li

    2016-01-01

    The MBW complex, comprised by R2R3-MYB, basic helix-loop-helix (bHLH) and WD40, is a single regulatory protein complex that drives the evolution of multiple traits such as flavonoid biosynthesis and epidermal cell differentiation in plants. In this study, two IIIf Clade-bHLH regulator genes, FhGL3L and FhTT8L, were isolated and functionally characterized from Freesia hybrida. Different spatio-temporal transcription patterns were observed showing diverse correlation with anthocyanin and proanthocyanidin accumulation. When overexpressed in Arabidopsis, FhGL3L could enhance the anthocyanin accumulation through up-regulating endogenous regulators and late structural genes. Unexpectedly, trichome formation was inhibited associating with the down-regulation of AtGL2. Comparably, only the accumulation of anthocyanins and proanthocyanidins was strengthened in FhTT8L transgenic lines. Furthermore, transient expression assays demonstrated that FhGL3L interacted with AtPAP1, AtTT2 and AtGL1, while FhTT8L only showed interaction with AtPAP1 and AtTT2. In addition, similar activation of the AtDFR promoter was found between AtPAP1-FhGL3L/FhTT8L and AtPAP1- AtGL3/AtTT8 combinations. When FhGL3L was fused with a strong activation domain VP16, it could activate the AtGL2 promoter when co-transfected with AtGL1. Therefore, it can be concluded that the functionality of bHLH factors may have diverged, and a sophisticated interaction and hierarchical network might exist in the regulation of flavonoid biosynthesis and trichome formation. PMID:27465838

  9. T-shaped trichome-specific expression of monoterpene synthase ADH2 using promoter-β-GUS fusion in transgenic Artemisia annua L.

    PubMed

    Fu, Xueqing; Shi, Pu; Shen, Qian; Jiang, Weimin; Tang, Yueli; Lv, Zongyou; Yan, Tingxiang; Li, Ling; Wang, Guofeng; Sun, Xiaofen; Tang, Kexuan

    2016-11-01

    Artemisinin, a sesquiterpene lactone isolated from Artemisia annua L. (sweet wormwood), is extensively used in the treatment of malaria. In order to better understand the metabolism of terpenes in A. annua and the influence of terpene synthases on artemisinin yield, the expression pattern of a monoterpene alcohol dehydrogenase (ADH2) has been studied using transgenic plants expressing promoter-β-glucuronidase (GUS) fusion. ADH2 played a major role in monoterpenoid biosynthesis including carveol, borneol, and artemisia ketone through in vitro biochemical analysis. In this study, the ADH2 promoter was cloned by the genome walking method. A number of putative cis-acting elements were predicted in promoter region, suggesting that the ADH2 is driven by a complex regulation mechanism. ADH2 gene was highly expressed in old leaves, whereas the artemisinin biosynthetic genes were mainly expressed in bud and young leaves. The expression of ADH2 gene increased quickly during leaf development, revealed by qRT-PCR. GUS expression analysis in different tissues of transgenic A. annua demonstrates that ADH2 expression is exclusively located to T-shaped trichome, not glandular secretory trichome.

  10. A Geranylfarnesyl Diphosphate Synthase Provides the Precursor for Sesterterpenoid (C25) Formation in the Glandular Trichomes of the Mint Species Leucosceptrum canum

    PubMed Central

    Luo, Shi-Hong; Schmidt, Axel; Sun, Gui-Ling; Kuang, Ce; Yang, Min-Jie; Jing, Shu-Xi; Li, Chun-Huan

    2016-01-01

    Plant sesterterpenoids, an important class of terpenoids, are widely distributed in various plants, including food crops. However, little is known about their biosynthesis. Here, we cloned and functionally characterized a plant geranylfarnesyl diphosphate synthase (Lc-GFDPS), the enzyme producing the C25 prenyl diphosphate precursor to all sesterterpenoids, from the glandular trichomes of the woody plant Leucosceptrum canum. GFDPS catalyzed the formation of GFDP after expression in Escherichia coli. Overexpressing GFDPS in Arabidopsis thaliana also gave an extract catalyzing GFDP formation. GFDPS was strongly expressed in glandular trichomes, and its transcript profile was completely in accordance with the sesterterpenoid accumulation pattern. GFDPS is localized to the plastids, and inhibitor studies indicated its use of isoprenyl diphosphate substrates supplied by the 2-C-methyl-d-erythritol 4-phosphate pathway. Application of a jasmonate defense hormone induced GFDPS transcript and sesterterpenoid accumulation, while reducing feeding and growth of the generalist insect Spodoptera exigua, suggesting that these C25 terpenoids play a defensive role. Phylogenetic analysis suggested that GFDPS probably evolved from plant geranylgeranyl diphosphate synthase under the influence of positive selection. The isolation of GFDPS provides a model for investigating sesterterpenoid formation in other species and a tool for manipulating the formation of this group in plants and other organisms. PMID:26941091

  11. Trichome-Related Mutants Provide a New Perspective on Multicellular Trichome Initiation and Development in Cucumber (Cucumis sativus L)

    PubMed Central

    Liu, Xingwang; Bartholomew, Ezra; Cai, Yanling; Ren, Huazhong

    2016-01-01

    Trichomes are specialized epidermal cells located in aerial parts of plants that function in plant defense against biotic and abiotic stresses. The simple unicellular trichomes of Arabidopsis serve as an excellent model to study the molecular mechanism of cell differentiation and pattern formation in plants. Loss-of-function mutations in Arabidopsis thaliana have suggested that the core genes GL1 (which encodes a MYB transcription factor) and TTG1 (which encodes a WD40 repeat-containing protein) are important for the initiation and spacing of leaf trichomes, while for normal trichome initiation, the genes GL3, and EGL3 (which encode a bHLH protein) are needed. However, the positive regulatory genes involved in multicellular trichrome development in cucumber remain unclear. This review focuses on the phenotype of mutants (csgl3, tril, tbh, mict, and csgl1) with disturbed trichomes in cucumber and then infers which gene(s) play key roles in trichome initiation and development in those mutants. Evidence indicates that MICT, TBH, and CsGL1 are allelic with alternative splicing. CsGL3 and TRIL are allelic and override the effect of TBH, MICT, and CsGL1 on the regulation of multicellular trichome development; and affect trichome initiation. CsGL3, TRIL, MICT, TBH, and CsGL1 encode HD-Zip proteins with different subfamilies. Genetic and molecular analyses have revealed that CsGL3, TRIL, MICT, TBH, and CsGL1 are responsible for the differentiation of epidermal cells and the development of trichomes. Based on current knowledge, a positive regulator pathway model for trichome development in cucumber was proposed and compared to a model in Arabidopsis. These data suggest that trichome development in cucumber may differ from that in Arabidopsis. PMID:27559338

  12. The identification of Cucumis sativus Glabrous 1 (CsGL1) required for the formation of trichomes uncovers a novel function for the homeodomain-leucine zipper I gene.

    PubMed

    Li, Qiang; Cao, Chenxing; Zhang, Cunjia; Zheng, Shuangshuang; Wang, Zenghui; Wang, Lina; Ren, Zhonghai

    2015-05-01

    The spines and bloom of cucumber (Cucumis sativus L.) fruit are two important quality traits related to fruit market value. However, until now, none of the genes involved in the formation of cucumber fruit spines and bloom trichomes has been identified. Here, the characterization of trichome development in wild-type (WT) cucumber and a spontaneous mutant, glabrous 1 (csgl1) controlled by a single recessive nuclear gene, with glabrous aerial organs, is reported. Via map-based cloning, CsGL1 was isolated and it was found that it encoded a member of the homeodomain-leucine zipper I (HD-Zip I) proteins previously identified to function mainly in the abiotic stress responses of plants. Tissue-specific expression analysis indicated that CsGL1 was strongly expressed in trichomes and fruit spines. In addition, CsGL1 was a nuclear protein with weak transcriptional activation activity in yeast. A comparative analysis of the digital gene expression (DGE) profile between csgl1 and WT leaves revealed that CsGL1 had a significant influence on the gene expression profile in cucumber, especially on genes related to cellular process, which is consistent with the phenotypic difference between csgl1 and the WT. Moreover, two genes, CsMYB6 and CsGA20ox1, possibly involved in the formation of cucumber trichomes and fruit spines, were characterized. Overall, the findings reveal a new function for the HD-Zip I gene subfamily, and provide some candidate genes for genetic engineering approaches to improve cucumber fruit external quality.

  13. Hydrophobic trichome layers and epicuticular wax powders in Bromeliaceae.

    PubMed

    Pierce, S; Maxwell, K; Griffiths, H; Winter, K

    2001-08-01

    The distinctive foliar trichome of Bromeliaceae has promoted the evolution of an epiphytic habit in certain taxa by allowing the shoot to assume a significant role in the uptake of water and mineral nutrients. Despite the profound ecophysiological and taxonomic importance of this epidermal structure, the functions of nonabsorbent trichomes in remaining Bromeliaceae are not fully understood. The hypothesis that light reflection from these trichome layers provides photoprotection was not supported by spectroradiometry and fluorimetry in the present study; the mean reflectance of visible light from trichome layers did not exceed 6.4% on the adaxial surfaces of species representing a range of ecophysiological types nor was significant photoprotection provided by their presence. Several reports suggesting water repellency in some terrestrial Bromeliaceae were investigated. Scanning electron microscopy (SEM) and a new technique-fluorographic dimensional imaging (FDI)-were used to assess the interaction between aqueous droplets and the leaf surfaces of 86 species from 25 genera. In the majority of cases a dense layer of overlapping, stellate or peltate trichomes held water off the leaf epidermis proper. In the case of hydrophobic tank-forming tillandsioideae, a powdery epicuticular wax layer provided water repellency. The irregular architecture of these indumenta resulted in relatively little contact with water droplets. Most mesic terrestrial Pitcairnioideae examined either possessed glabrous leaf blades or hydrophobic layers of confluent trichomes on the abaxial surface. Thus, the present study indicates that an important ancestral function of the foliar trichome in Bromeliaceae was water repellency. The ecophysiological consequences of hydrophobia are discussed.

  14. Characterization of the formation of branched short-chain fatty acid:CoAs for bitter acid biosynthesis in hop glandular trichomes.

    PubMed

    Xu, Haiyang; Zhang, Fengxia; Liu, Baoxiu; Huhman, David V; Sumner, Lloyd W; Dixon, Richard A; Wang, Guodong

    2013-07-01

    Bitter acids, known for their use as beer flavoring and for their diverse biological activities, are predominantly formed in hop (Humulus lupulus) glandular trichomes. Branched short-chain acyl-CoAs (e.g. isobutyryl-CoA, isovaleryl-CoA and 2-methylbutyryl-CoA), derived from the degradation of branched-chain amino acids (BCAAs), are essential building blocks for the biosynthesis of bitter acids in hops. However, little is known regarding what components are needed to produce and maintain the pool of branched short-chain acyl-CoAs in hop trichomes. Here, we present several lines of evidence that both CoA ligases and thioesterases are likely involved in bitter acid biosynthesis. Recombinant HlCCL2 (carboxyl CoA ligase) protein had high specific activity for isovaleric acid as a substrate (K cat /K m = 4100 s(-1) M(-1)), whereas recombinant HlCCL4 specifically utilized isobutyric acid (Kcat/K m = 1800 s(-1) M(-1)) and 2-methylbutyric acid (Kcat/K m = 6900 s(-1) M(-1)) as substrates. Both HlCCLs, like hop valerophenone synthase (HlVPS), were expressed strongly in glandular trichomes and localized to the cytoplasm. Co-expression of HlCCL2 and HlCCL4 with HlVPS in yeast led to significant production of acylphloroglucinols (the direct precursors for bitter acid biosynthesis), which further confirmed the biochemical function of these two HlCCLs in vivo. Functional identification of a thioesterase that catalyzed the reverse reaction of CCLs in mitochondria, together with the comprehensive analysis of genes involved BCAA catabolism, supported the idea that cytosolic CoA ligases are required for linking BCAA degradation and bitter acid biosynthesis in glandular trichomes. The evolution and other possible physiological roles of branched short-chain fatty acid:CoA ligases in planta are also discussed.

  15. A cold-tolerant evergreen interspecific hybrid of Ocimum kilimandscharicum and Ocimum basilicum: analyzing trichomes and molecular variations.

    PubMed

    Dhawan, Sunita Singh; Shukla, Preeti; Gupta, Pankhuri; Lal, R K

    2016-05-01

    Ocimum (Lamiaceae) is an important source of essential oils and aroma chemicals especially eugenol, methyl eugenol, linalool, methyl chavicol etc. An elite evergreen hybrid has been developed from Ocimum kilimandscharicum and Ocimum basilicum, which demonstrated adaptive behavior towards cold stress. A comparative molecular analysis has been done through RAPD, AFLP, and ISSR among O. basilicum and O. kilimandscharicum and their evergreen cold-tolerant hybrid. The RAPD and AFLP analyses demonstrated similar results, i.e., the hybrid of O. basilicum and O. kilimandscharicum shares the same cluster with O. kilimandscharicum, while O. basilicum behaves as an outgroup, whereas in ISSR analysis, the hybrid genotype grouped in the same cluster with O. basilicum. Ocimum genotypes were analyzed and compared for their trichome density. There were distinct differences on morphology, distribution, and structure between the two kinds of trichomes, i.e., glandular and non-glandular. Glandular trichomes contain essential oils, polyphenols, flavonoids, and acid polysaccharides. Hair-like trichomes, i.e., non-glandular trichomes, help in keeping the frost away from the living surface cells. O. basilicum showed less number of non-glandular trichomes on leaves compared to O. kilimandscharicum and the evergreen cold-tolerant hybrid. Trichomes were analyzed in O. kilimandscharicum, O. basilicum, and their hybrid. An increased proline content at the biochemical level represents a higher potential to survive in a stress condition like cold stress. In our analysis, the proline content is quite higher in tolerant variety O. kilimandscharicum, low in susceptible variety O. basilicum, and intermediate in the hybrid. Gene expression analysis was done in O. basilicum, O. kilimandscharicum and their hybrid for TTG1, GTL1, and STICHEL gene locus which regulates trichome development and its formation and transcription factors WRKY and MPS involved in the regulation of plant responses to freezing

  16. The ENHANCER OF TRY AND CPC1 gene acts redundantly with TRIPTYCHON and CAPRICE in trichome and root hair cell patterning in Arabidopsis.

    PubMed

    Kirik, Victor; Simon, Marissa; Huelskamp, Martin; Schiefelbein, John

    2004-04-15

    The development of trichomes and root hairs in Arabidopsis provide useful models for the study of cell fate determination in plants. A common network of putative transcriptional regulators, including the small MYB proteins TRIPTYCHON (TRY) and CAPRICE (CPC), is known to influence the patterning of both cell types. Here, we used an activation tagging strategy to identify a new regulator, ENHANCER OF TRY AND CPC 1 (ETC1). The ETC1 sequence is similar to TRY and CPC, and ETC1 overexpression causes a reduction in trichome formation and excessive root hair production. The etc1 single mutant has no significant phenotype, but it enhances the effect of cpc and try on trichome and root hair development, which shows that ETC1 function is partially redundant with TRY and CPC. In addition, the etc1 try cpc triple mutant has novel phenotypes, revealing previously unrecognized roles for these regulators in epidermis development. An ETC1 promoter-reporter gene fusion is expressed in the developing trichome and non-hair cells, similar to the expression of TRY and CPC. These results suggest that ETC1, TRY, and CPC act in concert to repress the trichome cell fate in the shoot epidermis and the non-hair cell fate in the root epidermis.

  17. Trichomes as dangerous lollipops

    PubMed Central

    Stork, William F. J.; Weinhold, Alexander; Baldwin, Ian T.

    2011-01-01

    When attacked by herbivores, plants produce toxic secondary metabolites that function as direct defenses, as well as indirect defenses that attract and reward predators of the offending herbivores. These indirect defenses include both nutritive rewards such as extra floral nectar, as well as informational rewards, such as the production and release of volatile compounds that betray the location of feeding herbivores to predators. Herbivory of Nicotiana attenuata by the tobacco hornworm (Manduca larvae) alters the volatile profiles of both the plant and larval headspace. Herbivory-elicited specific changes in the volatile profiles are detected by arthropod predators of Manduca larvae. The known predators that perceive volatile cues induced by Manduca herbivory of N. attenuata are insects that target Manduca at early developmental stages, when the larvae are still small; large, late-instar larvae may have outgrown these predation risks. However, here we offer evidence that branched chain aliphatic acids derived from the digestion of plant O-acyl sugars from trichomes may betray Manduca larvae to lizard predators during late developmental stages as well. PMID:22095147

  18. Trichome structure and evolution in Neotropical lianas

    PubMed Central

    Nogueira, Anselmo; El Ottra, Juliana Hanna Leite; Guimarães, Elza; Machado, Silvia Rodrigues; Lohmann, Lúcia G.

    2013-01-01

    Background and Aims Trichomes are epidermal outgrowths generally associated with protection against herbivores and/or desiccation that are widely distributed from ferns to angiosperms. Patterns of topological variation and morphological evolution of trichomes are still scarce in the literature, preventing valid comparisons across taxa. This study integrates detailed morphoanatomical data and the evolutionary history of the tribe Bignonieae (Bignoniaceae) in order to gain a better understanding of current diversity and evolution of trichome types. Methods Two sampling schemes were used to characterize trichome types: (1) macromorphological characterization of all 105 species currently included in Bignonieae; and (2) micromorphological characterization of 16 selected species. Individual trichome morphotypes were coded as binary in each vegetative plant part, and trichome density and size were coded as multistate. Ancestral character state reconstructions were conducted using maximum likelihood (ML) assumptions. Key Results Two main functional trichome categories were found: non-glandular and glandular. In glandular trichomes, three morphotypes were recognized: peltate (Pg), stipitate (Sg) and patelliform/cupular (P/Cg) trichomes. Non-glandular trichomes were uniseriate, uni- or multicellular and simple or branched. Pg and P/Cg trichomes were multicellular and non-vascularized with three clearly distinct cell layers. Sg trichomes were multicellular, uniseriate and long-stalked. ML ancestral character state reconstructions suggested that the most recent common ancestor (MRCA) of Bignonieae probably had non-glandular, Pg and P/Cg trichomes, with each trichome type presenting alternative histories of appearance on the different plant parts. For example, the MRCA of Bignonieae probably had non-glandular trichomes on the stems, prophylls, petiole, petiolule and leaflet veins while P/Cg trichomes were restricted to leaflet blades. Sg trichomes were not present in the MRCA

  19. Cotton AnnGh3 encoding an annexin protein is preferentially expressed in fibers and promotes initiation and elongation of leaf trichomes in transgenic Arabidopsis.

    PubMed

    Li, Bing; Li, Deng-Di; Zhang, Jie; Xia, Hui; Wang, Xiu-Lan; Li, Ying; Li, Xue-Bao

    2013-10-01

    The annexins are a multifamily of calcium-regulated phospholipid-binding proteins. To investigate the roles of annexins in fiber development, four genes encoding putative annexin proteins were isolated from cotton (Gossypium hirsutum) and designated AnnGh3, AnnGh4, AnnGh5, and AnnGh6. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) results indicated that AnnGh3, AnnGh4, and AnnGh5 were preferentially expressed in fibers, while the transcripts of AnnGh6 were predominantly accumulated in roots. During fiber development, the transcripts of AnnGh3/4/5 genes were mainly accumulated in rapidly elongating fibers. With fiber cells further developed, their expression activity was dramatically declined to a relatively low level. In situ hybridization results indicated that AnnGh3 and AnnGh5 were expressed in initiating fiber cells (0-2 DPA). Additionally, their expression in fibers was also regulated by phytohormones and [Ca(2+)]. Subcellular localization analysis discovered that AnnGh3 protein was localized in the cytoplasm. Overexpression of AnnGh3 in Arabidopsis resulted in a significant increase in trichome density and length on leaves of the transgenic plants, suggesting that AnnGh3 may be involved in fiber cell initiation and elongation of cotton.

  20. One, two, three...models for trichome patterning in Arabidopsis?

    PubMed

    Pesch, Marina; Hülskamp, Martin

    2009-10-01

    Trichome patterning in Arabidopsis rosette leaves serves as a model system to study how individual cells are determined in a regular spacing pattern from initially equivalent cells. A conserved gene cassette regulates this spacing pattern. bHLH, MYB, and WD40 factors are positive regulators of trichome development that are inhibited by R3 single-repeat MYB proteins. One positive regulator, the WD40-protein, and the negative regulators are mobile and are transported in opposite directions: the WD40 factor moves to the negative regulators away from trichome initials. This movement behavior and the genetic and molecular interactions between the contributing genes and proteins make it very difficult to derive the mechanistic core behind pattern formation. Attempts to develop mathematical models toward this end are critically discussed.

  1. Ectopic Expression of Capsicum-Specific Cell Wall Protein Capsicum annuum Senescence-Delaying 1 (CaSD1) Delays Senescence and Induces Trichome Formation in Nicotiana benthamiana

    PubMed Central

    Seo, Eunyoung; Yeom, Seon-In; Jo, SungHwan; Jeong, Heejin; Kang, Byoung-Cheorl; Choi, Doil

    2012-01-01

    Secreted proteins are known to have multiple roles in plant development, metabolism, and stress response. In a previous study to understand the roles of secreted proteins, Capsicum annuum secreted proteins (CaS) were isolated by yeast secretion trap. Among the secreted proteins, we further characterized Capsicum annuum senescence-delaying 1 (CaSD1), a gene encoding a novel secreted protein that is present only in the genus Capsicum. The deduced CaSD1 contains multiple repeats of the amino acid sequence KPPIHNHKPTDYDRS. Interestingly, the number of repeats varied among cultivars and species in the Capsicum genus. CaSD1 is constitutively expressed in roots, and Agrobacterium-mediated transient overexpression of CaSD1 in Nicotiana benthamiana leaves resulted in delayed senescence with a dramatically increased number of trichomes and enlarged epidermal cells. Furthermore, senescence- and cell division-related genes were differentially regulated by CaSD1-overexpressing plants. These observations imply that the pepper-specific cell wall protein CaSD1 plays roles in plant growth and development by regulating cell division and differentiation. PMID:22441673

  2. Ectopic expression of Capsicum-specific cell wall protein Capsicum annuum senescence-delaying 1 (CaSD1) delays senescence and induces trichome formation in Nicotiana benthamiana.

    PubMed

    Seo, Eunyoung; Yeom, Seon-In; Jo, Sunghwan; Jeong, Heejin; Kang, Byoung-Cheorl; Choi, Doil

    2012-04-01

    Secreted proteins are known to have multiple roles in plant development, metabolism, and stress response. In a previous study to understand the roles of secreted proteins, Capsicum annuum secreted proteins (CaS) were isolated by yeast secretion trap. Among the secreted proteins, we further characterized Capsicum annuum senescence-delaying 1 (CaSD1), a gene encoding a novel secreted protein that is present only in the genus Capsicum. The deduced CaSD1 contains multiple repeats of the amino acid sequence KPPIHNHKPTDYDRS. Interestingly, the number of repeats varied among cultivars and species in the Capsicum genus. CaSD1 is constitutively expressed in roots, and Agrobacterium-mediated transient overexpression of CaSD1 in Nicotiana benthamiana leaves resulted in delayed senescence with a dramatically increased number of trichomes and enlarged epidermal cells. Furthermore, senescence- and cell division-related genes were differentially regulated by CaSD1-overexpressing plants. These observations imply that the pepper-specific cell wall protein CaSD1 plays roles in plant growth and development by regulating cell division and differentiation.

  3. GLABROUS INFLORESCENCE STEMS3 (GIS3) regulates trichome initiation and development in Arabidopsis.

    PubMed

    Sun, Lili; Zhang, Aidong; Zhou, Zhongjing; Zhao, Yongqin; Yan, An; Bao, Shengjie; Yu, Hao; Gan, Yinbo

    2015-04-01

    Arabidopsis trichome formation is an excellent model for studying various aspects of plant cell development and cell differentiation. Our previous works have demonstrated that several C2H2 zinc finger proteins, including GIS, GIS2, ZFP5, ZFP6 and ZFP8, control trichome cell development through GA and cytokinin signalling in Arabidopsis. We identified a novel C2H2 zinc finger protein, GLABROUS INFLORESCENCE STEMS 3 (GIS3), which is a key factor in regulating trichome development in Arabidopsis. In comparison with wild-type plants, loss-of-function of GIS3 mutants exhibited a significantly decreased number of trichomes in cauline leaves, lateral branches, sepals of flowers, and main stems. Overexpression of GIS3 resulted in increased trichome densities in sepal, cauline leaves, lateral branches, main inflorescence stems and in the appearance of ectopic trichomes on carpels. The molecular and genetic analyses show that GIS3 acts upstream of GIS, GIS2, ZFP8 and the key trichome initiation factors, GL1 and GL3. Steroid-inducible gene expression analyses and chromatin immunoprecipitation (ChIP) experiments suggest that GIS and GIS2 are the direct target genes of GIS3.

  4. Promoting proximal formative assessment with relational discourse

    NASA Astrophysics Data System (ADS)

    Scherr, Rachel E.; Close, Hunter G.; McKagan, Sarah B.

    2012-02-01

    The practice of proximal formative assessment - the continual, responsive attention to students' developing understanding as it is expressed in real time - depends on students' sharing their ideas with instructors and on teachers' attending to them. Rogerian psychology presents an account of the conditions under which proximal formative assessment may be promoted or inhibited: (1) Normal classroom conditions, characterized by evaluation and attention to learning targets, may present threats to students' sense of their own competence and value, causing them to conceal their ideas and reducing the potential for proximal formative assessment. (2) In contrast, discourse patterns characterized by positive anticipation and attention to learner ideas increase the potential for proximal formative assessment and promote self-directed learning. We present an analysis methodology based on these principles and demonstrate its utility for understanding episodes of university physics instruction.

  5. Cu(II) promotes amyloid pore formation

    SciTech Connect

    Zhang, Hangyu; Rochet, Jean-Christophe; Stanciu, Lia A.

    2015-08-14

    The aggregation of α-synuclein is associated with dopamine neuron death in Parkinson's disease. There is controversy in the field over the question of which species of the aggregates, fibrils or protofibrils, are toxic. Moreover, compelling evidence suggested the exposure to heavy metals to be a risk of PD. Nevertheless, the mechanism of metal ions in promoting PD remains unclear. In this research, we investigated the structural basis of Cu(II) induced aggregation of α-synuclein. Using transmission electron microscopy experiments, Cu(II) was found to promote in vitro aggregation of α-synuclein by facilitating annular protofibril formation rather than fibril formation. Furthermore, neuroprotective baicalein disaggregated annular protofibrils accompanied by considerable decrease of β-sheet content. These results strongly support the hypothesis that annular protofibrils are the toxic species, rather than fibrils, thereby inspiring us to search novel therapeutic strategies for the suppression of the toxic annular protofibril formation. - Highlights: • Cu(II) promoted the annular protofibril formation of α-synuclein in vitro. • Cu(II) postponed the in vitro fibrillization of α-synuclein. • Neuroprotective baicalein disaggregated annular protofibrils.

  6. Semi-automated 3D leaf reconstruction and analysis of trichome patterning from light microscopic images.

    PubMed

    Failmezger, Henrik; Jaegle, Benjamin; Schrader, Andrea; Hülskamp, Martin; Tresch, Achim

    2013-04-01

    Trichomes are leaf hairs that are formed by single cells on the leaf surface. They are known to be involved in pathogen resistance. Their patterning is considered to emerge from a field of initially equivalent cells through the action of a gene regulatory network involving trichome fate promoting and inhibiting factors. For a quantitative analysis of single and double mutants or the phenotypic variation of patterns in different ecotypes, it is imperative to statistically evaluate the pattern reliably on a large number of leaves. Here we present a method that enables the analysis of trichome patterns at early developmental leaf stages and the automatic analysis of various spatial parameters. We focus on the most challenging young leaf stages that require the analysis in three dimensions, as the leaves are typically not flat. Our software TrichEratops reconstructs 3D surface models from 2D stacks of conventional light-microscope pictures. It allows the GUI-based annotation of different stages of trichome development, which can be analyzed with respect to their spatial distribution to capture trichome patterning events. We show that 3D modeling removes biases of simpler 2D models and that novel trichome patterning features increase the sensitivity for inter-accession comparisons.

  7. Semi-automated 3D Leaf Reconstruction and Analysis of Trichome Patterning from Light Microscopic Images

    PubMed Central

    Schrader, Andrea; Hülskamp, Martin; Tresch, Achim

    2013-01-01

    Trichomes are leaf hairs that are formed by single cells on the leaf surface. They are known to be involved in pathogen resistance. Their patterning is considered to emerge from a field of initially equivalent cells through the action of a gene regulatory network involving trichome fate promoting and inhibiting factors. For a quantitative analysis of single and double mutants or the phenotypic variation of patterns in different ecotypes, it is imperative to statistically evaluate the pattern reliably on a large number of leaves. Here we present a method that enables the analysis of trichome patterns at early developmental leaf stages and the automatic analysis of various spatial parameters. We focus on the most challenging young leaf stages that require the analysis in three dimensions, as the leaves are typically not flat. Our software TrichEratops reconstructs 3D surface models from 2D stacks of conventional light-microscope pictures. It allows the GUI-based annotation of different stages of trichome development, which can be analyzed with respect to their spatial distribution to capture trichome patterning events. We show that 3D modeling removes biases of simpler 2D models and that novel trichome patterning features increase the sensitivity for inter-accession comparisons. PMID:23637587

  8. Comparison of TRY and the closely related At1g01380 gene in controlling Arabidopsis trichome patterning.

    PubMed

    Esch, Jeffrey J; Chen, Margaret A; Hillestad, Matthew; Marks, M David

    2004-12-01

    A screen of activation-tagged Arabidopsis lines resulted in the identification of At1g01380, which encodes a small R3 single repeat MYB gene, as a negative regulator of trichome initiation. Plants that overexpress this gene have fewer trichomes. The gene is closely related to the previously identified negative regulator TRY, and has a similar pattern of expression as TRY in developing leaves. As previously shown for TRY, At1g01380 protein can inhibit the interaction between the positive trichome regulators GL1 and GL3, and likely limits trichome initiation via this inhibition. While TRY and At1g01380 are closely related, they are not completely functionally equivalent. When placed under the transcriptional control of the TRY promoter, At1g01380 can only partially rescue the try mutant. Interestingly, Atg01380 is highly expressed in gl3-sst trichomes, while TRY expression is greatly reduced. The mutation in gl3-sst causes a reduced interaction between the GL1 and GL3 proteins and results in fewer leaf trichomes that develop in clusters. The differential expression of TRY and At1g01380 in this mutant can be used to explain how its altered trichome pattern in gl3-sst [corrected] is generated.

  9. Terpene biosynthesis in glandular trichomes of hop.

    PubMed

    Wang, Guodong; Tian, Li; Aziz, Naveed; Broun, Pierre; Dai, Xinbin; He, Ji; King, Andrew; Zhao, Patrick X; Dixon, Richard A

    2008-11-01

    Hop (Humulus lupulus L. Cannabaceae) is an economically important crop for the brewing industry, where it is used to impart flavor and aroma to beer, and has also drawn attention in recent years due to its potential pharmaceutical applications. Essential oils (mono- and sesquiterpenes), bitter acids (prenylated polyketides), and prenylflavonoids are the primary phytochemical components that account for these traits, and all accumulate at high concentrations in glandular trichomes of hop cones. To understand the molecular basis for terpene accumulation in hop trichomes, a trichome cDNA library was constructed and 9,816 cleansed expressed sequence tag (EST) sequences were obtained from random sequencing of 16,152 cDNA clones. The ESTs were assembled into 3,619 unigenes (1,101 contigs and 2,518 singletons). Putative functions were assigned to the unigenes based on their homology to annotated sequences in the GenBank database. Two mono- and two sesquiterpene synthases identified from the EST collection were expressed in Escherichia coli. Hop MONOTERPENE SYNTHASE2 formed the linear monterpene myrcene from geranyl pyrophosphate, whereas hop SESQUITERPENE SYNTHASE1 (HlSTS1) formed both caryophyllene and humulene from farnesyl pyrophosphate. Together, these enzymes account for the production of the major terpene constituents of the hop trichomes. HlSTS2 formed the minor sesquiterpene constituent germacrene A, which was converted to beta-elemene on chromatography at elevated temperature. We discuss potential functions for other genes expressed at high levels in developing hop trichomes.

  10. Transcriptome profiling reveals roles of meristem regulators and polarity genes during fruit trichome development in cucumber (Cucumis sativus L.).

    PubMed

    Chen, Chunhua; Liu, Meiling; Jiang, Li; Liu, Xiaofeng; Zhao, Jianyu; Yan, Shuangshuang; Yang, Sen; Ren, Huazhong; Liu, Renyi; Zhang, Xiaolan

    2014-09-01

    Trichomes are epidermal hair-like structures that function in plant defence against biotic and abiotic stresses. Extensive studies have been performed on foliar trichomes development in Arabidopsis and tomato, but the molecular mechanism of fruit trichome formation remains elusive. Cucumber fruit is covered with trichomes (spines) that directly affect the appearance and quality of cucumber products. Here, we characterized the fruit spine development in wild-type (WT) cucumber and a spontaneous mutant, tiny branched hair (tbh). Our data showed that the cucumber trichome was multicellular and non-glandular, with malformed organelles and no endoreduplication. Fruit spine development was generally homogenous and marked by a rapid base expansion stage. Trichomes in the tbh mutant were tiny and branched, with increased density and aberrant cell shape. Transcriptome profiling indicated that meristem-related genes were highly enriched in the upregulated genes in the tbh versus the WT, as well as in WT spines after versus before base expansion, and that polarity regulators were greatly induced during spine base expansion. Quantitative reverse transcription PCR and in situ hybridization confirmed the differential expression of CUP-SHAPED COTYLEDON3 (CUC3) and SHOOT MERISTEMLESS (STM) during spine development. Therefore, cucumber trichomes are morphologically different from those of Arabidopsis and tomato, and their development may be regulated by a distinct pathway involving meristem genes and polarity regulators.

  11. Transcriptome profiling of trichome-less reveals genes associated with multicellular trichome development in Cucumis sativus.

    PubMed

    Zhao, Jun-Long; Wang, Yun-Li; Yao, Dan-Qing; Zhu, Wen-Ying; Chen, Long; He, Huan-Le; Pan, Jun-Song; Cai, Run

    2015-10-01

    Trichomes on plants, similar to fine hairs on animal and human bodies, play important roles in plant survival and development. They also represent a useful model for the study of cell differentiation. Although the regulatory gene network of unicellular trichome development in Arabidopsis thaliana has been well studied, the genes that regulate multicellular trichome development remain unclear. We confirmed that Cucumis sativus (cucumber) trichomes are multicellular and unbranched, but identified a spontaneous mutant, trichome-less (tril), which presented a completely glabrous phenotype. We compared the transcriptome profilings of the tril mutant and wild type using the Illumina HiSeq 2000 sequencing technology. A total of 991 genes exhibited differential expression: 518 were up-regulated and 473 were down-regulated. We further identified 62 differentially expressed genes that encoded crucial transcription factors and were subdivided into seven categories: homeodomain, MADS, MYB, and WRKY domains, ethylene-responsive, zinc finger, and other transcription factor genes. We further analyzed the tissue-expression profiles of two candidate genes, GLABRA2-like and ATHB51-like, using qRT-PCR and found that these two genes were specifically expressed in the epidermis and trichomes, respectively. These results and the tril mutant provide useful tools to study the molecular networks associated with multicellular trichome development.

  12. Multicellular Secretory Trichome Development on Soybean and Related Glycine Gynoecia

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Multicellular glandular trichomes form on gynoecia of wild Glycine annual species, annual soybean cultivars, and wild perennial species. These trichomes occur on gynoecia of annual taxa from ovary base to style base, and along style of perennial species. Trichomes form at least two days prior to ant...

  13. Comparative functional genomic analysis of Solanum glandular trichome types.

    PubMed

    McDowell, Eric T; Kapteyn, Jeremy; Schmidt, Adam; Li, Chao; Kang, Jin-Ho; Descour, Anne; Shi, Feng; Larson, Matthew; Schilmiller, Anthony; An, Lingling; Jones, A Daniel; Pichersky, Eran; Soderlund, Carol A; Gang, David R

    2011-01-01

    Glandular trichomes play important roles in protecting plants from biotic attack by producing defensive compounds. We investigated the metabolic profiles and transcriptomes to characterize the differences between different glandular trichome types in several domesticated and wild Solanum species: Solanum lycopersicum (glandular trichome types 1, 6, and 7), Solanum habrochaites (types 1, 4, and 6), Solanum pennellii (types 4 and 6), Solanum arcanum (type 6), and Solanum pimpinellifolium (type 6). Substantial chemical differences in and between Solanum species and glandular trichome types are likely determined by the regulation of metabolism at several levels. Comparison of S. habrochaites type 1 and 4 glandular trichomes revealed few differences in chemical content or transcript abundance, leading to the conclusion that these two glandular trichome types are the same and differ perhaps only in stalk length. The observation that all of the other species examined here contain either type 1 or 4 trichomes (not both) supports the conclusion that these two trichome types are the same. Most differences in metabolites between type 1 and 4 glands on the one hand and type 6 glands on the other hand are quantitative but not qualitative. Several glandular trichome types express genes associated with photosynthesis and carbon fixation, indicating that some carbon destined for specialized metabolism is likely fixed within the trichome secretory cells. Finally, Solanum type 7 glandular trichomes do not appear to be involved in the biosynthesis and storage of specialized metabolites and thus likely serve another unknown function, perhaps as the site of the synthesis of protease inhibitors.

  14. Glandular Trichomes and Essential Oil of Thymus quinquecostatus

    PubMed Central

    Jia, Ping; Liu, Hanzhu; Gao, Ting; Xin, Hua

    2013-01-01

    The distribution and types of glandular trichomes and essential oil chemistry of Thymus quinquecostatus were studied. The glandular trichomes are distributed on the surface of stem, leaf, rachis, calyx and corolla, except petiole, pistil and stamen. Three morphologically distinct types of glandular trichomes are described. Peltate trichomes, consisting of a basal cell, a stalk cell and a 12-celled head, are distributed on the stem, leaf, corolla and outer side of calyx. Capitate trichomes, consisting of a unicellular base, a 1–2-celled stalk and a unicellular head, are distributed more diffusely than peltate ones, existing on stem, leaf, rachis and calyx. Digitiform trichomes are just distributed on the outer side of corolla, consisting of 1 basal cell, 3 stalk cells and 1 head cell. All three types of glandular trichomes can secrete essential oil, and in small capitate trichomes of rachis, all peltate trichomes and digitiform trichomes, essential oil is stored in a large subcuticular space, released by cuticle rupture, whereas, in other capitate trichomes, essential oil crosses the thin cuticle. The essential oil of T. quinquecostatus is yellow, and its content is highest in the growth period. 68 constituents were identified in the essential oils. The main constituent is linalool. PMID:24250266

  15. TCS1, a Microtubule-Binding Protein, Interacts with KCBP/ZWICHEL to Regulate Trichome Cell Shape in Arabidopsis thaliana

    PubMed Central

    Tian, Juan; Wang, Xiaohong; Mao, Tonglin; Yuan, Ming; Li, Yunhai

    2016-01-01

    How cell shape is controlled is a fundamental question in developmental biology, but the genetic and molecular mechanisms that determine cell shape are largely unknown. Arabidopsis trichomes have been used as a good model system to investigate cell shape at the single-cell level. Here we describe the trichome cell shape 1 (tcs1) mutants with the reduced trichome branch number in Arabidopsis. TCS1 encodes a coiled-coil domain-containing protein. Pharmacological analyses and observations of microtubule dynamics show that TCS1 influences the stability of microtubules. Biochemical analyses and live-cell imaging indicate that TCS1 binds to microtubules and promotes the assembly of microtubules. Further results reveal that TCS1 physically associates with KCBP/ZWICHEL, a microtubule motor involved in the regulation of trichome branch number. Genetic analyses indicate that kcbp/zwi is epistatic to tcs1 with respect to trichome branch number. Thus, our findings define a novel genetic and molecular mechanism by which TCS1 interacts with KCBP to regulate trichome cell shape by influencing the stability of microtubules. PMID:27768706

  16. Taxonomic significance of trichomes micromorphology in cucurbits.

    PubMed

    Ali, Mohammad Ajmal; Al-Hemaid, Fahad M A

    2011-01-01

    Studies on trichomes micromorphology using Scanning Electron Microscope (SEM) were undertaken in 23 species with one variety under 13 genera of the family Cucurbitaceae (viz., Benincasa hispida (Thunb.) Cogn., Citrullus lanatus (Thunb.) Matsum. & Nakai, Cucumis melo var. agrestis Naudin, Cucumis sativus L., Diplocyclos palmatus (L.) C. Jeffrey, Edgaria dargeelingensis C.B. Clarke, Gynostemma burmanicum King ex Chakr., Gynostemma pentaphyllum (Thunb.) Makino, Gynostemma pubescens (Gagnep.) C.Y. Wu, Hemsleya dipterygia Kuang & A.M. Lu, Lagenaria siceraria (Molina) Standl., Luffa acutangula (L.) Roxb., Luffa cylindrica M. Roem., Luffa echinata Roxb., Melothria heterophylla (Lour.) Cogn., Melothria leucocarpa (Blume) Cogn., Melothria maderspatana (L.) Cogn., Sechium edule (Jacq.) Sw., Thladiantha cordifolia (Blume) Cogn., Trichosanthes cucumerina L., T. cucumerina var. anguina (L.) Haines, Trichosanthes dioica Roxb., Trichosanthes lepiniana (Naudin) Cogn. and T. tricuspidata Lour.). The trichomes in the family Cucurbitaceae vary from unicellular to multicellular, conical to elongated, smooth to ridges, with or without flattened disk at base and cyctolithic appendages, thin to thick walled, curved at apices to blunt. Trichomes micromorphology in the family Cucurbitaceae was found significant taxonomically.

  17. Micro-trichome as a class I homeodomain-leucine zipper gene regulates multicellular trichome development in Cucumis sativus.

    PubMed

    Zhao, Jun-Long; Pan, Jun-Song; Guan, Yuan; Zhang, Wei-Wei; Bie, Bei-Bei; Wang, Yun-Li; He, Huan-Le; Lian, Hong-Li; Cai, Run

    2015-11-01

    Plant trichomes serve as a highly suitable model for investigating cell differentiation at the single-cell level. The regulatory genes involved in unicellular trichome development in Arabidopsis thaliana have been intensively studied, but genes regulating multicellular trichome development in plants remain unclear. Here, we characterized Cucumis sativus (cucumber) trichomes as representative multicellular and unbranched structures, and identified Micro-trichome (Mict), using map-based cloning in an F2 segregating population of 7,936 individuals generated from a spontaneous mict mutant. In mict plants, trichomes in both leaves and fruits, are small, poorly developed, and denser than in the wild type. Sequence analysis revealed that a 2,649-bp genomic deletion, spanning the first and second exons, occurred in a plant-specific class I homeodomain-leucine zipper gene. Tissue-specific expression analysis indicated that Mict is strongly expressed in the trichome cells. Transcriptome profiling identified potential targets of Mict including putative homologs of genes known in other systems to regulate trichome development, meristem determinacy, and hormone responsiveness. Phylogenic analysis charted the relationships among putative homologs in angiosperms. Our paper represents initial steps toward understanding the development of multicellular trichomes.

  18. Zinc Finger Protein5 Is Required for the Control of Trichome Initiation by Acting Upstream of Zinc Finger Protein8 in Arabidopsis1[C][W][OA

    PubMed Central

    Zhou, Zhongjing; An, Lijun; Sun, Lili; Zhu, Shuijin; Xi, Wanyan; Broun, Pierre; Yu, Hao; Gan, Yinbo

    2011-01-01

    Arabidopsis (Arabidopsis thaliana) trichome development is a model system for studying cell development, cell differentiation, and the cell cycle. Our previous studies have shown that the GLABROUS INFLORESCENCE STEMS (GIS) family genes, GIS, GIS2, and ZINC FINGER PROTEIN8 (ZFP8), control shoot maturation and epidermal cell fate by integrating gibberellins (GAs) and cytokinin signaling in Arabidopsis. Here, we show that a new C2H2 zinc finger protein, ZFP5, plays an important role in controlling trichome cell development through GA signaling. Overexpression of ZFP5 results in the formation of ectopic trichomes on carpels and other inflorescence organs. zfp5 loss-of-function mutants exhibit a reduced number of trichomes on sepals, cauline leaves, paraclades, and main inflorescence stems in comparison with wild-type plants. More importantly, it is found that ZFP5 mediates the regulation of trichome initiation by GAs. These results are consistent with ZFP5 expression patterns and the regional influence of GA on trichome initiation. The molecular analyses suggest that ZFP5 functions upstream of GIS, GIS2, ZFP8, and the key trichome initiation regulators GLABROUS1 (GL1) and GL3. Using a steroid-inducible activation of ZFP5 and chromatin immunoprecipitation experiments, we further demonstrate that ZFP8 is the direct target of ZFP5 in controlling epidermal cell differentiation. PMID:21803862

  19. Are plant trichomes harmful to predatory insects and mites?

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Plants may use epidermal hairs (trichomes) to defend themselves from attack by herbivores. Predatory arthropods may serve as biocontrol agents against herbivores. Whether or not plant trichomes work in concert with predators is undocumented in many cases. We reviewed the peer-reviewed literature ...

  20. Plant trichomes have mixed impacts on predatory insects

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In response to our review article on the role of plant trichomes on insect predators (Riddick & Simmons 2014), Krimmel (2014) acknowledged the challenges that are faced in bringing together the varied publications on the subject of impact of plant trichomes on predatory insects. He also suggested t...

  1. Trichomes as sensors: detecting activity on the leaf surface.

    PubMed

    Tooker, John F; Peiffer, Michelle; Luthe, Dawn S; Felton, Gary W

    2010-01-01

    The dramatic movements of some carnivorous plants species are triggered by sensory structures derived from trichomes. While unusual plant species such as the Venus fly trap and sundews may be expected to have elaborate sensors to capture their insect prey, more modest plant species might not be expected to have similar sensory capabilities. Our recent work, however, has revealed that glandular trichomes on tomato (Solanum lycopersicum) appear to have a function similar to trigger hairs of carnivorous species, acting as "early warning" sensors. Using a combination of behavioral, molecular, and biochemical techniques, we determined that caterpillars, moths and mechanical disruption upregulate signaling molecules and defensive genes found in glandular trichomes. Importantly, we discovered that plants whose trichomes have been broken respond more vigorously when their defenses were induced. Taken together, our results suggest that glandular trichomes can act as sensors that detect activity on the leaf surface, and ready plants for herbivore attack.

  2. New Approaches for Studying and Exploiting an Old Protuberance, the Plant Trichome

    PubMed Central

    WAGNER, G. J.; WANG, E.; SHEPHERD, R. W.

    2004-01-01

    • Background and aims Much recent study of plant trichomes has focused on various aspects of glandular secreting trichomes (GSTs) and differentiation of simple trichomes. This Botanical Briefing will highlight: ‐ research on various aspects of, and manipulation of glandular secreting trichomes; ‐ molecular aspects of the differentiation and development of simple trichomes of arabidopsis and cotton; ‐ how methods for manipulation of model systems used in the above work can be applied to expand our understanding of less studied surface structures of plants. • Scope The Briefing will cover: ‐ established and suggested roles of simple and glandular secreting trichomes; ‐ recent results regarding solute and ion movement in trichomes; ‐ methods for isolating trichomes; ‐ recent studies of trichome differentiation and development; ‐ attempts to modify metabolism in secreting trichomes; ‐ efforts to exploit trichomes for commercial and agronomic purposes. PMID:14678941

  3. Glycerol metabolism promotes biofilm formation by Pseudomonas aeruginosa.

    PubMed

    Scoffield, Jessica; Silo-Suh, Laura

    2016-08-01

    Pseudomonas aeruginosa causes persistent infections in the airways of cystic fibrosis (CF) patients. Airway sputum contains various host-derived nutrients that can be utilized by P. aeruginosa, including phosphotidylcholine, a major component of host cell membranes. Phosphotidylcholine can be degraded by P. aeruginosa to glycerol and fatty acids to increase the availability of glycerol in the CF lung. In this study, we explored the role that glycerol metabolism plays in biofilm formation by P. aeruginosa. We report that glycerol metabolism promotes biofilm formation by both a chronic CF isolate (FRD1) and a wound isolate (PAO1) of P. aeruginosa. Moreover, loss of the GlpR regulator, which represses the expression of genes involved in glycerol metabolism, enhances biofilm formation in FRD1 through the upregulation of Pel polysaccharide. Taken together, our results suggest that glycerol metabolism may be a key factor that contributes to P. aeruginosa persistence by promoting biofilm formation.

  4. Trichoderma species form endophytic associations within Theobroma cacao trichomes.

    PubMed

    Bailey, Bryan A; Strem, Mary D; Wood, Delilah

    2009-12-01

    Trichoderma species are usually considered soil organisms that colonize plant roots, sometimes forming a symbiotic relationship. Recent studies demonstrate that Trichoderma species are also capable of colonizing the above ground tissues of Theobroma cacao (cacao) in what has been characterized as an endophytic relationship. Trichoderma species can be re-isolated from surface sterilized cacao stem tissue, including the bark and xylem, the apical meristem, and to a lesser degree from leaves. SEM analysis of cacao stems colonized by strains of four Trichoderma species (Trichoderma ovalisporum-DIS 70a, Trichoderma hamatum-DIS 219b, Trichoderma koningiopsis-DIS 172ai, or Trichoderma harzianum-DIS 219f) showed a preference for surface colonization of glandular trichomes versus non-glandular trichomes. The Trichoderma strains colonized the glandular trichome tips and formed swellings resembling appresoria. Hyphae were observed emerging from the glandular trichomes on surface sterilized stems from cacao seedlings that had been inoculated with each of the four Trichoderma strains. Fungal hyphae were observed under the microscope emerging from the trichomes as soon as 6h after their isolation from surface sterilized cacao seedling stems. Hyphae were also observed, in some cases, emerging from stalk cells opposite the trichome head. Repeated single trichome/hyphae isolations verified that the emerging hyphae were the Trichoderma strains with which the cacao seedlings had been inoculated. Strains of four Trichoderma species were able to enter glandular trichomes during the colonization of cacao stems where they survived surface sterilization and could be re-isolated. The penetration of cacao trichomes may provide the entry point for Trichoderma species into the cacao stem allowing systemic colonization of this tissue.

  5. The flavonoid biosynthetic enzyme chalcone isomerase modulates terpenoid production in glandular trichomes of tomato.

    PubMed

    Kang, Jin-Ho; McRoberts, John; Shi, Feng; Moreno, Javier E; Jones, A Daniel; Howe, Gregg A

    2014-03-01

    Flavonoids and terpenoids are derived from distinct metabolic pathways but nevertheless serve complementary roles in mediating plant interactions with the environment. Here, we show that glandular trichomes of the anthocyanin free (af) mutant of cultivated tomato (Solanum lycopersicum) fail to accumulate both flavonoids and terpenoids. This pleiotropic metabolic deficiency was associated with loss of resistance to native populations of coleopteran herbivores under field conditions. We demonstrate that Af encodes an isoform (SlCHI1) of the flavonoid biosynthetic enzyme chalcone isomerase (CHI), which catalyzes the conversion of naringenin chalcone to naringenin and is strictly required for flavonoid production in multiple tissues of tomato. Expression of the wild-type SlCHI1 gene from its native promoter complemented the anthocyanin deficiency in af. Unexpectedly, the SlCHI1 transgene also complemented the defect in terpenoid production in glandular trichomes. Our results establish a key role for SlCHI1 in flavonoid production in tomato and reveal a link between CHI1 and terpenoid production. Metabolic coordination of the flavonoid and terpenoid pathways may serve to optimize the function of trichome glands in dynamic environments.

  6. Morphology, Structure, and Ontogeny of Trichomes of the Grape Genus (Vitis, Vitaceae)

    PubMed Central

    Ma, Zhi-Yao; Wen, Jun; Ickert-Bond, Stefanie M.; Chen, Long-Qing; Liu, Xiu-Qun

    2016-01-01

    Trichomes are widely distributed on surfaces of different organs in the grape genus Vitis and are of taxonomic utility. To explore the morphology, structure and ontogeny of Vitis trichomes, we investigated the diversity and distribution of trichomes in 34 species of Vitis. Two main types of trichomes in Vitis are documented: non-glandular and glandular. Within non-glandular trichomes, ribbon and simple trichomes are found on different vegetative plant organs. The morphology and ontogeny of these types of trichomes are further examined with light microscopy and scanning electron microscopy. The ultrastructure of the glandular trichomes is explored with transmission electron microscopy. The ribbon trichomes are twisted, greatly elongated and unicellular, and this trichome type may be a morphological synapomorphy of Vitis and its closest tropical relative Ampelocissus and Pterisanthes in Vitaceae. The simple trichomes are documented in most species sampled in the genus. The glandular trichomes are multicellular, non-vascularized and composed of both epidermis and subjacent layers. We show that prickles occurring along the stems and petioles of Vitis davidii are modified glandular trichomes. We observed that glandular trichomes of V. romanetii secrete mucilage and volatile substances which trap insectes on the glands. Transmission electron microscopy indicates that metabolic products accumulate in vacuoles, the cytoplasm and intercellular spaces. We infer that glandular trichomes and young prickles are involved in the secretion of these metabolic products and the intercellular spaces may be the places of temporary storage of these secretions. PMID:27252720

  7. Trichomes control flower bud shape by linking together young petals.

    PubMed

    Tan, Jiafu; Walford, Sally-Anne; Dennis, Elizabeth S; Llewellyn, Danny

    2016-06-20

    Trichomes are widespread in plants and develop from surface cells on different tissues(1). They have many forms and functions, from defensive spines to physical barriers that trap layers of air to insulate against desiccation, but there is growing evidence that trichomes can also have developmental roles in regulating flower structure(2,3). We report here that the trichomes on petals of cotton, Gossypium hirsutum L., are essential for correct flower bud shape through a mechanical entanglement of the trichomes on adjacent petals that anchor the edges to counter the opposing force generated by asymmetric expansion of overlapping petals. Silencing a master regulator of petal trichomes, GhMYB-MIXTA-Like10 (GhMYBML10), by RNA interference (RNAi) suppressed petal trichome growth and resulted in flower buds forming into abnormal corkscrew shapes that exposed developing anthers and stigmas to desiccation damage. Artificially gluing petal edges together could partially restore correct bud shape and fertility. Such petal 'Velcro' is present in other Malvaceae and perhaps more broadly in other plant families, although it is not ubiquitous. This mechanism for physical association between separate organs to regulate flower shape and function is different from the usual organ shape control(4) exerted through cell-to-cell communication and differential cell expansion within floral tissues(5,6).

  8. Vanadium promotes hydroxyl radical formation by activated human neutrophils.

    PubMed

    Fickl, Heidi; Theron, Annette J; Grimmer, Heidi; Oommen, Joyce; Ramafi, Grace J; Steel, Helen C; Visser, Susanna S; Anderson, Ronald

    2006-01-01

    This study was undertaken to investigate the effects of vanadium in the +2, +3, +4, and +5 valence states on superoxide generation, myeloperoxidase (MPO) activity, and hydroxyl radical formation by activated human neutrophils in vitro, using lucigenin-enhanced chemiluminescence (LECL), autoiodination, and electron spin resonance with 5,5-dimethyl-l-pyrroline N-oxide as the spin trap, respectively. At concentrations of up to 25 microM, vanadium, in the four different valence states used, did not affect the LECL responses of neutrophils activated with either the chemoattractant, N-formyl-l-methionyl-l-leucyl-l-phenylalanine (1 microM), or the phorbol ester, phorbol 12-myristate 12-acetate (25 ng/ml). However, exposure to vanadium in the +2, +3, and +4, but not the +5, valence states was accompanied by significant augmentation of hydroxyl radical formation by activated neutrophils and attenuation of MPO-mediated iodination. With respect to hydroxyl radical formation, similar effects were observed using cell-free systems containing either hydrogen peroxide (100 microM) or xanthine/xanthine oxidase together with vanadium (+2, +3, +4), while the activity of purified MPO was inhibited by the metal in these valence states. These results demonstrate that vanadium in the +2, +3, and +4 valence states interacts prooxidatively with human neutrophils, competing effectively with MPO for hydrogen peroxide to promote formation of the highly toxic hydroxyl radical.

  9. Why stellar feedback promotes disc formation in simulated galaxies

    NASA Astrophysics Data System (ADS)

    Übler, Hannah; Naab, Thorsten; Oser, Ludwig; Aumer, Michael; Sales, Laura V.; White, Simon D. M.

    2014-09-01

    We study how feedback influences baryon infall on to galaxies using cosmological, zoom-in simulations of haloes with present mass Mvir = 6.9 × 1011 to 1.7 × 1012 M⊙. Starting at z = 4 from identical initial conditions, implementations of weak and strong stellar feedback produce bulge- and disc-dominated galaxies, respectively. Strong feedback favours disc formation: (1) because conversion of gas into stars is suppressed at early times, as required by abundance matching arguments, resulting in flat star formation histories and higher gas fractions; (2) because 50 per cent of the stars form in situ from recycled disc gas with angular momentum only weakly related to that of the z = 0 dark halo; (3) because late-time gas accretion is typically an order of magnitude stronger and has higher specific angular momentum, with recycled gas dominating over primordial infall; (4) because 25-30 per cent of the total accreted gas is ejected entirely before z ˜ 1, removing primarily low angular momentum material which enriches the nearby intergalactic medium. Most recycled gas roughly conserves its angular momentum, but material ejected for long times and to large radii can gain significant angular momentum before re-accretion. These processes lower galaxy formation efficiency in addition to promoting disc formation.

  10. Multi-protein Delivery by Nanodiamonds Promotes Bone Formation

    PubMed Central

    Moore, L.; Gatica, M.; Kim, H.; Osawa, E.; Ho, D.

    2013-01-01

    Bone morphogenetic proteins (BMPs) are well-studied regulators of cartilage and bone development that have been Food and Drug Administration (FDA)-approved for the promotion of bone formation in certain procedures. BMPs are seeing more use in oral and maxillofacial surgeries because of recent FDA approval of InFUSE® for sinus augmentation and localized alveolar ridge augmentation. However, the utility of BMPs in medical and dental applications is limited by the delivery method. Currently, BMPs are delivered to the surgical site by the implantation of bulky collagen sponges. Here we evaluate the potential of detonation nanodiamonds (NDs) as a delivery vehicle for BMP-2 and basic fibroblast growth factor (bFGF). Nanodiamonds are biocompatible, 4- to 5-nm carbon nanoparticles that have previously been used to deliver a wide variety of molecules, including proteins and peptides. We find that both BMP-2 and bFGF are readily loaded onto NDs by physisorption, forming a stable colloidal solution, and are triggered to release in slightly acidic conditions. Simultaneous delivery of BMP-2 and bFGF by ND induces differentiation and proliferation in osteoblast progenitor cells. Overall, we find that NDs provide an effective injectable alternative for the delivery of BMP-2 and bFGF to promote bone formation. PMID:24045646

  11. Multi-protein delivery by nanodiamonds promotes bone formation.

    PubMed

    Moore, L; Gatica, M; Kim, H; Osawa, E; Ho, D

    2013-11-01

    Bone morphogenetic proteins (BMPs) are well-studied regulators of cartilage and bone development that have been Food and Drug Administration (FDA)-approved for the promotion of bone formation in certain procedures. BMPs are seeing more use in oral and maxillofacial surgeries because of recent FDA approval of InFUSE(®) for sinus augmentation and localized alveolar ridge augmentation. However, the utility of BMPs in medical and dental applications is limited by the delivery method. Currently, BMPs are delivered to the surgical site by the implantation of bulky collagen sponges. Here we evaluate the potential of detonation nanodiamonds (NDs) as a delivery vehicle for BMP-2 and basic fibroblast growth factor (bFGF). Nanodiamonds are biocompatible, 4- to 5-nm carbon nanoparticles that have previously been used to deliver a wide variety of molecules, including proteins and peptides. We find that both BMP-2 and bFGF are readily loaded onto NDs by physisorption, forming a stable colloidal solution, and are triggered to release in slightly acidic conditions. Simultaneous delivery of BMP-2 and bFGF by ND induces differentiation and proliferation in osteoblast progenitor cells. Overall, we find that NDs provide an effective injectable alternative for the delivery of BMP-2 and bFGF to promote bone formation.

  12. Dirt and diarrhoea: formative research in hygiene promotion programmes.

    PubMed

    Curtis, V; Kanki, B; Cousens, S; Sanou, A; Diallo, I; Mertens, T

    1997-06-01

    Investment in the promotion of better hygiene for the prevention of diarrhoeal diseases and as a component of water and sanitation programmes is increasing. Before designing programmes capable of sustainably modifying hygiene behaviour in large populations, valid answers to a number of basic questions concerning the site and the intended beneficiaries have to be obtained. Such questions include 'what practices favour the transmission of enteric pathogens?', 'what advantages will be perceived by those who adopt safe practices?' and 'what channels of communication are currently employed by the target population?' A study of hygiene and diarrhoea in Bobo-Dioulasso, Burkina Faso, used a mixture of methods to address such questions. This paper draws on that experience to propose a plan of preliminary research using a variety of techniques which could be implemented over a period of a few months by planners of hygiene promotion programmes. The techniques discussed include structured observation, focus group discussions and behavioural trials. Modest investment in such systematic formative research with clear and limited goals is likely to be repaid many times over in the increased effectiveness of hygiene promotion programmes.

  13. Localization of Salvinorin A and Related Compounds in Glandular Trichomes of the Psychoactive Sage, Salvia divinorum

    PubMed Central

    SIEBERT, DANIEL J.

    2004-01-01

    • Background and Aims Salvia divinorum produces several closely related neoclerodane diterpenes. The most abundant of these, salvinorin A, is responsible for the psychoactive properties of the plant. To determine where these compounds occur in the plant, various organs, tissues and glandular secretions were chemically analysed. A microscopic survey of the S. divinorum plant was performed to examine the various types of trichomes present and to determine their distribution. • Methods Chemical analyses were performed using thin layer chromatographic and histochemical techniques. Trichomes were examined using conventional light microscopy and scanning electron microscopy. • Key Results It was found that neoclerodane diterpenes are secreted as components of a resin that accumulates in peltate glandular trichomes, specifically in the subcuticular space that exists between the trichome head cells and the cuticle that encloses them. Four main types of trichomes were observed: peltate glandular trichomes, short‐stalked capitate glandular trichomes, long‐stalked capitate glandular trichomes and non‐glandular trichomes. Their morphology and distribution is described. Peltate glandular trichomes were only found on the abaxial surfaces of the leaves, stems, rachises, bracts, pedicles and calyces. This was consistent with chemical analyses, which showed the presence of neoclerodane diterpenes in these organs, but not in parts of the plant where peltate glandular trichomes are absent. • Conclusions Salvinorin A and related compounds are secreted as components of a complex resin that accumulates in the subcuticular space of peltate glandular trichomes. PMID:15087301

  14. Differential expression of trichomes on the leaves of upland cotton (Gossypium hirsutum L)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This study evaluated and quantified trichome diversity on various cotton lines. Leaf trichomes were evaluated with scanning electron and light microscopy on fixed and live samples. Observations indicated that trichome complexity and quantity increased on the mainstem leaves as sampling progressed ...

  15. Microglia promote learning-dependent synapse formation through BDNF

    PubMed Central

    Parkhurst, Christopher N.; Yang, Guang; Ninan, Ipe; Savas, Jeffrey N.; Yates, John R.; Lafaille, Juan J.; Hempstead, Barbara L.; Littman, Dan R.; Gan, Wen-Biao

    2014-01-01

    SUMMARY Microglia are the resident macrophages of the central nervous system and their functions have been extensively studied in various brain pathologies. The physiological roles of microglia in brain plasticity and function, however, remain unclear. To address this question, we generated CX3CR1CreER mice expressing tamoxifen-inducible Cre recombinase that allow for specific manipulation of gene function in microglia. Using CX3CR1CreER to drive diphtheria toxin receptor expression in microglia, we found that microglia could be specifically depleted from the brain upon diphtheria toxin administration. Mice depleted of microglia show deficits in multiple learning tasks and a significant reduction in motor learning-dependent synapse formation. Furthermore, Cre-dependent removal of brain-derived neurotrophic factor (BDNF) from microglia largely recapitulated the effects of microglia depletion. Microglial BDNF increases neuronal TrkB phosphorylation, a key mediator of synaptic plasticity. Together, our findings reveal important physiological functions of microglia in learning and memory by promoting learning-related synapse formation through BDNF signaling. PMID:24360280

  16. Comparative Functional Genomic Analysis of Solanum Glandular Trichome Types1[W][OA

    PubMed Central

    McDowell, Eric T.; Kapteyn, Jeremy; Schmidt, Adam; Li, Chao; Kang, Jin-Ho; Descour, Anne; Shi, Feng; Larson, Matthew; Schilmiller, Anthony; An, Lingling; Jones, A. Daniel; Pichersky, Eran; Soderlund, Carol A.; Gang, David R.

    2011-01-01

    Glandular trichomes play important roles in protecting plants from biotic attack by producing defensive compounds. We investigated the metabolic profiles and transcriptomes to characterize the differences between different glandular trichome types in several domesticated and wild Solanum species: Solanum lycopersicum (glandular trichome types 1, 6, and 7), Solanum habrochaites (types 1, 4, and 6), Solanum pennellii (types 4 and 6), Solanum arcanum (type 6), and Solanum pimpinellifolium (type 6). Substantial chemical differences in and between Solanum species and glandular trichome types are likely determined by the regulation of metabolism at several levels. Comparison of S. habrochaites type 1 and 4 glandular trichomes revealed few differences in chemical content or transcript abundance, leading to the conclusion that these two glandular trichome types are the same and differ perhaps only in stalk length. The observation that all of the other species examined here contain either type 1 or 4 trichomes (not both) supports the conclusion that these two trichome types are the same. Most differences in metabolites between type 1 and 4 glands on the one hand and type 6 glands on the other hand are quantitative but not qualitative. Several glandular trichome types express genes associated with photosynthesis and carbon fixation, indicating that some carbon destined for specialized metabolism is likely fixed within the trichome secretory cells. Finally, Solanum type 7 glandular trichomes do not appear to be involved in the biosynthesis and storage of specialized metabolites and thus likely serve another unknown function, perhaps as the site of the synthesis of protease inhibitors. PMID:21098679

  17. Purification and Characterization of Polyphenol Oxidase from Glandular Trichomes of Solanum berthaultii.

    PubMed

    Kowalski, S P; Eannetta, N T; Hirzel, A T; Steffens, J C

    1992-10-01

    Type A glandular trichomes of the wild potato (Solanum berthaultii Hawkes) entrap insects by rapidly polymerizing the trichome contents after breakage by insect contact. Polymerization of trichome exudate appears to be driven by a soluble polyphenol oxidase (PPO). PPO constitutes up to 70% of the protein in individually collected trichomes and reaches a concentration approaching 200 mum in these organs. Trichome PPO has been purified and shown to be a monomeric copper metalloprotein with an isoelectric point of 5.5, possessing only o-diphenol oxygen oxido-reductase activity, and is larger than most other reported PPOs, with relative molecular weight of 59,000. Chlorogenic and caffeic acid were the most readily oxidized of 14 phenolic substrates tested. Polyclonal antibodies raised against the relative molecular weight 59,000 S. berthaultii trichome PPO were used to show that S. tuberosum L. trichomes express low levels of a cross-reactive protein that lacks detectable PPO activity.

  18. Vibration acceleration promotes bone formation in rodent models

    PubMed Central

    Uchida, Ryohei; Nakata, Ken; Kawano, Fuminori; Yonetani, Yasukazu; Ogasawara, Issei; Nakai, Naoya; Mae, Tatsuo; Matsuo, Tomohiko; Tachibana, Yuta; Yokoi, Hiroyuki; Yoshikawa, Hideki

    2017-01-01

    centrifuge acceleration group had no significant difference compared those in control-CA group. Union rate and BV in the low-magnitude group of RFH model were also significantly higher than those in the other groups (Union rate: 60% v.s. 0% in the high-magnitude group and 10% in the control-VA group, BV: 0.69±0.30mm3 v.s. 0.15±0.09mm3 in high-magnitude group and 0.22±0.17mm3 in control-VA group). BV/TV in the low-magnitude group of RFH model was significantly higher than that in control-VA group (59.4±14.9% v.s. 35.8±13.5%). On the other hand, radiographic union rate (10% in centrifuge acceleration group v.s. 20% in control-CA group) and micro-CT parameters in RFH model were not significantly different between two groups in the constant acceleration studies. Radiographic images of non-union rib fractures showed cartilage at the fracture site and poor new bone formation, whereas union samples showed only new bone. In conclusion, low-magnitude vibration acceleration promoted bone formation at the trunk in both BMP-induced ectopic bone formation and rib fracture healing models. However, the micro-CT parameters were not similar between two models, which suggested that there might be difference in the mechanism of effect by vibration between two models. PMID:28264058

  19. CHIP Regulates Osteoclast Formation through Promoting TRAF6 Protein Degradation

    PubMed Central

    Li, Shan; Shu, Bing; Zhang, Yanquan; Li, Jia; Guo, Junwei; Wang, Yinyin; Ren, Fangli; Xiao, Guozhi; Chang, Zhijie; Chen, Di

    2014-01-01

    Objective Carboxyl terminus of Hsp70-interacting protein (CHIP or STUB1) is an E3 ligase and regulates the stability of several proteins which are involved in tumor growth and metastasis. However, the role of CHIP in bone growth and bone remodeling in vivo has not been reported. The objective of this study is to investigate the role and mechanism of CHIP in regulation of bone mass and bone remodeling. Methods The bone phenotype of Chip−/− mice was examined by histology, histomorphometry and micro-CT analyses. The regulatory mechanism of CHIP on the degradation of TRAF6 and the inhibition of NF-κB signaling was examined by immunoprecipitation (IP), western blotting and luciferase reporter assays. Results In this study, we found that deletion of the Chip gene leads to osteopenic phenotype and increased osteoclast formation. We further found that TRAF6, as a novel substrate of CHIP, is up-regulated in Chip−/− osteoclasts. TRAF6 is critical for RANKL-induced osteoclastogenesis. TRAF6 is an adaptor protein which functions as an E3 ligase to regulate the activation of TAK1 and the I-κB kinase (IKK) and is a key regulator of NF-κB signaling. CHIP interacts with TRAF6 to promote TRAF6 ubiquitination and proteasome degradation. CHIP inhibits p65 nuclear translocation, leading to the repression of the TRAF6-mediated NF-κB transcription. Conclusion CHIP inhibits NF-κB signaling via promoting TRAF6 degradation and plays an important role in osteoclastogenesis and bone remodeling, suggesting that it may be a novel therapeutic target for the treatment of bone loss associated diseases. PMID:24578159

  20. Conduit speed limit promotes formation of explosive `super slugs'

    NASA Astrophysics Data System (ADS)

    Llewellin, E. W.; Burton, M. R.; Mader, H. M.; Polacci, M.

    2014-12-01

    Strombolian activity - characterized by discrete, impulsive explosions - is common at basaltic volcanoes. The explosions are caused by the arrival, at the volcanic vent, of over-pressured `slugs' of magmatic gas, which have ascended the volcanic conduit. We present data from large-scale analogue experiments which reveal a previously-undescribed process which promotes the formation of large, highly over-pressured `super slugs'. We propose that these large slugs may drive the most violent Strombolian eruptions. Experiments were performed at the Large Analogue Volcano Apparatus at Durham University, UK, which comprises a 13m tall, 24cm diameter transparent conduit, surmounting a reservoir of analogue magma held at constant pressure. We simulate a vesiculation event deep in the sub-volcanic plumbing system by fluxing gas into the reservoir. Magma analogues with a range of viscosities are used, giving slug Reynolds numbers in the range 16 to 140,000. At moderate-to-high Reynolds number, we find that the gas rapidly self-organizes to form a conduit-filling lead slug; this slug ascends the column slowly, at a velocity limited by the flux of the falling film of liquid around it. Trailing bubbles, which ascend through the wake of the lead slug, rise much more rapidly. As they catch and coalesce with the lead slug, it grows and becomes increasingly over-pressured. This mechanism causes large slugs to form rapidly and we propose that it underpins the formation of the very large slugs that are responsible for the most explosive strombolian eruptions.

  1. The Flavonoid Biosynthetic Enzyme Chalcone Isomerase Modulates Terpenoid Production in Glandular Trichomes of Tomato1[C][W][OPEN

    PubMed Central

    Kang, Jin-Ho; McRoberts, John; Shi, Feng; Moreno, Javier E.; Jones, A. Daniel; Howe, Gregg A.

    2014-01-01

    Flavonoids and terpenoids are derived from distinct metabolic pathways but nevertheless serve complementary roles in mediating plant interactions with the environment. Here, we show that glandular trichomes of the anthocyanin free (af) mutant of cultivated tomato (Solanum lycopersicum) fail to accumulate both flavonoids and terpenoids. This pleiotropic metabolic deficiency was associated with loss of resistance to native populations of coleopteran herbivores under field conditions. We demonstrate that Af encodes an isoform (SlCHI1) of the flavonoid biosynthetic enzyme chalcone isomerase (CHI), which catalyzes the conversion of naringenin chalcone to naringenin and is strictly required for flavonoid production in multiple tissues of tomato. Expression of the wild-type SlCHI1 gene from its native promoter complemented the anthocyanin deficiency in af. Unexpectedly, the SlCHI1 transgene also complemented the defect in terpenoid production in glandular trichomes. Our results establish a key role for SlCHI1 in flavonoid production in tomato and reveal a link between CHI1 and terpenoid production. Metabolic coordination of the flavonoid and terpenoid pathways may serve to optimize the function of trichome glands in dynamic environments. PMID:24424324

  2. Foliar trichomes, boundary layers, and gas exchange in 12 species of epiphytic Tillandsia (Bromeliaceae).

    PubMed

    Benz, Brett W; Martin, Craig E

    2006-04-01

    We examined the relationships between H2O and CO2 gas exchange parameters and leaf trichome cover in 12 species of Tillandsia that exhibit a wide range in trichome size and trichome cover. Previous investigations have hypothesized that trichomes function to enhance boundary layers around Tillandsioid leaves thereby buffering the evaporative demand of the atmosphere and retarding transpirational water loss. Data presented herein suggest that trichome-enhanced boundary layers have negligible effects on Tillandsia gas exchange, as indicated by the lack of statistically significant relationships in regression analyses of gas exchange parameters and trichome cover. We calculated trichome and leaf boundary layer components, and their associated effects on H2O and CO2 gas exchange. The results further indicate trichome-enhanced boundary layers do not significantly reduce transpirational water loss. We conclude that although the trichomes undoubtedly increase the thickness of the boundary layer, the increase due to Tillandsioid trichomes is inconsequential in terms of whole leaf boundary layers, and any associated reduction in transpirational water loss is also negligible within the whole plant gas exchange pathway.

  3. Low Reynolds number flow near tiny leaves, stems, and trichomes

    NASA Astrophysics Data System (ADS)

    Strickland, Christopher; Pasour, Virginia; Miller, Laura

    2016-11-01

    In terrestrial and aquatic environments such as forest canopies, grass fields, and seagrass beds, the density and shape of trunks, branches, stems, leaves and trichomes (the hairs or fine outgrowths on plants) can drastically alter both the average wind speed and profile through these environments and near each plant. While many studies of flow in these environments have focused on bulk properties of the flow at scales on the order of meters, the low Reynolds number flow close to vegetative structures is especially complex and relevant to nutrient exchange. Using three-dimensional immersed boundary simulations, we resolve the flow around trichomes and small leaves and quantify velocities, shear stresses, and mixing while varying the height and density of idealized structures. National Science Foundation Grant DMS-1127914 to the Statistical and Applied Mathematical Sciences Institute, and the Army Research Office.

  4. Roles of the GLABROUS1 and TRANSPARENT TESTA GLABRA Genes in Arabidopsis Trichome Development.

    PubMed Central

    Larkin, J. C.; Oppenheimer, D. G.; Lloyd, A. M.; Paparozzi, E. T.; Marks, M. D.

    1994-01-01

    Arabidopsis trichomes are branched, single-celled epidermal hairs. These specialized cells provide a convenient model for investigating the specification of cell fate in plants. Two key genes regulating the initiation of trichome development are GLABROUS1 (GL1) and TRANSPARENT TESTA GLABRA (TTG). GL1 is a member of the myb gene family. The maize R gene, which can functionally complement the Arabidopsis ttg mutation, encodes a basic helix-loop-helix protein. We used constitutively expressed copies of the GL1 and R genes to test hypotheses about the roles of GL1 and TTG in trichome development. The results support the hypothesis that TTG and GL1 cooperate at the same point in the trichome developmental pathway. Furthermore, the constitutive expression of both GL1 and R in the same plant caused trichomes to develop on all shoot epidermal surfaces. Results were also obtained indicating that TTG plays an additional role in inhibiting neighboring cells from becoming trichomes. PMID:12244266

  5. Variation in vegetative growth and trichomes in Cannabis sativa L. (Marihuana) in response to enviromental pollution

    SciTech Connect

    Sharma, G.K.; Mann, S.K.

    1984-07-01

    Four populations of Cannabis sativa L. (marihuana) growing in their native habitat and exposed to different levels of environmental pollution were studied for several leaf morphology and leaf trichome features. Leaf length, petiole length, length and width of central leaflet, and the number of teeth on leaf margin decreased with increase in pollution. Trichome length and trichome density values were found to be higher in populations exposed to higher levels of environmental pollution.

  6. Differential transcriptome analysis of glandular and filamentous trichomes in Artemisia annua

    PubMed Central

    2013-01-01

    Background The medicinal plant Artemisia annua is covered with filamentous trichomes and glandular, artemisinin producing trichomes. A high artemisinin supply is needed at a reduced cost for treating malaria. Artemisinin production in bioreactors can be facilitated if a better insight is obtained in the biosynthesis of artemisinin and other metabolites. Therefore, metabolic activities of glandular and filamentous trichomes were investigated at the transcriptome level. Results By laser pressure catapulting, glandular and filamentous trichomes as well as apical and sub-apical cells from glandular trichomes were collected and their transcriptome was sequenced using Illumina RNA-Seq. A de novo transcriptome was assembled (Trinity) and studied with a differential expression analysis (edgeR). A comparison of the transcriptome from glandular and filamentous trichomes shows that MEP, MVA, most terpene and lipid biosynthesis pathways are significantly upregulated in glandular trichomes. Conversely, some transcripts coding for specific sesquiterpenoid and triterpenoid enzymes such as 8-epi-cedrol synthase and an uncharacterized oxidosqualene cyclase were significantly upregulated in filamentous trichomes. All known artemisinin biosynthesis genes are upregulated in glandular trichomes and were detected in both the apical and sub-apical cells of the glandular trichomes. No significant differential expression could be observed between the apical and sub-apical cells. Conclusions Our results underscore the vast metabolic capacities of A. annua glandular trichomes but nonetheless point to the existence of specific terpene metabolic pathways in the filamentous trichomes. Candidate genes that might be involved in artemisinin biosynthesis are proposed based on their putative function and their differential expression level. PMID:24359620

  7. Brassica villosa, a system for studying non-glandular trichomes and genes in the Brassicas.

    PubMed

    Nayidu, Naghabushana K; Tan, Yifang; Taheri, Ali; Li, Xiang; Bjorndahl, Trent C; Nowak, Jacek; Wishart, David S; Hegedus, Dwayne; Gruber, Margaret Y

    2014-07-01

    Brassica villosa is a wild Brassica C genome species with very dense trichome coverage and strong resistance to many insect pests of Brassica oilseeds and vegetables. Transcriptome analysis of hairy B. villosa leaves indicated higher expression of several important trichome initiation genes compared with glabrous B. napus leaves and consistent with the Arabidopsis model of trichome development. However, transcripts of the TRY inhibitory gene in hairy B. villosa were surprisingly high relative to B. napus and relative transcript levels of SAD2, EGL3, and several XIX genes were low, suggesting potential ancillary or less important trichome-related roles for these genes in Brassica species compared with Arabidopsis. Several antioxidant, calcium, non-calcium metal and secondary metabolite genes also showed differential expression between these two species. These coincided with accumulation of two alkaloid-like compounds, high levels of calcium, and other metals in B. villosa trichomes that are correlated with the known tolerance of B. villosa to high salt and the calcium-rich natural habitat of this wild species. This first time report on the isolation of large amounts of pure B. villosa trichomes, on trichome content, and on relative gene expression differences in an exceptionally hairy Brassica species compared with a glabrous species opens doors for the scientific community to understand trichome gene function in the Brassicas and highlights the potential of B. villosa as a trichome research platform.

  8. Comparison of polyphenol oxidase expression in glandular trichomes of solanum and lycopersicon species.

    PubMed

    Yu, H; Kowalski, S P; Steffens, J C

    1992-12-01

    Tetralobulate glandular trichomes are present on the foliage of many solanaceous species. Resistance of many of these species to insects is conditioned by the ability of trichomes to rupture upon contact and to rapidly polymerize their contents, resulting in entrapment of insects in hardened trichome exudate. In the wild potato, Solanum berthaultii, polymerization of trichome exudate is initiated by a soluble M(r) 59,000 polyphenol oxidase (PPO), which is a dominant protein constituent of the organ. PPOs, although ubiquitous in angiosperms, typically display great heterogeneity in molecular weight and are found at low levels in plant cells. Because of the unusually high accumulation and tissue-specific expression of the M(r) 59,000 PPO in S. berthaultii glandular trichomes, we analyzed trichome proteins of a number of Lycopersicon and Solanum species to assess the extent to which possession of the M(r) 59,000 PPO is conserved. Trichomes were collected manually and examined for PPO activity, immuno-cross-reactivity with S. berthaultiiM(r) 59,000 PPO, and protein content. In addition, N-terminal amino acid sequences were obtained for five trichome PPOs. All species analyzed possessed trichome PPOs similar in structure and level of expression to that of S. berthaultii. The relationship between sequences and structures of these conserved PPOs and the variable PPOs of leaf is discussed.

  9. Roles of trichomes with silica particles on the surface of leaves in Aphananthe aspera

    NASA Astrophysics Data System (ADS)

    Takeda, Hiroyuki; Ito, Fuyu; Yamanaka, Shigeru; Takiyama, Naoyuki; Yoshino, Katsumi

    2013-03-01

    There exist many trichomes with silica particles on the surface of leaves in Aphananthe aspera (Muku tree). The size of the trichomes is on the order of some tens to hundreds of micrometers. Investigating the electric-field local densities of the states and distributions of electromagnetic wave propagation, we theoretically revealed that while trichomes play a role as waveguides for the far-infrared, silica particles help incident far-infrared light efficiently propagate inside the trichomes and leaves. These properties would be useful for efficiently warming plants, since absorbed far-infrared light is converted to heat.

  10. Comparison of Polyphenol Oxidase Expression in Glandular Trichomes of Solanum and Lycopersicon Species 1

    PubMed Central

    Yu, Haifeng; Kowalski, Stanley P.; Steffens, John C.

    1992-01-01

    Tetralobulate glandular trichomes are present on the foliage of many solanaceous species. Resistance of many of these species to insects is conditioned by the ability of trichomes to rupture upon contact and to rapidly polymerize their contents, resulting in entrapment of insects in hardened trichome exudate. In the wild potato, Solanum berthaultii, polymerization of trichome exudate is initiated by a soluble Mr 59,000 polyphenol oxidase (PPO), which is a dominant protein constituent of the organ. PPOs, although ubiquitous in angiosperms, typically display great heterogeneity in molecular weight and are found at low levels in plant cells. Because of the unusually high accumulation and tissue-specific expression of the Mr 59,000 PPO in S. berthaultii glandular trichomes, we analyzed trichome proteins of a number of Lycopersicon and Solanum species to assess the extent to which possession of the Mr 59,000 PPO is conserved. Trichomes were collected manually and examined for PPO activity, immuno-cross-reactivity with S. berthaultiiMr 59,000 PPO, and protein content. In addition, N-terminal amino acid sequences were obtained for five trichome PPOs. All species analyzed possessed trichome PPOs similar in structure and level of expression to that of S. berthaultii. The relationship between sequences and structures of these conserved PPOs and the variable PPOs of leaf is discussed. Images Figure 1 Figure 2 Figure 3 PMID:16653213

  11. Variation in the number of capitate glandular trichomes in wild and cultivated sunflower germplasm and potential for use in host plant resistance

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Capitate glandular trichomes of wild sunflower (Helianthus spp.) are considered an effective defense against the sunflower moth, Homoeosoma electellum (Hulst), but cultivated sunflowers are reportedly deficient in glandular trichomes. To investigate whether glandular trichomes have a role in protect...

  12. Correlated evolution of leaf shape and trichomes in Begonia dregei (Begoniaceae).

    PubMed

    McLellan, Tracy

    2005-10-01

    Structural features of leaves, including size, shape, and surfaces, vary greatly throughout the plant kingdom. In both functional and phylogenetic analyses of leaves, the various morphological aspects are often considered independently of each other, although it is likely that many combinations of features do not occur at random due to either functional constraint or genetic correlation. The distribution of variation in leaf morphology in the highly variable Begonia dregei species complex was examined in natural populations and in F(2) offspring from a cross between plants from two populations. Leaf shape was quantified using several morphometric measures, and trichomes on leaves were counted and measured. Correlations between leaf shape and the numbers and size of trichomes were examined. There were significant correlations between the shapes of leaves and the presence, number, and size of trichomes among populations and in hybrid plants. Deeply incised leaves had larger numbers of longer trichomes at the sinuses. Higher numbers of trichomes on upper leaf surfaces occurred together with trichomes at the petiole and on the abaxial surface. The potential for independent evolution of leaf shape and trichomes in this group is limited. Hypotheses to explain the correlated development of leaf shape and trichomes are discussed.

  13. Do plant trichomes cause more harm than good to predatory insects?

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Plants may use epidermal hairs (trichomes) as a morphological defense against attacks from some herbivores. Predatory arthropods can serve plants as biocontrol agents against herbivores. Whether or not plant trichomes work in concert with predators is equivocal. We reviewed the scientific literat...

  14. Fast neutron induced structural rearrangements at a soybean NAP1 locus result in gnarled trichomes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A soybean (Glycine max (L.) Merr.) gnarled trichome mutant, exhibiting stunted trichomes compared to wild-type, was identified in a fast neutron mutant population. Genetic mapping using whole genome sequence-based bulked segregant analysis identified a 26.6 megabase interval on chromosome 20 that ...

  15. Oxylipins produced by Pseudomonas aeruginosa promote biofilm formation and virulence

    PubMed Central

    Martínez, Eriel; Campos-Gómez, Javier

    2016-01-01

    The oxygenation of unsaturated fatty acids by dioxygenases occurs in all kingdoms of life and produces physiologically important lipids called oxylipins. The biological roles of oxylipins have been extensively studied in animals, plants, algae and fungi, but remain largely unidentified in prokaryotes. The bacterium Pseudomonas aeruginosa displays a diol synthase activity that transforms several monounsaturated fatty acids into mono- and di-hydroxylated derivatives. Here we show that oxylipins derived from this activity inhibit flagellum-driven motility and upregulate type IV pilus-dependent twitching motility of P. aeruginosa. Consequently, these oxylipins promote bacterial organization in microcolonies, increasing the ability of P. aeruginosa to form biofilms in vitro and in vivo (in Drosophila flies). We also demonstrate that oxylipins produced by P. aeruginosa promote virulence in Drosophila flies and lettuce. Our study thus uncovers a role for prokaryotic oxylipins in the physiology and pathogenicity of bacteria. PMID:27929111

  16. Induced dicentric chromosome formation promotes genomic rearrangements and tumorigenesis.

    PubMed

    Gascoigne, Karen E; Cheeseman, Iain M

    2013-07-01

    Chromosomal rearrangements can radically alter gene products and their function, driving tumor formation or progression. However, the molecular origins and evolution of such rearrangements are varied and poorly understood, with cancer cells often containing multiple, complex rearrangements. One mechanism that can lead to genomic rearrangements is the formation of a "dicentric" chromosome containing two functional centromeres. Indeed, such dicentric chromosomes have been observed in cancer cells. Here, we tested the ability of a single dicentric chromosome to contribute to genomic instability and neoplastic conversion in vertebrate cells. We developed a system to transiently and reversibly induce dicentric chromosome formation on a single chromosome with high temporal control. We find that induced dicentric chromosomes are frequently damaged and mis-segregated during mitosis, and that this leads to extensive chromosomal rearrangements including translocations with other chromosomes. Populations of pre-neoplastic cells in which a single dicentric chromosome is induced acquire extensive genomic instability and display hallmarks of cellular transformation including anchorage-independent growth in soft agar. Our results suggest that a single dicentric chromosome could contribute to tumor initiation.

  17. Methylglyoxal induces tau hyperphosphorylation via promoting AGEs formation.

    PubMed

    Li, Xiao-Hong; Xie, Jia-Zhao; Jiang, Xia; Lv, Bing-Ling; Cheng, Xiang-Shu; Du, Lai-Ling; Zhang, Jia-Yu; Wang, Jian-Zhi; Zhou, Xin-Wen

    2012-12-01

    The hyperphosphorylated tau is a major protein component of neurofibrillary tangle, which is one of hallmarks of Alzheimer's disease (AD). While the level of methylglyoxal (MG) is significantly increased in the AD brains, the role of MG in tau phosphorylation is still not reported. Here, we found that MG could induce tau hyperphosphorylation at multiple AD-related sites in neuroblastoma 2a cells under maintaining normal cell viability. MG treatment increased the level of advanced glycation end products (AGEs) and the receptor of AGEs (RAGE). Glycogen synthesis kinase-3β (GSK-3β) and p38 MAPK were activated, whereas the level and activity of JNK, Erk1/2, cdk5, and PP2A were not altered after MG treatment. Simultaneous inhibition of GSK-3β or p38 attenuated the MG-induced tau hyperphosphorylation. Aminoguanidine, a blocker of AGEs formation, could effectively reverse the MG-induced tau hyperphosphorylation. These data suggest that MG induces AD-like tau hyperphosphorylation through AGEs formation involving RAGE up-regulation and GSK-3β activation and p38 activation is also partially involved in MG-induced tau hyperphosphorylation. Thus, targeting MG may be a promising therapeutic strategy to prevent AD-like tau hyperphosphorylation.

  18. Mycobacterium abscessus cording prevents phagocytosis and promotes abscess formation

    PubMed Central

    Bernut, Audrey; Herrmann, Jean-Louis; Kissa, Karima; Dubremetz, Jean-François; Gaillard, Jean-Louis; Lutfalla, Georges; Kremer, Laurent

    2014-01-01

    Mycobacterium abscessus is a rapidly growing Mycobacterium causing a wide spectrum of clinical syndromes. It now is recognized as a pulmonary pathogen to which cystic fibrosis patients have a particular susceptibility. The M. abscessus rough (R) variant, devoid of cell-surface glycopeptidolipids (GPLs), causes more severe clinical disease than the smooth (S) variant, but the underlying mechanisms of R-variant virulence remain obscure. Exploiting the optical transparency of zebrafish embryos, we observed that the increased virulence of the M. abscessus R variant compared with the S variant correlated with the loss of GPL production. The virulence of the R variant involved the massive production of serpentine cords, absent during S-variant infection, and the cords initiated abscess formation leading to rapid larval death. Cording occurred within the vasculature and was highly pronounced in the central nervous system (CNS). It appears that M. abscessus is transported to the CNS within macrophages. The release of M. abscessus from apoptotic macrophages initiated the formation of cords that grew too large to be phagocytized by macrophages or neutrophils. This study is a description of the crucial role of cording in the in vivo physiopathology of M. abscessus infection and emphasizes cording as a mechanism of immune evasion. PMID:24567393

  19. GLABROUS INFLORESCENCE STEMS (GIS) is required for trichome branching through gibberellic acid signaling in Arabidopsis.

    PubMed

    An, Lijun; Zhou, Zhongjing; Su, Sha; Yan, An; Gan, Yinbo

    2012-02-01

    Cell differentiation generally corresponds to the cell cycle, typically forming a non-dividing cell with a unique differentiated morphology, and Arabidopsis trichome is an excellent model system to study all aspects of cell differentiation. Although gibberellic acid is reported to be involved in trichome branching in Arabidopsis, the mechanism for such signaling is unclear. Here, we demonstrated that GLABROUS INFLORESCENCE STEMS (GIS) is required for the control of trichome branching through gibberellic acid signaling. The phenotypes of a loss-of-function gis mutant and an overexpressor showed that GIS acted as a repressor to control trichome branching. Our results also show that GIS is not required for cell endoreduplication, and our molecular and genetic study results have shown that GIS functions downstream of the key regulator of trichome branching, STICHEL (STI), to control trichome branching through the endoreduplication-independent pathway. Furthermore, our results also suggest that GIS controls trichome branching in Arabidopsis through two different pathways and acts either upstream or downstream of the negative regulator of gibbellic acid signaling SPINDLY (SPY).

  20. PTH-IGF SIGNALING PROMOTES BONE FORMATION THROUGH GLYCOLYSIS

    PubMed Central

    Esen, Emel; Lee, Seung-Yon; Wice, Burton M; Long, Fanxin

    2016-01-01

    Teriparatide, a recombinant peptide corresponding to amino acids 1-34 of human parathyroid hormone (PTH), has been an effective bone anabolic drug for over a decade. However, the mechanism whereby PTH stimulates bone formation remains poorly understood. Here we report that in cultures of osteoblast-lineage cells, PTH stimulates glucose consumption and lactate production in the presence of oxygen, a hallmark of aerobic glycolysis, also known as Warburg effect. Experiments with radioactively labeled glucose demonstrate that PTH suppresses glucose entry into the tricarboxylic acid cycle (TCA cycle). Mechanistically, the increase in aerobic glycolysis is secondary to insulin-like growth factor (Igf) signaling induced by PTH, whereas the metabolic effect of Igf is dependent on activation of mammalian target of rapamycin complex 2 (mTORC2). Importantly, pharmacological perturbation of glycolysis suppresses the bone anabolic effect of intermittent PTH in the mouse. Thus, stimulation of aerobic glycolysis via Igf signaling contributes to bone anabolism in response to PTH. PMID:25990470

  1. Exercise promotes positive impression formation towards both men and women.

    PubMed

    Kanarek, Robin B; Mathes, Wendy Foulds; D'Anci, Kristen E

    2012-06-01

    Exercise is endorsed for its physiological and psychological benefits, and has been proposed to have positive effects on impression formation. To test this proposal, 62 female and 44 male college students read one of three brief descriptions of either a fictitious male or female "target" student. The descriptions varied only in exercise level: no exercise; moderate exercise and intensive exercise. Participants then rated the fictitious student on 38 personality traits. Ratings of characteristics that are associated with exercise (e.g. athletic; energetic) increased, while ratings associated with the lack of exercise (e.g. lazy; weak) decreased as a function of the reported level of exercise. Exercise level also positively influenced ratings of characteristics not related to exercise. These data show that even minimal information about exercise is an important component of first impressions in both men and women.

  2. Promoting cooperation by reputation-driven group formation

    NASA Astrophysics Data System (ADS)

    Yang, Han-Xin; Wang, Zhen

    2017-02-01

    In previous studies of the spatial public goods game, each player is able to establish a group. However, in real life, some players cannot successfully organize groups for various reasons. In this paper, we propose a mechanism of reputation-driven group formation, in which groups can only be organized by players whose reputation reaches or exceeds a threshold. We define a player’s reputation as the frequency of cooperation in the last T time steps. We find that the highest cooperation level can be obtained when groups are only established by pure cooperators who always cooperate in the last T time steps. Effects of the memory length T on cooperation are also studied.

  3. Morphology of foliar trichomes of the Chinese cork oak Quercus variabilis by electron microscopy and three-dimensional surface profiling.

    PubMed

    Kim, Ki Woo; Cho, Do-Hyun; Kim, Pan-Gi

    2011-06-01

    Morphology of foliar trichomes was analyzed in Quercus variabilis by electron microscopy and three-dimensional surface profiling. Leaves from suppressed or dominant sprouts of the oak species were collected after a forest fire to unravel the effects of the disturbance factor on sprouting of the oak species. Scanning electron microscopy revealed two types of trichomes depending on the leaf surface. The trichomes on the adaxial surface were branched and constricted, and possessed a single row of thin-walled cells with a collapsed morphology (glandular branched uniseriate trichomes). Meanwhile, the trichomes on the abaxial surface were star-shaped, unfused with each other, and had 6 to 10 rays (nonglandular simple stellate trichomes). An apparent proliferation of trichomes was evident on the adaxial surface of the dominant sprouts. Uniseriate trichomes could be discernable as an elevation from the surface by white light scanning interferometry. By transmission electron microscopy, thin and convoluted cell wall, degenerated cytoplasm, and a single row of cells were characteristic of the trichomes on the adaxial surface. The thick cell walls of the mature trichomes on the abaxial surface represented the nonglandular nature. This is the first report on the morphological and ultrastructural characterization of foliar trichomes of the oak species.

  4. Identification, functional characterization and developmental regulation of sesquiterpene synthases from sunflower capitate glandular trichomes

    PubMed Central

    Göpfert, Jens C; MacNevin, Gillian; Ro, Dae-Kyun; Spring, Otmar

    2009-01-01

    Background Sesquiterpene lactones are characteristic metabolites of Asteraceae (or Compositae) which often display potent bioactivities and are sequestered in specialized organs such as laticifers, resin ducts, and trichomes. For characterization of sunflower sesquiterpene synthases we employed a simple method to isolate pure trichomes from anther appendages which facilitated the identification of these genes and investigation of their enzymatic functions and expression patterns during trichome development. Results Glandular trichomes of sunflower (Helianthus annuus L.) were isolated, and their RNA was extracted to investigate the initial steps of sesquiterpene lactone biosynthesis. Reverse transcription-PCR experiments led to the identification of three sesquiterpene synthases. By combination of in vitro and in vivo characterization of sesquiterpene synthase gene products in Escherichia coli and Saccharomyces cerevisiae, respectively, two enzymes were identified as germacrene A synthases, the key enzymes of sesquiterpene lactone biosynthesis. Due to the very low in vitro activity, the third enzyme was expressed in vivo in yeast as a thioredoxin-fusion protein for functional characterization. In in vivo assays, it was identified as a multiproduct enzyme with the volatile sesquiterpene hydrocarbon δ-cadinene as one of the two main products with α-muuorlene, β-caryophyllene, α-humulene and α-copaene as minor products. The second main compound remained unidentified. For expression studies, glandular trichomes from the anther appendages of sunflower florets were isolated in particular developmental stages from the pre- to the post-secretory phase. All three sesquiterpene synthases were solely upregulated during the biosynthetically active stages of the trichomes. Expression in different aerial plant parts coincided with occurrence and maturity of trichomes. Young roots with root hairs showed expression of the sesquiterpene synthase genes as well. Conclusion This

  5. Tropoelastin coated PLLA-PLGA scaffolds promote vascular network formation.

    PubMed

    Landau, Shira; Szklanny, Ariel A; Yeo, Giselle C; Shandalov, Yulia; Kosobrodova, Elena; Weiss, Anthony S; Levenberg, Shulamit

    2017-04-01

    The robust repair of large wounds and tissue defects relies on blood flow. This vascularization is the major challenge faced by tissue engineering on the path to forming thick, implantable tissue constructs. Without this vasculature, oxygen and nutrients cannot reach the cells located far from host blood vessels. To make viable constructs, tissue engineering takes advantage of the mechanical properties of synthetic materials, while combining them with ECM proteins to create a natural environment for the tissue-specific cells. Tropoelastin, the precursor of the elastin, is the ECM protein responsible for elasticity in diverse tissues, including robust blood vessels. Here, we seeded endothelial cells with supporting cells on PLLA/PLGA scaffolds treated with tropoelastin, and examined the morphology, expansion and maturity of the newly formed vessels. Our results demonstrate that the treated scaffolds elicit a more expanded, complex and developed vascularization in comparison to the untreated group. Implantation of tropoelastin-treated scaffolds into mouse abdominal muscle resulted in enhanced perfusion of the penetrating vasculature and improved integration. This study points to the great potential of these combined materials in promoting the vascularization of implanted engineered constructs, which can be further exploited in the fabrication of clinically relevant engineered tissues.

  6. NADPH Oxidase Promotes Neutrophil Extracellular Trap Formation in Pulmonary Aspergillosis

    PubMed Central

    Röhm, Marc; Grimm, Melissa J.; D'Auria, Anthony C.; Almyroudis, Nikolaos G.

    2014-01-01

    NADPH oxidase is a crucial enzyme in antimicrobial host defense and in regulating inflammation. Chronic granulomatous disease (CGD) is an inherited disorder of NADPH oxidase in which phagocytes are defective in generation of reactive oxidant intermediates. Aspergillus species are ubiquitous, filamentous fungi, which can cause invasive aspergillosis, a major cause of morbidity and mortality in CGD, reflecting the critical role for NADPH oxidase in antifungal host defense. Activation of NADPH oxidase in neutrophils can be coupled to the release of proteins and chromatin that comingle in neutrophil extracellular traps (NETs), which can augment extracellular antimicrobial host defense. NETosis can be driven by NADPH oxidase-dependent and -independent pathways. We therefore undertook an analysis of whether NADPH oxidase was required for NETosis in Aspergillus fumigatus pneumonia. Oropharyngeal instillation of live Aspergillus hyphae induced neutrophilic pneumonitis in both wild-type and NADPH oxidase-deficient (p47phox−/−) mice which had resolved in wild-type mice by day 5 but progressed in p47phox−/− mice. NETs, identified by immunostaining, were observed in lungs of wild-type mice but were absent in p47phox−/− mice. Using bona fide NETs and nuclear chromatin decondensation as an early NETosis marker, we found that NETosis required a functional NADPH oxidase in vivo and ex vivo. In addition, NADPH oxidase increased the proportion of apoptotic neutrophils. Together, our results show that NADPH oxidase is required for pulmonary clearance of Aspergillus hyphae and generation of NETs in vivo. We speculate that dual modulation of NETosis and apoptosis by NADPH oxidase enhances antifungal host defense and promotes resolution of inflammation upon infection clearance. PMID:24549323

  7. Native low density lipoprotein promotes lipid raft formation in macrophages

    PubMed Central

    SONG, JIAN; PING, LING-YAN; DUONG, DUC M.; GAO, XIAO-YAN; HE, CHUN-YAN; WEI, LEI; WU, JUN-ZHU

    2016-01-01

    Oxidized low-density lipoprotein (LDL) has an important role in atherogenesis; however, the mechanisms underlying cell-mediated LDL oxidation remain to be elucidated. The present study investigated whether native-LDL induced lipid raft formation, in order to gain further insight into LDL oxidation. Confocal microscopic analysis revealed that lipid rafts were aggregated or clustered in the membrane, which were colocalized with myeloperoxidase (MPO) upon native LDL stimulation; however, in the presence of methyl-β-cyclodextrin (MβCD), LDL-stimulated aggregation, translocation, and colocalization of lipid rafts components was abolished.. In addition, lipid raft disruptors MβCD and filipin decreased malondialdehyde expression levels. Density gradient centrifugation coupled to label-free quantitative proteomic analysis identified 1,449 individual proteins, of which 203 were significantly upregulated following native-LDL stimulation. Functional classification of the proteins identified in the lipid rafts revealed that the expression levels of translocation proteins were upregulated. In conclusion, the results of the present study indicated that native-LDL induced lipid raft clustering in macrophages, and the expression levels of several proteins were altered in the stimulated macrophages, which provided novel insights into the mechanism underlying LDL oxidation. PMID:26781977

  8. Chlorogenic acid increased acrylamide formation through promotion of HMF formation and 3-aminopropionamide deamination.

    PubMed

    Cai, Yun; Zhang, Zhenhua; Jiang, Shanshan; Yu, Miao; Huang, Caihuan; Qiu, Ruixia; Zou, Yueyu; Zhang, Qirui; Ou, Shiyi; Zhou, Hua; Wang, Yong; Bai, Weibing; Li, Yiqun

    2014-03-15

    This research was aimed to investigate why chlorogenic acid, presents at high concentrations in some food raw material, influences acrylamide formation. In the asparagine/glucose Maillard reaction system (pH=6.8), addition of chlorogenic acid significantly increased acrylamide formation and inhibited its elimination. In contrast, the quinone derivative of chlorogenic acid decreased acrylamide formation. Three mechanisms may be involved for increasing acrylamide formation by chlorogenic acid. Firstly, it increased the formation of HMF, which acts as a more efficient precursor than glucose to form acrylamide. Secondly, it decreased activation energy for conversion of 3-aminopropionamide (3-APA) to acrylamide (from 173.2 to 136.6kJ/mol), and enhances deamination from 3-APA. And thirdly, it prevented attack of the produced acrylamide from free radicals by keeping high redox potential during the Maillard reaction.

  9. Boomeranging in structural defense: phytophagous insect uses cycad trichomes to defend against entomophagy.

    PubMed

    Marler, Thomas E

    2012-11-01

    Plant defensive behaviors that resist arthropod herbivory include trichome-mediated defenses, and variation in plant trichome morphology and abundance provides examples of the mechanistic complexities of insect-plant interactions. Trichomes were removed from Cycas revoluta cataphylls on the island of Guam to reveal Aulacaspis yasumatsui scale infestation, and predation of the newly exposed insects by pre-existing Rhyzobius lophanthae beetles commenced within one day. The quotient of predated/total scale insects was 0.5 by day 4 and stabilized at that found on adjacent glabrous leaves in about one week. The trichome phenotype covering the C. revoluta cataphyll complex offers the invasive A. yasumatsui armored scale effectual enemy-free space in this system. This pest and predator share no known evolutionary history with C. revoluta, therefore, the adaptive significance of this plant behavior in natural habitat is not yet known.

  10. Herbivore handling of a Plants trichome: the case of Heliconius charithonia (L.) (Lepidoptera: Nymphalidae) and Passiflora lobata (Killip) Hutch. (Passifloraceae).

    PubMed

    Cardoso, Márcio Z

    2008-01-01

    Trichomes reduce herbivore attack on plants by physically and/or chemically inhibiting movement or other activities. Despite evidence that herbivores are negatively affected by trichomes there also reports of insect counter-adaptations that circumvent the plants defense. This paper reports on a study that investigated the likely mechanisms employed by larvae of the nymphalid butterfly, Heliconius charithonia (L.), that allow it to feed on a host that is presumably protected by hooked trichomes (Passiflora lobata (Killip) Hutch). Evidence were gathered using data from direct observations of larval movement and behavior, faeces analysis, scanning electron microscopy of plant surface and experimental analysis of larval movement on plants with and without trichomes (manually removed). The latter involved a comparison with a non specialist congener, Heliconius pachinus Salvin. Observations showed that H. charithonia larvae are capable of freeing themselves from entrapment on trichome tips by physical force. Moreover, wandering larvae lay silk mats on the trichomes and remove their tips by biting. In fact, trichome tips were found in the faeces. Experimental removal of trichomes aided in the movement of the non specialist but had no noticeable effect on the specialist larvae. These results support the suggestion that trichomes are capable of deterring a non specialist herbivore (H. pachinus). The precise mechanisms that allow the success of H. charithonia are not known, but I suggest that a blend of behavioral as well as physical resistance mechanisms is involved. Future studies should ascertain whether larval integument provides physical resistance to trichomes.

  11. Influence of Environment on Glandular Trichomes and Composition of Essential Oil of Perovskia abrotanoides Karel.

    PubMed Central

    Oreizi, Elaheh; Rahiminejad, Mohammad Reza; Asghari, Gholamreza

    2014-01-01

    Background: Perovskia abrotanoides Karel. is a medicinal plant used in Iranian folk medicine as a pain killer. Forty-one components have been identified in P. abrotanoides samples collected from Baluchistan Province, and 29 components have been recognized in samples collected from Khorasan Province. The leaves of P. abrotanoides have glandular trichomes (capitates and peltate) on both sides of the lamina. Objectives: This study aimed to evaluate the variation of oil constituents of the plant and illustrate the glandular trichomes types and then show the influence of environment on oil constituents and glandular trichomes. Materials and Methods: The essential oil of the plant was obtained using hydrodistillation and the analysis of oils carried out using GC-MS. The anatomical analysis of leaves was done by fixing, coloring, and photoing the sections. Results: Glandular trichomes composed of capitates and peltate trichomes. The essential oil composition differs. Viridiflora and neryl acetate were not identified in yellow glandular trichomes. Conclusions: It seems that there is no relation between anatomical characteristics of the plant leaves and its essential oil composition. PMID:25625046

  12. Modeling Filamentous Cyanobacteria Reveals the Advantages of Long and Fast Trichomes for Optimizing Light Exposure

    PubMed Central

    Tamulonis, Carlos; Postma, Marten; Kaandorp, Jaap

    2011-01-01

    Cyanobacteria form a very large and diverse phylum of prokaryotes that perform oxygenic photosynthesis. Many species of cyanobacteria live colonially in long trichomes of hundreds to thousands of cells. Of the filamentous species, many are also motile, gliding along their long axis, and display photomovement, by which a trichome modulates its gliding according to the incident light. The latter has been found to play an important role in guiding the trichomes to optimal lighting conditions, which can either inhibit the cells if the incident light is too weak, or damage the cells if too strong. We have developed a computational model for gliding filamentous photophobic cyanobacteria that allows us to perform simulations on the scale of a Petri dish using over 105 individual trichomes. Using the model, we quantify the effectiveness of one commonly observed photomovement strategy—photophobic responses—in distributing large populations of trichomes optimally over a light field. The model predicts that the typical observed length and gliding speeds of filamentous cyanobacteria are optimal for the photophobic strategy. Therefore, our results suggest that not just photomovement but also the trichome shape itself improves the ability of the cyanobacteria to optimize their light exposure. PMID:21789215

  13. Plant Glandular Trichomes as Targets for Breeding or Engineering of Resistance to Herbivores

    PubMed Central

    Glas, Joris J.; Schimmel, Bernardus C. J.; Alba, Juan M.; Escobar-Bravo, Rocío; Schuurink, Robert C.; Kant, Merijn R.

    2012-01-01

    Glandular trichomes are specialized hairs found on the surface of about 30% of all vascular plants and are responsible for a significant portion of a plant’s secondary chemistry. Glandular trichomes are an important source of essential oils, i.e., natural fragrances or products that can be used by the pharmaceutical industry, although many of these substances have evolved to provide the plant with protection against herbivores and pathogens. The storage compartment of glandular trichomes usually is located on the tip of the hair and is part of the glandular cell, or cells, which are metabolically active. Trichomes and their exudates can be harvested relatively easily, and this has permitted a detailed study of their metabolites, as well as the genes and proteins responsible for them. This knowledge now assists classical breeding programs, as well as targeted genetic engineering, aimed to optimize trichome density and physiology to facilitate customization of essential oil production or to tune biocide activity to enhance crop protection. We will provide an overview of the metabolic diversity found within plant glandular trichomes, with the emphasis on those of the Solanaceae, and of the tools available to manipulate their activities for enhancing the plant’s resistance to pests. PMID:23235331

  14. Insight into the mechanism of Sb promoted Cu(In,Ga)Se{sub 2} formation

    SciTech Connect

    Xiang, Yong Zhang, Xiaokun; Zhang, Shu

    2013-08-15

    Sb-doping has been demonstrated to be a new approach to promote Cu(In,Ga)Se{sub 2} (CIGS) thin film formation. To study the mechanism of Sb-promoted CIGS formation, we investigated the influence of Sb on the evolution of the critical intermediate Cu–Se phases, and found that Cu{sub 3}SbSe{sub 3} species was formed. Phase change of the as-prepared Cu–Se compounds at elevated temperature was determined using the differential scanning calorimetry analysis. For samples containing Sb, the melting decomposition occurred at 507.1 °C, along with aggregation of nanocrystals into a bulk, while the morphology of the sample without Sb barely changed after heating. This result suggests that the mobile Cu{sub 3}SbSe{sub 3} is likely the key intermediate responsible for Sb-promoted CIGS formation. Furthermore, we extended the scope of Sb-doping approach to solvothermal synthesis and CIGS nanocrystals were synthesized with significantly promoted kinetics in the presence of Sb. Based on these results, we propose the mechanism of Sb promoted CIGS formation. - Graphical abstract: Cu{sub 3}SbSe{sub 3} mobile phase is likely the key species to promote the formation of Cu(In,Ga)Se{sub 2}, and significantly promoting effect by Sb is also found in the synthesis of Cu(In,Ga)Se{sub 2} nanocrystals. Highlights: • In the presence of Sb, Cu{sub 3}SbSe{sub 3} is formed while synthesizing the essential intermediate Cu–Se compounds for Cu(In,Ga)Se{sub 2} materials. • Cu{sub 3}SbSe{sub 3} shows high mobility at elevated temperature. • Cu{sub 3}SbSe{sub 3} mobile phase is likely the key species to improve Cu(In,Ga)Se{sub 2} thin film formation. • A synthesis methodology is developed to produce Cu(In,Ga)Se{sub 2} nanocrystals with significantly promoted reaction kinetics.

  15. Structural Analysis of G-Quadruplex Formation at the Human MEST Promoter

    PubMed Central

    Kennedy, Martin A.

    2017-01-01

    The promoter region of the imprinted gene MEST contains several motifs capable of forming G-quadruplex (G4) structures, which appear to contribute to consistent allelic dropout during polymerase chain reaction (PCR) analysis of this region. Here, we extend our previous analysis of MEST G4 structures by applying fluorescent footprinting techniques to assess non B-DNA structure and topology in dsDNA from the full MEST promoter region, under conditions that mimic PCR. We demonstrate that the buffer used for PCR provides an extremely favourable milieu for G4 formation, and that cytosine methylation helps maintain G4 structures during PCR. Additionally, we demonstrate G4 formation at motifs not previously identified through bioinformatic analysis of the MEST promoter, and provide nucleotide level resolution for topological reconstruction of these structures. These observations increase our understanding of the mechanisms through which methylation and G4 contribute towards allelic drop-out during PCR. PMID:28052120

  16. Circadian glucocorticoid oscillations promote learning-dependent synapse formation and maintenance.

    PubMed

    Liston, Conor; Cichon, Joseph M; Jeanneteau, Freddy; Jia, Zhengping; Chao, Moses V; Gan, Wen-Biao

    2013-06-01

    Excessive glucocorticoid exposure during chronic stress causes synapse loss and learning impairment. Under normal physiological conditions, glucocorticoid activity oscillates in synchrony with the circadian rhythm. Whether and how endogenous glucocorticoid oscillations modulate synaptic plasticity and learning is unknown. Here we show that circadian glucocorticoid peaks promote postsynaptic dendritic spine formation in the mouse cortex after motor skill learning, whereas troughs are required for stabilizing newly formed spines that are important for long-term memory retention. Conversely, chronic and excessive exposure to glucocorticoids eliminates learning-associated new spines and disrupts previously acquired memories. Furthermore, we show that glucocorticoids promote rapid spine formation through a non-transcriptional mechanism by means of the LIM kinase-cofilin pathway and increase spine elimination through transcriptional mechanisms involving mineralocorticoid receptor activation. Together, these findings indicate that tightly regulated circadian glucocorticoid oscillations are important for learning-dependent synaptic formation and maintenance. They also delineate a new signaling mechanism underlying these effects.

  17. Sleep promotes branch-specific formation of dendritic spines after learning.

    PubMed

    Yang, Guang; Lai, Cora Sau Wan; Cichon, Joseph; Ma, Lei; Li, Wei; Gan, Wen-Biao

    2014-06-06

    How sleep helps learning and memory remains unknown. We report in mouse motor cortex that sleep after motor learning promotes the formation of postsynaptic dendritic spines on a subset of branches of individual layer V pyramidal neurons. New spines are formed on different sets of dendritic branches in response to different learning tasks and are protected from being eliminated when multiple tasks are learned. Neurons activated during learning of a motor task are reactivated during subsequent non-rapid eye movement sleep, and disrupting this neuronal reactivation prevents branch-specific spine formation. These findings indicate that sleep has a key role in promoting learning-dependent synapse formation and maintenance on selected dendritic branches, which contribute to memory storage.

  18. The key role of peltate glandular trichomes in symbiota comprising clavicipitaceous fungi of the genus periglandula and their host plants.

    PubMed

    Steiner, Ulrike; Kucht, Sabine Hellwig neé; Ahimsa-Müller, Mahalia A; Grundmann, Nicola; Li, Shu-Ming; Drewke, Christel; Leistner, Eckhard

    2015-04-16

    Clavicipitaceous fungi producing ergot alkaloids were recently discovered to be epibiotically associated with peltate glandular trichomes of Ipomoea asarifolia and Turbina corymbosa, dicotyledonous plants of the family Convolvulaceae. Mediators of the close association between fungi and trichomes may be sesquiterpenes, main components in the volatile oil of different convolvulaceous plants. Molecular biological studies and microscopic investigations led to the observation that the trichomes do not only secrete sesquiterpenes and palmitic acid but also seem to absorb ergot alkaloids from the epibiotic fungal species of the genus Periglandula. Thus, the trichomes are likely to have a dual and key function in a metabolic dialogue between fungus and host plant.

  19. The Key Role of Peltate Glandular Trichomes in Symbiota Comprising Clavicipitaceous Fungi of the Genus Periglandula and Their Host Plants

    PubMed Central

    Steiner, Ulrike; Hellwig, Sabine; Ahimsa-Müller, Mahalia A.; Grundmann, Nicola; Li, Shu-Ming; Drewke, Christel; Leistner, Eckhard

    2015-01-01

    Clavicipitaceous fungi producing ergot alkaloids were recently discovered to be epibiotically associated with peltate glandular trichomes of Ipomoea asarifolia and Turbina corymbosa, dicotyledonous plants of the family Convolvulaceae. Mediators of the close association between fungi and trichomes may be sesquiterpenes, main components in the volatile oil of different convolvulaceous plants. Molecular biological studies and microscopic investigations led to the observation that the trichomes do not only secrete sesquiterpenes and palmitic acid but also seem to absorb ergot alkaloids from the epibiotic fungal species of the genus Periglandula. Thus, the trichomes are likely to have a dual and key function in a metabolic dialogue between fungus and host plant. PMID:25894995

  20. Thyroid dysfunction, either hyper or hypothyroidism, promotes gallstone formation by different mechanisms*

    PubMed Central

    Wang, Yong; Yu, Xing; Zhao, Qun-zi; Zheng, Shu; Qing, Wen-jie; Miao, Chun-di; Sanjay, Jaiswal

    2016-01-01

    We have investigated comprehensively the effects of thyroid function on gallstone formation in a mouse model. Gonadectomized gallstone-susceptible male C57BL/6 mice were randomly distributed into three groups each of which received an intervention to induce hyperthyroidism, hypothyroidism, or euthyroidism. After 5 weeks of feeding a lithogenic diet of 15% (w/w) butter fat, 1% (w/w) cholesterol, and 0.5% (w/w) cholic acid, mice were killed for further experiments. The incidence of cholesterol monohydrate crystal formation was 100% in mice with hyperthyroidism, 83% in hypothyroidism, and 33% in euthyroidism, the differences being statistically significant. Among the hepatic lithogenic genes, Trβ was found to be up-regulated and Rxr down-regulated in the mice with hypothyroidism. In contrast, Lxrα, Rxr, and Cyp7α1 were up-regulated and Fxr down-regulated in the mice with hyperthyroidism. In conclusion, thyroid dysfunction, either hyperthyroidism or hypothyroidism, promotes the formation of cholesterol gallstones in C57BL/6 mice. Gene expression differences suggest that thyroid hormone disturbance leads to gallstone formation in different ways. Hyperthyroidism induces cholesterol gallstone formation by regulating expression of the hepatic nuclear receptor genes such as Lxrα and Rxr, which are significant in cholesterol metabolism pathways. However, hypothyroidism induces cholesterol gallstone formation by promoting cholesterol biosynthesis. PMID:27381728

  1. Protein tyrosine phosphatase SHP2 promotes invadopodia formation through suppression of Rho signaling

    PubMed Central

    Tsai, Wan-Chen; Chen, Chien-Lin; Chen, Hong-Chen

    2015-01-01

    Invadopodia are actin-enriched membrane protrusions that are important for extracellular matrix degradation and invasive cell motility. Src homolog domain-containing phosphatase 2 (SHP2), a non-receptor protein tyrosine phosphatase, has been shown to play an important role in promoting cancer metastasis, but the underlying mechanism is unclear. In this study, we found that depletion of SHP2 by short-hairpin RNA suppressed invadopodia formation in several cancer cell lines, particularly in the SAS head and neck squamous cell line. In contrast, overexpression of SHP2 promoted invadopodia formation in the CAL27 head and neck squamous cell line, which expresses low levels of endogenous SHP2. The depletion of SHP2 in SAS cells significantly decreased their invasive motility. The suppression of invadopodia formation by SHP2 depletion was restored by the Clostridium botulinum C3 exoenzyme (a Rho GTPase inhibitor) or Y27632 (a specific inhibitor for Rho-associated kinase). Together, our results suggest that SHP2 may promote invadopodia formation through inhibition of Rho signaling in cancer cells. PMID:26204488

  2. The calcium-dependent ribonuclease XendoU promotes ER network formation through local RNA degradation

    PubMed Central

    Schwarz, Dianne S.

    2014-01-01

    How cells shape and remodel organelles in response to cellular signals is a poorly understood process. Using Xenopus laevis egg extract, we found that increases in cytosolic calcium lead to the activation of an endogenous ribonuclease, XendoU. A fraction of XendoU localizes to the endoplasmic reticulum (ER) and is required for nuclear envelope assembly and ER network formation in a catalysis-dependent manner. Using a purified vesicle fusion assay, we show that XendoU functions on the surface of ER membranes to promote RNA cleavage and ribonucleoprotein (RNP) removal. Additionally, RNA removal from the surface of vesicles by RNase treatment leads to increased ER network formation. Using human tissue culture cells, we found that hEndoU localizes to the ER, where it promotes the formation of ER tubules in a catalysis-dependent manner. Together, these results demonstrate that calcium-activated removal of RNA from membranes by XendoU promotes and refines ER remodeling and the formation of tubular ER. PMID:25287301

  3. Autofluorescence as a Signal to Sort Developing Glandular Trichomes by Flow Cytometry

    PubMed Central

    Bergau, Nick; Navarette Santos, Alexander; Henning, Anja; Balcke, Gerd U.; Tissier, Alain

    2016-01-01

    The industrial relevance of a number of metabolites produced in plant glandular trichomes (GTs) has spurred research on these specialized organs for a number of years. Most of the research, however, has focused on the elucidation of secondary metabolite pathways and comparatively little has been undertaken on the development and differentiation of GTs. One way to gain insight into these developmental processes is to generate stage-specific transcriptome and metabolome data. The difficulty for this resides in the isolation of early stages of development of the GTs. Here we describe a method for the separation and isolation of intact young and mature type VI trichomes from the wild tomato species Solanum habrochaites. The final and key step of the method uses cell sorting based on distinct autofluorescence signals of the young and mature trichomes. We demonstrate that sorting by flow cytometry allows recovering pure fractions of young and mature trichomes. Furthermore, we show that the sorted trichomes can be used for transcript and metabolite analyses. Because many plant tissues or cells have distinct autofluorescence components, the principles of this method can be generally applicable for the isolation of specific cell types without prior labeling. PMID:27446176

  4. Terpene Biosynthesis in Glandular Trichomes of Hop12[W][OA

    PubMed Central

    Wang, Guodong; Tian, Li; Aziz, Naveed; Broun, Pierre; Dai, Xinbin; He, Ji; King, Andrew; Zhao, Patrick X.; Dixon, Richard A.

    2008-01-01

    Hop (Humulus lupulus L. Cannabaceae) is an economically important crop for the brewing industry, where it is used to impart flavor and aroma to beer, and has also drawn attention in recent years due to its potential pharmaceutical applications. Essential oils (mono- and sesquiterpenes), bitter acids (prenylated polyketides), and prenylflavonoids are the primary phytochemical components that account for these traits, and all accumulate at high concentrations in glandular trichomes of hop cones. To understand the molecular basis for terpene accumulation in hop trichomes, a trichome cDNA library was constructed and 9,816 cleansed expressed sequence tag (EST) sequences were obtained from random sequencing of 16,152 cDNA clones. The ESTs were assembled into 3,619 unigenes (1,101 contigs and 2,518 singletons). Putative functions were assigned to the unigenes based on their homology to annotated sequences in the GenBank database. Two mono- and two sesquiterpene synthases identified from the EST collection were expressed in Escherichia coli. Hop MONOTERPENE SYNTHASE2 formed the linear monterpene myrcene from geranyl pyrophosphate, whereas hop SESQUITERPENE SYNTHASE1 (HlSTS1) formed both caryophyllene and humulene from farnesyl pyrophosphate. Together, these enzymes account for the production of the major terpene constituents of the hop trichomes. HlSTS2 formed the minor sesquiterpene constituent germacrene A, which was converted to β-elemene on chromatography at elevated temperature. We discuss potential functions for other genes expressed at high levels in developing hop trichomes. PMID:18775972

  5. Transcriptional profiling unravels potential metabolic activities of the olive leaf non-glandular trichome

    PubMed Central

    Koudounas, Konstantinos; Manioudaki, Maria E.; Kourti, Anna; Banilas, Georgios; Hatzopoulos, Polydefkis

    2015-01-01

    The olive leaf trichomes are multicellular peltate hairs densely distributed mainly at the lower leaf epidermis. Although, non-glandular, they have gained much attention since they significantly contribute to abiotic and biotic stress tolerance of olive leaves. The exact mechanisms by which olive trichomes achieve these goals are not fully understood. They could act as mechanical barrier but they also accumulate high amounts of flavonoids among other secondary metabolites. However, little is currently known about the exact compounds they produce and the respective metabolic pathways. Here we present the first EST analysis from olive leaf trichomes by using 454-pyrosequencing. A total of 5368 unigenes were identified out of 7258 high quality reads with an average length of 262 bp. Blast search revealed that 27.5% of them had high homologies to known proteins. By using Blast2GO, 1079 unigenes (20.1%) were assigned at least one Gene Ontology (GO) term. Most of the genes were involved in cellular and metabolic processes and in binding functions followed by catalytic activity. A total of 521 transcripts were mapped to 67 KEGG pathways. Olive trichomes represent a tissue of highly unique transcriptome as per the genes involved in developmental processes and the secondary metabolism. The results indicate that mature olive trichomes are trancriptionally active, mainly through the potential production of enzymes that contribute to phenolic compounds with important roles in biotic and abiotic stress responses. PMID:26322070

  6. Intercellular transfer along the trichomes of the invasive terminal heterocyst forming cyanobacterium Cylindrospermopsis raciborskii CS-505.

    PubMed

    Plominsky, Álvaro M; Delherbe, Nathalie; Mandakovic, Dinka; Riquelme, Brenda; González, Karen; Bergman, Birgitta; Mariscal, Vicente; Vásquez, Mónica

    2015-03-01

    Cylindrospermopsis raciborskii CS-505 is an invasive freshwater filamentous cyanobacterium that when grown diazotrophically may develop trichomes of up to 100 vegetative cells while differentiating only two end heterocysts, the sole sites for their N2-fixation process. We examined the diazotrophic growth and intercellular transfer mechanisms in C. raciborskii CS-505. Subjecting cultures to a combined-nitrogen-free medium to elicit N2 fixation, the trichome length remained unaffected while growth rates decreased. The structures and proteins for intercellular communication showed that while a continuous periplasmic space was apparent along the trichomes, the putative septal junction sepJ gene is divided into two open reading frames and lacks several transmembrane domains unlike the situation in Anabaena, differentiating a 5-fold higher frequency of heterocysts. FRAP analyses also showed that the dyes calcein and 5-CFDA were taken up by heterocysts and vegetative cells, and that the transfer from heterocysts and 'terminal' vegetative cells showed considerably higher transfer rates than that from vegetative cells located in the middle of the trichomes. The data suggest that C. raciborskii CS-505 compensates its low-frequency heterocyst phenotype by a highly efficient transfer of the fixed nitrogen towards cells in distal parts of the trichomes (growing rapidly) while cells in central parts suffers (slow growth).

  7. Transcriptome analysis in Cucumis sativus identifies genes involved in multicellular trichome development.

    PubMed

    Zhao, Jun-Long; Pan, Jun-Song; Guan, Yuan; Nie, Jing-Tao; Yang, Jun-Jun; Qu, Mei-Ling; He, Huan-Le; Cai, Run

    2015-05-01

    The regulatory gene network of unicellular trichome development in Arabidopsis thaliana has been studied intensively, but that of multicellular remains unclear. In the present study, we characterized cucumber trichomes as representative multicellular and unbranched structures, but in a spontaneous mutant, mict (micro-trichome), all trichomes showed a micro-size and stunted morphologies. We revealed the transcriptome profile using Illumina HiSeq 2000 sequencing technology, and determined that a total of 1391 genes exhibited differential expression. We further validated the accuracy of the transcriptome data by RT-qPCR and found that 43 genes encoding critical transcription factors were likely involved in multicellular trichome development. These 43 candidate genes were subdivided into seven groups: homeodomain, MYB-domain, WRKY-domain, bHLH-domain, ethylene-responsive, zinc finger and other transcription factor genes. Our findings also serve as a powerful tool to further study the relevant molecular networks, and provide a new perspective for investigating this complex and species-specific developmental process.

  8. Entrapment of bed bugs by leaf trichomes inspires microfabrication of biomimetic surfaces.

    PubMed

    Szyndler, Megan W; Haynes, Kenneth F; Potter, Michael F; Corn, Robert M; Loudon, Catherine

    2013-06-06

    Resurgence in bed bug infestations and widespread pesticide resistance have greatly renewed interest in the development of more sustainable, environmentally friendly methods to manage bed bugs. Historically, in Eastern Europe, bed bugs were entrapped by leaves from bean plants, which were then destroyed; this purely physical entrapment was related to microscopic hooked hairs (trichomes) on the leaf surfaces. Using scanning electron microscopy and videography, we documented the capture mechanism: the physical impaling of bed bug feet (tarsi) by these trichomes. This is distinct from a Velcro-like mechanism of non-piercing entanglement, which only momentarily holds the bug without sustained capture. Struggling, trapped bed bugs are impaled by trichomes on several legs and are unable to free themselves. Only specific, mechanically vulnerable locations on the bug tarsi are pierced by the trichomes, which are located at effective heights and orientations for bed bug entrapment despite a lack of any evolutionary association. Using bean leaves as templates, we microfabricated surfaces indistinguishable in geometry from the real leaves, including the trichomes, using polymers with material properties similar to plant cell walls. These synthetic surfaces snag the bed bugs temporarily but do not hinder their locomotion as effectively as real leaves.

  9. Effect of trichome density on soybean pod feeding by adult bean leaf beetles (Coleoptera: Chrysomelidae).

    PubMed

    Lam, W K; Pedigo, L P

    2001-12-01

    The role of soybean, Glycine max (L.) Merrill, pod trichomes on feeding by adult bean leaf beetles. Cerotoma trifurcata (Forster), was evaluated under laboratory and field conditions during 1997 and 1998. Three Clark isolines and 'Corsoy 79' were used to compare the feeding preference of the beetle on pods with different trichome densities. The three Clark isolines, including densely pubescent, sparsely pubescent, and glabrous, were isogenic except for trichome density. The trichome densities on the pods of Clark densely pubescent, Clark sparsely pubescent, and Corsoy 79 were significantly different. In no-choice tests, under both laboratory and field conditions, the feeding on pods of Clark densely pubescent was significantly lower than that on the other isolines. In the choice test of Corsoy 79 conducted in the laboratory and field, when the trichomes on one of the two pods were shaven off, the feeding on the shaven pods was significantly higher than that of the intact ones. In the choice test among Clark isolines under laboratory condition, the result was significant, with the lowest feeding on the densely pubescent pods. This study demonstrates that densely pubescent soybean has the potential to resist bean leaf beetle feeding on pods.

  10. Biological activity of acyl glucose esters from Datura wrightii glandular trichomes against three native insect herbivores.

    PubMed

    Hare, J Daniel

    2005-07-01

    Datura wrightii is dimorphic for leaf trichome type in southern California. "Sticky" plants produce glandular trichomes that secrete acylsugars, whereas velvety plants produce nonglandular trichomes. Glandular trichomes confer resistance to some potential insect herbivores and are associated with reduced feeding in the field by two native coleopteran herbivores: the tobacco flea beetle, Epitrix hirtipennis, and a weevil, Trichobaris compacta. In contrast, another native beetle, Lema daturaphila, damages sticky and velvety plants similarly in the field. A series of choice and no-choice "ester removal" and "ester addition" feeding experiments were performed in the laboratory to evaluate the role of acylsugars in feeding by all three insect species. Consumption of sticky leaves after their esters were removed by washing was compared to consumption of unwashed sticky leaves and velvety leaves in ester removal experiments. Consumption of velvety leaves was measured after acylsugars were applied to those leaves in controlled amounts in the ester addition experiments. Consumption by E. hirtipennis was reduced by acylsugars in all experiments. Consumption by T. compacta was reduced by acylsugars in the ester removal experiments, but not in the ester addition experiments. The location of the acylsugars at the tip of a long trichome, rather than simply on the leaf surface, may be an important component of the biological activity of acylsugars against T. compacta in nature. Consumption by L. daturaphila was not significantly reduced by acylsugars in any experiment. The acylsugars caused no significant mortality of any of the three insect species.

  11. Transcriptional inhibition of the bacteriophage T7 early promoter region by oligonucleotide triple helix formation.

    PubMed

    Ross, C; Samuel, M; Broitman, S L

    1992-12-30

    We have identified a purine-rich triplex binding sequence overlapping a -35 transcriptional early promoter region of the bacteriophage T7. Triplex-forming oligonucleotide designed to bind this target was annealed to T7 templates and introduced into in vitro transcription systems under conditions favoring specific initiation from this promoter. These templates demonstrated significant transcriptional inhibition relative to naked genomic templates and templates mixed with non-triplex-forming oligonucleotide. It is suggested that triplex formation along this target interferes with transcriptional initiation, and this mechanism may hold potential to disrupt bacteriophage T7 early transcription in vivo.

  12. SCCmec-associated psm-mec mRNA promotes Staphylococcus epidermidis biofilm formation.

    PubMed

    Yang, Yongchang; Zhang, Xuemei; Huang, Wenfang; Yin, Yibing

    2016-10-01

    Biofilm formation is considered the major pathogenic mechanism of Staphylococcus epidermidis-associated nosocomial infections. Reports have shown that SCCmec-associated psm-mec regulated methicillin-resistant Staphylococcus aureus virulence and biofilm formation. However, the role of psm-mec in S. epidermidis remains unclear. To this purpose, we analysed 165 clinical isolates of S. epidermidis to study the distribution, mutation and expression of psm-mec and the relationship between this gene and biofilm formation. Next, we constructed three psm-mec deletion mutants, one psm-mec transgene expression strain (p221) and two psm-mec point mutant strains (pM, pAG) to explore its effects on S. epidermidis biofilm formation. Then, the amount of biofilm formation, extracellular DNA (eDNA) and Triton X-100-induced autolysis of the constructed strains was measured. Results of psm-mec deletion and transgene expression showed that the gene regulated S. epidermidis biofilm formation. Compared with the control strains, the ability to form biofilm, Triton X-100-induced autolysis and the amount of eDNA increased in the p221 strain and the two psm-mec mutants pM and pAG expressed psm-mec mRNA without its protein, whereas no differences were observed among the three constructed strains, illustrating that psm-mec mRNA promoted S. epidermidis biofilm formation through up-regulation of bacterial autolysis and the release of eDNA. Our results reveal that acquisition of psm-mec promotes S. epidermidis biofilm formation.

  13. Effect of gibberellic acid and calliterpenone on plant growth attributes, trichomes, essential oil biosynthesis and pathway gene expression in differential manner in Mentha arvensis L.

    PubMed

    Bose, Subir K; Yadav, Ritesh Kumar; Mishra, Smrati; Sangwan, Rajender S; Singh, A K; Mishra, B; Srivastava, A K; Sangwan, Neelam S

    2013-05-01

    Extensive research is going on throughout the world to find out new molecules from natural sources to be used as plant growth promoter. Mentha arvensis L. is the main source of menthol rich essential oil used commercially in various food, pharmaceutical and other preparations. Experiments were conducted on field grown plants for understanding the effect of calliterpenone (CA), a stereo-isomer of abbeokutone, in comparison to gibberellic acid (GA3) on growth attributes, trichomes, essential oil biosynthesis and expression of some oil biosynthetic pathway genes. The exogenous application of CA (1 μM, 10 μM and 100 μM) was found to be better in improving plant biomass and stolon yield, leaf area, branching and leaf stem ratio than with counterpart GA3 at the same concentrations. CA treated plants showed higher glandular trichome number, density and diameter and also correlated with enhanced oil biogenetic capacity as revealed by feeding labeled (14)C-sucrose for 72 h to excised shoots. Semi-quantitative PCR analysis of key pathway genes revealed differential up regulation under CA treatments. Transcript level of menthol dehydrogenase/menthone reductase was found highly up regulated in CA treated plants with increased content of menthone and menthol in oil. These findings demonstrate that CA positively regulated the yields by enhanced branching and higher density of trichomes resulting into higher accumulation of essential oil. The results suggest CA as a novel plant derived diterpenoid with growth promoting action and opens up new possibilities for improving the crop yields and essential oil biosynthesis in qualitative and quantitative manner.

  14. Rapid formation of N-Glycopeptides via Cu(II)-promoted glycosylative ligation.

    PubMed

    Joseph, Ryan; Dyer, Frank Brock; Garner, Philip

    2013-02-15

    Herein is described the chemoselective Cu(II)-HOBt promoted chemical ligation of glycosylamines and peptide thioacids to give N-glycosylated peptides. The method is distinguished from other chemical approaches to peptide N-glycosylation in that (1) it can be employed in the presence of unprotected N-terminal and Lys side chain amines; (2) it is remarkably fast, going to completion in under 30 min; and (3) it produces glycopeptides without attendant aspartimide formation.

  15. A new quantitative classification of ecological types in the bromeliad genus Tillandsia (Bromeliaceae) based on trichomes.

    PubMed

    Stefano, Mosti; Papini, Alessio; Brighigna, Luigi

    2008-03-01

    Using collection specimens, we measured the density and wing area of trichomes in 37 species of the bromeliad genus Tillandsia, specifically the abaxial proximal, abaxial distal, adaxial proximal and adaxial distal parts of the leaf. The product of the trichome "wing" area by the number of trichomes (means) produced a pure number (T) that was correlated to ecological features. The correlation was positive with respect to arid environments (xeric Tillands) and negative with respect to humid environments (mesic Tillands). Bulbous, and particularly myrmecophytic species and species with tanks, represented particular categories. Other intermediate types were identified based on the T number, totalling five ecological types. In comparison with other systems of ecological typification for Tillands and other Bromeliaceae, the present system offers measurable data whose analysis is reproducible.

  16. Effect of heat shock on ultrastructure and calcium distribution in Lavandula pinnata L. glandular trichomes.

    PubMed

    Huang, S S; Kirchoff, B K; Liao, J P

    2013-02-01

    The effects of heat shock (HS) on the ultrastructure and calcium distribution of Lavandula pinnata secretory trichomes are examined using transmission electron microscopy and potassium antimonate precipitation. After 48-h HS at 40°C, plastids become distorted and lack stroma and osmiophilic deposits, the cristae of the mitochondria become indistinct, the endoplasmic reticulum acquires a chain-like appearance with ribosomes prominently attached to the lamellae, and the plasma and organelle membranes become distorted. Heat shock is associated with a decrease in calcium precipitates in the trichomes, while the number of precipitates increases in the mesophyll cells. Prolonged exposure to elevated calcium levels may be toxic to the mesophyll cells, while the lack of calcium in the glands cell may deprive them of the normal protective advantages of elevated calcium levels. The inequality in calcium distribution may result not only from uptake from the transpiration stream, but also from redistribution of calcium from the trichomes to the mesophyll cells.

  17. Cell wall maturation of Arabidopsis trichomes is dependent on exocyst subunit EXO70H4 and involves callose deposition.

    PubMed

    Kulich, Ivan; Vojtíková, Zdeňka; Glanc, Matouš; Ortmannová, Jitka; Rasmann, Sergio; Žárský, Viktor

    2015-05-01

    Arabidopsis (Arabidopsis thaliana) leaf trichomes are single-cell structures with a well-studied development, but little is understood about their function. Developmental studies focused mainly on the early shaping stages, and little attention has been paid to the maturation stage. We focused on the EXO70H4 exocyst subunit, one of the most up-regulated genes in the mature trichome. We uncovered EXO70H4-dependent development of the secondary cell wall layer, highly autofluorescent and callose rich, deposited only in the upper part of the trichome. The boundary is formed between the apical and the basal parts of mature trichome by a callose ring that is also deposited in an EXO70H4-dependent manner. We call this structure the Ortmannian ring (OR). Both the secondary cell wall layer and the OR are absent in the exo70H4 mutants. Ecophysiological aspects of the trichome cell wall thickening include interference with antiherbivore defense and heavy metal accumulation. Ultraviolet B light induces EXO70H4 transcription in a CONSTITUTIVE PHOTOMORPHOGENIC1-dependent way, resulting in stimulation of trichome cell wall thickening and the OR biogenesis. EXO70H4-dependent trichome cell wall hardening is a unique phenomenon, which may be conserved among a variety of the land plants. Our analyses support a concept that Arabidopsis trichome is an excellent model to study molecular mechanisms of secondary cell wall deposition.

  18. Photosynthetic Trichomes Contain a Specific Rubisco with a Modified pH-Dependent Activity1[OPEN

    PubMed Central

    Laterre, Raphaëlle; Remacle, Claire

    2017-01-01

    Ribulose-1,5-biphosphate carboxylase/oxygenase (Rubisco) is the most abundant enzyme in plants and is responsible for CO2 fixation during photosynthesis. This enzyme is assembled from eight large subunits (RbcL) encoded by a single chloroplast gene and eight small subunits (RbcS) encoded by a nuclear gene family. Rubisco is primarily found in the chloroplasts of mesophyll (C3 plants), bundle-sheath (C4 plants), and guard cells. In certain species, photosynthesis also takes place in the secretory cells of glandular trichomes, which are epidermal outgrowths (hairs) involved in the secretion of specialized metabolites. However, photosynthesis and, in particular, Rubisco have not been characterized in trichomes. Here, we show that tobacco (Nicotiana tabacum) trichomes contain a specific Rubisco small subunit, NtRbcS-T, which belongs to an uncharacterized phylogenetic cluster (T). This cluster contains RbcS from at least 33 species, including monocots, many of which are known to possess glandular trichomes. Cluster T is distinct from the cluster M, which includes the abundant, functionally characterized RbcS isoforms expressed in mesophyll or bundle-sheath cells. Expression of NtRbcS-T in Chlamydomonas reinhardtii and purification of the full Rubisco complex showed that this isoform conferred higher Vmax and Km values as well as higher acidic pH-dependent activity than NtRbcS-M, an isoform expressed in the mesophyll. This observation was confirmed with trichome extracts. These data show that an ancient divergence allowed for the emergence of a so-far-uncharacterized RbcS cluster. We propose that secretory trichomes have a particular Rubisco uniquely adapted to secretory cells where CO2 is released by the active specialized metabolism. PMID:28250069

  19. Cofilin-mediated actin dynamics promotes actin bundle formation during Drosophila bristle development

    PubMed Central

    Wu, Jing; Wang, Heng; Guo, Xuan; Chen, Jiong

    2016-01-01

    The actin bundle is an array of linear actin filaments cross-linked by actin-bundling proteins, but its assembly and dynamics are not as well understood as those of the branched actin network. Here we used the Drosophila bristle as a model system to study actin bundle formation. We found that cofilin, a major actin disassembly factor of the branched actin network, promotes the formation and positioning of actin bundles in the developing bristles. Loss of function of cofilin or AIP1, a cofactor of cofilin, each resulted in increased F-actin levels and severe defects in actin bundle organization, with the defects from cofilin deficiency being more severe. Further analyses revealed that cofilin likely regulates actin bundle formation and positioning by the following means. First, cofilin promotes a large G-actin pool both locally and globally, likely ensuring rapid actin polymerization for bundle initiation and growth. Second, cofilin limits the size of a nonbundled actin-myosin network to regulate the positioning of actin bundles. Third, cofilin prevents incorrect assembly of branched and myosin-associated actin filament into bundles. Together these results demonstrate that the interaction between the dynamic dendritic actin network and the assembling actin bundles is critical for actin bundle formation and needs to be closely regulated. PMID:27385345

  20. Nitrite promotes protein carbonylation and Strecker aldehyde formation in experimental fermented sausages: are both events connected?

    PubMed

    Villaverde, A; Ventanas, J; Estévez, M

    2014-12-01

    The role played by curing agents (nitrite, ascorbate) on protein oxidation and Strecker aldehyde formation is studied. To fulfill this objective, increasing concentrations of nitrite (0, 75 and 150ppm) and ascorbate (0, 250 and 500ppm) were added to sausages subjected to a 54day drying process. The concurrence of intense proteolysis, protein carbonylation and formation of Strecker aldehydes during processing of sausages suggests that α-aminoadipic semialdehyde (AAS) and γ-glutamic semialdehyde (GGS) may be implicated in the formation of Strecker aldehydes. The fact that nitrite (150ppm, ingoing amount) significantly promoted the formation of protein carbonyls at early stages of processing and the subsequent formation of Strecker aldehydes provides strength to this hypothesis. Ascorbate (125 and 250ppm) controlled the overall extent of protein carbonylation in sausages without declining the formation of Strecker aldehydes. These results may contribute to understanding the chemistry fundamentals of the positive influence of nitrite on the flavor and overall acceptability of cured muscle foods.

  1. HMGA2 regulates CD44 expression to promote gastric cancer cell motility and sphere formation

    PubMed Central

    Sun, Junying; Sun, Baocun; Zhu, Dongwang; Zhao, Xiulan; Zhang, Yanhui; Dong, Xueyi; Che, Na; Li, Jing; Liu, Fang; Zhao, Nan; Zhang, Danfang; Liu, Tieju; Lin, Xian

    2017-01-01

    High mobility group AT-hook 2 (HMGA2) is a transcriptional modulator that mediates motility and self-renewal in cancer stem cells. Gastric cancer (GC) is the third leading cause of cancer-related deaths worldwide. GC contains a population of stem-like cells that promote tumor invasion and resistance to therapy. In the current study, we investigated the expression of HMGA2 and the cancer stem cell marker CD44 in 200 GC samples and found that HMGA2 and CD44 were significantly associated with distant metastasis, histological differentiation and poor prognosis in GC patients. Positive clinical correlations of HMGA2 with CD44 were also observed in tissue sections. In vitro, overexpression of HMGA2 promoted GC sphere formation and migration in MKN74/MKN28 cells, whereas downregulation of HMGA2 decreased GC sphere formation and migration in MKN45/MGC803 cells. In addition, western blot and immunofluorescent analyses showed that HMGA2 increased the expression of the stem cell markers CD44, ALDH1, Sox2, and Oct4 and the EMT-related factors Snail and β-catenin. In a xenograft mouse model, overexpression of HMGA2 promoted tumor growth. Further immunohistochemical (IHC) analysis showed that HMGA2 increased the expression of CD44 and β-catenin, resulting in the promotion of tumor growth. Taken together, our findings indicate that HMGA2 promotes GC cancer stem cell induction and cell motility by regulating the expression of CD44. Therefore, targeting HMGA2 in GC may be therapeutically beneficial. PMID:28337375

  2. SH2B1 and IRSp53 proteins promote the formation of dendrites and dendritic branches.

    PubMed

    Chen, Chien-Jen; Shih, Chien-Hung; Chang, Yu-Jung; Hong, Shao-Jing; Li, Tian-Neng; Wang, Lily Hui-Ching; Chen, Linyi

    2015-03-06

    SH2B1 is an adaptor protein known to enhance neurite outgrowth. In this study, we provide evidence suggesting that the SH2B1 level is increased during in vitro culture of hippocampal neurons, and the β isoform (SH2B1β) is the predominant isoform. The fact that formation of filopodia is prerequisite for neurite initiation suggests that SH2B1 may regulate filopodium formation and thus neurite initiation. To investigate whether SH2B1 may regulate filopodium formation, the effect of SH2B1 and a membrane and actin regulator, IRSp53 (insulin receptor tyrosine kinase substrate p53), is investigated. Overexpressing both SH2B1β and IRSp53 significantly enhances filopodium formation, neurite outgrowth, and branching. Both in vivo and in vitro data show that SH2B1 interacts with IRSp53 in hippocampal neurons. This interaction depends on the N-terminal proline-rich domains of SH2B1. In addition, SH2B1 and IRSp53 co-localize at the plasma membrane, and their levels increase in the Triton X-100-insoluble fraction of developing neurons. These findings suggest that SH2B1-IRSp53 complexes promote the formation of filopodia, neurite initiation, and neuronal branching.

  3. SH2B1 and IRSp53 Proteins Promote the Formation of Dendrites and Dendritic Branches*

    PubMed Central

    Chen, Chien-Jen; Shih, Chien-Hung; Chang, Yu-Jung; Hong, Shao-Jing; Li, Tian-Neng; Wang, Lily Hui-Ching; Chen, Linyi

    2015-01-01

    SH2B1 is an adaptor protein known to enhance neurite outgrowth. In this study, we provide evidence suggesting that the SH2B1 level is increased during in vitro culture of hippocampal neurons, and the β isoform (SH2B1β) is the predominant isoform. The fact that formation of filopodia is prerequisite for neurite initiation suggests that SH2B1 may regulate filopodium formation and thus neurite initiation. To investigate whether SH2B1 may regulate filopodium formation, the effect of SH2B1 and a membrane and actin regulator, IRSp53 (insulin receptor tyrosine kinase substrate p53), is investigated. Overexpressing both SH2B1β and IRSp53 significantly enhances filopodium formation, neurite outgrowth, and branching. Both in vivo and in vitro data show that SH2B1 interacts with IRSp53 in hippocampal neurons. This interaction depends on the N-terminal proline-rich domains of SH2B1. In addition, SH2B1 and IRSp53 co-localize at the plasma membrane, and their levels increase in the Triton X-100-insoluble fraction of developing neurons. These findings suggest that SH2B1-IRSp53 complexes promote the formation of filopodia, neurite initiation, and neuronal branching. PMID:25586189

  4. Nickel Promotes Biofilm Formation by Escherichia coli K-12 Strains That Produce Curli▿

    PubMed Central

    Perrin, Claire; Briandet, Romain; Jubelin, Gregory; Lejeune, Philippe; Mandrand-Berthelot, Marie-Andrée; Rodrigue, Agnès; Dorel, Corinne

    2009-01-01

    The survival of bacteria exposed to toxic compounds is a multifactorial phenomenon, involving well-known molecular mechanisms of resistance but also less-well-understood mechanisms of tolerance that need to be clarified. In particular, the contribution of biofilm formation to survival in the presence of toxic compounds, such as nickel, was investigated in this study. We found that a subinhibitory concentration of nickel leads Escherichia coli bacteria to change their lifestyle, developing biofilm structures rather than growing as free-floating cells. Interestingly, whereas nickel and magnesium both alter the global cell surface charge, only nickel promotes biofilm formation in our system. Genetic evidence indicates that biofilm formation induced by nickel is mediated by the transcriptional induction of the adhesive curli-encoding genes. Biofilm formation induced by nickel does not rely on efflux mechanisms using the RcnA pump, as these require a higher concentration of nickel to be activated. Our results demonstrate that the nickel-induced biofilm formation in E. coli is an adaptational process, occurring through a transcriptional effect on genes coding for adherence structures. The biofilm lifestyle is obviously a selective advantage in the presence of nickel, but the means by which it improves bacterial survival needs to be investigated. PMID:19168650

  5. Dapper1 promotes autophagy by enhancing the Beclin1-Vps34-Atg14L complex formation.

    PubMed

    Ma, Benyu; Cao, Weipeng; Li, Wenxia; Gao, Chan; Qi, Zhen; Zhao, Yan; Du, Jun; Xue, Hua; Peng, Junya; Wen, Jun; Chen, Hua; Ning, Yuanheng; Huang, Lei; Zhang, Hong; Gao, Xiang; Yu, Li; Chen, Ye-Guang

    2014-08-01

    Autophagy is an intracellular degradation process to clear up aggregated proteins or aged and damaged organelles. The Beclin1-Vps34-Atg14L complex is essential for autophagosome formation. However, how the complex formation is regulated is unclear. Here, we show that Dapper1 (Dpr1) acts as a critical regulator of the Beclin1-Vps34-Atg14L complex to promote autophagy. Dpr1 ablation in the central nervous system results in motor coordination defect and accumulation of p62 and ubiquitinated proteins. Dpr1 increases autophagosome formation as indicated by elevated puncta formation of LC3, Atg14L and DFCP1 (Double FYVE-containing protein 1). Conversely, loss of Dpr1 impairs LC3 lipidation and causes p62/SQSTM1 accumulation. Dpr1 directly interacts with Beclin1 and Atg14L and enhances the Beclin1-Vps34 interaction and Vps34 activity. Together, our findings suggest that Dpr1 enhances the Atg14L-Beclin1-Vps34 complex formation to drive autophagy.

  6. Nickel promotes biofilm formation by Escherichia coli K-12 strains that produce curli.

    PubMed

    Perrin, Claire; Briandet, Romain; Jubelin, Gregory; Lejeune, Philippe; Mandrand-Berthelot, Marie-Andrée; Rodrigue, Agnès; Dorel, Corinne

    2009-03-01

    The survival of bacteria exposed to toxic compounds is a multifactorial phenomenon, involving well-known molecular mechanisms of resistance but also less-well-understood mechanisms of tolerance that need to be clarified. In particular, the contribution of biofilm formation to survival in the presence of toxic compounds, such as nickel, was investigated in this study. We found that a subinhibitory concentration of nickel leads Escherichia coli bacteria to change their lifestyle, developing biofilm structures rather than growing as free-floating cells. Interestingly, whereas nickel and magnesium both alter the global cell surface charge, only nickel promotes biofilm formation in our system. Genetic evidence indicates that biofilm formation induced by nickel is mediated by the transcriptional induction of the adhesive curli-encoding genes. Biofilm formation induced by nickel does not rely on efflux mechanisms using the RcnA pump, as these require a higher concentration of nickel to be activated. Our results demonstrate that the nickel-induced biofilm formation in E. coli is an adaptational process, occurring through a transcriptional effect on genes coding for adherence structures. The biofilm lifestyle is obviously a selective advantage in the presence of nickel, but the means by which it improves bacterial survival needs to be investigated.

  7. Mesenchymal Deletion of Histone Demethylase NO66 in Mice Promotes Bone Formation

    PubMed Central

    Chen, Qin; Sinha, Krishna; Deng, Jian Min; Yasuda, Hideyo; Krahe, Ralf; Behringer, Richard R.; de Crombrugghe, Benoit

    2016-01-01

    Our previous studies indicated that the Jumonji C (JmjC)-domain-containing NO66 is a histone demethylase with specificity for methylated histone H3K4 and H3K36. NO66 binds to the transcription factor Osterix (Osx) and inhibits its transcriptional activity in promoter assays. However, the physiological role of NO66 in formation of mammalian bones is unknown. Here, using a genetically engineered mouse model, we show that during early skeletal development, Prx1-Cre dependent mesenchymal deletion of NO66 promotes osteogenesis and formation of both endochondral as well as intramembranous skeletal elements, leading to a larger skeleton and a high bone mass phenotype in adult mice. The excess bone formation in mice where NO66 was deleted in cells of mesenchymal origin is associated with an increase in the number of preosteoblasts and osteoblasts. Further analysis revealed that in the embryonic limbs and adult calvaria of mice with deletion of NO66 in cells of mesenchymal origin, expression of several genes including bone morphogenetic protein 2, insulin-like growth factor 1 and osteoclast inhibitor osteoprotegerin was increased, concurrent with an increase in expression of bone formation markers such as Osx, type I collagen and bone sialoprotein. Taken together, our results provide the first in vivo evidence that NO66 histone demethylase plays an important role in mammalian osteogenesis during early development as well as in adult bone homeostasis. We postulate that NO66 regulates bone formation, at least in part, via regulating the number of bone-forming cells and expression of multiple genes that are critical for these processes. PMID:25736226

  8. Kartogenin with PRP promotes the formation of fibrocartilage zone in the tendon-bone interface.

    PubMed

    Zhou, Yiqin; Zhang, Jianying; Yang, Jinsong; Narava, Manoj; Zhao, Guangyi; Yuan, Ting; Wu, Haishan; Zheng, Nigel; Hogan, MaCalus V; Wang, James H-C

    2017-01-27

    Treatment of tendon-bone junction injuries is a challenge because tendon-bone interface often heals poorly and the fibrocartilage zone, which reduces stress concentration, at the interface is not formed. In this study, we used a compound called kartogenin (KGN) with platelet-rich plasma (PRP) to induce the formation of fibrocartilage zone in a rat tendon graft-bone tunnel model. The experimental rats received KGN-PRP or PRP injections in the tendon graft-bone tunnel interface. The control group received saline. After 4, 8 and 12 weeks, Safranin O staining of the tendon graft-bone tunnels revealed abundant proteoglycans in the KGN-PRP group indicating the formation of cartilage-like transition zone. Immunohistochemical and immuno-fluorescence staining revealed collagen types I (Col-I) and II (Col-II) in the newly formed fibrocartilage zone. Both fibrocartilage zone formation and maturation were healing time dependent. In contrast, the PRP and saline control groups had no cartilage-like tissues and minimal Col-I and Col-II staining. Some gaps were also present in the saline control group. Finally, pull-out strength in the KGN-PRP-treated group at 8 weeks was 1.4-fold higher than the PRP-treated group and 1.6-fold higher than the saline control group. These findings indicate that KGN, with PRP as a carrier, promotes the formation of fibrocartilage zone between the tendon graft and bone interface. Thus, KGN-PRP may be used as a convenient cell-free therapy in clinics to promote fibrocartilage zone formation in rotator calf repair and anterior cruciate ligament reconstruction, thereby enhancing the mechanical strength of the tendon-bone interface and hence the clinical outcome of these procedures. Copyright © 2017 John Wiley & Sons, Ltd.

  9. Erythropoietin promotes bone formation through EphrinB2/EphB4 signaling.

    PubMed

    Li, C; Shi, C; Kim, J; Chen, Y; Ni, S; Jiang, L; Zheng, C; Li, D; Hou, J; Taichman, R S; Sun, H

    2015-03-01

    Recent studies have demonstrated that erythropoietin (EPO) has extensive nonhematopoietic biological functions. However, little is known about how EPO regulates bone formation, although several studies suggested that EPO can affect bone homeostasis. In this study, we investigated the effects of EPO on the communication between osteoclasts and osteoblasts through the ephrinB2/EphB4 signaling pathway. We found that EPO slightly promotes osteoblastic differentiation with the increased expression of EphB4 in ST2 cells. However, EPO increased the expression of Nfatc1 and ephrinB2 but decreased the expression of Mmp9 in RAW264.7 cells, resulting in an increase of ephrinB2-expressing osteoclasts and a decrease in resorption activity. The stimulation of ephrinB2/EphB4 signaling via ephrinB2-Fc significantly promoted EPO-mediated osteoblastic differentiation in ST2 cells. EphB4 knockdown through EphB4 shRNA inhibited EPO-mediated osteoblastic phenotypes. Furthermore, in vivo assays clearly demonstrated that EPO efficiently induces new bone formation in the alveolar bone regeneration model. Taken together, these results suggest that ephrinB2/EphB4 signaling may play an important role in EPO-mediated bone formation.

  10. Semaphorin3A regulates neuronal polarization by suppressing axon formation and promoting dendrite growth.

    PubMed

    Shelly, Maya; Cancedda, Laura; Lim, Byung Kook; Popescu, Andrei T; Cheng, Pei-lin; Gao, Hongfeng; Poo, Mu-ming

    2011-08-11

    Semaphorin 3A (Sema3A) is a secreted factor known to guide axon/dendrite growth and neuronal migration. We found that it also acts as a polarizing factor for axon/dendrite development in cultured hippocampal neurons. Exposure of the undifferentiated neurite to localized Sema3A suppressed its differentiation into axon and promoted dendrite formation, resulting in axon formation away from the Sema3A source, and bath application of Sema3A to polarized neurons promoted dendrite growth but suppressed axon growth. Fluorescence resonance energy transfer (FRET) imaging showed that Sema3A elevated the cGMP but reduced cAMP and protein kinase A (PKA) activity, and its axon suppression is attributed to the downregulation of PKA-dependent phosphorylation of axon determinants LKB1 and GSK-3β. Downregulating Sema3A signaling in rat embryonic cortical progenitors via in utero electroporation of siRNAs against the Sema3A receptor neuropilin-1 also resulted in polarization defects in vivo. Thus, Sema3A regulates the earliest step of neuronal morphogenesis by polarizing axon/dendrite formation.

  11. Erythropoietin Promotes Bone Formation through EphrinB2/EphB4 Signaling

    PubMed Central

    Li, C.; Shi, C.; Kim, J.; Chen, Y.; Ni, S.; Jiang, L.; Zheng, C.; Li, D.; Hou, J.; Sun, H.

    2015-01-01

    Recent studies have demonstrated that erythropoietin (EPO) has extensive nonhematopoietic biological functions. However, little is known about how EPO regulates bone formation, although several studies suggested that EPO can affect bone homeostasis. In this study, we investigated the effects of EPO on the communication between osteoclasts and osteoblasts through the ephrinB2/EphB4 signaling pathway. We found that EPO slightly promotes osteoblastic differentiation with the increased expression of EphB4 in ST2 cells. However, EPO increased the expression of Nfatc1 and ephrinB2 but decreased the expression of Mmp9 in RAW264.7 cells, resulting in an increase of ephrinB2-expressing osteoclasts and a decrease in resorption activity. The stimulation of ephrinB2/EphB4 signaling via ephrinB2-Fc significantly promoted EPO-mediated osteoblastic differentiation in ST2 cells. EphB4 knockdown through EphB4 shRNA inhibited EPO-mediated osteoblastic phenotypes. Furthermore, in vivo assays clearly demonstrated that EPO efficiently induces new bone formation in the alveolar bone regeneration model. Taken together, these results suggest that ephrinB2/EphB4 signaling may play an important role in EPO-mediated bone formation. PMID:25586589

  12. Expression of Brassica napus TTG2, a regulator of trichome development, increases plant sensitivity to salt stress by suppressing the expression of auxin biosynthesis genes.

    PubMed

    Li, Qingyuan; Yin, Mei; Li, Yongpeng; Fan, Chuchuan; Yang, Qingyong; Wu, Jian; Zhang, Chunyu; Wang, Hong; Zhou, Yongming

    2015-09-01

    WRKY transcription factors (TFs) are plant specific and play important roles in regulating diverse biological processes. To identify TFs with broad-spectrum effects on various stress responses in Brassica napus, an important oil crop grown across diverse ecological regions worldwide, we functionally characterized Bna.TTG2 genes, which are homologous to the Arabidopsis AtTTG2 (WRKY44) gene. Four Bna.TTG2 genes were capable of rescuing the trichome phenotypes of Arabidopsis ttg2 mutants. Overexpressing one Bna.TTG2 family member, BnaA.TTG2.a.1, remarkably increased trichome numbers in Arabidopsis and B. napus plants. Interestingly, the BnaA.TTG2.a.1-overexpressing plants of both species exhibited increased sensitivity to salt stress. In BnaA.TTG2.a.1-overexpressing Arabidopsis under salt stress, the endogenous indole-3-acetic acid (IAA) content was reduced, and the expression of two auxin biosynthesis genes, TRYPTOPHAN BIOSYNTHESIS 5 (TRP5) and YUCCA2 (YUC2), was downregulated. The results from yeast one-hybrid, electrophoretic mobility shift, and dual-luciferase reporter assays revealed that BnaA.TTG2.a.1 is able to bind to the promoters of TRP5 and YUC2. These data indicated that BnaA.TTG2.a.1 confers salt sensitivity to overexpressing plants by suppressing the expression of IAA synthesis genes and thus lowering IAA levels. Transgenic Arabidopsis plants with an N-terminus-deleted BnaA.TTG2.a.1 no longer showed hypersensitivity to salt stress, suggesting that the N terminus of BnaA.TTG2.a.1 plays a critical role in salt stress responses. Therefore, in addition to its classical function in trichome development, our study reveals a novel role for Bna.TTG2 genes in salt stress responses.

  13. Phenylpropanoid biosynthesis in leaves and glandular trichomes of basil (Ocimum basilicum L.).

    PubMed

    Deschamps, Cícero; Simon, James E

    2010-01-01

    Basil (Ocimum basilicum L.) essential oil phenylpropenes are synthesized and accumulate in peltate glandular trichomes and their content and composition depend on plant developmental stage. Studies on gene expression and enzymatic activity indicate that the phenylpropene biosynthetic genes are developmentally regulated. In this study, the methylchavicol accumulation in basil leaves and the enzyme activities and gene expression of both chavicol O-methyltransferase (CVOMT) and eugenol O-methyltransferase (EOMT) were investigated in all leaves at four plant developmental stages. Methylchavicol accumulation decreased over time as leaves matured. There was a significant correlation between methylchavicol accumulation and CVOMT (r(2) = 0.88) enzyme activity, suggesting that the levels of biosynthetic enzymes control the essential oil content. CVOMT and EOMT transcript expression levels, which decreased with leaf age, followed the same pattern in both whole leaves and isolated glandular trichomes, providing evidence that CVOMT transcript levels are developmentally regulated in basil glandular trichomes themselves and that differences in CVOMT expression observed in whole leaves are not solely the result of differences in glandular trichome density.

  14. Biochemical and histochemical localization of monoterpene biosynthesis in the glandular trichomes of spearmint (Mentha spicata)

    SciTech Connect

    Gershenzon, J.; Maffei, M.; Croteau, R. )

    1989-04-01

    The primary monoterpene accumulated in the glandular trichomes of spearmint (Mentha spicata) is the ketone (-)-carvone which is formed by cyclization of the C{sub 10} isoprenoid intermediate geranyl pyrophosphate to the olefin (-)-limonene, hydroxylation to (-)-trans-carveol and subsequent dehydrogenation. Selective extraction of the contents of the glandular trichomes indicated that essentially all of the cyclase and hydroxylase activities resided in these structures, whereas only about 30% of the carveol dehydrogenase was located here with the remainder located in the rest of the leaf. This distribution of carveol dehydrogenase activity was confirmed by histochemical methods. Electrophoretic analysis of the partially purified carveol dehydrogenase from extracts of both the glands and the leaves following gland removal indicated the presence of a unique carveol dehydrogenase species in the glandular trichomes, suggesting that the other dehydrogenase found throughout the leaf probably utilizes carveol only as an adventitious substrate. These results demonstrate that carvone biosynthesis takes place exclusively in the glandular trichomes in which this natural product accumulates.

  15. Phytotoxicity of Constituents of Glandular Trichomes and the Leaf Surface of Camphorweed, Heterotheca subaxillaris

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Camphorweed, Heterotheca subaxillaris (Lam.) Britt. & Rusby has a camphor-like odor, and its leaf surfaces contain glandular trichomes of the type shown to contain high levels of isoprenoids in other species. Phytotoxic calamenene-type sesquiterpenes (1-4, 8-10), borneol (11) and methylated flavone...

  16. Properties of synthetic ferrihydrite as an amino acid adsorbent and a promoter of peptide bond formation.

    PubMed

    Matrajt, G; Blanot, D

    2004-03-01

    Ferrihydrite, an iron oxide hydroxide, is found in all kinds of environments, from hydrothermal hot springs to extraterrestrial materials. It has been shown that this material is nanoporous, and because of its high surface area, it has outstanding adsorption properties and in some cases catalysis properties. In this work we studied the adsorption properties of ferrihydrite with respect to amino acids. Samples of pure ferrihydrite were synthesised and exposed to solutions of amino acids including both proteinaceous and non-proteinaceous species. These experiments revealed important characteristics of this mineral as both an adsorbent of amino acids and a promoter of peptide bond formation.

  17. Chibby promotes ciliary vesicle formation and basal body docking during airway cell differentiation.

    PubMed

    Burke, Michael C; Li, Feng-Qian; Cyge, Benjamin; Arashiro, Takeshi; Brechbuhl, Heather M; Chen, Xingwang; Siller, Saul S; Weiss, Matthew A; O'Connell, Christopher B; Love, Damon; Westlake, Christopher J; Reynolds, Susan D; Kuriyama, Ryoko; Takemaru, Ken-Ichi

    2014-10-13

    Airway multiciliated epithelial cells play crucial roles in the mucosal defense system, but their differentiation process remains poorly understood. Mice lacking the basal body component Chibby (Cby) exhibit impaired mucociliary transport caused by defective ciliogenesis, resulting in chronic airway infection. In this paper, using primary cultures of mouse tracheal epithelial cells, we show that Cby facilitates basal body docking to the apical cell membrane through proper formation of ciliary vesicles at the distal appendage during the early stages of ciliogenesis. Cby is recruited to the distal appendages of centrioles via physical interaction with the distal appendage protein CEP164. Cby then associates with the membrane trafficking machinery component Rabin8, a guanine nucleotide exchange factor for the small guanosine triphosphatase Rab8, to promote recruitment of Rab8 and efficient assembly of ciliary vesicles. Thus, our study identifies Cby as a key regulator of ciliary vesicle formation and basal body docking during the differentiation of airway ciliated cells.

  18. Collagen-derived matricryptins promote inhibitory nerve terminal formation in the developing neocortex

    PubMed Central

    Su, Jianmin; Chen, Jiang; Lippold, Kumiko; Monavarfeshani, Aboozar; Carrillo, Gabriela Lizana; Jenkins, Rachel

    2016-01-01

    Inhibitory synapses comprise only ∼20% of the total synapses in the mammalian brain but play essential roles in controlling neuronal activity. In fact, perturbing inhibitory synapses is associated with complex brain disorders, such as schizophrenia and epilepsy. Although many types of inhibitory synapses exist, these disorders have been strongly linked to defects in inhibitory synapses formed by Parvalbumin-expressing interneurons. Here, we discovered a novel role for an unconventional collagen—collagen XIX—in the formation of Parvalbumin+ inhibitory synapses. Loss of this collagen results not only in decreased inhibitory synapse number, but also in the acquisition of schizophrenia-related behaviors. Mechanistically, these studies reveal that a proteolytically released fragment of this collagen, termed a matricryptin, promotes the assembly of inhibitory nerve terminals through integrin receptors. Collectively, these studies not only identify roles for collagen-derived matricryptins in cortical circuit formation, but they also reveal a novel paracrine mechanism that regulates the assembly of these synapses. PMID:26975851

  19. Formation of a seed germination promoter from carbohydrates and amino acids.

    PubMed

    Light, Marnie E; Burger, Barend V; van Staden, Johannes

    2005-07-27

    The ability of plant-derived smoke to act as a germination cue in many species has led to widespread interest in this aspect of seed biology. Recently, 3-methyl-2H-furo[2,3-c]pyran-2-one was identified as the main germination cue from smoke. Here, we report on the formation of this compound from reactions of sugars with amino acids. Heating proteins or amino acids with sugars at 180 degrees C for 30 min produces water soluble extracts that promote germination. High-performance liquid chromatography indicated that the active compound(s) derived from these reactions coeluted with the active fraction from a smoke solution. Gas chromatography-mass spectroscopy showed that the active constituent is identical to the germination cue from plant-derived smoke. The results presented in this paper provide evidence for the formation of the major germination cue found in smoke from ubiquitously occurring organic compounds.

  20. Differential regulation of a MYB transcription factor is correlated with transgenerational epigenetic inheritance of trichome density in Mimulus guttatus

    PubMed Central

    Scoville, Alison G.; Barnett, Laryssa L.; Bodbyl-Roels, Sarah; Kelly, John K.; Hileman, Lena C.

    2011-01-01

    Summary Epigenetic inheritance, transgenerational transmission of traits not proximally determined by DNA sequence, has been linked to transmission of chromatin modifications and gene regulation, which are known to be sensitive to environmental factors. Mimulus guttatus increases trichome (plant hair) density in response to simulated herbivore damage. Increased density is expressed in progeny even if progeny do not experience damage. To better understand epigenetic inheritance of trichome production, we tested the hypothesis that candidate gene expression states are inherited in response to parental damage.Using M. guttatus recombinant inbred lines, offspring of leaf-damaged and control plants were raised without damage. Relative expression of candidate trichome development genes was measured in offspring. Line and parental damage effects on trichome density were measured. Associations between gene expression, trichome density, and response to parental damage were determined.We identified M. guttatus MYB MIXTA-like 8 as a possible negative regulator of trichome development. We found that parental leaf damage induces down-regulation of MYB MIXTA-like 8 in progeny, which is associated with epigenetically inherited increased trichome density.Our results link epigenetic transmission of an ecologically important trait with differential gene expression states – providing insight into a mechanism underlying environmentally induced ‘soft inheritance’. PMID:21352232

  1. Genome-Wide Survey Reveals Transcriptional Differences Underlying the Contrasting Trichome Phenotypes of Two Sister Desert Poplars

    PubMed Central

    Ma, Jianchao; He, Xiaodong; Bai, Xiaotao; Niu, Zhimin; Duan, Bingbing; Chen, Ningning; Shao, Xuemin; Wan, Dongshi

    2016-01-01

    Trichomes, which are widely used as an important diagnostic characteristic in plant species delimitation, play important roles in plant defense and adaptation to adverse environments. In this study, we used two sister poplar species, Populus pruinosa and Populus euphratica—which have, respectively, dense and sparse trichomes—to examine the genetic differences associated with these contrasting phenotypes. The results showed that 42 and 45 genes could be identified as candidate genes related to trichomes in P. pruinosa and P. euphratica, respectively; most of these genes possessed high degrees of diversification in their coding sequences, but they were similar in intron/exon structure in the two species. We also found that most of the candidate trichome genes were expressed at higher levels in P. pruinosa, which has dense trichomes, than in P. euphratica, where there are few trichomes. Based on analyses of transcriptional profiles, a total of 195 genes, including many transcription factors, were found to show distinct differences in expression. The results of gene function annotation suggested that the genes identified as having contrasting levels of expression level are mainly associated with trichome elongation, ATPase activity, and hormone transduction. Changes in the expression of these and other related genes with high sequence diversification may have contributed to the contrast in the pattern of trichome phenotypes between the two species. PMID:27916935

  2. Natural allelic variation defines a role for ATMYC1: trichome cell fate determination.

    PubMed

    Symonds, V Vaughan; Hatlestad, Greg; Lloyd, Alan M

    2011-06-01

    The molecular nature of biological variation is not well understood. Indeed, many questions persist regarding the types of molecular changes and the classes of genes that underlie morphological variation within and among species. Here we have taken a candidate gene approach based on previous mapping results to identify the gene and ultimately a polymorphism that underlies a trichome density QTL in Arabidopsis thaliana. Our results show that natural allelic variation in the transcription factor ATMYC1 alters trichome density in A. thaliana; this is the first reported function for ATMYC1. Using site-directed mutagenesis and yeast two-hybrid experiments, we demonstrate that a single amino acid replacement in ATMYC1, discovered in four ecotypes, eliminates known protein-protein interactions in the trichome initiation pathway. Additionally, in a broad screen for molecular variation at ATMYC1, including 72 A. thaliana ecotypes, a high-frequency block of variation was detected that results in >10% amino acid replacement within one of the eight exons of the gene. This sequence variation harbors a strong signal of divergent selection but has no measurable effect on trichome density. Homologs of ATMYC1 are pleiotropic, however, so this block of variation may be the result of natural selection having acted on another trait, while maintaining the trichome density role of the gene. These results show that ATMYC1 is an important source of variation for epidermal traits in A. thaliana and indicate that the transcription factors that make up the TTG1 genetic pathway generally may be important sources of epidermal variation in plants.

  3. Interferon-β promotes macrophage foam cell formation by altering both cholesterol influx and efflux mechanisms.

    PubMed

    Boshuizen, Marieke C S; Hoeksema, Marten A; Neele, Annette E; van der Velden, Saskia; Hamers, Anouk A J; Van den Bossche, Jan; Lutgens, Esther; de Winther, Menno P J

    2016-01-01

    Foam cell formation is a crucial event in atherogenesis. While interferon-β (IFNβ) is known to promote atherosclerosis in mice, studies on the role of IFNβ on foam cell formation are minimal and conflicting. We therefore extended these studies using both in vitro and in vivo approaches and examined IFNβ's function in macrophage foam cell formation. To do so, murine bone marrow-derived macrophages (BMDMs) and human monocyte-derived macrophages were loaded with acLDL overnight, followed by 6h IFNβ co-treatment. This increased lipid content as measured by Oil red O staining. We next analyzed the lipid uptake pathways of IFNβ-stimulated BMDMs and observed increased endocytosis of DiI-acLDL as compared to controls. These effects were mediated via SR-A, as its gene expression was increased and inhibition of SR-A with Poly(I) blocked the IFNβ-induced increase in Oil red O staining and DiI-acLDL endocytosis. The IFNβ-induced increase in lipid content was also associated with decreased ApoA1-mediated cholesterol efflux, in response to decreased ABCA1 protein and gene expression. To validate our findings in vivo, LDLR(-/-) mice were put on chow or a high cholesterol diet for 10weeks. 24 and 8h before sacrifice mice were injected with IFNβ or PBS, after which thioglycollate-elicited peritoneal macrophages were collected and analyzed. In accordance with the in vitro data, IFNβ increased lipid accumulation. In conclusion, our experimental data support the pro-atherogenic role of IFNβ, as we show that IFNβ promotes macrophage foam cell formation by increasing SR-A-mediated cholesterol influx and decreasing ABCA1-mediated efflux mechanisms.

  4. Cisplatin upregulates mitochondrial nitric oxide synthase and peroxynitrite formation to promote renal injury.

    PubMed

    Jung, Michaela; Hotter, Georgina; Viñas, Jose Luis; Sola, Anna

    2009-01-15

    The mitochondria are a critical target for cisplatin-associated nephrotoxicity. Though nitric oxide formation has been implicated in the toxicity of cisplatin, this formation has not so far been related to a possible activation of mitochondrial nitric oxide synthase (mNOS). We show here that the upregulation of oxide mNOS and peroxynitrite formation in cisplatin treatment are key events that influence the development of the harmful parameters described in cisplatin-associated kidney failure. We confirm this by isolating the mitochondrial fraction of the kidney and across different access routes such as the use of a specific inhibitor of neuronal NOS, L-NPA, a peroxynitrite scavenger, FeTMPyP, and a peroxynitrite donor, SIN-1. The in vitro studies corroborated the information obtained in the in vivo experiments. The administration of cisplatin reveals a clear upregulation in the transcription of neuronal NOS and an increase in the levels of nitrites in the mitochondrial fractions of the kidneys. The upregulated transcription directly affects the cytoskeleton structure and the apoptosis. The inhibition of neuronal NOS reduces the levels of nitrites, cell death, and cytoskeleton derangement. Peroxynitrite is involved in the mechanism promoting the NOS transcription. In addition, in controls SIN-1 imitates the effects of cisplatin. In summary, we demonstrate that upregulation of mNOS in cisplatin treatment is a key component in both the initiation and the spread of cisplatin-associated damage in the kidney. Furthermore, peroxynitrite formation is directly involved in this process.

  5. Cisplatin upregulates mitochondrial nitric oxide synthase and peroxynitrite formation to promote renal injury

    SciTech Connect

    Jung, Michaela; Sola, Anna

    2009-01-15

    The mitochondria are a critical target for cisplatin-associated nephrotoxicity. Though nitric oxide formation has been implicated in the toxicity of cisplatin, this formation has not so far been related to a possible activation of mitochondrial nitric oxide synthase (mNOS). We show here that the upregulation of oxide mNOS and peroxynitrite formation in cisplatin treatment are key events that influence the development of the harmful parameters described in cisplatin-associated kidney failure. We confirm this by isolating the mitochondrial fraction of the kidney and across different access routes such as the use of a specific inhibitor of neuronal NOS, L-NPA, a peroxynitrite scavenger, FeTMPyP, and a peroxynitrite donor, SIN-1. The in vitro studies corroborated the information obtained in the in vivo experiments. The administration of cisplatin reveals a clear upregulation in the transcription of neuronal NOS and an increase in the levels of nitrites in the mitochondrial fractions of the kidneys. The upregulated transcription directly affects the cytoskeleton structure and the apoptosis. The inhibition of neuronal NOS reduces the levels of nitrites, cell death, and cytoskeleton derangement. Peroxynitrite is involved in the mechanism promoting the NOS transcription. In addition, in controls SIN-1 imitates the effects of cisplatin. In summary, we demonstrate that upregulation of mNOS in cisplatin treatment is a key component in both the initiation and the spread of cisplatin-associated damage in the kidney. Furthermore, peroxynitrite formation is directly involved in this process.

  6. Biosurfactant as a Promoter of Methane Hydrate Formation: Thermodynamic and Kinetic Studies

    PubMed Central

    Arora, Amit; Cameotra, Swaranjit Singh; Kumar, Rajnish; Balomajumder, Chandrajit; Singh, Anil Kumar; Santhakumari, B.; Kumar, Pushpendra; Laik, Sukumar

    2016-01-01

    Natural gas hydrates (NGHs) are solid non-stoichiometric compounds often regarded as a next generation energy source. Successful commercialization of NGH is curtailed by lack of efficient and safe technology for generation, dissociation, storage and transportation. The present work studied the influence of environment compatible biosurfactant on gas hydrate formation. Biosurfactant was produced by Pseudomonas aeruginosa strain A11 and was characterized as rhamnolipids. Purified rhamnolipids reduced the surface tension of water from 72 mN/m to 36 mN/m with Critical Micelle Concentration (CMC) of 70 mg/l. Use of 1000 ppm rhamnolipids solution in C type silica gel bed system increased methane hydrate formation rate by 42.97% and reduced the induction time of hydrate formation by 22.63% as compared to water saturated C type silica gel. Presence of rhamnolipids also shifted methane hydrate formation temperature to higher values relative to the system without biosurfactant. Results from thermodynamic and kinetic studies suggest that rhamnolipids can be applied as environment friendly methane hydrate promoter. PMID:26869357

  7. Biosurfactant as a Promoter of Methane Hydrate Formation: Thermodynamic and Kinetic Studies.

    PubMed

    Arora, Amit; Cameotra, Swaranjit Singh; Kumar, Rajnish; Balomajumder, Chandrajit; Singh, Anil Kumar; Santhakumari, B; Kumar, Pushpendra; Laik, Sukumar

    2016-02-12

    Natural gas hydrates (NGHs) are solid non-stoichiometric compounds often regarded as a next generation energy source. Successful commercialization of NGH is curtailed by lack of efficient and safe technology for generation, dissociation, storage and transportation. The present work studied the influence of environment compatible biosurfactant on gas hydrate formation. Biosurfactant was produced by Pseudomonas aeruginosa strain A11 and was characterized as rhamnolipids. Purified rhamnolipids reduced the surface tension of water from 72 mN/m to 36 mN/m with Critical Micelle Concentration (CMC) of 70 mg/l. Use of 1000 ppm rhamnolipids solution in C type silica gel bed system increased methane hydrate formation rate by 42.97% and reduced the induction time of hydrate formation by 22.63% as compared to water saturated C type silica gel. Presence of rhamnolipids also shifted methane hydrate formation temperature to higher values relative to the system without biosurfactant. Results from thermodynamic and kinetic studies suggest that rhamnolipids can be applied as environment friendly methane hydrate promoter.

  8. Biosurfactant as a Promoter of Methane Hydrate Formation: Thermodynamic and Kinetic Studies

    NASA Astrophysics Data System (ADS)

    Arora, Amit; Cameotra, Swaranjit Singh; Kumar, Rajnish; Balomajumder, Chandrajit; Singh, Anil Kumar; Santhakumari, B.; Kumar, Pushpendra; Laik, Sukumar

    2016-02-01

    Natural gas hydrates (NGHs) are solid non-stoichiometric compounds often regarded as a next generation energy source. Successful commercialization of NGH is curtailed by lack of efficient and safe technology for generation, dissociation, storage and transportation. The present work studied the influence of environment compatible biosurfactant on gas hydrate formation. Biosurfactant was produced by Pseudomonas aeruginosa strain A11 and was characterized as rhamnolipids. Purified rhamnolipids reduced the surface tension of water from 72 mN/m to 36 mN/m with Critical Micelle Concentration (CMC) of 70 mg/l. Use of 1000 ppm rhamnolipids solution in C type silica gel bed system increased methane hydrate formation rate by 42.97% and reduced the induction time of hydrate formation by 22.63% as compared to water saturated C type silica gel. Presence of rhamnolipids also shifted methane hydrate formation temperature to higher values relative to the system without biosurfactant. Results from thermodynamic and kinetic studies suggest that rhamnolipids can be applied as environment friendly methane hydrate promoter.

  9. Autophagy is induced by anti-neutrophil cytoplasmic Abs and promotes neutrophil extracellular traps formation.

    PubMed

    Sha, Li-Li; Wang, Huan; Wang, Chen; Peng, Hong-Ying; Chen, Min; Zhao, Ming-Hui

    2016-11-01

    Dysregulated neutrophil extracellular traps (NETs) formation contributes to the pathogenesis of anti-neutrophil cytoplasmic Ab (ANCA)-associated vasculitis (AAV). Increasing evidence indicates that autophagy is involved in the process of NETs formation. In this study, we aimed to investigate whether ANCA could induce autophagy in the process of NETs formation. Autophagy was detected using live cell imaging, microtubule-associated protein light chain 3B (LC3B) accumulation and Western blotting. The results showed that autophagy vacuolization was detected in neutrophils treated with ANCA-positive IgG by live cell imaging. This effect was enhanced by rapamycin, the autophagy inducer, and weakened by 3-methyladenine (3-MA), the autophagy inhibitor. In line with these results, the autophagy marker, LC3B, showed a punctate distribution pattern in the neutrophils stimulated with ANCA-positive IgG. In the presence of rapamycin, LC3B accumulation was further increased; however, this effect was attenuated by 3-MA. Moreover, incubated with ANCA-positive IgG, the NETosis rate significantly increased compared with the unstimulated group. And, the rate significantly increased or decreased in the neutrophils pretreated with rapamycin or 3-MA, respectively, as compared with the cells incubated with ANCA-positive IgG. Overall, this study demonstrates that autophagy is induced by ANCA and promotes ANCA-induced NETs formation.

  10. Incorporation of RANKL promotes osteoclast formation and osteoclast activity on β-TCP ceramics.

    PubMed

    Choy, John; Albers, Christoph E; Siebenrock, Klaus A; Dolder, Silvia; Hofstetter, Wilhelm; Klenke, Frank M

    2014-12-01

    β-Tricalcium phosphate (β-TCP) ceramics are approved for the repair of osseous defects. In large defects, however, the substitution of the material by authentic bone is inadequate to provide sufficient long-term mechanical stability. We aimed to develop composites of β-TCP ceramics and receptor activator of nuclear factor κ-B ligand (RANKL) to enhance the formation of osteoclasts and promote cell mediated calcium phosphate resorption. RANKL was adsorbed superficially onto β-TCP ceramics or incorporated into a crystalline layer of calcium phosphate by the use of a co-precipitation technique. Murine osteoclast precursors were seeded onto the ceramics. After 15 days, the formation of osteoclasts was quantified cytologically and colorimetrically with tartrate-resistant acidic phosphatase (TRAP) staining and TRAP activity measurements, respectively. Additionally, the expression of transcripts encoding the osteoclast gene products cathepsin K, calcitonin receptor, and of the sodium/hydrogen exchanger NHA2 were quantified by real-time PCR. The activity of newly formed osteoclasts was evaluated by means of a calcium phosphate resorption assay. Superficially adsorbed RANKL did not induce the formation of osteoclasts on β-TCP ceramics. When co-precipitated onto β-TCP ceramics RANKL supported the formation of mature osteoclasts. The development of osteoclast lineage cells was further confirmed by the increased expression of cathepsin K, calcitonin receptor, and NHA2. Incorporated RANKL stimulated the cells to resorb crystalline calcium phosphate. Our in vitro study shows that RANKL incorporated into β-TCP ceramics induces the formation of active, resorbing osteoclasts on the material surface. Once formed, osteoclasts mediate the release of RANKL thereby perpetuating their differentiation and activation. In vivo, the stimulation of osteoclast-mediated resorption may contribute to a coordinated sequence of material resorption and bone formation. Further in vivo studies

  11. The costs of anti-herbivore defense traits in agricultural crop plants: a case study involving leafhoppers and trichomes.

    PubMed

    Kaplan, Ian; Dively, Galen P; Denno, Robert F

    2009-06-01

    The expression of plant defenses is thought to entail costs (e.g., the allocation of resources away from growth or reproduction) that constrain the evolution of plant genotypes maximally defended against herbivores. Although central to the ecological theory underlying plant-insect interactions at large, the concept of defense costs is particularly evident in agricultural crops where plants may be under simultaneous selection for enhanced growth and/or reproduction (i.e., yield) and anti-herbivore resistance traits that deter pests. In this study we investigate the role of trichomes as a resistance mechanism against a sap-feeding insect (the leafhopper, Empoasca fabae) on potato. Natural variation in trichome density among 17 potato cultivars was used to test for the role of trichomes as a putative defense against leafhoppers, and evidence of costs in trichome expression. Two different types of costs were explored: (1) allocation costs (i.e., the relationship between trichomes and yield), and (2) costs involving trade-offs with alternative defense strategies (e.g., tolerance). Although leafhopper abundance did not decrease as trichome density increased, leafhopper injury to potato plants (foliar necrosis) was negatively correlated with trichome density. As a result, the per capita effect of leafhopper adults and nymphs on foliar damage was lower on plants with high trichome densities. We found no evidence, however, for costs of expressing this resistance trait; trichomes were not correlated with either potato yield or tolerance to herbivory. Thus, selection for multiple plant defenses to alleviate the impact of pests in agronomic crops may indeed be possible without inherent losses in plant yield.

  12. Analysis of cannabinoids in laser-microdissected trichomes of medicinal Cannabis sativa using LCMS and cryogenic NMR.

    PubMed

    Happyana, Nizar; Agnolet, Sara; Muntendam, Remco; Van Dam, Annie; Schneider, Bernd; Kayser, Oliver

    2013-03-01

    Trichomes, especially the capitate-stalked glandular hairs, are well known as the main sites of cannabinoid and essential oil production of Cannabis sativa. In this study the distribution and density of various types of Cannabis sativa L. trichomes, have been investigated by scanning electron microscopy (SEM). Furthermore, glandular trichomes were isolated over the flowering period (8 weeks) by laser microdissection (LMD) and the cannabinoid profile analyzed by LCMS. Cannabinoids were detected in extracts of 25-143 collected cells of capitate-sessile and capitate stalked trichomes and separately in the gland (head) and the stem of the latter. Δ(9)-Tetrahydrocannabinolic acid [THCA (1)], cannabidiolic acid [CBDA (2)], and cannabigerolic acid [CBGA (3)] were identified as most-abundant compounds in all analyzed samples while their decarboxylated derivatives, Δ(9)-tetrahydrocannabinol [THC (4)], cannabidiol [CBD (5)], and cannabigerol [CBG (6)], co-detected in all samples, were present at significantly lower levels. Cannabichromene [CBC (8)] along with cannabinol (CBN (9)) were identified as minor compounds only in the samples of intact capitate-stalked trichomes and their heads harvested from 8-week old plants. Cryogenic nuclear magnetic resonance spectroscopy (NMR) was used to confirm the occurrence of major cannabinoids, THCA (1) and CBDA (2), in capitate-stalked and capitate-sessile trichomes. Cryogenic NMR enabled the additional identification of cannabichromenic acid [CBCA (7)] in the dissected trichomes, which was not possible by LCMS as standard was not available. The hereby documented detection of metabolites in the stems of capitate-stalked trichomes indicates a complex biosynthesis and localization over the trichome cells forming the glandular secretion unit.

  13. Promotion

    PubMed Central

    Alam, Hasan B.

    2013-01-01

    This article gives an overview of the promotion process in an academic medical center. A description of different promotional tracks, tenure and endowed chairs, and the process of submitting an application is provided. Finally, some practical advice about developing skills and attributes that can help with academic growth and promotion is dispensed. PMID:24436683

  14. The foliar trichomes of Hypoestes aristata (Vahl) Sol. ex Roem. & Schult var aristata (Acanthaceae) a widespread medicinal plant species in tropical sub-Saharan Africa: with comments on its possible phylogenetic significance.

    PubMed

    Bhatt, A; Naidoo, Y; Nicholas, A

    2010-01-01

    The micromorphology of foliar trichomes of Hypoestes aristata var. aristata was studied using stereo, light and scanning microscopy (SEM). This genus belongs to the advanced angiosperm family Acanthaceae, for which few micromorphological leaf studies exist. Results revealed both glandular and non-glandular trichomes, the latter being more abundant on leaf veins, particularly on the abaxial surface of very young leaves. With leaf maturity, the density of non-glandular trichomes decreased. Glandular trichomes were rare and of two types: long-stalked capitate and globose-like peltate trichomes. Capitate trichomes were observed only on the abaxial leaf surface, while peltate trichomes were distributed on both adaxial and abaxial leaf surfaces.

  15. Mechanical strain promotes osteoblast ECM formation and improves its osteoinductive potential

    PubMed Central

    2012-01-01

    Background The extracellular matrix (ECM) provides a supportive microenvironment for cells, which is suitable as a tissue engineering scaffold. Mechanical stimulus plays a significant role in the fate of osteoblast, suggesting that it regulates ECM formation. Therefore, we investigated the influence of mechanical stimulus on ECM formation and bioactivity. Methods Mouse osteoblastic MC3T3-E1 cells were cultured in cell culture dishes and stimulated with mechanical tensile strain. After removing the cells, the ECMs coated on dishes were prepared. The ECM protein and calcium were assayed and MC3T3-E1 cells were re-seeded on the ECM-coated dishes to assess osteoinductive potential of the ECM. Results The cyclic tensile strain increased collagen, bone morphogenetic protein 2 (BMP-2), BMP-4, and calcium levels in the ECM. Compared with the ECM produced by unstrained osteoblasts, those of mechanically stimulated osteoblasts promoted alkaline phosphatase activity, elevated BMP-2 and osteopontin levels and mRNA levels of runt-related transcriptional factor 2 (Runx2) and osteocalcin (OCN), and increased secreted calcium of the re-seeded MC3T3-E1 cells. Conclusion Mechanical strain promoted ECM production of osteoblasts in vitro, increased BMP-2/4 levels, and improved osteoinductive potential of the ECM. This study provided a novel method to enhance bioactivity of bone ECM in vitro via mechanical strain to osteoblasts. PMID:23098360

  16. Antheraea pernyi silk sericin mediating biomimetic nucleation and growth of hydroxylapatite crystals promoting bone matrix formation.

    PubMed

    Jiayao, Zhuang; Guanshan, Zhou; Jinchi, Zhang; Yuyin, Chen; Yongqiang, Zhu

    2017-03-01

    Bone biomineralization is well-regulated processes mediated by extracellular matrix proteins. The materials that can direct nucleation of hydroxylapatite (HAp) crystals and assembly of well-structured material-minerals complex are the key to mimicking the natural mineralization. This study used sericin from Antheraea pernyi (A.pernyi), non-mulberry silkworm cocoon as template to mediate nucleation of HAp crystals. Here we find out that AS (Antheraea pernyi sericin) can nucleate the formation HAp crystals in simulated body fluid verified by XRD and FTIR observations. The HAp crystals are organized into nano-rods oriented with c-axis preferentially parallel to the long axis of AS due to hydrogen bonds and electrostatic interaction and finally aggregated into HAp globule. The cell culture of human bone marrow-derived mesenchymal stem cells (BMSCs) showed that the HAp crystals mediated by AS not only stimulate cell adhesion and proliferation but also promote 0f osteogenic differentiation, suggesting that the resultant mineralized AS biomaterial has potential in promoting bone formation. Thus our work will provide significant implication on biomineralization of A. pernyi silk sericin as a potential scaffold for tissue engineering.

  17. Interruptions between the triple helix peptides can promote the formation of amyloid-like fibrils

    NASA Astrophysics Data System (ADS)

    Parmar, Avanish; Hwang, Eileen; Brodsky, Barbara

    2010-03-01

    It has been reported that collagen can initiate or accelerate the formation of amyloid fibrils. Non-fibrillar collagen types have sites where the repeating (Gly-Xaa-Yaa)n sequences are interrupted by non- Gly-Xaa-Yaa sequences, and we are investigating the hypothesis that some of these interruptions can promote amyloid formation. Our experimental data show that model peptides containing an 8 or 9 residue interruption sequence between (Gly-Pro-Hyp)n domains have a strong propensity for self association to form fibrous structures. A peptide containing only the 9-residue interruption sequence forms amyloid like fibrils with anti-parallel β sheet. Computational analysis predicts that 33 out of 374 naturally occurring human non-fibrillar collagen sequences within or between triple-helical sequences have significant cross-β aggregation potential, including the 8 and 9 residue sequences studied in peptides. Further studies are in progress to investigate whether a triple-helix peptide promotes amyloidogenesis and whether amyloid interferes with collagen fibrillogenesis.

  18. An invasive podosome-like structure promotes fusion pore formation during myoblast fusion

    PubMed Central

    Sens, Kristin L.; Zhang, Shiliang; Jin, Peng; Duan, Rui; Zhang, Guofeng; Luo, Fengbao; Parachini, Lauren

    2010-01-01

    Recent studies in Drosophila have implicated actin cytoskeletal remodeling in myoblast fusion, but the cellular mechanisms underlying this process remain poorly understood. Here we show that actin polymerization occurs in an asymmetric and cell type–specific manner between a muscle founder cell and a fusion-competent myoblast (FCM). In the FCM, a dense F-actin–enriched focus forms at the site of fusion, whereas a thin sheath of F-actin is induced along the apposing founder cell membrane. The FCM-specific actin focus invades the apposing founder cell with multiple finger-like protrusions, leading to the formation of a single-channel macro fusion pore between the two muscle cells. Two actin nucleation–promoting factors of the Arp2/3 complex, WASP and Scar, are required for the formation of the F-actin foci, whereas WASP but not Scar promotes efficient foci invasion. Our studies uncover a novel invasive podosome-like structure (PLS) in a developing tissue and reveal a previously unrecognized function of PLSs in facilitating cell membrane juxtaposition and fusion. PMID:21098115

  19. Subinhibitory concentrations of triclosan promote Streptococcus mutans biofilm formation and adherence to oral epithelial cells.

    PubMed

    Bedran, Telma Blanca Lombardo; Grignon, Louis; Spolidorio, Denise Palomari; Grenier, Daniel

    2014-01-01

    Triclosan is a general membrane-active agent with a broad-spectrum antimicrobial activity that is commonly used in oral care products. In this study, we investigated the effect of sub-minimum inhibitory concentrations (MICs) of triclosan on the capacity of the cariogenic bacterium Streptococcus mutans to form biofilm and adhere to oral epithelial cells. As quantified by crystal violet staining, biofilm formation by two reference strains of S. mutans was dose-dependently promoted, in the range of 2.2- to 6.2-fold, by 1/2 and 1/4 MIC of triclosan. Observations by scanning electron microscopy revealed the presence of a dense biofilm attached to the polystyrene surface. Growth of S. mutans in the presence of triclosan at sub-MICs also increased its capacity to adhere to a monolayer of gingival epithelial cells. The expression of several genes involved in adherence and biofilm formation in S. mutans was investigated by quantitative RT-PCR. It was found that sub-MICs of triclosan significantly increased the expression of comD, gtfC, and luxS, and to a lesser extent of gtfB and atlA genes. These findings stress the importance of maintaining effective bactericidal concentrations of therapeutic triclosan since sub-MICs may promote colonization of the oral cavity by S. mutans.

  20. Examining the Factors That Promote Long-Term Change in Elementary Teachers' Instructional Practices: Sustaining Formative Assessment Reform

    ERIC Educational Resources Information Center

    Sherbinko, Thomas, Jr.

    2011-01-01

    The purpose of this holistic case study with a mixed methods approach was two-fold: first, to investigate the extent to which elementary teachers in the BTSD sustained the use of formative assessment reform to regulate instruction, and second, to explore the factors that promoted fidelity to formative assessment reform. Although this study…

  1. Amyloid Aβ 42, a promoter of magnetite nanoparticle formation in Alzheimer’s disease

    NASA Astrophysics Data System (ADS)

    Bogachan Tahirbegi, Islam; Pardo, Wilmer Alfonso; Alvira, Margarita; Mir, Mònica; Samitier, Josep

    2016-11-01

    The accumulation of iron oxides—mainly magnetite—with amyloid peptide is a key process in the development of Alzheimer’s disease (AD). However, the mechanism for biogeneration of magnetite inside the brain of someone with AD is still unclear. The iron-storing protein ferritin has been identified as the main magnetite-storing molecule. However, accumulations of magnetite in AD are not correlated with an increase in ferritin, leaving this question unresolved. Here we demonstrate the key role of amyloid peptide Aβ 42, one of the main hallmarks of AD, in the generation of magnetite nanoparticles in the absence of ferritin. The capacity of amyloid peptide to bind and concentrate iron hydroxides, the basis for the formation of magnetite, benefits the spontaneous synthesis of these nanoparticles, even under unfavorable conditions for their formation. Using scanning and transmission electron microscopy, electron energy loss spectroscopy and magnetic force microscopy we characterized the capacity of amyloid peptide Aβ 42 to promote magnetite formation.

  2. WNT7B promotes bone formation in part through mTORC1.

    PubMed

    Chen, Jianquan; Tu, Xiaolin; Esen, Emel; Joeng, Kyu Sang; Lin, Congxin; Arbeit, Jeffrey M; Rüegg, Markus A; Hall, Michael N; Ma, Liang; Long, Fanxin

    2014-01-01

    WNT signaling has been implicated in both embryonic and postnatal bone formation. However, the pertinent WNT ligands and their downstream signaling mechanisms are not well understood. To investigate the osteogenic capacity of WNT7B and WNT5A, both normally expressed in the developing bone, we engineered mouse strains to express either protein in a Cre-dependent manner. Targeted induction of WNT7B, but not WNT5A, in the osteoblast lineage dramatically enhanced bone mass due to increased osteoblast number and activity; this phenotype began in the late-stage embryo and intensified postnatally. Similarly, postnatal induction of WNT7B in Runx2-lineage cells greatly stimulated bone formation. WNT7B activated mTORC1 through PI3K-AKT signaling. Genetic disruption of mTORC1 signaling by deleting Raptor in the osteoblast lineage alleviated the WNT7B-induced high-bone-mass phenotype. Thus, WNT7B promotes bone formation in part through mTORC1 activation.

  3. Ethylene promotes induction of aerenchyma formation and ethanolic fermentation in waterlogged roots of Dendranthema spp.

    PubMed

    Yin, Dongmei; Chen, Sumei; Chen, Fadi; Jiang, Jiafu

    2013-07-01

    The role of ethylene in induction of aerenchyma formation and ethanolic fermentation in waterlogged roots of Dendranthema zawadskii and D. nankingense, two species that differ with respect to waterlogging tolerance, was examined. In the more tolerant D. zawadskii, but not in D. nankingense, ethylene accelerated programmed cell death and promoted formation of lysigenous aerenchyma, both of which were inhibited by treatment with the ethylene inhibitor 1-methylcyclopropene. Waterlogged D. zawadskii roots generated a higher quantity of endogenous ethylene than did those of D. nankingense. In waterlogged D. zawadskii roots, transcription of the genes encoding alcohol dehydrogenase (EC 1.1.1.1) and pyruvate decarboxylase (EC 4.1.1.1) increased rapidly but transiently, whereas expression of these genes in D. nankingense increased gradually and over a longer period. In D. nankingense, waterlogging elevated both alcohol dehydrogenase and pyruvate decarboxylase activity, and the production of ethanol and acetaldehyde was increased in the presence of exogenous ethylene and inhibited by 1-methylcyclopropene. In D. zawadskii, in contrast, after a prolonged episode of waterlogging stress, exogenous supply of ethylene suppressed the production of ethanol and acetaldehyde, whereas exogenous 1-methylcyclopropene enhanced their production. In the more tolerant Dendranthema species, ethylene appeared to signal an acceleration of both waterlogging-induced programmed cell death and aerenchyma formation and to alleviate ethanolic fermentation, whereas in the more sensitive species ethylene activated fermentation and increased the release of ethanol and acetaldehyde, which are by-products probably responsible for the collapse of the waterlogging-damaged root.

  4. Promoter-Specific Effects of DREADD Modulation on Hippocampal Synaptic Plasticity and Memory Formation

    PubMed Central

    López, Alberto J.; Kramár, Enikö; Matheos, Dina P.; White, André O.; Kwapis, Janine; Vogel-Ciernia, Annie; Sakata, Keith; Espinoza, Monica

    2016-01-01

    Designer receptors exclusively activated by designer drug (DREADDs) are a novel tool with the potential to bidirectionally drive cellular, circuit, and ultimately, behavioral changes. We used DREADDs to evaluate memory formation in a hippocampus-dependent task in mice and effects on synaptic physiology in the dorsal hippocampus. We expressed neuron-specific (hSyn promoter) DREADDs that were either excitatory (HM3D) or inhibitory (HM4D) in the dorsal hippocampus. As predicted, hSyn–HM3D was able to transform a subthreshold learning event into long-term memory (LTM), and hSyn–HM4D completely impaired LTM formation. Surprisingly, the opposite was observed during experiments examining the effects on hippocampal long-term potentiation (LTP). hSyn–HM3D impaired LTP and hSyn–HM4D facilitated LTP. Follow-up experiments indicated that the hSyn–HM3D-mediated depression of fEPSP appears to be driven by presynaptic activation of inhibitory currents, whereas the hSyn–HM4D-mediated increase of fEPSP is induced by a reduction in GABAA receptor function. To determine whether these observations were promoter specific, we next examined the effects of using the CaMKIIα promoter that limits expression to forebrain excitatory neurons. CaMKIIα–HM3D in the dorsal hippocampus led to the transformation of a subthreshold learning event into LTM, whereas CaMKIIα–HM4D blocked LTM formation. Consistent with these findings, baseline synaptic transmission and LTP was increased in CaMKIIα–HM3D hippocampal slices, whereas slices from CaMKIIα–HM4D mice produced expected decreases in baseline synaptic transmission and LTP. Together, these experiments further demonstrate DREADDs as being a robust and reliable means of modulating neuronal function to manipulate long-term changes in behavior, while providing evidence for specific dissociations between LTM and LTP. SIGNIFICANCE STATEMENT This study evaluates the efficacy of designer receptors exclusively activated by designer

  5. Biosynthesis of a new tobacco alkaloid, hydroxy-N-acylnornicotine in the trichomes of Nicotiana stocktonii. [Manduca sexta

    SciTech Connect

    Zador, E.; Jones, D.

    1986-04-01

    A new tobacco alkaloid from section Repandae is highly toxic to an insect (Manduca sexta) unsusceptible to previously described nicotine alkaloids (1). They have localized the alkaloid, HO-N-acylnornicotine (HO-NAN) nearly entirely to the exudate secreted by the epidermal trichomes of N. stocktonii. Only the nicotine and nornicotine were found in abundance inside the trichomes, while primarily nicotine was present inside the aerial vegetative parts and root. These results suggest that the HO-NAN is synthesized by the trichomes. When unlabelled nicotine was fed to isolated leaves there was an increase in internal nicotine, nornicotine and secretion of HO-NAN. Feeding leaves with 2'-C/sup 14/ nicotine resulted in labelling of both nornicotine and HO-NAN. These data strongly suggest synthesis of HO-NAN from nicotine via nornicotine in the trichomes, followed by rapid secretion. The possible evolutionary significance of this pathway of synthesis and secretion is discussed.

  6. Tetrahydrocannabinolic acid synthase, the enzyme controlling marijuana psychoactivity, is secreted into the storage cavity of the glandular trichomes.

    PubMed

    Sirikantaramas, Supaart; Taura, Futoshi; Tanaka, Yumi; Ishikawa, Yu; Morimoto, Satoshi; Shoyama, Yukihiro

    2005-09-01

    Tetrahydrocannabinolic acid (THCA) synthase is the enzyme responsible for the production of tetrahydrocannabinol (THC), the psychoactive component of marijuana (Cannabis sativa L.). We suggest herein that THCA is biosynthesized in the storage cavity of the glandular trichomes based on the following observations. (i) The exclusive expression of THCA synthase was confirmed in the secretory cells of glandular trichomes by reverse transcription-PCR (RT-PCR) analysis. (ii) THCA synthase activity was detected in the storage cavity content. (iii) Transgenic tobacco expressing THCA synthase fused to green fluorescent protein showed fluorescence in the trichome head corresponding to the storage cavity. These results also showed that secretory cells of the glandular trichomes secrete not only metabolites but also biosynthetic enzyme.

  7. Effect of plant trichomes on the vertical migration of Haemonchus contortus infective larvae on five tropical forages.

    PubMed

    Oliveira, Aruaque L F; Costa, Ciniro; Rodella, Roberto A; Silva, Bruna F; Amarante, Alessandro F T

    2009-06-01

    The influence of trichomes on vertical migration and survival of Haemonchus contortus infective larvae (L3) on different forages was investigated. Four different forages showing different distributions of trichomes (Brachiaria brizantha cv. Marandu, Brachiaria brizantha cv. Xaraes, Andropogon gayanus, and Stylosanthes spp.), and one forage species without trichomes (Panicum maximum cv. Tanzania), were used. Forages cut at the post-grazing height were contaminated with faeces containing L3. Samples of different grass strata (0-10, 10-20, >20 cm) and faeces were collected for L3 quantification once per week over four weeks. In all forages studied, the highest L3 recovery occurred seven days after contamination, with the lowest recovery on A. gayanus. In general, larvae were found on all forages' strata. However, most of the larvae were at the lower stratum. There was no influence of trichomes on migration and survival of H. contortus L3 on the forages.

  8. Plasticity-related Gene 5 Promotes Spine Formation in Murine Hippocampal Neurons*

    PubMed Central

    Coiro, Pierluca; Stoenica, Luminita; Strauss, Ulf; Bräuer, Anja Ursula

    2014-01-01

    The transmembrane protein plasticity-related genes 3 and 5 (PRG3 and PRG5) increase filopodial formation in various cell lines, independently of Cdc42. However, information on the effects of PRG5 during neuronal development is sparse. Here, we present several lines of evidence for the involvement of PRG5 in the genesis and stabilization of dendritic spines. First, PRG5 was strongly expressed during mouse brain development from embryonic day 14 (E14), peaked around the time of birth, and remained stable at least until early adult stages (i.e. P30). Second, on a subcellular level, PRG5 expression shifted from an equal distribution along all neurites toward accumulation only along dendrites during hippocampal development in vitro. Third, overexpression of PRG5 in immature hippocampal neurons induced formation of spine-like structures ahead of time. Proper amino acid sequences in the extracellular domains (D1 to D3) of PRG5 were a prerequisite for trafficking and induction of spine-like structures, as shown by mutation analysis. Fourth, at stages when spines are present, knockdown of PRG5 reduced the number but not the length of protrusions. This was accompanied by a decrease in the number of excitatory synapses and, consequently, by a reduction of miniature excitatory postsynaptic current frequencies, although miniature excitatory postsynaptic current amplitudes remained similar. In turn, overexpressing PRG5 in mature neurons not only increased Homer-positive spine numbers but also augmented spine head diameters. Mechanistically, PRG5 interacts with phosphorylated phosphatidylinositols, phospholipids involved in dendritic spine formation by different lipid-protein assays. Taken together, our data propose that PRG5 promotes spine formation. PMID:25074937

  9. Testosterone promotes tube formation of endothelial cells isolated from veins via activation of Smad1 protein.

    PubMed

    Liu, Pei; Li, Xiaosa; Song, Fuhu; Li, Ping; Wei, Jinzhi; Yan, Qing; Xu, Xingyan; Yang, Jun; Li, Chuanxiang; Fu, Xiaodong

    2017-05-05

    Testosterone (T) deficiency is positively correlated with the increased incidence of cardiovascular disease. However, the effects of T on vascular endothelial cells remain obscure. Tube formation capacity is critical for vascular regeneration/repair and Smad1 plays an important role in these events. In this study, we investigated the effects of T on Smad1 activation and tube formation of cultured human umbilical endothelial cells (HUVECs). Our results showed that T rapidly increased endothelial Smad1 phosphorylation. This effect was mimicked by cell-impermeable T-BSA conjugates and was not altered by transcriptional inhibitor actinomycin D or translational inhibitor cycloheximide. T-induced Smad1 phosphorylation was blocked by ERK1/2 and c-Src inhibitors or their specific siRNAs, while it was reinforced by ERK1/2 or c-Src overexpression. Indeed, T rapidly activated ERK1/2 and c-Src signalings and c-Src was confirmed as the upstream of ERK1/2. Moreover, caveolae disruptor methyl-β-cyclodextrin (β-MCD) blocked Smad1 activation induced by T. The association of caveolin-1 with androgen receptor (AR) or c-Src was detected by immunoprecipitation and it was significantly increased by rapid T stimulation. Furthermore, fractional analysis showed that AR and c-Src were expressed in caveolae-enriched membrane fractions. T promoted tube formation of HUVECs, which was blocked by c-Src and ERK1/2 inhibitors or by the knockdown of Smad1. In conclusion, T increased tube formation of endothelial cells isolated from veins by stimulating Smad1 phosphorylation in a nongenomic manner, which was mediated by signals from AR/c-Src located in caveolae to ERK1/2 cascade. These findings may shed new light on the relevance of T to its vascular functions.

  10. Non-glandular trichomes of Solanum carolinense deter feeding by Manduca sexta caterpillars and cause damage to the gut peritrophic matrix.

    PubMed

    Kariyat, Rupesh R; Smith, Jason D; Stephenson, Andrew G; De Moraes, Consuelo M; Mescher, Mark C

    2017-02-22

    Plant trichomes constitute a first line of defence against insect herbivores. The pre- and post-ingestive defensive functions of glandular trichomes are well documented and include direct toxicity, adhesion, antinutrition and defence gene induction. By contrast, the defensive functions of non-glandular trichomes are less well characterized, although these structures are thought to serve as physical barriers that impede herbivore feeding and movement. We experimentally varied the density of stellate non-glandular trichomes in several ways to explore their pre- and post-ingestive effects on herbivores. Larvae of Manduca sexta (Sphingidae) initiated feeding faster and gained more weight on Solanum carolinense (Solanaceae) leaves having lower trichome densities (or experimentally removed trichomes) than on leaves having higher trichome densities. Adding trichomes to artificial diet also deterred feeding and adversely affected caterpillar growth relative to controls. Scanning electron and light microscopy revealed that the ingestion of stellate trichomes by M. sexta caterpillars caused extensive damage to the peritrophic membrane, a gut lining that is essential to digestion and pathogen isolation. These findings suggest that, in addition to acting as a physical barrier to deter feeding, trichomes can inhibit caterpillar growth and development via post-ingestive effects.

  11. A pro-chelator triggered by hydrogen peroxide inhibits iron-promoted hydroxyl radical formation.

    PubMed

    Charkoudian, Louise K; Pham, David M; Franz, Katherine J

    2006-09-27

    The synthesis and structural characterization of a new pro-chelating agent, isonicotinic acid [2-(4,4,5,5-tetramethyl-[1,3,2]dioxaborolan-2-yl)-benzylidene]-hydrazide (BSIH), are presented. BSIH only weakly interacts with iron unless hydrogen peroxide (H2O2) is present to remove the boronic ester protecting group to reveal a phenol that is a key metal-binding group of tridentate salicylaldehyde isonicotinoyl hydrazone (SIH). BSIH prevents deoxyribose degradation caused by hydroxyl radicals that are generated from H2O2 and redox-active iron by sequestering Fe3+ and preventing iron-promoted hydroxyl radical formation. The rate-determining step for iron sequestration is conversion of BSIH to SIH, followed by rapid Fe3+ complexation. The pro-chelate approach of BSIH represents a promising strategy for chelating a specific pool of detrimental metal ions without disturbing healthy metal ion distribution.

  12. Communication between nitric oxide synthase and positively-charged surface and bone formation promotion.

    PubMed

    Zhang, Wei; Liu, Jun; Shi, Haigang; Yang, Kun; Wang, Pingli; Wang, Gexia; Liu, Na; Wang, Huaiyu; Ji, Junhui; Chu, Paul K

    2016-12-01

    Despite the effects on physiology of bone marrow mesenchymal stem cells (BMSCs) and bone tissue, biological signal communication between bone implants and them is seldom employed as a guidance to create an osteo-inductive interface. Herein, the positively-charged surface is constructed on bone implant from the perspective of mediation of nitric oxide synthase (NOS) expression to signal BMSCs osteo-differentiation. In vitro and in vivo results indicate that the proper surface potential on the positively-charged surface affects NOS to express a high level of inducible nitric oxide synthase (iNOS) in three NOS isoforms of the contacted BMSCs, upregulates their osteogenetic expression, and ultimately foster new bone growth. However, an excessively high surface potential produces substantial immunomodulatory effects thereby offsetting the aforementioned advantages. This study demonstrates that fine-tuning of the positively-charged surface and proper utilization of the communication between NOS and bone implants promote bone formation.

  13. Monoallelic Loss of the Imprinted Gene Grb10 Promotes Tumor Formation in Irradiated Nf1+/- Mice

    PubMed Central

    Mroue, Rana; Huang, Brian; Braunstein, Steve; Firestone, Ari J.; Nakamura, Jean L.

    2015-01-01

    Imprinted genes are expressed from only one parental allele and heterozygous loss involving the expressed allele is sufficient to produce complete loss of protein expression. Genetic alterations are common in tumorigenesis but the role of imprinted genes in this process is not well understood. In earlier work we mutagenized mice heterozygous for the Neurofibromatosis I tumor suppressor gene (NF1) to model radiotherapy-associated second malignant neoplasms that arise in irradiated NF1 patients. Expression analysis of tumor cell lines established from our mouse models identified Grb10 expression as widely absent. Grb10 is an imprinted gene and polymorphism analysis of cell lines and primary tumors demonstrates that the expressed allele is commonly lost in diverse Nf1 mutant tumors arising in our mouse models. We performed functional studies to test whether Grb10 restoration or loss alter fundamental features of the tumor growth. Restoring Grb10 in Nf1 mutant tumors decreases proliferation, decreases soft agar colony formation and downregulates Ras signaling. Conversely, Grb10 silencing in untransformed mouse embryo fibroblasts significantly increased cell proliferation and increased Ras-GTP levels. Expression of a constitutively activated MEK rescued tumor cells from Grb10-mediated reduction in colony formation. These studies reveal that Grb10 loss can occur during in vivo tumorigenesis, with a functional consequence in untransformed primary cells. In tumors, Grb10 loss independently promotes Ras pathway hyperactivation, which promotes hyperproliferation, an early feature of tumor development. In the context of a robust Nf1 mutant mouse model of cancer this work identifies a novel role for an imprinted gene in tumorigenesis. PMID:26000738

  14. Topical hemostatic powder promotes reepithelialization and reduces scar formation after extensive esophageal mucosal resection.

    PubMed

    Beye, B; Barret, M; Alatawi, A; Beuvon, F; Nicco, C; Pratico, C A; Chereau, C; Chaussade, S; Batteux, F; Prat, F

    2016-08-01

    The development of techniques for endoscopic resection has provided new strategies for radical conservative treatment of superficial esophageal neoplasms, even those that are circumferential, such as Barrett's neoplasia. However, it is necessary to prevent the formation of scar tissue that can be responsible for esophageal strictures following circumferential resection. Preliminary data have suggested the possible efficacy of a hemostatic powder in the promotion of wound healing. The study aims to assess the effectiveness of Hemospray (Cook Medical) in a swine model of post-endoscopic esophageal stricture. Our prospective controlled study included 21 pigs. A 6-cm circumferential submucosal dissection of the esophagus (CESD) was performed in each pig. Group 1 (n = 11) only underwent CESD and Group 2 (n = 10) had repeated Hemospray applications after CESD. Clinical, endoscopic, and radiological monitoring were performed, blood levels of four inflammatory or pro-fibrotic cytokines were assessed, and histological analysis was performed. Median esophageal diameter was greater in the group treated with Hemospray (2 mm [1-3] vs. 3 mm [2-4], P = 0.01), and the rate of symptomatic esophageal stricture was 100% and 60% in Groups 1 and 2, respectively (P = 0.09). The thicknesses of esophageal fibrosis and inflammatory cell infiltrate were significantly lower in Group 2 than in Group 1 (P = 0.002 and 0.0003, respectively). The length of the neoepithelium was greater in Group 2 than in Group 1 (P = 0.0004). Transforming growth factor-β levels were significantly lower in Group 2 than in Group 1 (P = 0.01). The application of Hemospray after esophageal CESD reduces scar tissue formation and promotes reepithelialization, and therefore is a promising therapeutic approach in the prevention of post-endoscopic esophageal stricture.

  15. Src and cortactin promote lamellipodia protrusion and filopodia formation and stability in growth cones

    PubMed Central

    He, Yingpei; Ren, Yuan; Wu, Bingbing; Decourt, Boris; Lee, Aih Cheun; Taylor, Aaron; Suter, Daniel M.

    2015-01-01

    Src tyrosine kinases have been implicated in axonal growth and guidance; however, the underlying cellular mechanisms are not well understood. Specifically, it is unclear which aspects of actin organization and dynamics are regulated by Src in neuronal growth cones. Here, we investigated the function of Src2 and one of its substrates, cortactin, in lamellipodia and filopodia of Aplysia growth cones. We found that up-regulation of Src2 activation state or cortactin increased lamellipodial length, protrusion time, and actin network density, whereas down-regulation had opposite effects. Furthermore, Src2 or cortactin up-regulation increased filopodial density, length, and protrusion time, whereas down-regulation promoted lateral movements of filopodia. Fluorescent speckle microscopy revealed that rates of actin assembly and retrograde flow were not affected in either case. In summary, our results support a model in which Src and cortactin regulate growth cone motility by increasing actin network density and protrusion persistence of lamellipodia by controlling the state of actin-driven protrusion versus retraction. In addition, both proteins promote the formation and stability of actin bundles in filopodia. PMID:26224308

  16. Structure and biomechanics of trapping flower trichomes and their role in the pollination biology of Aristolochia plants (Aristolochiaceae).

    PubMed

    Oelschlägel, Birgit; Gorb, Stanislav; Wanke, Stefan; Neinhuis, Christoph

    2009-12-01

    *Catching insects to ensure pollination is one of the most elaborate and specialized mechanisms of insect-plant interactions. Phylogenetically, Aristolochiaceae represent the first angiosperm lineage that developed trap flowers. Here we report the structure and function of specific trichomes contributing to the highly specialized trapping devices. *Investigations were carried out on six Mediterranean Aristolochia species. The morphology and arrangement of the trapping trichomes were investigated by scanning electron microscopy (SEM) and cryo-SEM. To demonstrate frictional anisotropy of the trapping trichome array, a microtribological approach was used. *The results of our experiments support a hypothesis long proposed, but never tested, regarding the trapping mechanism in proterogynous Aristolochia flowers: that an array of highly specialized trichomes arranged eccentrically to the underlying surface is responsible for the easy entrance of insects into flowers but impedes their escape. As they enter the male stage of anthesis, flowers significantly modify their inner surface characteristics, allowing insects to leave. *We have demonstrated the substantial contribution of trapping trichomes to the capture, retention and release of pollinators, an important prerequisite for making cross-pollination possible in most Aristolochia species. Finally, we compare trapping trichomes of Aristolochia with similar structures found in other trapping flowers as well as in pitchers of carnivorous plants not optimized for insect release.

  17. Detergent-Mediated Formation of β-Hematin: Heme Crystallization Promoted by Detergents Implicates Nanostructure Formation for Use as a Biological Mimic

    PubMed Central

    2016-01-01

    Hemozoin is a unique biomineral that results from the sequestration of toxic free heme liberated as a consequence of hemoglobin degradation in the malaria parasite. Synthetic neutral lipid droplets (SNLDs) and phospholipids were previously shown to support the rapid formation of β-hematin, abiological hemozoin, under physiologically relevant pH and temperature, though the mechanism by which heme crystallization occurs remains unclear. Detergents are particularly interesting as a template because they are amphiphilic molecules that spontaneously organize into nanostructures and have been previously shown to mediate β-hematin formation. Here, 11 detergents were investigated to elucidate the physicochemical properties that best recapitulate crystal formation in the parasite. A strong correlation between the detergent’s molecular structure and the corresponding kinetics of β-hematin formation was observed, where higher molecular weight polar chains promoted faster reactions. The larger hydrophilic chains correlated to the detergent’s ability to rapidly sequester heme into the lipophilic core, allowing for crystal nucleation to occur. The data presented here suggest that detergent nanostructures promote β-hematin formation in a similar manner to SNLDs and phospholipids. Through understanding mediator properties that promote optimal crystal formation, we are able to establish an in vitro assay to probe this drug target pathway. PMID:27175104

  18. Urokinase Receptor Promotes Skin Tumor Formation by Preventing Epithelial Cell Activation of Notch1.

    PubMed

    Mazzieri, Roberta; Pietrogrande, Giovanni; Gerasi, Laura; Gandelli, Alessandro; Colombo, Piergiuseppe; Moi, Davide; Brombin, Chiara; Ambrosi, Alessandro; Danese, Silvio; Mignatti, Paolo; Blasi, Francesco; D'Alessio, Silvia

    2015-11-15

    The urokinase-type plasminogen activator receptor (uPAR) has a well-established role in cancer progression, but it has been little studied at earlier stages of cancer initiation. Here, we show that uPAR deficiency in the mouse dramatically reduces susceptibility to the classical two-stage protocol of inflammatory skin carcinogenesis. uPAR genetic deficiency decreased papilloma formation and accelerated keratinocyte differentiation, effects mediated by Notch1 hyperactivation. Notably, Notch1 inhibition in uPAR-deficient mice rescued their susceptibility to skin carcinogenesis. Clinically, we found that human differentiated keratoacanthomas expressed low levels of uPAR and high levels of activated Notch1, with opposite effects in proliferating tumors, confirming the relevance of the observations in mice. Furthermore, we found that TACE-dependent activation of Notch1 in basal kerantinocytes was modulated by uPAR. Mechanistically, uPAR sequestered TACE within lipid rafts to prevent Notch1 activation, thereby promoting cell proliferation and tumor formation. Given that uPAR signaling is nonessential for normal epidermal homeostasis, our results argue that uPAR may present a promising disease-specific target for preventing skin cancer development.

  19. Myeloid STAT3 promotes formation of colitis-associated colorectal cancer in mice

    PubMed Central

    Pathria, Paulina; Gotthardt, Dagmar; Prchal-Murphy, Michaela; Putz, Eva-Maria; Holcmann, Martin; Schlederer, Michaela; Grabner, Beatrice; Crncec, Ilija; Svinka, Jasmin; Musteanu, Monica; Hoffmann, Thomas; Filipits, Martin; Berger, Walter; Poli, Valeria; Kenner, Lukas; Bilban, Martin; Casanova, Emilio; Müller, Mathias; Strobl, Birgit; Bayer, Editha; Mohr, Thomas; Sexl, Veronika; Eferl, Robert

    2015-01-01

    Myeloid cells lacking STAT3 promote antitumor responses of NK and T cells but it is unknown if this crosstalk affects development of autochthonous tumors. We deleted STAT3 in murine myeloid cells (STAT3Δm) and examined the effect on the development of autochthonous colorectal cancers (CRCs). Formation of Azoxymethane/Dextransulfate (AOM/DSS)-induced CRCs was strongly suppressed in STAT3Δm mice. Gene expression profiling showed strong activation of T cells in the stroma of STAT3Δm CRCs. Moreover, STAT3Δm host mice were better able to control the growth of transplanted MC38 colorectal tumor cells which are known to be killed in a T cell-dependent manner. These data suggest that myeloid cells lacking STAT3 control formation of CRCs mainly via cross activation of T cells. Interestingly, the few CRCs that formed in STAT3Δm mice displayed enhanced stromalization but appeared normal in size indicating that they have acquired ways to escape enhanced tumor surveillance. We found that CRCs in STAT3Δm mice consistently activate STAT3 signaling which is implicated in immune evasion and might be a target to prevent tumor relapse. PMID:26137415

  20. The transcription factor OBP4 controls root growth and promotes callus formation.

    PubMed

    Ramirez-Parra, Elena; Perianez-Rodriguez, Juan; Navarro-Neila, Sara; Gude, Inmaculada; Moreno-Risueno, Miguel A; Del Pozo, Juan C

    2017-03-01

    Plant growth and development require a continuous balance between cell division and differentiation. In root meristems, differentiated cells acquire specialized functions, losing their mitotic potential. Some plant cells, such as pericycle cells, have a remarkable plasticity to regenerate new organs. The molecular mechanisms underlying cell reprogramming are not completely known. In this work, a functional screening of transcription factors identified Arabidopsis OBP4 (OBF Binding Protein 4) as a novel regulator of root growth and cell elongation and differentiation. Overexpression of OBP4 regulates the levels of a large number of transcripts in roots, many involved in hormonal signaling and callus formation. OBP4 controls cell elongation and differentiation in root cells. OBP4 does not induce cell division in the root meristem, but promotes pericycle cell proliferation, forming callus-like structures at the root tip, as shown by the expression of stem cell markers. Callus formation is enhanced by ectopic expression of OBP4 in the wild-type or alf4-1, but is significantly reduced in roots that have lower levels of OBP4. Our data provide molecular insights into how differentiated root cells acquire the potential to generate callus, a pluripotent mass of cells that can regenerate fully functional plant organs.

  1. Microglia promote learning-dependent synapse formation through brain-derived neurotrophic factor.

    PubMed

    Parkhurst, Christopher N; Yang, Guang; Ninan, Ipe; Savas, Jeffrey N; Yates, John R; Lafaille, Juan J; Hempstead, Barbara L; Littman, Dan R; Gan, Wen-Biao

    2013-12-19

    Microglia are the resident macrophages of the CNS, and their functions have been extensively studied in various brain pathologies. The physiological roles of microglia in brain plasticity and function, however, remain unclear. To address this question, we generated CX3CR1(CreER) mice expressing tamoxifen-inducible Cre recombinase that allow for specific manipulation of gene function in microglia. Using CX3CR1(CreER) to drive diphtheria toxin receptor expression in microglia, we found that microglia could be specifically depleted from the brain upon diphtheria toxin administration. Mice depleted of microglia showed deficits in multiple learning tasks and a significant reduction in motor-learning-dependent synapse formation. Furthermore, Cre-dependent removal of brain-derived neurotrophic factor (BDNF) from microglia largely recapitulated the effects of microglia depletion. Microglial BDNF increases neuronal tropomyosin-related kinase receptor B phosphorylation, a key mediator of synaptic plasticity. Together, our findings reveal that microglia serve important physiological functions in learning and memory by promoting learning-related synapse formation through BDNF signaling.

  2. Sfp-type PPTase inactivation promotes bacterial biofilm formation and ability to enhance wheat drought tolerance

    PubMed Central

    Timmusk, Salme; Kim, Seong-Bin; Nevo, Eviatar; Abd El Daim, Islam; Ek, Bo; Bergquist, Jonas; Behers, Lawrence

    2015-01-01

    Paenibacillus polymyxa is a common soil bacterium with broad range of practical applications. An important group of secondary metabolites in P. polymyxa are non-ribosomal peptide and polyketide derived metabolites (NRPs/PKs). Modular non-ribosomal peptide synthetases catalyze main steps in the biosynthesis of the complex secondary metabolites. Here we report on the inactivation of an A26 Sfp-type 4'-phosphopantetheinyl transferase (Sfp-type PPTase). The inactivation of the gene resulted in loss of NRPs/PKs production. In contrast to the former Bacillus spp. model the mutant strain compared to wild type showed greatly enhanced biofilm formation ability. A26Δsfp biofilm promotion is directly mediated by NRPs/PKs, as exogenous addition of the wild type metabolite extracts restores its biofilm formation level. Wheat inoculation with bacteria that had lost their Sfp-type PPTase gene resulted in two times higher plant survival and about three times increased biomass under severe drought stress compared to wild type. Challenges with P. polymyxa genetic manipulation are discussed. PMID:26052312

  3. BipA Is Associated with Preventing Autoagglutination and Promoting Biofilm Formation in Bordetella holmesii

    PubMed Central

    Hiramatsu, Yukihiro; Saito, Momoko; Otsuka, Nao; Suzuki, Eri; Watanabe, Mineo; Shibayama, Keigo; Kamachi, Kazunari

    2016-01-01

    Bordetella holmesii causes both invasive and respiratory diseases in humans. Although the number of cases of pertussis-like respiratory illnesses due to B. holmesii infection has increased in the last decade worldwide, little is known about the virulence factors of the organism. Here, we analyzed a B. holmesii isolate that forms large aggregates and precipitates in suspension, and subsequently demonstrated that the autoagglutinating isolate is deficient in Bordetella intermediate protein A (BipA) and that this deletion is caused by a frame-shift mutation in the bipA gene. A BipA-deficient mutant generated by homologous recombination also exhibited the autoagglutination phenotype. Moreover, the BipA mutant adhered poorly to an abiotic surface and failed to form biofilms, as did two other B. holmesii autoagglutinating strains, ATCC 51541 and ATCC 700053, which exhibit transcriptional down-regulation of bipA gene expression, indicating that autoagglutination indirectly inhibits biofilm formation. In a mouse intranasal infection model, the BipA mutant showed significantly lower levels of initial lung colonization than did the parental strain (P < 0.01), suggesting that BipA might be a critical virulence factor in B. holmesii respiratory infection. Together, our findings suggest that BipA production plays an essential role in preventing autoagglutination and indirectly promoting biofilm formation by B. holmesii. PMID:27448237

  4. Hand2 inhibits kidney specification while promoting vein formation within the posterior mesoderm

    PubMed Central

    Perens, Elliot A; Garavito-Aguilar, Zayra V; Guio-Vega, Gina P; Peña, Karen T; Schindler, Yocheved L; Yelon, Deborah

    2016-01-01

    Proper organogenesis depends upon defining the precise dimensions of organ progenitor territories. Kidney progenitors originate within the intermediate mesoderm (IM), but the pathways that set the boundaries of the IM are poorly understood. Here, we show that the bHLH transcription factor Hand2 limits the size of the embryonic kidney by restricting IM dimensions. The IM is expanded in zebrafish hand2 mutants and is diminished when hand2 is overexpressed. Within the posterior mesoderm, hand2 is expressed laterally adjacent to the IM. Venous progenitors arise between these two territories, and hand2 promotes venous development while inhibiting IM formation at this interface. Furthermore, hand2 and the co-expressed zinc-finger transcription factor osr1 have functionally antagonistic influences on kidney development. Together, our data suggest that hand2 functions in opposition to osr1 to balance the formation of kidney and vein progenitors by regulating cell fate decisions at the lateral boundary of the IM. DOI: http://dx.doi.org/10.7554/eLife.19941.001 PMID:27805568

  5. VEGF expression in mesenchymal stem cells promotes bone formation of tissue-engineered bones.

    PubMed

    Liu, Boling; Li, Xihai; Liang, Guiqing; Liu, Xianxiang

    2011-01-01

    The purpose of this study was to investigate the in vivo vascularization and bone formation activity of tissue-engineered bone constructed using bone marrow mesenchymal stem cells (MSCs) transfected with vascular endothelial growth factor (VEGF). The expression of VEGF165 in rat bone marrow MSCs was confirmed using RT-PCR and immunohistochemistry. The MSCs were cultured together with nano-hydroxyapatite/collagen (NHAC) to form tissue-engineered bone. Untransfected MSCs were used as controls. The mice were sacrificed, and the bone xenografts were analyzed using immunohistochemistry and quantified for the degree of vascularization and new bone formation. Based on our results, expression of the VEGF165 gene was detected using RT-PCR and immunohistochemistry following transfection and 4 weeks of selection. The co-cultured NHAC- and VEGF-transfected MSCs had significantly higher alkaline phosphatase (AP) activity compared to the controls (P<0.05). In the mice that received the tissue-engineered bone xenografts, clumps of cartilage cells, irregular bone-like tissue and microvessels were observed. The growth of these structures progressed with time. In the control mice, however, only small amounts of bone-like and fibrotic tissue were observed. The differences between the control and experimental groups were statistically significant (P<0.05). In conclusion, VEGF165‑transfected bone marrow MSCs promotes vascularization of tissue-engineered bone and ectopic osteogenesis.

  6. The MinCDJ System in Bacillus subtilis Prevents Minicell Formation by Promoting Divisome Disassembly

    PubMed Central

    van Baarle, Suey; Bramkamp, Marc

    2010-01-01

    Background Cell division in Bacillus subtilis takes place precisely at midcell, through the action of Noc, which prevents division from occurring over the nucleoids, and the Min system, which prevents cell division from taking place at the poles. Originally it was thought that the Min system acts directly on FtsZ, preventing the formation of a Z-ring and, therefore, the formation of a complete cytokinetic ring at the poles. Recently, a new component of the B. subtilis Min system was identified, MinJ, which acts as a bridge between DivIVA and MinCD. Methodology/Principal Findings We used fluorescence microscopy and molecular genetics to examine the molecular role of MinJ. We found that in the absence of a functional Min system, FtsA, FtsL and PBP-2B remain associated with completed division sites. Evidence is provided that MinCDJ are responsible for the failure of these proteins to localize properly, indicating that MinCDJ can act on membrane integral components of the divisome. Conclusions/Significance Taken together, we postulate that the main function of the Min system is to prevent minicell formation adjacent to recently completed division sites by promoting the disassembly of the cytokinetic ring, thereby ensuring that cell division occurs only once per cell cycle. Thus, the role of the Min system in rod-shaped bacteria seems not to be restricted to an inhibitory function on FtsZ polymerization, but can act on different levels of the divisome. PMID:20352045

  7. Ezrin/Radixin/Moesin proteins and flotillins cooperate to promote uropod formation in T cells.

    PubMed

    Martinelli, Sibylla; Chen, Emily J H; Clarke, Fiona; Lyck, Ruth; Affentranger, Sarah; Burkhardt, Janis K; Niggli, Verena

    2013-01-01

    T cell uropods are enriched in specific proteins including adhesion receptors such as P-selectin glycoprotein ligand-1 (PSGL-1), lipid raft-associated proteins such as flotillins and ezrin/radixin/moesin (ERM) proteins which associate with cholesterol-rich raft domains and anchor adhesion receptors to the actin cytoskeleton. Using dominant mutants and siRNA technology we have tested the interactions among these proteins and their role in shaping the T cell uropod. Expression of wild type (WT) ezrin-EGFP failed to affect the morphology of human T cells or chemokine-induced uropod recruitment of PSGL-1 and flotillin-1 and -2. In contrast, expression of constitutively active T567D ezrin-EGFP induced a motile, polarized phenotype in some of the transfected T cells, even in the absence of chemokine. These cells featured F-actin-rich ruffles in the front and uropod enrichment of PSGL-1 and flotillins. T567D ezrin-EGFP was itself strongly enriched in the rear of the polarized T cells. Uropod formation induced by T567D ezrin-EGFP was actin-dependent as it was attenuated by inhibition of Rho-kinase or myosin II, and abolished by disruption of actin filaments. While expression of constitutively active ezrin enhanced cell polarity, expression of a dominant-negative deletion mutant of ezrin, 1-310 ezrin-EGFP, markedly reduced uropod formation induced by the chemokine SDF-1, T cell front-tail polarity, and capping of PSGL-1 and flotillins. Transfection of T cells with WT or T567D ezrin did not affect chemokine-mediated chemotaxis whereas 1-310 ezrin significantly impaired spontaneous 2D migration and chemotaxis. siRNA-mediated downregulation of flotillins in murine T cells attenuated moesin capping and uropod formation, indicating that ERM proteins and flotillins cooperate in uropod formation. In summary, our results indicate that activated ERM proteins function together with flotillins to promote efficient chemotaxis of T cells by structuring the uropod of migrating T cells.

  8. Monoterpenes in the glandular trichomes of tomato are synthesized from a neryl diphosphate precursor rather than geranyl diphosphate.

    PubMed

    Schilmiller, Anthony L; Schauvinhold, Ines; Larson, Matthew; Xu, Richard; Charbonneau, Amanda L; Schmidt, Adam; Wilkerson, Curtis; Last, Robert L; Pichersky, Eran

    2009-06-30

    We identified a cis-prenyltransferase gene, neryl diphosphate synthase 1 (NDPS1), that is expressed in cultivated tomato (Solanum lycopersicum) cultivar M82 type VI glandular trichomes and encodes an enzyme that catalyzes the formation of neryl diphosphate from isopentenyl diphosphate and dimethylallyl diphosphate. mRNA for a terpene synthase gene, phellandrene synthase 1 (PHS1), was also identified in these glands. It encodes an enzyme that uses neryl diphosphate to produce beta-phellandrene as the major product as well as a variety of other monoterpenes. The profile of monoterpenes produced by PHS1 is identical with the monoterpenes found in type VI glands. PHS1 and NDPS1 map to chromosome 8, and the presence of a segment of chromosome 8 derived from Solanum pennellii LA0716 causes conversion from the M82 gland monoterpene pattern to that characteristic of LA0716 plants. The data indicate that, contrary to the textbook view of geranyl diphosphate as the "universal" substrate of monoterpene synthases, in tomato glands neryl diphosphate serves as a precursor for the synthesis of monoterpenes.

  9. Hydroxyl radical induced by lipid in Maillard reaction model system promotes diet-derived N(ε)-carboxymethyllysine formation.

    PubMed

    Han, Lipeng; Li, Lin; Li, Bing; Zhao, Di; Li, Yuting; Xu, Zhenbo; Liu, Guoqin

    2013-10-01

    N(ε)-carboxymethyllysine (CML) is commonly found in food, and is considered as a potential hazard to human health. However, the effect of lipids on CML formation in Maillard reaction is still not clarified. In this study, the content of diet-derived CML and its key intermediates, epsilon-fructoselysine (FL) and glyoxal (GO), is determined with high performance liquid chromatography mass spectrum (HPLC-MS) in model system containing lipid compounds. According to the results, hydroxyl radical (OH) induced by Fenton reagent can promote the three pathways of CML formation. Moreover, in the Maillard reaction system, linoleic acid (Lin), oleic acid (Ole) and glycerol trioleate (Tri) can induce more OH·, which promotes CML formation. Their level of promoting CML formation is in the order of Ole>Lin>Tri. On the contrary, glycerol (Gly) can scavenge OH·, which inhibit the CML formation. Finally, it is proved that FL content and GO content decreases with heating time in model system, while CML content increases with heating time. Thus, it is concluded that in the Maillard reaction system lipids can induce more OH·, which promotes the conversion from FL and GO to CML. Our research may contribute to the development of inhibitory methods for diet-derived CML by scavenging OH·.

  10. EPO Promotes Bone Repair through Enhanced Cartilaginous Callus Formation and Angiogenesis

    PubMed Central

    Wan, Lin; Zhang, Fengjie; He, Qiling; Tsang, Wing Pui; Lu, Li; Li, Qingnan; Wu, Zhihong; Qiu, Guixing; Zhou, Guangqian; Wan, Chao

    2014-01-01

    Erythropoietin (EPO)/erythropoietin receptor (EPOR) signaling is involved in the development and regeneration of several non-hematopoietic tissues including the skeleton. EPO is identified as a downstream target of the hypoxia inducible factor-α (HIF-α) pathway. It is shown that EPO exerts a positive role in bone repair, however, the underlying cellular and molecular mechanisms remain unclear. In the present study we show that EPO and EPOR are expressed in the proliferating, pre-hypertrophic and hypertrophic zone of the developing mouse growth plates as well as in the cartilaginous callus of the healing bone. The proliferation rate of chondrocytes is increased under EPO treatment, while this effect is decreased following siRNA mediated knockdown of EPOR in chondrocytes. EPO treatment increases biosynthesis of proteoglycan, accompanied by up-regulation of chondrogenic marker genes including SOX9, SOX5, SOX6, collagen type 2, and aggrecan. The effects are inhibited by knockdown of EPOR. Blockage of the endogenous EPO in chondrocytes also impaired the chondrogenic differentiation. In addition, EPO promotes metatarsal endothelial sprouting in vitro. This coincides with the in vivo data that local delivery of EPO increases vascularity at the mid-stage of bone healing (day 14). In a mouse femoral fracture model, EPO promotes cartilaginous callus formation at days 7 and 14, and enhances bone healing at day 28 indexed by improved X-ray score and micro-CT analysis of microstructure of new bone regenerates, which results in improved biomechanical properties. Our results indicate that EPO enhances chondrogenic and angiogenic responses during bone repair. EPO's function on chondrocyte proliferation and differentiation is at least partially mediated by its receptor EPOR. EPO may serve as a therapeutic agent to facilitate skeletal regeneration. PMID:25003898

  11. EPO promotes bone repair through enhanced cartilaginous callus formation and angiogenesis.

    PubMed

    Wan, Lin; Zhang, Fengjie; He, Qiling; Tsang, Wing Pui; Lu, Li; Li, Qingnan; Wu, Zhihong; Qiu, Guixing; Zhou, Guangqian; Wan, Chao

    2014-01-01

    Erythropoietin (EPO)/erythropoietin receptor (EPOR) signaling is involved in the development and regeneration of several non-hematopoietic tissues including the skeleton. EPO is identified as a downstream target of the hypoxia inducible factor-α (HIF-α) pathway. It is shown that EPO exerts a positive role in bone repair, however, the underlying cellular and molecular mechanisms remain unclear. In the present study we show that EPO and EPOR are expressed in the proliferating, pre-hypertrophic and hypertrophic zone of the developing mouse growth plates as well as in the cartilaginous callus of the healing bone. The proliferation rate of chondrocytes is increased under EPO treatment, while this effect is decreased following siRNA mediated knockdown of EPOR in chondrocytes. EPO treatment increases biosynthesis of proteoglycan, accompanied by up-regulation of chondrogenic marker genes including SOX9, SOX5, SOX6, collagen type 2, and aggrecan. The effects are inhibited by knockdown of EPOR. Blockage of the endogenous EPO in chondrocytes also impaired the chondrogenic differentiation. In addition, EPO promotes metatarsal endothelial sprouting in vitro. This coincides with the in vivo data that local delivery of EPO increases vascularity at the mid-stage of bone healing (day 14). In a mouse femoral fracture model, EPO promotes cartilaginous callus formation at days 7 and 14, and enhances bone healing at day 28 indexed by improved X-ray score and micro-CT analysis of microstructure of new bone regenerates, which results in improved biomechanical properties. Our results indicate that EPO enhances chondrogenic and angiogenic responses during bone repair. EPO's function on chondrocyte proliferation and differentiation is at least partially mediated by its receptor EPOR. EPO may serve as a therapeutic agent to facilitate skeletal regeneration.

  12. WHI-131 Promotes Osteoblast Differentiation and Prevents Osteoclast Formation and Resorption in Mice.

    PubMed

    Cheon, Yoon-Hee; Kim, Ju-Young; Baek, Jong Min; Ahn, Sung-Jun; Jun, Hong Young; Erkhembaatar, Munkhsoyol; Kim, Min Seuk; Lee, Myeung Su; Oh, Jaemin

    2016-02-01

    levels of genes related to osteoblast differentiation, and induced the phosphorylation of Akt, p38, and Smad1/5/8. Furthermore, 5-week-old ICR mice treated with WHI-131 exhibited antiresorbing effects in a lipopolysaccharide-induced calvaria bone loss model in vivo and increased bone-forming activity in a calvarial bone formation model. Therefore, the results of this study show that WHI-131 plays a dual role by inhibiting osteoclast differentiation and promoting osteoblast differentiation. Thus, WHI-131 could be a useful pharmacological agent to treat osteoporosis by promoting bone growth and inhibiting resorption.

  13. Ecological distribution of leaf stomata and trichomes among tree species in a Malaysian lowland tropical rain forest.

    PubMed

    Ichie, Tomoaki; Inoue, Yuta; Takahashi, Narumi; Kamiya, Koichi; Kenzo, Tanaka

    2016-07-01

    The vertical structure of a tropical rain forest is complex and multilayered, with strong variation of micro-environment with height up to the canopy. We investigated the relation between morphological traits of leaf surfaces and tree ecological characteristics in a Malaysian tropical rain forest. The shapes and densities of stomata and trichomes on the abaxial leaf surfaces and their relation with leaf characteristics such as leaf area and leaf mass per area (LMA) were studied in 136 tree species in 35 families with different growth forms in the tropical moist forest. Leaf physiological properties were also measured in 50 canopy and emergent species. Most tree species had flat type (40.4 %) or mound type (39.7 %) stomata. In addition, 84 species (61.76 %) in 22 families had trichomes, including those with glandular (17.65 %) and non-glandular trichomes (44.11 %). Most leaf characteristics significantly varied among the growth form types: species in canopy and emergent layers and canopy gap conditions had higher stomatal density, stomatal pore index (SPI), trichome density and LMA than species in understory and subcanopy layers, though the relation of phylogenetically independent contrasts to each characteristic was not statistically significant, except for leaf stomatal density, SPI and LMA. Intrinsic water use efficiency in canopy and emergent tree species with higher trichome densities was greater than in species with lower trichome densities. These results suggest that tree species in tropical rain forests adapt to a spatial difference in their growth forms, which are considerably affected by phylogenetic context, by having different stomatal and trichome shapes and/or densities.

  14. Promotion of experimental thrombus formation by the procoagulant activity of breast cancer cells

    NASA Astrophysics Data System (ADS)

    Berny-Lang, M. A.; Aslan, J. E.; Tormoen, G. W.; Patel, I. A.; Bock, P. E.; Gruber, A.; McCarty, O. J. T.

    2011-02-01

    The routine observation of tumor emboli in the peripheral blood of patients with carcinomas raises questions about the clinical relevance of these circulating tumor cells. Thrombosis is a common clinical manifestation of cancer, and circulating tumor cells may play a pathogenetic role in this process. The presence of coagulation-associated molecules on cancer cells has been described, but the mechanisms by which circulating tumor cells augment or alter coagulation remains unclear. In this study we utilized suspensions of a metastatic adenocarcinoma cell line, MDA-MB-231, and a non-metastatic breast epithelial cell line, MCF-10A, as models of circulating tumor cells to determine the thromobogenic activity of these blood-foreign cells. In human plasma, both metastatic MDA-MB-231 cells and non-metastatic MCF-10A cells significantly enhanced clotting kinetics. The effect of MDA-MB-231 and MCF-10A cells on clotting times was cell number-dependent and inhibited by a neutralizing antibody to tissue factor (TF) as well as inhibitors of activated factor X and thrombin. Using fluorescence microscopy, we found that both MDA-MB-231 and MCF-10A cells supported the binding of fluorescently labeled thrombin. Furthermore, in a model of thrombus formation under pressure-driven flow, MDA-MB-231 and MCF-10A cells significantly decreased the time to occlusion. Our findings indicate that the presence of breast epithelial cells in blood can stimulate coagulation in a TF-dependent manner, suggesting that tumor cells that enter the circulation may promote the formation of occlusive thrombi under shear flow conditions.

  15. Arsenic promotes centrosome abnormalities and cell colony formation in p53 compromised human lung cells

    SciTech Connect

    Liao Weiting; Lin Pinpin; Cheng, T.-S.; Yu, H.-S.; Chang, Louis W.

    2007-12-01

    Epidemiological evidence indicated that residents, especially cigarette smokers, in arseniasis areas had significantly higher lung cancer risk than those living in non-arseniasis areas. Thus, an interaction between arsenic and cigarette smoking in lung carcinogenesis was suspected. p53 dysfunction or mutation in lung epithelial cells was frequently observed in cigarette smokers. Our present study was to explore the differential effects by arsenic on H1355 cells (human lung adenocarcinoma cell line with mutation in p53), BEAS-2B (immortalized lung epithelial cell with functional p53) and pifithrin-{alpha}-treated BEAS-2B cells (p53-inhibited cells). These cells were treated with different doses of sodium arsenite (0, 0.1, 1, 5 and 10 {mu}M) for 48 h. A greater reduction in cell viability was observed in the BEAS-2B cells vs. p53 compromised cells (H1355 or p53-inhibited BEAS-2B). Similar observation was also made on 7-day cell survival (growth) study. TUNEL analysis confirmed that there was indeed a significantly reduced arsenite-induced apoptosis found in p53-compromised cells. Centrosomal abnormality has been attributed to eventual chromosomal missegregation, aneuploidy and tumorigenesis. In our present study, reduced p21 and Gadd45a expressions and increased centrosomal abnormality (atopic and multiple centrosomes) were observed in both arsenite-treated H1355 and p53-inhibited BEAS-2B cells as compared with similarly treated BEAS-2B cells. Increased anchorage-independent growth (colony formation) of BEAS-2B cells co-treated with pifithrin-{alpha} and 5 {mu}M sodium arsenite was also observed in soft agar. Our present investigation demonstrated that arsenic would act specifically on p53 compromised cells (either with p53 dysfunction or inhibited) to induce centrosomal abnormality and colony formation. These findings provided strong evidence on the carcinogenic promotional role of arsenic, especially under the condition of p53 dysfunction.

  16. GLUTAMATERGIC SYNAPSE FORMATION IS PROMOTED BY α7-CONTAINING NICOTINIC ACETYLCHOLINE RECEPTORS

    PubMed Central

    Lozada, Adrian F.; Wang, Xulong; Gounko, Natalia V.; Massey, Kerri A.; Duan, Jingjing; Liu, Zhaoping; Berg, Darwin K.

    2012-01-01

    Glutamate is the primary excitatory transmitter in adult brain, acting through synapses on dendritic spines and shafts. Early in development, however, when glutamatergic synapses are only beginning to form, nicotinic cholinergic excitation is already widespread; it is mediated by acetylcholine activating nicotinic acetylcholine receptors (nAChRs) that generate waves of activity across brain regions. A major class of nAChRs contributing at this time is a species containing α7 subunits (α7-nAChRs). These receptors are highly permeable to calcium, influence a variety of calcium-dependent events, and are diversely distributed throughout the developing CNS. Here we show that α7-nAChRs unexpectedly promote formation of glutamatergic synapses during development. The dependence on α7-nAChRs becomes clear when comparing wild-type mice with mice constitutively lacking the α7-nAChR gene. Ultrastructural analysis, immunostaining, and patch-clamp recording all reveal synaptic deficits when α7-nAChR input is absent. Similarly, nicotinic activation of α7-nAChRs in wild-type organotypic culture, as well as cell culture, increases the number of glutamatergic synapses. RNA interference demonstrates that the α7-nAChRs must be expressed in the neuron being innervated for normal innervation to occur. Moreover the deficits persist throughout the developmental period of major de novo synapse formation and are still fully apparent in the adult. GABAergic synapses, in contrast, are undiminished in number under such conditions. As a result, mice lacking α7-nAChRs have an altered balance in the excitatory/inhibitory input they receive. This ratio represents a fundamental feature of neural networks and shows for the first time that endogenous nicotinic cholinergic signaling plays a key role in network construction. PMID:22649244

  17. Absence of keratin 8 or 18 promotes antimitochondrial autoantibody formation in aging male mice

    PubMed Central

    Toivola, Diana M.; Habtezion, Aida; Misiorek, Julia O.; Zhang, Linxing; Nyström, Joel H.; Sharpe, Orr; Robinson, William H.; Kwan, Raymond; Omary, M. Bishr

    2015-01-01

    Human mutations in keratin 8 (K8) and keratin 18 (K18), the intermediate filament proteins of hepatocytes, predispose to several liver diseases. K8-null mice develop chronic liver injury and fragile hepatocytes, dysfunctional mitochondria, and Th2-type colitis. We tested the hypothesis that autoantibody formation accompanies the liver damage that associates with K8/K18 absence. Sera from wild-type control, K8-null, and K18-null mice were analyzed by immunoblotting and immunofluorescence staining of cell and mouse tissue homogenates. Autoantibodies to several antigens were identified in 81% of K8-null male mice 8 mo or older. Similar autoantibodies were detected in aging K18-null male mice that had a related liver phenotype but normal colon compared with K8-null mice, suggesting that the autoantibodies are linked to liver rather than colonic disease. However, these autoantibodies were not observed in nontransgenic mice subjected to 4 chronic injury models. The autoantigens are ubiquitous and partition with mitochondria. Mass spectrometry and purified protein analysis identified, mitochondrial HMG-CoA synthase, aldehyde dehydrogenase, and catalase as the primary autoantigens, and glutamate dehydrogenase and epoxide hydrolase-2 as additional autoantigens. Therefore, absence of the hepatocyte keratins results in production of anti-mitochondrial autoantibodies (AMA) that recognize proteins involved in energy metabolism and oxidative stress, raising the possibility that AMA may be found in patients with keratin mutations that associate with liver and other diseases.—Toivola, D. M., Habtezion, A., Misiorek, J. O., Zhang, L., Nyström, J. H., Sharpe, O., Robinson, W. H., Kwan, R., Omary, M. B. Absence of keratin 8 or 18 promotes antimitochondrial autoantibody formation in aging male mice. PMID:26399787

  18. Spp1, a member of the Set1 Complex, promotes meiotic DSB formation in promoters by tethering histone H3K4 methylation sites to chromosome axes.

    PubMed

    Sommermeyer, Vérane; Béneut, Claire; Chaplais, Emmanuel; Serrentino, Maria Elisabetta; Borde, Valérie

    2013-01-10

    Meiotic chromosomes are organized into arrays of loops that are anchored to the chromosome axis structure. Programmed DNA double-strand breaks (DSBs) that initiate meiotic recombination, catalyzed by Spo11 and accessory DSB proteins, form in loop sequences in promoters, whereas the DSB proteins are located on chromosome axes. Mechanisms bridging these two chromosomal regions for DSB formation have remained elusive. Here we show that Spp1, a conserved member of the histone H3K4 methyltransferase Set1 complex, is required for normal levels of DSB formation and is associated with chromosome axes during meiosis, where it physically interacts with the Mer2 DSB protein. The PHD finger module of Spp1, which reads H3K4 methylation close to promoters, promotes DSB formation by tethering these regions to chromosome axes and activating cleavage by the DSB proteins. This paper provides the molecular mechanism linking DSB sequences to chromosome axes and explains why H3K4 methylation is important for meiotic recombination.

  19. Polymethylated Myricetin in Trichomes of the Wild Tomato Species Solanum habrochaites and Characterization of Trichome-Specific 3′/5′- and 7/4′-Myricetin O-Methyltransferases1[C][W][OA

    PubMed Central

    Schmidt, Adam; Li, Chao; Shi, Feng; Jones, A. Daniel; Pichersky, Eran

    2011-01-01

    Flavonoids are a class of metabolites found in many plant species. They have been reported to serve several physiological roles, such as in defense against herbivores and pathogens and in protection against harmful ultraviolet radiation. They also serve as precursors of pigment compounds found in flowers, leaves, and seeds. Highly methylated, nonglycosylated derivatives of the flavonoid myricetin flavonoid, have been previously reported from a variety of plants, but O-methyltransferases responsible for their synthesis have not yet been identified. Here, we show that secreting glandular trichomes (designated types 1 and 4) and storage glandular trichomes (type 6) on the leaf surface of wild tomato (Solanum habrochaites accession LA1777) plants contain 3,7,3′-trimethyl myricetin, 3,7,3′,5′-tetramethyl myricetin, and 3,7,3′,4′,5′-pentamethyl myricetin, with gland types 1 and 4 containing severalfold more of these compounds than type 6 glands and with the tetramethylated compound predominating in all three gland types. We have also identified transcripts of two genes expressed in the glandular trichomes and showed that they encode enzymes capable of methylating myricetin at the 3′ and 5′ and the 7 and 4′ positions, respectively. Both genes are preferentially expressed in secreting glandular trichome types 1 and 4 and to a lesser degree in storage trichome type 6, and the levels of the proteins they encode are correspondingly higher in types 1 and 4 glands compared with type 6 glands. PMID:21343428

  20. Soybean NAC transcription factors promote abiotic stress tolerance and lateral root formation in transgenic plants.

    PubMed

    Hao, Yu-Jun; Wei, Wei; Song, Qing-Xin; Chen, Hao-Wei; Zhang, Yu-Qin; Wang, Fang; Zou, Hong-Feng; Lei, Gang; Tian, Ai-Guo; Zhang, Wan-Ke; Ma, Biao; Zhang, Jin-Song; Chen, Shou-Yi

    2011-10-01

    NAC transcription factors play important roles in plant growth, development and stress responses. Previously, we identified multiple NAC genes in soybean (Glycine max). Here, we identify the roles of two genes, GmNAC11 and GmNAC20, in stress responses and other processes. The two genes were differentially induced by multiple abiotic stresses and plant hormones, and their transcripts were abundant in roots and cotyledons. Both genes encoded proteins that localized to the nucleus and bound to the core DNA sequence CGT[G/A]. In the protoplast assay system, GmNAC11 acts as a transcriptional activator, whereas GmNAC20 functions as a mild repressor; however, the C-terminal end of GmANC20 has transcriptional activation activity. Over-expression of GmNAC20 enhances salt and freezing tolerance in transgenic Arabidopsis plants; however, GmNAC11 over-expression only improves salt tolerance. Over-expression of GmNAC20 also promotes lateral root formation. GmNAC20 may regulate stress tolerance through activation of the DREB/CBF-COR pathway, and may control lateral root development by altering auxin signaling-related genes. GmNAC11 probably regulates DREB1A and other stress-related genes. The roles of the two GmNAC genes in stress tolerance were further analyzed in soybean transgenic hairy roots. These results provide a basis for genetic manipulation to improve the agronomic traits of important crops.

  1. MEKK2 mediates an alternative β-catenin pathway that promotes bone formation

    PubMed Central

    Greenblatt, Matthew Blake; Shin, Dong Yeon; Oh, Hwanhee; Lee, Ki-Young; Zhai, Bo; Gygi, Steven P.; Lotinun, Sutada; Baron, Roland; Liu, Dou; Su, Bing; Glimcher, Laurie H.; Shim, Jae-Hyuck

    2016-01-01

    Proper tuning of β-catenin activity in osteoblasts is required for bone homeostasis, because both increased and decreased β-catenin activity have pathologic consequences. In the classical pathway for β-catenin activation, stimulation with WNT ligands suppresses constitutive phosphorylation of β-catenin by glycogen synthase kinase 3β, preventing β-catenin ubiquitination and proteasomal degradation. Here, we have found that mitogen-activated protein kinase kinase kinase 2 (MAP3K2 or MEKK2) mediates an alternative pathway for β-catenin activation in osteoblasts that is distinct from the canonical WNT pathway. FGF2 activates MEKK2 to phosphorylate β-catenin at serine 675, promoting recruitment of the deubiquitinating enzyme, ubiquitin-specific peptidase 15 (USP15). USP15 in turn prevents the basal turnover of β-catenin by inhibiting its ubiquitin-dependent proteasomal degradation, thereby enhancing WNT signaling. Analysis of MEKK2-deficient mice and genetic interaction studies between Mekk2- and β-catenin–null alleles confirm that this pathway is an important physiologic regulator of bone mass in vivo. Thus, an FGF2/MEKK2 pathway mediates an alternative nonclassical pathway for β-catenin activation, and this pathway is a key regulator of bone formation by osteoblasts. PMID:26884171

  2. NGF promotes long-term memory formation by activating poly(ADP-ribose)polymerase-1.

    PubMed

    Wang, Shao-Hui; Liao, Xiao-Mei; Liu, Dan; Hu, Juan; Yin, Yang-Yang; Wang, Jian-Zhi; Zhu, Ling-Qiang

    2012-11-01

    Nerve growth factor (NGF) is a critical secreted protein that plays an important role in development, survival, and function of the mammalian nervous system. Previously reports suggest that endogenous NGF is essential for the hippocampal plasticity/memory and NGF deprivation induces the impairment of hippocampus-related memory and synaptic plasticity. However, whether exogenous supplement of NGF could promote the hippocampus-dependent synaptic plasticity/memory and the possible underlying mechanisms are not clear. In this study we found that NGF administration facilitates the hippocampus-dependent long-term memory and synaptic plasticity by increasing the activity of PARP-1, a polymerase mediating the PolyADP-ribosylation and important for the memory formation. Co-application of 3-Aminobenzamide (3-AB), a specific inhibitor of PARP-1, distinctly blocked the boosting effect of NGF on memory and synaptic plasticity, and the activation of downstream PKA-CREB signal pathway. Our data provide the first evidence that NGF supplement facilitates synaptic plasticity and the memory ability through PARP-1-mediated protein polyADP-ribosylation and activation of PKA-CREB pathway.

  3. Unfractionated Heparin Promotes Osteoclast Formation in Vitro by Inhibiting Osteoprotegerin Activity.

    PubMed

    Li, Binghan; Lu, Dan; Chen, Yuqing; Zhao, Minghui; Zuo, Li

    2016-04-22

    Heparin has been proven to enhance bone resorption and induce bone loss. Since osteoclasts play a pivotal role in bone resorption, the effect of heparin on osteoclastogenesis needs to be clarified. Since osteocytes are the key modulator during osteoclastogenesis, we evaluated heparin's effect on osteoclastogenesis in vitro by co-culturing an osteocyte cell line (MLO-Y4) and pre-osteoclasts (RAW264.7). In this co-culture system, heparin enhanced osteoclastogenesis and osteoclastic bone resorption while having no influence on the production of RANKL (receptor activator of NFκB ligand), M-CSF (macrophage colony-stimulating factor), and OPG (osteoprotegerin), which are three main regulatory factors derived from osteocytes. According to previous studies, heparin could bind specifically to OPG and inhibit its activity, so we hypothesized that this might be a possible mechanism of heparin activity. To test this hypothesis, osteoclastogenesis was induced using recombinant RANKL or MLO-Y4 supernatant. We found that heparin has no effect on RANKL-induced osteoclastogenesis (contains no OPG). However, after incubation with OPG, the capacity of MLO-Y4 supernatant for supporting osteoclast formation was increased. This effect disappeared after OPG was neutralized and reappeared after OPG was replenished. These results strongly suggest that heparin promotes osteocyte-modulated osteoclastogenesis in vitro, at least partially, through inhibiting OPG activity.

  4. Phytotoxicity of constituents of glandular trichomes and the leaf surface of camphorweed, Heterotheca subaxillaris.

    PubMed

    Morimoto, Masanori; Cantrell, Charles L; Libous-Bailey, Lynn; Duke, Stephen O

    2009-01-01

    Camphorweed, Heterotheca subaxillaris (Lam.) Britt. & Rusby, has a camphor-like odor, and its leaf surfaces contain glandular trichomes of the type shown to contain high levels of isoprenoids in other species. Borneol (1), the phytotoxic calamenene-type sesquiterpenes (2-5, 9-11), and methylated flavones (12-15) were isolated from the dichloromethane rinsate of camphorweed aerial tissues. The strongest plant growth inhibitor against Agrostis stolonifera and Lactuca sativa seedlings, as well as duckweed (Lemna pausicostata), was 2-methoxy-calamenene-14-carboxylic acid (2). Esterification of calamenene carboxylic acids decreased their biological activity.

  5. Trichome-derived O-acyl sugars are a first meal for caterpillars that tags them for predation

    PubMed Central

    Weinhold, Alexander; Baldwin, Ian Thomas

    2011-01-01

    Plant glandular trichomes exude secondary metabolites with defensive functions, but these epidermal protuberances are surprisingly the first meal of Lepidopteran herbivores on Nicotiana attenuata. O-acyl sugars, the most abundant metabolite of glandular trichomes, impart a distinct volatile profile to the body and frass of larvae that feed on them. The headspace composition of Manduca sexta larvae is dominated by the branched chain aliphatic acids hydrolyzed from ingested O-acyl sugars, which waxes and wanes rapidly with trichome ingestion. In native habitats a ground-hunting predator, the omnivorous ant Pogonomyrmex rugosus, but not the big-eyed bug Geocoris spp., use these volatile aliphatic acids to locate their prey. PMID:21518882

  6. Trichomes and chemical composition of the volatile oil of Trichogonia cinerea (Gardner) R. M. King & H. Rob. (Eupatorieae, Asteraceae).

    PubMed

    Fernandes, Yanne S; Trindade, Luma M P; Rezende, Maria Helena; Paula, José R; Gonçalves, Letícia A

    2016-03-01

    Trichogonia cinerea is endemic to Brazil and occurs in areas of cerrado and campo rupestre. In this study, we characterized the glandular and non-glandular trichomes on the aerial parts of this species, determined the principal events in the development of the former, and identified the main constituents of the volatile oil produced in its aerial organs. Fully expanded leaves, internodes, florets, involucral bracts, and stem apices were used for the characterization of trichomes. Leaves, internodes, florets, and involucral bracts were examined by light microscopy and scanning electron microscopy, whereas stem apices were examined only by light microscopy. Branches in the reproductive phase were used for the extraction and determination of the composition of the volatile oil. The species has three types of glandular trichomes, biseriate vesicular, biseriate pedunculate, and multicellular uniseriate, which secrete volatile oils and phenolic compounds. The major components identified in the volatile oil were 3,5-muuroladiene (39.56%) and butylated hydroxytoluene (13.07%).

  7. Effect of external stress on density and size of glandular trichomes in full-grown Artemisia annua, the source of anti-malarial artemisinin

    PubMed Central

    Kjær, Anders; Grevsen, Kai; Jensen, Martin

    2012-01-01

    Background and aims Glandular trichomes (GT) of Artemisia annua produce valuable compounds for pharmaceutical and industrial uses, most notably the anti-malarial artemisinin. Our aim was to find out whether the density, number and size of GT can be manipulated to advantage by environmental stress. A range of external stress treatments, including stress response regulators, was therefore given to fully grown plants under field and greenhouse conditions. Methodology In a field experiment (Ex1), seed-grown plants were subjected to chemical or physical stress and plants analysed after 5 weeks. In a greenhouse experiment (Ex2), three groups of clonally derived plants were stressed at weekly intervals for 5 weeks. Stress treatments included sandblasting, leaf cutting and spraying with jasmonic acid, salicylic acid, chitosan oligosaccharide (COS), H2O2 (HP) and NaCl (SC)at different concentrations. Leaves from an upper and a lower position on the plants were analysed by fluorescence microscopy to determine the density and size of GT. Principal results Densities of GT on upper leaves of full-grown A. annua plants generally showed no response to external stress and only plants from one clone of Ex2 supported the hypothesis that increased density of GT was inducible in upper leaves by stress (significant for SC, HP and COS). The density of GT on lower leaves was not affected by stress in any experiment. Glandular trichomes were significantly smaller on the lower leaves in response to stress in Ex2, and a similar non-significant trend was observed in Ex1. Conclusions The results indicate a dynamic system in which stress treatments of large A. annua plants had a minor promoting effect on the initiation of GT in developing leaves, and a maturing effect of GT later in the lifetime of the individual GT. The hypothesis that applying stress can induce larger GT or more numerous GT was rejected. PMID:22833781

  8. Sucrose esters of carboxylic acids in glandular trichomes ofSolanum berthaultii deter settling and probing by green peach aphid.

    PubMed

    Neal, J J; Tingey, W M; Steffens, J C

    1990-02-01

    Removal of type B trichome exudate fromSolanum berthaultii leaflets leads to a decrease in tarsal gumming and mortality and an increase in feeding by the green peach aphid,Myzus persicae. Type B trichome exudate of theS. berthaultii accession PI 473331 is composed of a complex of 3',3,4,6-tetra-O-acyl sucroses containing primarily short-chain branched carboxylic acids. The acyl constituents are primarily derived from 2-methylpropanoic, 2-methylbutyric, and 8-methylnonanoic acids but constituents derived fromn-decanoic and dodecanoic acids are also present. Sucrose esters inhibit settling and probing by aphids in glass feeding cages.

  9. Capitate glandular trichomes of Helianthus annuus (Asteraceae): ultrastructure and cytological development.

    PubMed

    Amrehn, Evelyn; Heller, Annerose; Spring, Otmar

    2014-01-01

    Previous studies have shown that capitate glandular trichomes (CGT) of the common sunflower, Helianthus annuus, produce sesquiterpene lactones (STL) and flavonoids, which are sequestered and accumulated between the apical cuticle and the wall of the tip cells. To explore the cellular structures required and putatively involved in the STL biosynthesis and secretion, the present study was focused on the development of CGT and the comparison of the ultrastructure of its different cell types. Gradual maturation of flowers in the capitulum of the sunflower provided the possibility to study the simultaneous differentiation from the primordial to the secretory stage of CGT located by light microscopy (bright field, differential interference contrast and fluorescence) as well as transmission electron microscopy. It was shown that the CGT of sunflower anthers had a biseriate structure with up to 14 cell pairs. In mature trichomes, the apical cells called secretory cells were covered entirely by a large cuticle globe, which enclosed the resinous terpenoids and was specialised in thickness and structure. The secretory cells lacked chloroplasts and contained mainly smooth endoplasmic reticulum (sER). Conspicuous cell wall protuberances and an accumulation of mitochondria nearby occurred in the horizontally oriented cell walls. The cytological differences between stalk cells and secretory cells indicate a different function. The dominance of sER suggests its involvement in STL biosynthesis and cell wall protuberances enlarge the surface of the plasmamembrane of secretory cells and may be involved in the secretion processes of STL into the subcuticular space.

  10. Localization of sesquiterpene lactone biosynthesis in cells of capitate glandular trichomes of Helianthus annuus (Asteraceae).

    PubMed

    Amrehn, Evelyn; Aschenbrenner, Anna-Katharina; Heller, Annerose; Spring, Otmar

    2016-03-01

    Capitate glandular trichomes (CGT) of sunflower, Helianthus annuus, synthesize bioactive sesquiterpene lactones (STLs) within a short period of only a few days during trichome development. In the current project, the subcellular localization of H. annuus germacrene A monooxygenase (HaGAO), a key enzyme of the STL biosynthesis in sunflower CGT, was investigated. A polyclonal antibody raised against this enzyme was used for immunolabelling. HaGAO was found in secretory and stalk cells of CGT. This correlated with the appearance of smooth endoplasmic reticulum in both cell types. Stalk cells and secretory cells differed in form, size and types of plastids, but both had structures necessary for secretion. No HaGAO-specific immunoreaction was found in sunflower leaf tissue outside of CGT or in developing CGT before the secretory phase had started. Our results indicated that not only secretory cells but also nearly all cells of the CGT were involved in the biosynthesis of STL and that this process was not linked to the presence or absence of a specific type of plastid.

  11. Proteomic snapshot of spearmint (Mentha spicata L.) leaf trichomes: a genuine terpenoid factory.

    PubMed

    Champagne, Antoine; Boutry, Marc

    2013-11-01

    Peltate glandular trichomes from Mentha spicata were purified on a Percoll gradient and soluble and membrane proteins were trypsinized and the peptides were separated by nano-LC fractionation and analyzed by MALDI-MS/MS. The vast majority of the 1666 proteins identified were housekeeping proteins or involved in the primary metabolism. However, 57 were predicted to be involved in the secondary metabolism. Of these, 21 were involved in the synthesis of phenylpropanoids and phenolics and 32 in terpenoid synthesis. Of the 14 membrane transporters identified, the 11 ATP-binding cassette transporters provide good material for assessing whether active transport is required for the transfer of monoterpenoid intermediates between cellular compartments and for the secretion of the final products into the subcuticular storage cavity. In conclusion, this proteome analysis of M. spicata peltate trichomes has identified several candidate proteins that might be involved in terpenoid synthesis and transport. The data have been deposited to the ProteomeXchange with identifier PXD000352 (http://proteomecentral.proteomexchange.org/dataset/PXD000352).

  12. Filamentous Trichomic Prokaryotes in Carbonaceous Meteorites: Indigenous Microfossils, Minerals, or Modern Bio-Contaminants?

    NASA Technical Reports Server (NTRS)

    Hoover, Richard B.; Rozanov, Alexei Yu.

    2011-01-01

    Large complex filaments have been detected in freshly fractured interior surfaces of a variety of carbonaceous meteorites. Many exhibit the detailed morphological and morphometric characteristics of known filamentous trichomic prokaryotic microorganisms. In this paper we review prior studies of filamentous microstructures encountered in the meteorites along with the elemental compositions and characteristics of the, fibrous evaporite minerals and filamentous cyanobacteria and homologous trichomic sulfur bacteria. The meteorite images and elemental compositions will compared with data obtained with the same instruments for abiotic microstructures and living and fossil microorganisms in order to evaluate the relative merits of the alternate hypotheses that have been advanced to explain the nature and characteristics of the meteorite filaments. The possibiility that the filaments found in the meteorites may be comprise modern bio-contaminants will be evaluated in light of their observed elemental compositions and data by other researchers on the detection of indigenous complex organic biosignatures, and extraterrestrial amino acids and nucleobases found in the Murchison CM2 and the Orgueil CI1 carbonaceous meteorites.

  13. Leaf trichome responses to herbivory in willows: induction, relaxation and costs.

    PubMed

    Björkman, Christer; Dalin, Peter; Ahrné, Karin

    2008-01-01

    To circumvent the inherent problem of discriminating between the cost of losing photosynthetic tissue and the cost of producing an inducible defence, the growth response of herbivore-damaged plants was compared with plants damaged mechanically to the same extent but without eliciting the defence. Two experiments were conducted, studying the response of willows (Salix cinerea) to damage by adult leaf beetles (Phratora vulgatissima). In the first experiment, willows produced new leaves with an enhanced leaf trichome density 10-20 d after damage, coinciding in time with the feeding of beetle offspring. The response was relaxed in foliage produced 30-40 d after damage. In the second experiment, which also included mechanical damage, willows exposed to beetle feeding showed an increase in leaf trichome density of the same magnitude (> 70%) as in the first experiment. The cost of producing the defence was a 20% reduction in shoot length growth and biomass production. Willows exposed to mechanical damage had an 8% reduction in shoot length growth compared with control plants, that is, a cost of leaf area removal. The results are the first quantitative estimates of the cost of a plant defence induced by natural and low amounts (3.3%) of herbivory.

  14. Cloning of a sesquiterpene synthase from Lavandula x intermedia glandular trichomes.

    PubMed

    Sarker, Lukman S; Demissie, Zerihun A; Mahmoud, Soheil S

    2013-11-01

    The essential oil (EO) of Lavandula is dominated by monoterpenes, but can also contain small amounts of sesquiterpenes, depending on species and environmental conditions. For example, the sesquiterpene 9-epi-caryophyllene can make up to 8 % of the EO in a few species, including those commercially propagated for EO production. Here, we report the cloning and functional characterization of 9-epi-caryophyllene synthase (LiCPS) from the glandular trichomes of Lavandula x intermedia, cv. Grosso. The 1,617 bp open reading frame of LiCPS, which did not encode a transit peptide, was expressed in Escherichia coli and the recombinant protein purified by Ni-NTA agarose affinity chromatography. The ca. 60 kDa recombinant protein specifically converted farnesyl diphosphate to 9-epi-caryophyllene. LiCPS also produced a few monoterpenes when assayed with the monoterpene precursor geranyl diphosphate (GPP), but--unlike most monoterpene synthases--was not able to derive detectable amounts of any products from the cis isomer of GPP, neryl diphosphate. The LiCPS transcripts accumulated in developing L. x intermedia flowers and were highly enriched in glandular trichomes, but were not detected in leaves suggesting that the transcriptional expression of this gene is spatially and developmentally regulated.

  15. Molecular cloning and functional characterization of borneol dehydrogenase from the glandular trichomes of Lavandula x intermedia.

    PubMed

    Sarker, Lukman S; Galata, Mariana; Demissie, Zerihun A; Mahmoud, Soheil S

    2012-12-15

    Several varieties of Lavandula x intermedia (lavandins) are cultivated for their essential oils (EOs) for use in cosmetic, hygiene and personal care products. These EOs are mainly constituted of monoterpenes including camphor, which contributes an off odor reducing the olfactory appeal of the oil. We have recently constructed a cDNA library from the glandular trichomes (the sites of EO synthesis) of L. x intermedia plants. Here, we describe the cloning of a borneol dehydrogenase cDNA (LiBDH) from this library. The 780 bp open reading frame of the cDNA encoded a 259 amino acid short chain alcohol dehydrogenase with a predicted molecular mass of ca. 27.5 kDa. The recombinant LiBDH was expressed in Escherichia coli, purified by Ni-NTA agarose affinity chromatography, and functionally characterized in vitro. The bacterially produced enzyme specifically converted borneol to camphor as the only product with K(m) and k(cat) values of 53 μM and 4.0 × 10(-4) s(-1), respectively. The LiBDH transcripts were specifically expressed in glandular trichomes of mature flowers indicating that like other Lavandula monoterpene synthases the expression of this gene is regulated in a tissue-specific manner. The cloning of LiBDH has far reaching implications in improving the quality of Lavandula EOs through metabolic engineering.

  16. Trichomes related to an unusual method of water retention and protection of the stem apex in an arid zone perennial species

    PubMed Central

    Lusa, Makeli Garibotti; Cardoso, Elaine Cristina; Machado, Silvia Rodrigues; Appezzato-da-Glória, Beatriz

    2015-01-01

    It is well known that trichomes protect plant organs, and several studies have investigated their role in the adaptation of plants to harsh environments. Recent studies have shown that the production of hydrophilic substances by glandular trichomes and the deposition of this secretion on young organs may facilitate water retention, thus preventing desiccation and favouring organ growth until the plant develops other protective mechanisms. Lychnophora diamantinana is a species endemic to the Brazilian ‘campos rupestres’ (rocky fields), a region characterized by intense solar radiation and water deficits. This study sought to investigate trichomes and the origin of the substances observed on the stem apices of L. diamantinana. Samples of stem apices, young and expanded leaves were studied using standard techniques, including light microscopy and scanning and transmission electron microscopy. Histochemical tests were used to identify the major groups of metabolites present in the trichomes and the hyaline material deposited on the apices. Non-glandular trichomes and glandular trichomes were observed. The material deposited on the stem apices was hyaline, highly hydrophilic and viscous. This hyaline material primarily consists of carbohydrates that result from the partial degradation of the cell wall of uniseriate trichomes. This degradation occurs at the same time that glandular trichomes secrete terpenoids, phenolic compounds and proteins. These results suggest that the non-glandular trichomes on the leaves of L. diamantinana help protect the young organ, particularly against desiccation, by deposition of highly hydrated substances on the apices. Furthermore, the secretion of glandular trichomes probably repels herbivore and pathogen attacks. PMID:25527474

  17. In vitro oxidation of fibrinogen promotes functional alterations and formation of advanced oxidation protein products, an inflammation mediator.

    PubMed

    Torbitz, Vanessa Dorneles; Bochi, Guilherme Vargas; de Carvalho, José Antônio Mainardi; de Almeida Vaucher, Rodrigo; da Silva, José Edson Paz; Moresco, Rafael Noal

    2015-01-01

    Fibrinogen (FB) is a soluble blood plasma protein and is a key molecule involved in coagulation. Oxidative modification of proteins, such as the formation of advanced oxidation protein products (AOPP), a heterogeneous family of protein compounds structurally modified and derived from oxidative stress, may be associated with the pathophysiology of a number of chronic inflammatory diseases. Therefore, the aim of this study was to determine whether the formation of this mediator of inflammation occurs from FB and whether its generation is associated with structural changes. Results of the present study suggest that the oxidation of FB may provoke the formation of AOPP, which in turn, may promote functional alterations in FB, thus causing changes in its structural domains and increasing its procoagulant activity.

  18. Terrific Trichomes (and Other Specialised Cells) in African Violets: How to Get a Lot from One Plant in the Classroom

    ERIC Educational Resources Information Center

    Cottrell, Vicki M.

    2013-01-01

    African violet (genus "Saintpaulia") was identified as a particularly suitable genus for the study of specialised plant cells in the classroom using microscopes. The techniques described here involve simple preparation without staining. The cells and structures that can be investigated include: trichomes (hairs); stomata; guard cells and…

  19. Identification and characterization of two bisabolene synthases from linear glandular trichomes of sunflower (Helianthus annuus L., Asteraceae).

    PubMed

    Aschenbrenner, Anna-Katharina; Kwon, Moonhyuk; Conrad, Jürgen; Ro, Dae-Kyun; Spring, Otmar

    2016-04-01

    Sunflower is known to produce a variety of bisabolene-type sesquiterpenes and accumulates these substances in trichomes of leaves, stems and flowering parts. A bioinformatics approach was used to identify the enzyme responsible for the initial step in the biosynthesis of these compounds from its precursor farnesyl pyrophosphate. Based on sequence similarity with a known bisabolene synthases from Arabidopsis thaliana AtTPS12, candidate genes of Helianthus were searched in EST-database and used to design specific primers. PCR experiments identified two candidates in the RNA pool of linear glandular trichomes of sunflower. Their sequences contained the typical motifs of sesquiterpene synthases and their expression in yeast functionally characterized them as bisabolene synthases. Spectroscopic analysis identified the stereochemistry of the product of both enzymes as (Z)-γ-bisabolene. The origin of the two sunflower bisabolene synthase genes from the transcripts of linear trichomes indicates that they may be involved in the synthesis of sesquiterpenes produced in these trichomes. Comparison of the amino acid sequences of the sunflower bisabolene synthases showed high similarity with sesquiterpene synthases from other Asteracean species and indicated putative evolutionary origin from a β-farnesene synthase.

  20. Opposing Roles of Foliar and Glandular Trichome Volatile Components in Cultivated Nightshade Interaction with a Specialist Herbivore

    PubMed Central

    Murungi, Lucy Kananu; Kirwa, Hillary; Salifu, Daisy; Torto, Baldwyn

    2016-01-01

    Plant chemistry is an important contributor to the interaction with herbivores. Here, we report on a previously unknown role for foliar and glandular trichome volatiles in their interaction with the specialist herbivore of solanaceous plants, the tomato red spider mite Tetranychus evansi. We used various bioassays and chemical analyses including coupled gas chromatography-mass spectrometry (GC/MS) and liquid chromatography coupled to quadrupole time of flight mass spectrometry (LC-QToF-MS) to investigate this interaction between cultivated African nightshades and T. evansi. We show that, whereas morphologically different cultivated African nightshade species released similar foliar volatile organic compounds (VOCs) that attracted T. evansi, VOCs released from exudates of ruptured glandular trichomes of one nightshade species influenced local defense on the leaf surface. VOCs from ruptured glandular trichomes comprising mainly saturated and unsaturated fatty acids deterred T. evansi oviposition. Of the fatty acids, the unsaturated fatty acids accounted for >40% of the oviposition deterrent activity. Our findings point to a defense strategy in a plant, based on opposing roles for volatiles released by foliar and glandular trichomes in response to attack by a specialist herbivore. PMID:27556560

  1. Opposing Roles of Foliar and Glandular Trichome Volatile Components in Cultivated Nightshade Interaction with a Specialist Herbivore.

    PubMed

    Murungi, Lucy Kananu; Kirwa, Hillary; Salifu, Daisy; Torto, Baldwyn

    2016-01-01

    Plant chemistry is an important contributor to the interaction with herbivores. Here, we report on a previously unknown role for foliar and glandular trichome volatiles in their interaction with the specialist herbivore of solanaceous plants, the tomato red spider mite Tetranychus evansi. We used various bioassays and chemical analyses including coupled gas chromatography-mass spectrometry (GC/MS) and liquid chromatography coupled to quadrupole time of flight mass spectrometry (LC-QToF-MS) to investigate this interaction between cultivated African nightshades and T. evansi. We show that, whereas morphologically different cultivated African nightshade species released similar foliar volatile organic compounds (VOCs) that attracted T. evansi, VOCs released from exudates of ruptured glandular trichomes of one nightshade species influenced local defense on the leaf surface. VOCs from ruptured glandular trichomes comprising mainly saturated and unsaturated fatty acids deterred T. evansi oviposition. Of the fatty acids, the unsaturated fatty acids accounted for >40% of the oviposition deterrent activity. Our findings point to a defense strategy in a plant, based on opposing roles for volatiles released by foliar and glandular trichomes in response to attack by a specialist herbivore.

  2. Azelnidipine, Not Amlodipine, Induces Secretion of Vascular Endothelial Growth Factor From Smooth Muscle Cells and Promotes Endothelial Tube Formation

    PubMed Central

    Kawamura, Akira; Miura, Shin-ichiro; Matsuo, Yoshino; Tanigawa, Hiroyuki; Saku, Keijiro

    2014-01-01

    Background We previously reported that the calcium channel blocker (CCB) nifedipine-induced secretion of vascular endothelial growth factor (VEGF) from human coronary smooth muscle cells (HCSMCs) promoted human coronary endothelial cell (HCEC) tube formation. Therefore, we analyzed whether other CCBs, azelnidipine and amlodipine, also induced the secretion of VEGF and promoted HCEC tube formation, and the underlying molecular mechanisms. Methods To evaluate the tube formation, HCECs were grown on Matrigel for 18 hours in the supernatants from HCSMCs that had been treated with different kinds of reagents. Concentrations of VEGF in cultured HCSMCs were determined by specific enzyme immunoassays. Nuclear extracts from HCSMCs were prepared, and nuclear factor-kappa B (NF-κB) activation was measured by EZ-DetectTM Transcription Factor Kits for NF-κB p50 or p65. Results Although azelnidipine dose-dependently stimulated the significant secretion of VEGF from HCSMCs and this stimulation was abolished by a protein kinase C inhibitor, amlodipine-induced secretion of VEGF was significantly lower than that induced by azelnidipine. The medium derived from azelnidipine (at up to 2 μM)-treated HCSMCs led to HCEC tube formation, whereas that obtained with amlodipine did not. Azelnidipine-induced tube formation was blocked by an inhibitor of kinase insert domain-containing receptor/fetal liver kinase-1 tyrosine kinase. Azelnidipine at up to 2 μM induced NF-κB activation. Conclusions Azelnidipine, but not amlodipine, stimulated the secretion of VEGF from HCSMCs and induced HCEC tube formation. This secretion is mediated at least in part via the activation of NF-κB. Azelnidipine may have a novel beneficial effect in improving coronary microvascular blood flow in addition to its main effect of lowering blood pressure.

  3. A New Glabrous Gene (csgl3) Identified in Trichome Development in Cucumber (Cucumis sativus L.)

    PubMed Central

    Ding, Li-Hong; Wehner, Todd C.; Liu, Pan-Na; Wang, Ye; Zhang, Sheng-Ping; Gu, Xing-Fang

    2016-01-01

    Spines or trichomes on the fruit of cucumbers enhance their commercial value in China. In addition, glabrous mutants exhibit resistance to aphids and therefore their use by growers can reduce pesticide residues. Previous studies have reported two glabrous mutant plants containing the genes, csgl1 and csgl2. In the present study, a new glabrous mutant, NCG157, was identified showing a gene interaction effect with csgl1 and csgl2. This mutant showed the glabrous character on stems, leaves, tendrils, receptacles and ovaries, and there were no spines or tumors on the fruit surface. Inheritance analysis showed that a single recessive gene, named csgl3, determined the glabrous trait. An F2 population derived from the cross of two inbred lines 9930 (a fresh market type from Northern China that exhibits trichomes) and NCG157 (an American processing type with glabrous surfaces) was used for genetic mapping of the csgl3 gene. By combining bulked segregant analysis (BAS) with molecular markers, 18 markers, including two simple sequence repeats (SSR), nine insertion deletions (InDel) and seven derived cleaved amplified polymorphism sequences (dCAPs), were identified to link to the csgl3 gene. All of the linked markers were used as anchor loci to locate the csgl3 gene on cucumber chromosome 6. The csgl3 gene was mapped between the dCAPs markers dCAPs-21 and dCAPs-19, at genetic distances of 0.05 cM and 0.15 cM, respectively. The physical distance of this region was 19.6 kb. Three markers, InDel-19, dCAPs-2 and dCAPs-11, co-segregated with csgl3. There were two candidate genes in the region, Csa6M514860 and Csa6M514870. Quantitative real-time PCR showed that the expression of Csa6M514870 was higher in the tissues of 9930 than that of NCG157, and this was consistent with their phenotypic characters. Csa6M514870 is therefore postulated to be the candidate gene for the development of trichomes in cucumber. This study will facilitate marker-assisted selection (MAS) of the smooth

  4. A New Glabrous Gene (csgl3) Identified in Trichome Development in Cucumber (Cucumis sativus L.).

    PubMed

    Cui, Jin-Ying; Miao, Han; Ding, Li-Hong; Wehner, Todd C; Liu, Pan-Na; Wang, Ye; Zhang, Sheng-Ping; Gu, Xing-Fang

    2016-01-01

    Spines or trichomes on the fruit of cucumbers enhance their commercial value in China. In addition, glabrous mutants exhibit resistance to aphids and therefore their use by growers can reduce pesticide residues. Previous studies have reported two glabrous mutant plants containing the genes, csgl1 and csgl2. In the present study, a new glabrous mutant, NCG157, was identified showing a gene interaction effect with csgl1 and csgl2. This mutant showed the glabrous character on stems, leaves, tendrils, receptacles and ovaries, and there were no spines or tumors on the fruit surface. Inheritance analysis showed that a single recessive gene, named csgl3, determined the glabrous trait. An F2 population derived from the cross of two inbred lines 9930 (a fresh market type from Northern China that exhibits trichomes) and NCG157 (an American processing type with glabrous surfaces) was used for genetic mapping of the csgl3 gene. By combining bulked segregant analysis (BAS) with molecular markers, 18 markers, including two simple sequence repeats (SSR), nine insertion deletions (InDel) and seven derived cleaved amplified polymorphism sequences (dCAPs), were identified to link to the csgl3 gene. All of the linked markers were used as anchor loci to locate the csgl3 gene on cucumber chromosome 6. The csgl3 gene was mapped between the dCAPs markers dCAPs-21 and dCAPs-19, at genetic distances of 0.05 cM and 0.15 cM, respectively. The physical distance of this region was 19.6 kb. Three markers, InDel-19, dCAPs-2 and dCAPs-11, co-segregated with csgl3. There were two candidate genes in the region, Csa6M514860 and Csa6M514870. Quantitative real-time PCR showed that the expression of Csa6M514870 was higher in the tissues of 9930 than that of NCG157, and this was consistent with their phenotypic characters. Csa6M514870 is therefore postulated to be the candidate gene for the development of trichomes in cucumber. This study will facilitate marker-assisted selection (MAS) of the smooth

  5. DNA structural variation affects complex formation and promoter melting in ribosomal RNA transcription.

    PubMed

    Marilley, M; Radebaugh, C A; Geiss, G K; Laybourn, P J; Paule, M R

    2002-08-01

    Eukaryotic ribosomal RNA promoters exhibit an unusual conservation of non-canonical DNA structure (curvature, twist angle and duplex stability) despite a lack of primary sequence conservation. This raises the possibility that rRNA transcription factors might utilize structural anomalies in their sequence recognition process. We have analyzed in detail the interaction of the polymerase I transcription factor TIF-IB from Acanthmoeba castellanii with the CORE promoter. TIF-IB interacts primarily with the minor groove of the promoter. By correlating the effects on transcription and on DNA structure of promoter point mutations, we show that the TIF-IB interaction is strongly inhibited by increases in minor groove width. This suggests that a particular DNA structure is required for interaction with the transcription factor. In addition, TIF-IB induces a small bend in the promoter upon binding. Modeling of this bend reveals that it requires an additional narrowing of the minor groove, which would favor binding to mutants with narrower grooves. We also discuss how this narrowing would induce a small destabilization of the helix upstream of the transcription start site. Telestability predicts this would result in destabilization of the sequence that melts during initiation, suggesting that TIF-IB may have a role in stimulating melting.

  6. Surfactant process for promoting gas hydrate formation and application of the same

    DOEpatents

    Rogers, Rudy E.; Zhong, Yu

    2002-01-01

    This invention relates to a method of storing gas using gas hydrates comprising forming gas hydrates in the presence of a water-surfactant solution that comprises water and surfactant. The addition of minor amounts of surfactant increases the gas hydrate formation rate, increases packing density of the solid hydrate mass and simplifies the formation-storage-decomposition process of gas hydrates. The minor amounts of surfactant also enhance the potential of gas hydrates for industrial storage applications.

  7. Genetic elaborations of glandular and non-glandular trichomes in Mentha arvensis genotypes: assessing genotypic and phenotypic correlations along with gene expressions.

    PubMed

    Mishra, Anand; Lal, R K; Chanotiya, C S; Dhawan, Sunita Singh

    2017-03-01

    Mentha arvensis (corn mint) is well known for the production of menthol, a widely used commodity in pharma and flavoring industries and provides natural fragrances and products. Glandular trichomes are specialized hairs found on the aerial surface of vascular plants species producing specific secondary metabolite chemistry. Correlations were established among trichomes, oil yield, and major secondary metabolites. Nine improved, elite cultivars representing different M. arvensis genotypes were used for analysis. Phenotypic coefficient of variation (PCV) and genotypic coefficient of variation (GCV) were estimated; results indicated the presence of considerable amount of genetic variability, thereby emphasizing wide scope of selection. Positive and significant associations were found among glandular trichomes, oil yield, essential oil constituents, and leaf morphology itself, whereas morphological parameters of leaf show positive and negative correlations to average number of trichome and essential oil constituents. Average number of glandular, non-glandular trichomes, their ratios, menthol content, and trichome number showed a good heritability. Trichomes were studied microscopically in leaf parts in all varieties for analyzing their distribution pattern. The trichome number variations showed significant correlation throughout the genotypes with essential oil yield and monoterpenoid constituents. Differential changes were analyzed for Glutathione S-transferases, Glutathione reductase, Malondialdehyde, phenolics, and chlorophyll content. Gene expressions were analyzed for biosynthesis genes and selected transcription factors TRANSPARENT TESTA GLABRA 1 (TTG1), ENOLASE 1, GLABRA 3, GTL 1, NUCLEAR TRANSCRIPTION FACTOR Y SUBUNIT B-6, WRKY transcription factor 22, putative WRKY 33, WRKY 17, WRKY 1, and WRKY 65-like for harnessing their relation with trichome development in M. arvensis genotypes.

  8. Capitate glandular trichomes of Paragutzlaffia henryi harbor new phytotoxic labdane diterpenoids.

    PubMed

    Wang, Ying; Luo, Shi-Hong; Hua, Juan; Liu, Yan; Jing, Shu-Xi; Li, Xiao-Nian; Li, Sheng-Hong

    2015-11-18

    The morphology and chemical profile of the capitate glandular trichomes (CGTs) of Paragutzlaffia henryi (Acanthaceae) were investigated. Four new labdane diterpenoids named paraguhenryisins A-D (1-4), together with the known physacoztomatin (5), were localized to the CGTs using laser microdissection coupled with cryogenic (1)H NMR and HPLC analyses and were traced and isolated from the CGT extract of inflorescences. Their structures were determined by spectroscopic methods and single-crystal X-ray diffraction. Bioassays indicated significant inhibitory effect for these diterpenoids on Arabidopsis thaliana seed germination and seedling root elongation. The most potent inhibitor, paraguhenryisin C (3), was interestingly detected in both the rhizosphere soil and water rinsed inflorescences extract of P. henryi but not the roots, with average contents in the rhizosphere soil relevant to its phytotoxic EC50 values. These results suggested that phytotoxic labdane diterpenoids in the CGTs might be released into the environment as a defensive measure for P. henryi against other competitive plants.

  9. The "Balance Intervention" for Promoting Caloric Compensatory Behaviours in Response to Overeating: A Formative Evaluation

    ERIC Educational Resources Information Center

    Wammes, Birgitte; Breedveld, Boudewijn; Kremers, Stef; Brug, Johannes

    2006-01-01

    To help people prevent weight gain, the Netherlands Nutrition Centre initiated the "balance intervention", which promotes moderation of food intake and/or increased physical activity in response to occasions of overeating. The aim of this study was to determine whether intervention materials were appreciated, encouraged information…

  10. A Vygotskian Approach to Promote and Formatively Assess Academic Concept Learning

    ERIC Educational Resources Information Center

    Edens, Kellah; Shields, Carol

    2015-01-01

    Promoting students' conceptual understanding and academic language development is a primary goal of instructors in all subject areas. These goals, however, are sometimes hindered by the way students' learning is assessed. In many college-level courses, knowledge-level tests that assess concrete thinking associated with superficial approaches to…

  11. A Rac/Cdc42 exchange factor complex promotes formation of lateral filopodia and blood vessel lumen morphogenesis

    PubMed Central

    Abraham, Sabu; Scarcia, Margherita; Bagshaw, Richard D.; McMahon, Kathryn; Grant, Gary; Harvey, Tracey; Yeo, Maggie; Esteves, Filomena O.G.; Thygesen, Helene H.; Jones, Pamela F.; Speirs, Valerie; Hanby, Andrew M.; Selby, Peter J.; Lorger, Mihaela; Dear, T. Neil; Pawson, Tony; Marshall, Christopher J.; Mavria, Georgia

    2015-01-01

    During angiogenesis, Rho-GTPases influence endothelial cell migration and cell–cell adhesion; however it is not known whether they control formation of vessel lumens, which are essential for blood flow. Here, using an organotypic system that recapitulates distinct stages of VEGF-dependent angiogenesis, we show that lumen formation requires early cytoskeletal remodelling and lateral cell–cell contacts, mediated through the RAC1 guanine nucleotide exchange factor (GEF) DOCK4 (dedicator of cytokinesis 4). DOCK4 signalling is necessary for lateral filopodial protrusions and tubule remodelling prior to lumen formation, whereas proximal, tip filopodia persist in the absence of DOCK4. VEGF-dependent Rac activation via DOCK4 is necessary for CDC42 activation to signal filopodia formation and depends on the activation of RHOG through the RHOG GEF, SGEF. VEGF promotes interaction of DOCK4 with the CDC42 GEF DOCK9. These studies identify a novel Rho-family GTPase activation cascade for the formation of endothelial cell filopodial protrusions necessary for tubule remodelling, thereby influencing subsequent stages of lumen morphogenesis. PMID:26129894

  12. Ubiquilin/Dsk2 promotes inclusion body formation and vacuole (lysosome)-mediated disposal of mutated huntingtin.

    PubMed

    Chuang, Kun-Han; Liang, Fengshan; Higgins, Ryan; Wang, Yanchang

    2016-07-01

    Ubiquilin proteins contain a ubiquitin-like domain (UBL) and ubiquitin-associated domain(s) that interact with the proteasome and ubiquitinated substrates, respectively. Previous work established the link between ubiquilin mutations and neurodegenerative diseases, but the function of ubiquilin proteins remains elusive. Here we used a misfolded huntingtin exon I containing a 103-polyglutamine expansion (Htt103QP) as a model substrate for the functional study of ubiquilin proteins. We found that yeast ubiquilin mutant (dsk2Δ) is sensitive to Htt103QP overexpression and has a defect in the formation of Htt103QP inclusion bodies. Our evidence further suggests that the UBL domain of Dsk2 is critical for inclusion body formation. Of interest, Dsk2 is dispensable for Htt103QP degradation when Htt103QP is induced for a short time before noticeable inclusion body formation. However, when the inclusion body forms after a long Htt103QP induction, Dsk2 is required for efficient Htt103QP clearance, as well as for autophagy-dependent delivery of Htt103QP into vacuoles (lysosomes). Therefore our data indicate that Dsk2 facilitates vacuole-mediated clearance of misfolded proteins by promoting inclusion body formation. Of importance, the defect of inclusion body formation in dsk2 mutants can be rescued by human ubiquilin 1 or 2, suggesting functional conservation of ubiquilin proteins.

  13. A Rac/Cdc42 exchange factor complex promotes formation of lateral filopodia and blood vessel lumen morphogenesis.

    PubMed

    Abraham, Sabu; Scarcia, Margherita; Bagshaw, Richard D; McMahon, Kathryn; Grant, Gary; Harvey, Tracey; Yeo, Maggie; Esteves, Filomena O G; Thygesen, Helene H; Jones, Pamela F; Speirs, Valerie; Hanby, Andrew M; Selby, Peter J; Lorger, Mihaela; Dear, T Neil; Pawson, Tony; Marshall, Christopher J; Mavria, Georgia

    2015-07-01

    During angiogenesis, Rho-GTPases influence endothelial cell migration and cell-cell adhesion; however it is not known whether they control formation of vessel lumens, which are essential for blood flow. Here, using an organotypic system that recapitulates distinct stages of VEGF-dependent angiogenesis, we show that lumen formation requires early cytoskeletal remodelling and lateral cell-cell contacts, mediated through the RAC1 guanine nucleotide exchange factor (GEF) DOCK4 (dedicator of cytokinesis 4). DOCK4 signalling is necessary for lateral filopodial protrusions and tubule remodelling prior to lumen formation, whereas proximal, tip filopodia persist in the absence of DOCK4. VEGF-dependent Rac activation via DOCK4 is necessary for CDC42 activation to signal filopodia formation and depends on the activation of RHOG through the RHOG GEF, SGEF. VEGF promotes interaction of DOCK4 with the CDC42 GEF DOCK9. These studies identify a novel Rho-family GTPase activation cascade for the formation of endothelial cell filopodial protrusions necessary for tubule remodelling, thereby influencing subsequent stages of lumen morphogenesis.

  14. The Rho-guanine nucleotide exchange factor Trio controls leukocyte transendothelial migration by promoting docking structure formation.

    PubMed

    van Rijssel, Jos; Kroon, Jeffrey; Hoogenboezem, Mark; van Alphen, Floris P J; de Jong, Renske J; Kostadinova, Elena; Geerts, Dirk; Hordijk, Peter L; van Buul, Jaap D

    2012-08-01

    Leukocyte transendothelial migration involves the active participation of the endothelium through the formation of apical membrane protrusions that embrace adherent leukocytes, termed docking structures. Using live-cell imaging, we find that prior to transmigration, endothelial docking structures form around 80% of all neutrophils. Previously we showed that endothelial RhoG and SGEF control leukocyte transmigration. In this study, our data reveal that both full-length Trio and the first DH-PH (TrioD1) domain of Trio, which can activate Rac1 and RhoG, interact with ICAM-1 and are recruited to leukocyte adhesion sites. Moreover, upon clustering of ICAM-1, the Rho-guanine nucleotide exchange factor Trio activates Rac1, prior to activating RhoG, in a filamin-dependent manner. We further show that docking structure formation is initiated by ICAM-1 clustering into ring-like structures, which is followed by apical membrane protrusion. Interestingly, we find that Rac1 is required for ICAM-1 clustering, whereas RhoG controls membrane protrusion formation. Finally, silencing endothelial Trio expression or reducing TrioD1 activity without affecting SGEF impairs both docking structure formation and leukocyte transmigration. We conclude that Trio promotes leukocyte transendothelial migration by inducing endothelial docking structure formation in a filamin-dependent manner through the activation of Rac1 and RhoG.

  15. Acylsugar Acylhydrolases: Carboxylesterase-Catalyzed Hydrolysis of Acylsugars in Tomato Trichomes1[OPEN

    PubMed Central

    Gilgallon, Karin; Ghosh, Banibrata

    2016-01-01

    Glandular trichomes of cultivated tomato (Solanum lycopersicum) and many other species throughout the Solanaceae produce and secrete mixtures of sugar esters (acylsugars) on the plant aerial surfaces. In wild and cultivated tomato, these metabolites consist of a sugar backbone, typically glucose or sucrose, and two to five acyl chains esterified to various positions on the sugar core. The aliphatic acyl chains vary in length and branching and are transferred to the sugar by a series of reactions catalyzed by acylsugar acyltransferases. A phenotypic screen of a set of S. lycopersicum M82 × Solanum pennellii LA0716 introgression lines identified a dominant genetic locus on chromosome 5 from the wild relative that affected total acylsugar levels. Genetic mapping revealed that the reduction in acylsugar levels was consistent with the presence and increased expression of two S. pennellii genes (Sopen05g030120 and Sopen05g030130) encoding putative carboxylesterase enzymes of the α/β-hydrolase superfamily. These two enzymes, named ACYLSUGAR ACYLHYDROLASE1 (ASH1) and ASH2, were shown to remove acyl chains from specific positions of certain types of acylsugars in vitro. A survey of related genes in M82 and LA0716 identified another trichome-expressed ASH gene on chromosome 9 (M82, Solyc09g075710; LA0716, Sopen09g030520) encoding a protein with similar activity. Characterization of the in vitro activities of the SpASH enzymes showed reduced activities with acylsugars produced by LA0716, presumably contributing to the high-level production of acylsugars in the presence of highly expressed SpASH genes. PMID:26811191

  16. The yeast telomere-binding protein RAP1 binds to and promotes the formation of DNA quadruplexes in telomeric DNA.

    PubMed Central

    Giraldo, R; Rhodes, D

    1994-01-01

    The protein RAP1 is essential for the maintenance of the telomeres of Saccharomyces cerevisiae and binds in vitro to multiple sites found within the TG1-3 telomeric repeats. We show here that, in addition to its known binding activity for double-stranded DNA, RAP1 binds sequence-specifically to the GT-strands. This indicates that RAP1 is the protein that binds to the telomeric terminal GT-tails. Furthermore, we have found that RAP1 binds to and promotes the formation of G-tetrads, i.e. DNA quadruplexes, in GT-strand oligonucleotides at nanomolar concentrations. The formation of DNA quadruplexes appears to involve the intermolecular association of GT-strands. The minimal DNA-binding domain of RAP1 (DBD) binds only to double-stranded DNA, so that the novel DNA-binding activity we have found involves regions of the protein located outside of the DBD. The finding that a telomeric protein promotes the formation of G-tetrads argues for the use of DNA quadruplexes in telomere association. Images PMID:8194531

  17. Rad51 and Rad54 promote noncrossover recombination between centromere repeats on the same chromatid to prevent isochromosome formation

    PubMed Central

    Onaka, Atsushi T.; Toyofuku, Naoko; Inoue, Takahiro; Okita, Akiko K.; Sagawa, Minami; Su, Jie; Shitanda, Takeshi; Matsuyama, Rei; Zafar, Faria; Takahashi, Tatsuro S.; Masukata, Hisao; Nakagawa, Takuro

    2016-01-01

    Centromeres consist of DNA repeats in many eukaryotes. Non-allelic homologous recombination (HR) between them can result in gross chromosomal rearrangements (GCRs). In fission yeast, Rad51 suppresses isochromosome formation that occurs between inverted repeats in the centromere. However, how the HR enzyme prevents homology-mediated GCRs remains unclear. Here, we provide evidence that Rad51 with the aid of the Swi/Snf-type motor protein Rad54 promotes non-crossover recombination between centromere repeats to prevent isochromosome formation. Mutations in Rad51 and Rad54 epistatically increased the rates of isochromosome formation and chromosome loss. In sharp contrast, these mutations decreased gene conversion between inverted repeats in the centromere. Remarkably, analysis of recombinant DNAs revealed that rad51 and rad54 increase the proportion of crossovers. In the absence of Rad51, deletion of the structure-specific endonuclease Mus81 decreased both crossovers and isochromosomes, while the cdc27/pol32-D1 mutation, which impairs break-induced replication, did not. We propose that Rad51 and Rad54 promote non-crossover recombination between centromere repeats on the same chromatid, thereby suppressing crossover between non-allelic repeats on sister chromatids that leads to chromosomal rearrangements. Furthermore, we found that Rad51 and Rad54 are required for gene silencing in centromeres, suggesting that HR also plays a role in the structure and function of centromeres. PMID:27697832

  18. Oxygen promotes biofilm formation of Shewanella putrefaciens CN32 through a diguanylate cyclase and an adhesin

    PubMed Central

    Wu, Chao; Cheng, Yuan-Yuan; Yin, Hao; Song, Xiang-Ning; Li, Wen-Wei; Zhou, Xian-Xuan; Zhao, Li-Ping; Tian, Li-Jiao; Han, Jun-Cheng; Yu, Han-Qing

    2013-01-01

    Although oxygen has been reported to regulate biofilm formation by several Shewanella species, the exact regulatory mechanism mostly remains unclear. Here, we identify a direct oxygen-sensing diguanylate cyclase (DosD) and reveal its regulatory role in biofilm formation by Shewanella putrefaciens CN32 under aerobic conditions. In vitro and in vivo analyses revealed that the activity of DosD culminates to synthesis of cyclic diguanylate (c-di-GMP) in the presence of oxygen. DosD regulates the transcription of bpfA operon which encodes seven proteins including a large repetitive adhesin BpfA and its cognate type I secretion system (TISS). Regulation of DosD in aerobic biofilms is heavily dependent on an adhesin BpfA and the TISS. This study offers an insight into the molecular mechanism of oxygen-stimulated biofilm formation by S. putrefaciens CN32. PMID:23736081

  19. Sam37 is crucial for formation of the mitochondrial TOM–SAM supercomplex, thereby promoting β-barrel biogenesis

    PubMed Central

    Wenz, Lena-Sophie; Ellenrieder, Lars; Qiu, Jian; Bohnert, Maria; Zufall, Nicole; van der Laan, Martin; Becker, Thomas

    2015-01-01

    Biogenesis of mitochondrial β-barrel proteins requires two preprotein translocases, the general translocase of the outer membrane (TOM) and the sorting and assembly machinery (SAM). TOM and SAM form a supercomplex that promotes transfer of β-barrel precursors. The SAM core complex contains the channel protein Sam50, which cooperates with Sam35 in precursor recognition, and the peripheral membrane protein Sam37. The molecular function of Sam37 has been unknown. We report that Sam37 is crucial for formation of the TOM–SAM supercomplex. Sam37 interacts with the receptor domain of Tom22 on the cytosolic side of the mitochondrial outer membrane and links TOM and SAM complexes. Sam37 thus promotes efficient transfer of β-barrel precursors to the SAM complex. We conclude that Sam37 functions as a coupling factor of the translocase supercomplex of the mitochondrial outer membrane. PMID:26416958

  20. The C-Terminal of Nucleolin Promotes the Formation of the c-MYC G-Quadruplex and Inhibits c-MYC Promoter Activity†

    PubMed Central

    González, Verónica; Hurley, Laurence H.

    2010-01-01

    Nucleolin, the most abundant nucleolar phosphoprotein of eukaryotic cells, is known primarily for its role in ribosome biogenesis and cell proliferation. It is, however, a multifunctional protein that, depending on the cellular context, can drive either cell proliferation or apoptosis. Our laboratory recently demonstrated that nucleolin can function as a repressor of c-MYC transcription by binding to and stabilizing the formation of a G-quadruplex structure in a region of the c-MYC promoter responsible for controlling 85–90% of c-MYC’s transcriptional activity. In this study, we investigate the structural elements of nucleolin that are required for c-MYC repression. The effect of nucleolin deletion mutants on the formation and stability of the c-MYC G-quadruplex, as well as c-MYC transcriptional activity, was assessed by circular dichroic spectropolarimetry (CD),1 thermal stability, and in vitro transcription. Here we report that nucleolin’s RNA binding domains (RBDs) 3 and 4, as well as the arginine-glycine-glycine (RGG) domain, are required to repress c-MYC transcription. PMID:20932061

  1. Internalized Tau sensitizes cells to stress by promoting formation and stability of stress granules

    PubMed Central

    Brunello, Cecilia A.; Yan, Xu; Huttunen, Henri J.

    2016-01-01

    Stress granules are membrane-less RNA- and RNA-binding protein-containing complexes that are transiently assembled in stressful conditions to promote cell survival. Several stress granule-associated RNA-binding proteins have been associated with neurodegenerative diseases. In addition, a close link was recently identified between the stress granule core-nucleating protein TIA-1 and Tau. Tau is a central pathological protein in Alzheimer’s disease and other tauopathies, and misfolded, aggregated Tau is capable of propagating pathology via cell-to-cell transmission. Here we show that following internalization hyperphosphorylated extracellular Tau associates with stress granules in a TIA-1 dependent manner. Cytosolic Tau normally only weakly interacts with TIA-1 but mutations mimicking abnormal phosphorylation promote this interaction. We show that internalized Tau significantly delays normal clearance of stress granules in the recipient cells sensitizing them to secondary stress. These results suggest that secreted Tau species may have properties, likely related to its hyperphosphorylation and oligomerization, which promote pathological association of internalized Tau with stress granules altering their dynamics and reducing cell viability. We suggest that stress granules and TIA-1 play a central role in the cell-to-cell transmission of Tau pathology. PMID:27460788

  2. Induction of mutation and differentiation in mammalian cells by chemicals which initiate or promote tumor formation

    SciTech Connect

    Huberman, E.

    1980-01-01

    A cell-mediated mutagenesis assay was developed to predict the potential carcinogenic hazard of some environmental chemicals. In this assay, cells with appropriate markers for mutagenesis, such as the Chinese hamster V79 cells, are co-cultivated with cells capable of metabolizing chemical carcinogens. Use of this assay made it possible to demonstrate a relationship between the degree of carcinogenicity and mutagenicity of a series of polycyclic hydrocarbons, nitrosamines and aflatoxins, and to establish means to study organ specificity of some chemical carcinogens. However, most short term in vitro assays are designed to detect mutagenic activity and therefore do not detect tumor promoting agents which are devoid of this activity. By analyzing various markers of terminal differentiation in cultured human melanoma and myeloid leukemia cells, we have established a relationship between the activity of a series of tumor promoting phorbol diesters in the mouse skin and their ability to induce terminal differentiation. We suggest that measuring alterations in the differentiation characteristics of some cultured cells may represent an approach by which environmental tumor promoting agents can be studied and detected.

  3. Control of mammalian cell mutagenesis and differentiation by chemicals which initiate or promote tumor formation

    SciTech Connect

    Jones, C. A.; Huberman, E.

    1980-01-01

    A cell-mediated mutagenesis assay was developed to predict the potential carcinogenic hazard of some environmental chemicals. In this assay, Chinese hamster V79 cells, which are susceptible to mutagenesis, are co-cultivated with cells capable of metabolizing chemical carcinogens. Use of this assay made it possible to demonstrate a relationship between the degree of carcinogenicity and mutagenicity of a series of polycyclic hydrocarbons and nitrosamines and to study the organ specificity exhibited by some chemical carcinogens. However, most short-term in vitro assays are designed to detect mutagenic activity and therefore do not detect tumor promoting agents which are devoid of this activity. By analyzing various markers of terminal differentiation in cultured human melanoma and myeloid leukemia cells, we have established a relationship between the activity of a series of tumor promoting phorbol diesters in the mouse skin and their ability to induce terminal differentiation. We suggest that measuring alterations in the differentiation characteristics of some cultured cells may represent an approach by which environmental tumor promoting agents can be studied and detected.

  4. An Undergraduate Study of Two Transcription Factors that Promote Lateral Root Formation

    ERIC Educational Resources Information Center

    Bargmann, Bastiaan O. R.; Birnbaum, Kenneth D.; Brenner, Eric D.

    2014-01-01

    We present a lab that enables students to test the role of genes involved in the regulation of lateral roots growth in the model plant "Arabidopsis thaliana." Here, students design an experiment that follows the effects of the hormone auxin on the stimulation of genes involved in the formation of lateral root initials. These genes, known…

  5. Using Common Formative Assessments to Promote Student Achievement: A Case Study of Practice, Leadership, and Culture

    ERIC Educational Resources Information Center

    Wall, Patricia T. C.

    2012-01-01

    It is the moral responsibility of educators to work diligently to provide every student with rich, challenging coursework in efforts to prepare them for post high school careers and education. The use of common formative assessments provides teachers with the valuable, timely information they need to make instructional decisions that will better…

  6. Embedded Formative Assessment and Classroom Process Quality: How Do They Interact in Promoting Science Understanding?

    ERIC Educational Resources Information Center

    Decristan, Jasmin; Klieme, Eckhard; Kunter, Mareike; Hochweber, Jan; Büttner, Gerhard; Fauth, Benjamin; Hondrich, A. Lena; Rieser, Svenja; Hertel, Silke; Hardy, Ilonca

    2015-01-01

    In this study we examine the interplay between curriculum-embedded formative assessment--a well-known teaching practice--and general features of classroom process quality (i.e., cognitive activation, supportive climate, classroom management) and their combined effect on elementary school students' understanding of the scientific concepts of…

  7. Acetoacetate promotes the formation of fluorescent advanced glycation end products (AGEs).

    PubMed

    Bohlooli, Mousa; Ghaffari-Moghaddam, Mansour; Khajeh, Mostafa; Aghashiri, Zohre; Sheibani, Nader; Moosavi-Movahedi, Ali Akbar

    2016-12-01

    Acetoacetate (AA) is an important ketone body, which produces reactive oxygen species (ROS). Advanced glycation end products (AGEs) are defined as final products of glycation process whose production is influenced by the levels of ROS. The accumulation of AGEs in the body contributes to pathogenesis of many diseases including complications of diabetes, and Alzheimer's and Parkinson's disease. Here, we evaluated the impact of AA on production of AGEs upon incubation of human serum albumin (HSA) with glucose. The effect of AA on the AGEs formation of HSA was studied under physiological conditions after incubation with glucose for 35 days. The physical techniques including circular dichroism (CD) and fluorescence spectroscopy were used to assess the impact of AA on formation and structural changes of glycated HSA (GHSA). Our results indicated that the secondary and tertiary structural changes of GHSA were increased in the presence of AA. The fluorescence intensity measurements of AGEs also showed an increase in AGEs formation. Acetoacetate has an activator effect in formation of AGEs through ROS production. The presence of AA may result in enhanced glycation in the presence of glucose and severity of complications associated with accumulation of AGEs.

  8. A Needs Assessment, Development, and Formative Evaluation of a Health Promotion Smartphone Application for College Students

    ERIC Educational Resources Information Center

    Miller, Tiffany; Chandler, Laura; Mouttapa, Michele

    2015-01-01

    Background: Approximately half of college students who completed the National College Health Assessment 2013 indicated a greater need for health-related information. University-based smartphone applications may help students better access this information. Purpose: This study describes the needs assessment, development, and formative evaluation of…

  9. Promoting Prospective Elementary Teachers' Learning to Use Formative Assessment for Life Science Instruction

    NASA Astrophysics Data System (ADS)

    Sabel, Jaime L.; Forbes, Cory T.; Zangori, Laura

    2015-06-01

    To support elementary students' learning of core, standards-based life science concepts highlighted in the Next Generation Science Standards, prospective elementary teachers should develop an understanding of life science concepts and learn to apply their content knowledge in instructional practice to craft elementary science learning environments grounded in students' thinking. To do so, teachers must learn to use high-leverage instructional practices, such as formative assessment, to engage students in scientific practices and connect instruction to students' ideas. However, teachers may not understand formative assessment or possess sufficient science content knowledge to effectively engage in related instructional practices. To address these needs, we developed and conducted research within an innovative course for preservice elementary teachers built upon two pillars—life science concepts and formative assessment. An embedded mixed methods study was used to evaluate the effect of the intervention on preservice teachers' (n = 49) content knowledge and ability to engage in formative assessment practices for science. Findings showed that increased life content knowledge over the semester helped preservice teachers engage more productively in anticipating and evaluating students' ideas, but not in identifying effective instructional strategies to respond to those ideas.

  10. Identification of Genes That Promote or Inhibit Olfactory Memory Formation in Drosophila

    PubMed Central

    Walkinshaw, Erica; Gai, Yunchao; Farkas, Caitlin; Richter, Daniel; Nicholas, Eric; Keleman, Krystyna; Davis, Ronald L.

    2015-01-01

    Genetic screens in Drosophila melanogaster and other organisms have been pursued to filter the genome for genetic functions important for memory formation. Such screens have employed primarily chemical or transposon-mediated mutagenesis and have identified numerous mutants including classical memory mutants, dunce and rutabaga. Here, we report the results of a large screen using panneuronal RNAi expression to identify additional genes critical for memory formation. We identified >500 genes that compromise memory when inhibited (low hits), either by disrupting the development and normal function of the adult animal or by participating in the neurophysiological mechanisms underlying memory formation. We also identified >40 genes that enhance memory when inhibited (high hits). The dunce gene was identified as one of the low hits and further experiments were performed to map the effects of the dunce RNAi to the α/β and γ mushroom body neurons. Additional behavioral experiments suggest that dunce knockdown in the mushroom body neurons impairs memory without significantly affecting acquisition. We also characterized one high hit, sickie, to show that RNAi knockdown of this gene enhances memory through effects in dopaminergic neurons without apparent effects on acquisition. These studies further our understanding of two genes involved in memory formation, provide a valuable list of genes that impair memory that may be important for understanding the neurophysiology of memory or neurodevelopmental disorders, and offer a new resource of memory suppressor genes that will aid in understanding restraint mechanisms employed by the brain to optimize resources. PMID:25644700

  11. The CUC1 and CUC2 genes promote carpel margin meristem formation during Arabidopsis gynoecium development

    PubMed Central

    Kamiuchi, Yuri; Yamamoto, Kayo; Furutani, Masahiko; Tasaka, Masao; Aida, Mitsuhiro

    2014-01-01

    Carpel margin meristems (CMMs), a pair of meristematic tissues present along the margins of two fused carpel primordia of Arabidopsis thaliana, are essential for the formation of ovules and the septum, two major internal structures of the gynoecium. Although a number of regulatory factors involved in shoot meristem activity are known to be required for the formation of these gynoecial structures, their direct roles in CMM development have yet to be addressed. Here we show that the CUP-SHAPED COTYLEDON genes CUC1 and CUC2, which are essential for shoot meristem initiation, are also required for formation and stable positioning of the CMMs. Early in CMM formation, CUC1 and CUC2 are also required for expression of the SHOOT MERISTEMLESS gene, a central regulator for stem cell maintenance in the shoot meristem. Moreover, plants carrying miR164-resistant forms of CUC1 and CUC2 resulted in extra CMM activity with altered positioning. Our results thus demonstrate that the two regulatory proteins controlling shoot meristem activity also play critical roles in elaboration of the female reproductive organ through the control of meristematic activity. PMID:24817871

  12. Down-regulation of MUC1 in cancer cells inhibits cell migration by promoting E-cadherin/catenin complex formation

    SciTech Connect

    Yuan Zhenglong; Wong, Sandy; Borrelli, Alexander; Chung, Maureen A.

    2007-10-26

    MUC1, a tumor associated glycoprotein, is over-expressed in most cancers and can promote proliferation and metastasis. The objective of this research was to study the role of MUC1 in cancer metastasis and its potential mechanism. Pancreatic (PANC1) and breast (MCF-7) cancer cells with stable 'knockdown' of MUC1 expression were created using RNA interference. {beta}-Catenin and E-cadherin protein expression were upregulated in PANC1 and MCF-7 cells with decreased MUC1 expression. Downregulation of MUC1 expression also induced {beta}-catenin relocation from the nucleus to the cytoplasm, increased E-cadherin/{beta}-catenin complex formation and E-cadherin membrane localization in PANC1 cells. PANC1 cells with 'knockdown' MUC1 expression had decreased in vitro cell invasion. This study suggested that MUC1 may affect cancer cell migration by increasing E-cadherin/{beta}-catenin complex formation and restoring E-cadherin membrane localization.

  13. Down-regulation of MUC1 in cancer cells inhibits cell migration by promoting E-cadherin/catenin complex formation.

    PubMed

    Yuan, Zhenglong; Wong, Sandy; Borrelli, Alexander; Chung, Maureen A

    2007-10-26

    MUC1, a tumor associated glycoprotein, is over-expressed in most cancers and can promote proliferation and metastasis. The objective of this research was to study the role of MUC1 in cancer metastasis and its potential mechanism. Pancreatic (PANC1) and breast (MCF-7) cancer cells with stable 'knockdown' of MUC1 expression were created using RNA interference. beta-Catenin and E-cadherin protein expression were upregulated in PANC1 and MCF-7 cells with decreased MUC1 expression. Downregulation of MUC1 expression also induced beta-catenin relocation from the nucleus to the cytoplasm, increased E-cadherin/beta-catenin complex formation and E-cadherin membrane localization in PANC1 cells. PANC1 cells with 'knockdown' MUC1 expression had decreased in vitro cell invasion. This study suggested that MUC1 may affect cancer cell migration by increasing E-cadherin/beta-catenin complex formation and restoring E-cadherin membrane localization.

  14. Preformed template fluctuations promote fibril formation: Insights from lattice and all-atom models

    SciTech Connect

    Kouza, Maksim Kolinski, Andrzej; Co, Nguyen Truong; Nguyen, Phuong H.; Li, Mai Suan

    2015-04-14

    Fibril formation resulting from protein misfolding and aggregation is a hallmark of several neurodegenerative diseases such as Alzheimer’s and Parkinson’s diseases. Despite the fact that the fibril formation process is very slow and thus poses a significant challenge for theoretical and experimental studies, a number of alternative pictures of molecular mechanisms of amyloid fibril formation have been recently proposed. What seems to be common for the majority of the proposed models is that fibril elongation involves the formation of pre-nucleus seeds prior to the creation of a critical nucleus. Once the size of the pre-nucleus seed reaches the critical nucleus size, its thermal fluctuations are expected to be small and the resulting nucleus provides a template for sequential (one-by-one) accommodation of added monomers. The effect of template fluctuations on fibril formation rates has not been explored either experimentally or theoretically so far. In this paper, we make the first attempt at solving this problem by two sets of simulations. To mimic small template fluctuations, in one set, monomers of the preformed template are kept fixed, while in the other set they are allowed to fluctuate. The kinetics of addition of a new peptide onto the template is explored using all-atom simulations with explicit water and the GROMOS96 43a1 force field and simple lattice models. Our result demonstrates that preformed template fluctuations can modulate protein aggregation rates and pathways. The association of a nascent monomer with the template obeys the kinetics partitioning mechanism where the intermediate state occurs in a fraction of routes to the protofibril. It was shown that template immobility greatly increases the time of incorporating a new peptide into the preformed template compared to the fluctuating template case. This observation has also been confirmed by simulation using lattice models and may be invoked to understand the role of template fluctuations in

  15. Preformed template fluctuations promote fibril formation: insights from lattice and all-atom models.

    PubMed

    Kouza, Maksim; Co, Nguyen Truong; Nguyen, Phuong H; Kolinski, Andrzej; Li, Mai Suan

    2015-04-14

    Fibril formation resulting from protein misfolding and aggregation is a hallmark of several neurodegenerative diseases such as Alzheimer's and Parkinson's diseases. Despite the fact that the fibril formation process is very slow and thus poses a significant challenge for theoretical and experimental studies, a number of alternative pictures of molecular mechanisms of amyloid fibril formation have been recently proposed. What seems to be common for the majority of the proposed models is that fibril elongation involves the formation of pre-nucleus seeds prior to the creation of a critical nucleus. Once the size of the pre-nucleus seed reaches the critical nucleus size, its thermal fluctuations are expected to be small and the resulting nucleus provides a template for sequential (one-by-one) accommodation of added monomers. The effect of template fluctuations on fibril formation rates has not been explored either experimentally or theoretically so far. In this paper, we make the first attempt at solving this problem by two sets of simulations. To mimic small template fluctuations, in one set, monomers of the preformed template are kept fixed, while in the other set they are allowed to fluctuate. The kinetics of addition of a new peptide onto the template is explored using all-atom simulations with explicit water and the GROMOS96 43a1 force field and simple lattice models. Our result demonstrates that preformed template fluctuations can modulate protein aggregation rates and pathways. The association of a nascent monomer with the template obeys the kinetics partitioning mechanism where the intermediate state occurs in a fraction of routes to the protofibril. It was shown that template immobility greatly increases the time of incorporating a new peptide into the preformed template compared to the fluctuating template case. This observation has also been confirmed by simulation using lattice models and may be invoked to understand the role of template fluctuations in

  16. Biofuel-Promoted Polychlorinated Dibenzodioxin/furan Formation in an Iron-Catalyzed Diesel Particle Filter.

    PubMed

    Heeb, Norbert V; Rey, Maria Dolores; Zennegg, Markus; Haag, Regula; Wichser, Adrian; Schmid, Peter; Seiler, Cornelia; Honegger, Peter; Zeyer, Kerstin; Mohn, Joachim; Bürki, Samuel; Zimmerli, Yan; Czerwinski, Jan; Mayer, Andreas

    2015-08-04

    Iron-catalyzed diesel particle filters (DPFs) are widely used for particle abatement. Active catalyst particles, so-called fuel-borne catalysts (FBCs), are formed in situ, in the engine, when combusting precursors, which were premixed with the fuel. The obtained iron oxide particles catalyze soot oxidation in filters. Iron-catalyzed DPFs are considered as safe with respect to their potential to form polychlorinated dibenzodioxins/furans (PCDD/Fs). We reported that a bimetallic potassium/iron FBC supported an intense PCDD/F formation in a DPF. Here, we discuss the impact of fatty acid methyl ester (FAME) biofuel on PCDD/F emissions. The iron-catalyzed DPF indeed supported a PCDD/F formation with biofuel but remained inactive with petroleum-derived diesel fuel. PCDD/F emissions (I-TEQ) increased 23-fold when comparing biofuel and diesel data. Emissions of 2,3,7,8-TCDD, the most toxic congener [toxicity equivalence factor (TEF) = 1.0], increased 90-fold, and those of 2,3,7,8-TCDF (TEF = 0.1) increased 170-fold. Congener patterns also changed, indicating a preferential formation of tetra- and penta-chlorodibenzofurans. Thus, an inactive iron-catalyzed DPF becomes active, supporting a PCDD/F formation, when operated with biofuel containing impurities of potassium. Alkali metals are inherent constituents of biofuels. According to the current European Union (EU) legislation, levels of 5 μg/g are accepted. We conclude that risks for a secondary PCDD/F formation in iron-catalyzed DPFs increase when combusting potassium-containing biofuels.

  17. HGF expression induced by HIF-1α promote the proliferation and tube formation of endothelial progenitor cells.

    PubMed

    Yu, Feng; Lin, Yuhua; Zhan, Tao; Chen, Lili; Guo, Sufen

    2015-03-01

    Hypoxia-inducible factor-1α (HIF-1α) and hepatocyte growth factor (HGF) play important roles in postnatal neovascularization. However, the interaction of these two pathways is not fully understood. The present study utilized CoCl(2) treated-endothelial progenitor cells (EPCs) (EPCs exposure to CoCl(2) are under mimic hypoxia) to examine the expressions of HIF-1α and HGF and futher to assess whether or not the inhibitor (2-methoxyestradiol [2ME2]) of HIF-1α decrease the HGF expression. In addition, to investigate the effects of HGF on the proliferation and tube formation of EPCs under mimic hypoxia, EPCs were transfected with NK4 (HGF antagonist) plasmid and exposed to CoCl(2), then the proliferation of these EPCs was assayed by MTS and the tube formation capacity of these EPCs on Matrigel was detected. The analysis indicated that CoCl(2) treatment induced HIF-1α expression of EPCs, and futher promoted HGF expression. While after 2ME2 was used in CoCl(2) treated-EPCs, HGF expression was markedly inhibited compared with non-pretreated EPCs with 2ME2, which also showed that HGF expression in EPCs was mediated by HIF-1α. Further, the results showed that after EPCs were transfected with NK4 in spite of being exposed to CoCl(2), their proliferation activity and tube formation capacity were weakened, which in turn indicated that HGF could promote the proliferation and the tube formation of EPCs, and this process might be regulated by HIF-1α.

  18. Crmp4 deletion promotes recovery from spinal cord injury by neuroprotection and limited scar formation

    PubMed Central

    Nagai, Jun; Kitamura, Yoshiteru; Owada, Kazuki; Yamashita, Naoya; Takei, Kohtaro; Goshima, Yoshio; Ohshima, Toshio

    2015-01-01

    Axonal outgrowth inhibitors and scar formation are two major obstacles to central nervous system (CNS) repair. No target molecule that regulates both axonal growth and scarring has been identified. Here we identified collapsin response mediator protein 4 (CRMP4), a common mediator of inhibitory signals after neural injury, as a crucial factor that contributes to both axonal growth inhibition and scarring after spinal cord injury (SCI). We found increases in the inhibitory and toxic forms of CRMP4 in injured spinal cord. Notably, CRMP4 expression was evident in inflammatory cells as well as in neurons after spinal cord transection. Crmp4−/− mice displayed neuroprotection against SCI and reductions in inflammatory response and scar formation. This permissive environment for axonal growth due to CRMP4 deletion restored locomotor activity at an unusually early phase of healing. These results suggest that deletion of CRMP4 is a unique therapeutic strategy that overcomes two obstacles to CNS repair after SCI. PMID:25652774

  19. Reactive oxidation products promote secondary organic aerosol formation from green leaf volatiles

    NASA Astrophysics Data System (ADS)

    Hamilton, J. F.; Lewis, A. C.; Carey, T. J.; Wenger, J. C.; Garcia, E. Borrás. I.; Muñoz, A.

    2009-02-01

    Green leaf volatiles (GLVs) are an important group of chemicals released by vegetation which have emission fluxes that can be significantly increased when plants are damaged or stressed. A series of simulation chamber experiments has been conducted at the European Photoreactor in Valencia, Spain, to investigate secondary organic aerosol (SOA) formation from the atmospheric oxidation of the major GLVs cis-3-hexenylacetate and cis-3-hexen-1-ol. Liquid chromatography-ion trap mass spectrometry was used to identify chemical species present in the SOA. Cis-3-hexen-1-ol proved to be a more efficient SOA precursor due to the high reactivity of its first generation oxidation product, 3-hydroxypropanal, which can hydrate and undergo further reactions with other aldehydes resulting in SOA dominated by higher molecular weight oligomers. The lower SOA yields produced from cis-3-hexenylacetate are attributed to the acetate functionality, which inhibits oligomer formation in the particle phase. Based on observed SOA yields and best estimates of global emissions, these compounds may be calculated to be a substantial unidentified global source of SOA, contributing 1-5 TgC yr-1, equivalent to around a third of that predicted from isoprene. Molecular characterization of the SOA, combined with organic mechanistic information, has provided evidence that the formation of organic aerosols from GLVs is closely related to the reactivity of their first generation atmospheric oxidation products, and indicates that this may be a simple parameter that could be used in assessing the aerosol formation potential for other unstudied organic compounds in the atmosphere.

  20. Reactive oxidation products promote secondary organic aerosol formation from green leaf volatiles

    NASA Astrophysics Data System (ADS)

    Hamilton, J. F.; Lewis, A. C.; Carey, T. J.; Wenger, J. C.; Garcia, E. Borrás. I.; Muñoz, A.

    2009-06-01

    Green leaf volatiles (GLVs) are an important group of chemicals released by vegetation which have emission fluxes that can be significantly increased when plants are damaged or stressed. A series of simulation chamber experiments has been conducted at the European Photoreactor in Valencia, Spain, to investigate secondary organic aerosol (SOA) formation from the atmospheric oxidation of the major GLVs cis-3-hexenylacetate and cis-3-hexen-1-ol. Liquid chromatography-ion trap mass spectrometry was used to identify chemical species present in the SOA. Cis-3-hexen-1-ol proved to be a more efficient SOA precursor due to the high reactivity of its first generation oxidation product, 3-hydroxypropanal, which can hydrate and undergo further reactions with other aldehydes resulting in SOA dominated by higher molecular weight oligomers. The lower SOA yields produced from cis-3-hexenylacetate are attributed to the acetate functionality, which inhibits oligomer formation in the particle phase. Based on observed SOA yields and best estimates of global emissions, these compounds may be calculated to be a substantial unidentified global source of SOA, contributing 1-5 TgC yr-1, equivalent to around a third of that predicted from isoprene. Molecular characterization of the SOA, combined with organic mechanistic information, has provided evidence that the formation of organic aerosols from GLVs is closely related to the reactivity of their first generation atmospheric oxidation products, and indicates that this may be a simple parameter that could be used in assessing the aerosol formation potential for other unstudied organic compounds in the atmosphere.

  1. Neuraminidase A-Exposed Galactose Promotes Streptococcus pneumoniae Biofilm Formation during Colonization

    PubMed Central

    Blanchette, Krystle A.; Shenoy, Anukul T.; Milner, Jeffrey; Gilley, Ryan P.; McClure, Erin; Hinojosa, Cecilia A.; Kumar, Nikhil; Daugherty, Sean C.; Tallon, Luke J.; Ott, Sandra; King, Samantha J.; Ferreira, Daniela M.; Gordon, Stephen B.; Tettelin, Hervé

    2016-01-01

    Streptococcus pneumoniae is an opportunistic pathogen that colonizes the nasopharynx. Herein we show that carbon availability is distinct between the nasopharynx and bloodstream of adult humans: glucose is absent from the nasopharynx, whereas galactose is abundant. We demonstrate that pneumococcal neuraminidase A (NanA), which cleaves terminal sialic acid residues from host glycoproteins, exposed galactose on the surface of septal epithelial cells, thereby increasing its availability during colonization. We observed that S. pneumoniae mutants deficient in NanA and β-galactosidase A (BgaA) failed to form biofilms in vivo despite normal biofilm-forming abilities in vitro. Subsequently, we observed that glucose, sucrose, and fructose were inhibitory for biofilm formation, whereas galactose, lactose, and low concentrations of sialic acid were permissive. Together these findings suggested that the genes involved in biofilm formation were under some form of carbon catabolite repression (CCR), a regulatory network in which genes involved in the uptake and metabolism of less-preferred sugars are silenced during growth with preferred sugars. Supporting this notion, we observed that a mutant deficient in pyruvate oxidase, which converts pyruvate to acetyl-phosphate under non-CCR-inducing growth conditions, was unable to form biofilms. Subsequent comparative transcriptome sequencing (RNA-seq) analyses of planktonic and biofilm-grown pneumococci showed that metabolic pathways involving the conversion of pyruvate to acetyl-phosphate and subsequently leading to fatty acid biosynthesis were consistently upregulated during diverse biofilm growth conditions. We conclude that carbon availability in the nasopharynx impacts pneumococcal biofilm formation in vivo. Additionally, biofilm formation involves metabolic pathways not previously appreciated to play an important role. PMID:27481242

  2. Bovine binder-of-sperm protein BSP1 promotes protrusion and nanotube formation from liposomes

    SciTech Connect

    Lafleur, Michel; Courtemanche, Lesley; Karlsson, Goeran; Edwards, Katarina; Schwartz, Jean-Louis; Manjunath, Puttaswamy

    2010-08-27

    Research highlights: {yields} Binder-of-sperm protein 1 (BSP1) modifies the morphology of lipidic vesicles inducing bead necklace-like and thread-like structures. {yields} In the presence of multilamellar liposomes, BSP1 leads to the formation of long nanotubes. {yields} The insertion of BSP1 in the external lipid leaflet of membranes induces local changes in bilayer curvature. -- Abstract: Binder-of-sperm (BSP) proteins interact with sperm membranes and are proposed to extract selectively phosphatidylcholine and cholesterol from these. This change in lipid composition is a key step in sperm capacitation. The present work demonstrates that the interactions between the protein BSP1 and model membranes composed with phosphatidylcholine lead to drastic changes in the morphology of the lipidic self-assemblies. Using cryo-electron microscopy and fluorescence microscopy, we show that, in the presence of the protein, the lipid vesicles elongate, and form bead necklace-like structures that evolve toward small vesicles or thread-like structures. In the presence of multilamellar vesicles, where a large reservoir of lipid is available, the presence of BSP proteins lead to the formation of long nanotubes. Long spiral-like threads, associated with lipid/protein complexes, are also observed. The local curvature of lipid membranes induced by the BSP proteins may be involved in lipid domain formation and the extraction of some lipids during the sperm maturation process.

  3. Augmin promotes meiotic spindle formation and bipolarity in Xenopus egg extracts.

    PubMed

    Petry, Sabine; Pugieux, Céline; Nédélec, François J; Vale, Ronald D

    2011-08-30

    Female meiotic spindles in many organisms form in the absence of centrosomes, the organelle typically associated with microtubule (MT) nucleation. Previous studies have proposed that these meiotic spindles arise from RanGTP-mediated MT nucleation in the vicinity of chromatin; however, whether this process is sufficient for spindle formation is unknown. Here, we investigated whether a recently proposed spindle-based MT nucleation pathway that involves augmin, an 8-subunit protein complex, also contributes to spindle morphogenesis. We used an assay system in which hundreds of meiotic spindles can be observed forming around chromatin-coated beads after introduction of Xenopus egg extracts. Spindles forming in augmin-depleted extracts showed reduced rates of MT formation and were predominantly multipolar, revealing a function of augmin in stabilizing the bipolar shape of the acentrosomal meiotic spindle. Our studies also have uncovered an apparent augmin-independent MT nucleation process from acentrosomal poles, which becomes increasingly active over time and appears to partially rescue the spindle defects that arise from augmin depletion. Our studies reveal that spatially and temporally distinct MT generation pathways from chromatin, spindle MTs, and acentrosomal poles all contribute to robust bipolar spindle formation in meiotic extracts.

  4. Translation suppression promotes stress granule formation and cell survival in response to cold shock

    PubMed Central

    Hofmann, Sarah; Cherkasova, Valeria; Bankhead, Peter; Bukau, Bernd; Stoecklin, Georg

    2012-01-01

    Cells respond to different types of stress by inhibition of protein synthesis and subsequent assembly of stress granules (SGs), cytoplasmic aggregates that contain stalled translation preinitiation complexes. Global translation is regulated through the translation initiation factor eukaryotic initiation factor 2α (eIF2α) and the mTOR pathway. Here we identify cold shock as a novel trigger of SG assembly in yeast and mammals. Whereas cold shock–induced SGs take hours to form, they dissolve within minutes when cells are returned to optimal growth temperatures. Cold shock causes eIF2α phosphorylation through the kinase PERK in mammalian cells, yet this pathway is not alone responsible for translation arrest and SG formation. In addition, cold shock leads to reduced mitochondrial function, energy depletion, concomitant activation of AMP-activated protein kinase (AMPK), and inhibition of mTOR signaling. Compound C, a pharmacological inhibitor of AMPK, prevents the formation of SGs and strongly reduces cellular survival in a translation-dependent manner. Our results demonstrate that cells actively suppress protein synthesis by parallel pathways, which induce SG formation and ensure cellular survival during hypothermia. PMID:22875991

  5. Hydroxyapatite bioactivated bacterial cellulose promotes osteoblast growth and the formation of bone nodules

    PubMed Central

    2012-01-01

    The goal of this study was to investigate the feasibility of bacterial cellulose (BC) scaffold to support osteoblast growth and bone formation. BC was produced by culturing Acetobacter xylinum supplemented with hydroxyapatite (HA) to form BC membranes (without HA) and BC/HA membranes. Membranes were subjected to X-ray photoelectron spectroscopy (XPS) analysis to determine surface element composition. The membranes were further used to evaluate osteoblast growth, alkaline phosphatase activity and bone nodule formation. BC was free of calcium and phosphate. However, XPS analysis revealed the presence of both calcium (10%) and phosphate (10%) at the surface of the BC/HA membrane. Osteoblast culture showed that BC alone was non-toxic and could sustain osteoblast adhesion. Furthermore, osteoblast adhesion and growth were significantly (p ≤0.05) increased on BC/HA membranes as compared to BC alone. Both BC and BC/HA membranes improved osteoconductivity, as confirmed by the level of alkaline phosphatase (ALP) activity that increased from 2.5 mM with BC alone to 5.3 mM with BC/HA. BC/HA membranes also showed greater nodule formation and mineralization than the BC membrane alone. This was confirmed by Alizarin red staining (ARS) and energy dispersive X-ray spectroscopy (EDX). This work demonstrates that both BC and BC/HA may be useful in bone tissue engineering. PMID:23174338

  6. Abluminal stimulation of sphingosine 1-phosphate receptors 1 and 3 promotes and stabilizes endothelial sprout formation.

    PubMed

    Das, Anusuya; Lenz, Steven M; Awojoodu, Anthony O; Botchwey, Edward A

    2015-01-01

    Local delivery of lipid mediators has become a promising new approach for therapeutic angiogenesis and regenerative medicine. In this study, we investigated how gradient stimulation (either abluminal/distal or luminal/proximal) of engineered microvessels with sphingosine 1-phosphate (S1P) receptor-subtype-targeted molecules affects endothelial sprout growth using a microfluidic device. Our studies show that distal stimulation of microvessels with FTY720, an S1P1/3 selective agonist, promotes both arterial and venular sprout growth, whereas proximal stimulation does not. Using novel pharmacological antagonists of S1P receptor subtypes, we further show that S1P3 functionality is necessary for VEGF-induced sprouting, and confirmed these findings ex vivo using a murine aortic ring assay from S1P3-deficient mice. S1P3 agonist stimulation enhanced vascular stability in both cell types via upregulation of the interendothelial junction protein VE-cadherin. Lastly, S1P3 activation under flow promoted endothelial sprouting and branching while decreasing migratory cell fate in the microfluidic device. We used an in vivo murine dorsal skinfold window chamber model to confirm S1P3's role in neovascular branching. Together, these data suggest that a distal transendothelial gradient of S1P1/3-targeted drugs is an effective technique for both enhancing and stabilizing capillary morphogenesis in angiogenic applications.

  7. Institute of social justice and medicine: developing a think tank to promote policy formation.

    PubMed

    Boozary, Andrew; Dugani, Sagar B

    2011-10-01

    The World Health Organization (WHO) defines health as a "resource for everyday living, not the objective of living"; however, worldwide, there remains an unmistakable inequity in level of health and access to healthcare. The WHO has published documents on financing health systems towards universal health coverage [1], promoting healthy life [2], improving performance of health systems [3], and enriching humanity [4], highlighting our shared responsibility towards improving both national and global health and access to healthcare. These documents also recognize that, despite our local and regional priorities, there is a global desire to develop international strategies to improve healthcare. [1] WHO Report. Health systems financing and the path to universal coverage. 2010. http://www.who.int/bulletin/health_financing/en/index.html [2] WHO Report. Reducing risks, promoting healthy life. 2002. http://www.who.int/whr/2002/en/index.html [3] WHO Bulletin. Health systems: improving performance. 2000. http://www.who.int/whr/2000/en/index.html [4] WHO Bulletin. Conquering suffering, enriching humanity 1997. http://www.who.int/whr/1997/en/index.html.

  8. Vinca alkaloid drugs promote stress-induced translational repression and stress granule formation

    PubMed Central

    Szaflarski, Witold; Fay, Marta M.; Kedersha, Nancy; Zabel, Maciej; Anderson, Paul; Ivanov, Pavel

    2016-01-01

    Resistance to chemotherapy drugs is a serious therapeutic problem and its underlying molecular mechanisms are complex. Stress granules (SGs), cytoplasmic ribonucleoprotein complexes assembled in cells exposed to stress, are implicated in various aspects of cancer cell metabolism and survival. SGs promote the survival of stressed cells by reprogramming gene expression and inhibiting pro-apoptotic signaling cascades. We show that the vinca alkaloid (VA) class of anti-neoplastic agents potently activates a SG-mediated stress response program. VAs inhibit translation initiation by simultaneous activation of eIF4E-BP1 and phosphorylation of eIF2α, causing polysome disassembly and SG assembly. VA-induced SGs contain canonical SG components but lack specific signaling molecules. Blocking VA-induced SG assembly by inactivating eIF4EBP1 or inhibiting eIF2α phosphorylation decreases cancer cell viability and promotes apoptosis. Our data describe previously unappreciated effects of VAs on cellular RNA metabolism and illuminate the roles of SGs in cancer cell survival. PMID:27083003

  9. Abluminal Stimulation of Sphingosine 1-Phosphate Receptors 1 and 3 Promotes and Stabilizes Endothelial Sprout Formation

    PubMed Central

    Lenz, Steven M.; Awojoodu, Anthony O.

    2015-01-01

    Local delivery of lipid mediators has become a promising new approach for therapeutic angiogenesis and regenerative medicine. In this study, we investigated how gradient stimulation (either abluminal/distal or luminal/proximal) of engineered microvessels with sphingosine 1-phosphate (S1P) receptor-subtype-targeted molecules affects endothelial sprout growth using a microfluidic device. Our studies show that distal stimulation of microvessels with FTY720, an S1P1/3 selective agonist, promotes both arterial and venular sprout growth, whereas proximal stimulation does not. Using novel pharmacological antagonists of S1P receptor subtypes, we further show that S1P3 functionality is necessary for VEGF-induced sprouting, and confirmed these findings ex vivo using a murine aortic ring assay from S1P3-deficient mice. S1P3 agonist stimulation enhanced vascular stability in both cell types via upregulation of the interendothelial junction protein VE-cadherin. Lastly, S1P3 activation under flow promoted endothelial sprouting and branching while decreasing migratory cell fate in the microfluidic device. We used an in vivo murine dorsal skinfold window chamber model to confirm S1P3's role in neovascular branching. Together, these data suggest that a distal transendothelial gradient of S1P1/3-targeted drugs is an effective technique for both enhancing and stabilizing capillary morphogenesis in angiogenic applications. PMID:25315888

  10. Protonation switching to the least-basic heteroatom of carbamate through cationic hydrogen bonding promotes the formation of isocyanate cations.

    PubMed

    Kurouchi, Hiroaki; Sumita, Akinari; Otani, Yuko; Ohwada, Tomohiko

    2014-07-07

    We found that phenethylcarbamates that bear ortho-salicylate as an ether group (carbamoyl salicylates) dramatically accelerate OC bond dissociation in strong acid to facilitate generation of isocyanate cation (N-protonated isocyanates), which undergo subsequent intramolecular aromatic electrophilic cyclization to give dihydroisoquinolones. To generate isocyanate cations from carbamates in acidic media as electrophiles for aromatic substitution, protonation at the ether oxygen, the least basic heteroatom, is essential to promote CO bond cleavage. However, the carbonyl oxygen of carbamates, the most basic site, is protonated exclusively in strong acids. We found that the protonation site can be shifted to an alternative basic atom by linking methyl salicylate to the ether oxygen of carbamate. The methyl ester oxygen ortho to the phenolic (ether) oxygen of salicylate is as basic as the carbamate carbonyl oxygen, and we found that monoprotonation at the methyl ester oxygen in strong acid resulted in the formation of an intramolecular cationic hydrogen bond (>CO(+) H⋅⋅⋅O<) with the phenolic ether oxygen. This facilitates OC bond dissociation of phenethylcarbamates, thereby promoting isocyanate cation formation. In contrast, superacid-mediated diprotonation at the methyl ester oxygen of the salicylate and the carbonyl oxygen of the carbamate afforded a rather stable dication, which did not readily undergo CO bond dissociation. This is an unprecedented and unknown case in which the monocation has greater reactivity than the dication.

  11. LGR5 promotes the proliferation and tumor formation of cervical cancer cells through the Wnt/β-catenin signaling pathway

    PubMed Central

    Chen, Qing; Cao, Hao-Zhe; Zheng, Peng-Sheng

    2014-01-01

    Leucine-rich repeat-containing G protein-coupled receptor 5 (LGR5), a seven transmembrane receptor known as a potential stem cell marker for intestinal crypts and hair follicles, has recently been found to be overexpressed in some types of human cancers. However, the role of LGR5 in cervical cancer remains unclear. In this study, the expression of LGR5 gradually increases from normal cervix to cervical cancer in situ and to cervical cancers as revealed by immunohistochemistry and western blot analyses. Through knocking down or overexpressing LGR5 in SiHa and HeLa cells, the expression level of LGR5 was found to be positively related to cell proliferation in vitro and to tumor formation in vivo. Further investigation indicated that LGR5 protein could significantly promote the acceleration of cell cycle. Moreover, the TOP-Flash reporter assay and western blot for β-catenin, cyclinD1, and c-myc proteins, target genes of the Wnt/β-catenin pathway, indicated that LGR5 significantly activated Wnt/β-catenin signaling. Additionally, the blockage of Wnt/β-catenin pathway resulted in a significant inhibition of cell proliferation induced by LGR5. Taken together, these results demonstrate that LGR5 can promote proliferation and tumor formation in cervical cancer cells by activating the Wnt/β-catenin pathway. PMID:25193857

  12. Appalachian mountaintop mining particulate matter induces neoplastic transformation of human bronchial epithelial cells and promotes tumor formation.

    PubMed

    Luanpitpong, Sudjit; Chen, Michael; Knuckles, Travis; Wen, Sijin; Luo, Juhua; Ellis, Emily; Hendryx, Michael; Rojanasakul, Yon

    2014-11-04

    Epidemiological studies suggest that living near mountaintop coal mining (MTM) activities is one of the contributing factors for high lung cancer incidence. The purpose of this study was to investigate the long-term carcinogenic potential of MTM particulate matter (PMMTM) exposure on human bronchial epithelial cells. Our results show that chronic exposure (3 months) to noncytotoxic, physiological relevant concentration (1 μg/mL) of PMMTM, but not control particle PMCON, induced neoplastic transformation, accelerated cell proliferation, and enhanced cell migration of the exposed lung cells. Xenograft transplantation of the PMMTM-exposed cells in mice caused no apparent tumor formation, but promoted tumor growth of human lung carcinoma H460 cells, suggesting the tumor-promoting effect of PMMTM. Chronic exposure to the main inorganic chemical constituent of PMMTM, molybdenum but not silica, similarly induced cell transformation and tumor promotion, suggesting the contribution of molybdenum, at least in part, in the PMMTM effects. These results provide new evidence for the carcinogenic potential of PMMTM and support further risk assessment and implementation of exposure control for PMMTM.

  13. Cbln family proteins promote synapse formation by regulating distinct neurexin signaling pathways in various brain regions.

    PubMed

    Matsuda, Keiko; Yuzaki, Michisuke

    2011-04-01

    Cbln1 (a.k.a. precerebellin) is a unique bidirectional synaptic organizer that plays an essential role in the formation and maintenance of excitatory synapses between granule cells and Purkinje cells in the mouse cerebellum. Cbln1 secreted from cerebellar granule cells directly induces presynaptic differentiation and indirectly serves as a postsynaptic organizer by binding to its receptor, the δ2 glutamate receptor. However, it remains unclear how Cbln1 binds to the presynaptic sites and interacts with other synaptic organizers. Furthermore, although Cbln1 and its family members Cbln2 and Cbln4 are expressed in brain regions other than the cerebellum, it is unknown whether they regulate synapse formation in these brain regions. In this study, we showed that Cbln1 and Cbln2, but not Cbln4, specifically bound to its presynaptic receptor -α and β isoforms of neurexin carrying the splice site 4 insert [NRXs(S4+)] - and induced synaptogenesis in cerebellar, hippocampal and cortical neurons in vitro. Cbln1 competed with synaptogenesis mediated by neuroligin 1, which lacks the splice sites A and B, but not leucine-rich repeat transmembrane protein 2, possibly by sharing the presynaptic receptor NRXs(S4+). However, unlike neurexins/neuroligins or neurexins/leucine-rich repeat transmembrane proteins, the interaction between NRX1β(S4+) and Cbln1 was insensitive to extracellular Ca(2+) concentrations. These findings revealed the unique and general roles of Cbln family proteins in mediating the formation and maintenance of synapses not only in the cerebellum but also in various other brain regions.

  14. Condensin II Promotes the Formation of Chromosome Territories by Inducing Axial Compaction of Polyploid Interphase Chromosomes

    PubMed Central

    Bauer, Christopher R.; Hartl, Tom A.; Bosco, Giovanni

    2012-01-01

    The eukaryotic nucleus is both spatially and functionally partitioned. This organization contributes to the maintenance, expression, and transmission of genetic information. Though our ability to probe the physical structure of the genome within the nucleus has improved substantially in recent years, relatively little is known about the factors that regulate its organization or the mechanisms through which specific organizational states are achieved. Here, we show that Drosophila melanogaster Condensin II induces axial compaction of interphase chromosomes, globally disrupts interchromosomal interactions, and promotes the dispersal of peri-centric heterochromatin. These Condensin II activities compartmentalize the nucleus into discrete chromosome territories and indicate commonalities in the mechanisms that regulate the spatial structure of the genome during mitosis and interphase. PMID:22956908

  15. Calcium Promotes the Formation of Syntaxin 1 Mesoscale Domains through Phosphatidylinositol 4,5-Bisphosphate*

    PubMed Central

    Platen, Mitja; Junius, Meike; Diederichsen, Ulf; Schaap, Iwan A. T.; Honigmann, Alf; Jahn, Reinhard; van den Bogaart, Geert

    2016-01-01

    Phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) is a minor component of total plasma membrane lipids, but it has a substantial role in the regulation of many cellular functions, including exo- and endocytosis. Recently, it was shown that PI(4,5)P2 and syntaxin 1, a SNARE protein that catalyzes regulated exocytosis, form domains in the plasma membrane that constitute recognition sites for vesicle docking. Also, calcium was shown to promote syntaxin 1 clustering in the plasma membrane, but the molecular mechanism was unknown. Here, using a combination of superresolution stimulated emission depletion microscopy, FRET, and atomic force microscopy, we show that Ca2+ acts as a charge bridge that specifically and reversibly connects multiple syntaxin 1/PI(4,5)P2 complexes into larger mesoscale domains. This transient reorganization of the plasma membrane by physiological Ca2+ concentrations is likely to be important for Ca2+-regulated secretion. PMID:26884341

  16. Curcumin promotes fibril formation in F isomer of human serum albumin via amorphous aggregation.

    PubMed

    Mothi, Nivin; Muthu, Shivani A; Kale, Avinash; Ahmad, Basir

    2015-12-01

    We here describe the amyloid fibrils promoting behavior of curcumin, which ability to inhibit amyloid fibrillization of several globular proteins is well documented. Transmission electron microscopy (TEM), 90° light scattering (RLS), thioflavine T (ThT) and Congo red (CR) binding studies demonstrated that both F (pH3.4) and E (pH1.8) isomers of human serum albumin (HSA) in the absence and presence of curcumin initially converted into amorphous aggregates. Interestingly, only the sample containing F isomer preincubated with curcumin formed fibrils on incubation for longer period. We also found that curcumin strongly bind to the F isomer, alter its secondary, tertiary structures and thermal stability. We conclude that the conversion of intermediate states into amorphous aggregate to fibrils is dictated by its conformation. This study provides unique insights into ligand-controlled HSA aggregation pathway and should provide a useful model system to study both amorphous and the fibrillar aggregation of multidomain proteins.

  17. Natural products for treatment of osteoporosis: The effects and mechanisms on promoting osteoblast-mediated bone formation.

    PubMed

    An, Jing; Yang, Hao; Zhang, Qian; Liu, Cuicui; Zhao, Jingjing; Zhang, Lingling; Chen, Bo

    2016-02-15

    Osteoporosis is a systemic metabolic bone disease characterized by a reduction in bone mass, bone quality, and microarchitectural deterioration. An imbalance in bone remodeling that is caused by more osteoclast-mediated bone resorption than osteoblast-mediated bone formation results in such pathologic bone disorder. Traditional Chinese medicines (TCM) have long been used to prevent and treat osteoporosis and have received extensive attentions and researches at home and abroad, because they have fewer adverse reactions and are more suitable for long-term use compared with chemically synthesized medicines. Here, we put the emphasis on osteoblasts, summarized the detailed research progress on the active compounds derived from TCM with potential anti-osteoporosis effects and their molecular mechanisms on promoting osteoblast-mediated bone formation. It could be concluded that TCM with kidney-tonifying, spleen-tonifying, and stasis-removing effects all have the potential effects on treating osteoporosis. The active ingredients derived from TCM that possess effects on promoting osteoblasts proliferation and differentiation include flavonoids, glycosides, coumarins, terpenoids (sesquiterpenoids, monoterpenoids, diterpenoids), phenolic acids, phenols and others (tetrameric stilbene, anthraquinones, diarylheptanoids). And it was confirmed that the bone formation effect induced by the above natural products was regulated by the expressions of bone specific matrix proteins (ALP, BSP, OCN, OPN, COL I), transcription factor (Runx2, Cbfa1, Osx), signal pathways (MAPK, BMP), local factors (ROS, NO), OPG/RANKL system of osteoblasts and estrogen-like biological activities. All the studies provided theoretical basis for clinical application, as well as new drug research and development on treating osteoporosis.

  18. Vaccination with DKK1-derived peptides promotes bone formation and bone mass in an aged mouse osteoporosis model.

    PubMed

    Wu, Qiong; Li, Rui-Shu; Zhao, Yue; Wang, Zhi-Xia; Tang, Yan-Chun; Zhang, Jing; Liu, Jian-Ning; Tan, Xiang-Yang

    2014-08-01

    The investigation of agents for the treatment of osteoporosis has been a long-standing effort. The Wnt pathway plays an important role in bone formation and regeneration, and expression of Wnt pathway inhibitors, Dickkopf-1 (DKK1), appears to be associated with changes in bone mass. Inactivation of DKK1 leads to substantially increased bone mass in genetically manipulated animals. DKK1-derived peptides (DDPs) were added to BMP2-stimulated MC3T3-E1 preosteoblastic cells in vitro to evaluate inhibitory activity of DDPs in MC3T3-E1 cell differentiation. Study was extended in vivo on old female mice to show whether or not inhibition of endogenous DKK1 biological activity using DDPs vaccination approach leads to increase of bone formation, bone density, and improvement of bone microstructure. We reported that synthetic DDPs were able to reduce alkaline phosphatase activity, prevent mineralization and inhibit the differentiation of MC3T3-E1 cells in vitro. Furthermore, vaccination with these DDPs in aged female mice 4 times for a total period of 22 weeks promoted bone mass and bone microstructure. 3D microCT and histomorphometric analysis showed that there were significant increase in bone mineral densities, improvement of bone microstructure and promotion of bone formation in the vaccinated mice, especially in the mice vaccinated with DDP-A and DDP-C. Histological and scanning electron microscopy image analysis also indicated that vaccination increased trabecular bone mass and significantly decreased fragmentation of bone fibers. Taken together, these preclinical results suggest that vaccination with DDPs represents a promising new therapeutic approach for the treatment of bone-related disorders, such as osteoporosis.

  19. Telomeres and centromeres have interchangeable roles in promoting meiotic spindle formation

    PubMed Central

    Fennell, Alex; Fernández-Álvarez, Alfonso; Tomita, Kazunori

    2015-01-01

    Telomeres and centromeres have traditionally been considered to perform distinct roles. During meiotic prophase, in a conserved chromosomal configuration called the bouquet, telomeres gather to the nuclear membrane (NM), often near centrosomes. We found previously that upon disruption of the fission yeast bouquet, centrosomes failed to insert into the NM at meiosis I and nucleate bipolar spindles. Hence, the trans-NM association of telomeres with centrosomes during prophase is crucial for efficient spindle formation. Nonetheless, in approximately half of bouquet-deficient meiocytes, spindles form properly. Here, we show that bouquet-deficient cells can successfully undergo meiosis using centromere–centrosome contact instead of telomere–centrosome contact to generate spindle formation. Accordingly, forced association between centromeres and centrosomes fully rescued the spindle defects incurred by bouquet disruption. Telomeres and centromeres both stimulate focal accumulation of the SUN domain protein Sad1 beneath the centrosome, suggesting a molecular underpinning for their shared spindle-generating ability. Our observations demonstrate an unanticipated level of interchangeability between the two most prominent chromosomal landmarks. PMID:25688135

  20. The extracellular matrix component laminin promotes gap junction formation in the rat anterior pituitary gland.

    PubMed

    Horiguchi, Kotaro; Kouki, Tom; Fujiwara, Ken; Kikuchi, Motoshi; Yashiro, Takashi

    2011-03-01

    Folliculo-stellate (FS) cells in the anterior pituitary gland are believed to have multifunctional properties. FS cells connect to each other not only by mechanical means, but also by gap junctional cell-to-cell communication. Using transgenic rats that express green fluorescent protein (GFP) specifically in FS cells in the anterior pituitary gland (S100b-GFP rats), we recently revealed that FS cells in primary culture markedly change their shape, and form numerous interconnections with neighboring FS cells in the presence of laminin, an extracellular matrix (ECM) component of the basement membrane. Morphological and functional changes in cells are believed to be partly modified by matricrine signaling, by which ECM components function as cellular signals. In the present study, we examined whether gap junction formation between FS cells is affected by matricrine cues. A cell sorter was used to isolate FS cells from male S100b-GFP rat anterior pituitary for primary culture. We observed that mRNA and protein levels of connexin 43 in gap junction channels were clearly higher in the presence of laminin. In addition, we confirmed the formation of gap junctions between FS cells in primary culture by electron microscopy. Interestingly, we also observed that FS cells in the presence of laminin displayed well-developed rough endoplasmic reticulum and Golgi apparatus. Our findings suggest that, in anterior pituitary gland, FS cells may facilitate functional roles such as gap junctional cell-to-cell communication by matricrine signaling.

  1. Multiple endothelial cells constitute the tip of developing blood vessels and polarize to promote lumen formation.

    PubMed

    Pelton, John C; Wright, Catherine E; Leitges, Michael; Bautch, Victoria L

    2014-11-01

    Blood vessel polarization in the apical-basal axis is important for directed secretion of proteins and lumen formation; yet, when and how polarization occurs in the context of angiogenic sprouting is not well understood. Here, we describe a novel topology for endothelial cells at the tip of angiogenic sprouts in several mammalian vascular beds. Two cells that extend filopodia and have significant overlap in space and time were present at vessel tips, both in vitro and in vivo. The cell overlap is more extensive than predicted for tip cell switching, and it sets up a longitudinal cell-cell border that is a site of apical polarization and lumen formation, presumably via a cord-hollowing mechanism. The extent of cell overlap at the tip is reduced in mice lacking aPKCζ, and this is accompanied by reduced distal extension of both the apical border and patent lumens. Thus, at least two polarized cells occupy the distal tip of blood vessel sprouts, and topology, polarization and lumenization along the longitudinal border of these cells are influenced by aPKCζ.

  2. Multiple endothelial cells constitute the tip of developing blood vessels and polarize to promote lumen formation

    PubMed Central

    Pelton, John C.; Wright, Catherine E.; Leitges, Michael; Bautch, Victoria L.

    2014-01-01

    Blood vessel polarization in the apical-basal axis is important for directed secretion of proteins and lumen formation; yet, when and how polarization occurs in the context of angiogenic sprouting is not well understood. Here, we describe a novel topology for endothelial cells at the tip of angiogenic sprouts in several mammalian vascular beds. Two cells that extend filopodia and have significant overlap in space and time were present at vessel tips, both in vitro and in vivo. The cell overlap is more extensive than predicted for tip cell switching, and it sets up a longitudinal cell-cell border that is a site of apical polarization and lumen formation, presumably via a cord-hollowing mechanism. The extent of cell overlap at the tip is reduced in mice lacking aPKCζ, and this is accompanied by reduced distal extension of both the apical border and patent lumens. Thus, at least two polarized cells occupy the distal tip of blood vessel sprouts, and topology, polarization and lumenization along the longitudinal border of these cells are influenced by aPKCζ. PMID:25336741

  3. INF2 promotes the formation of detyrosinated microtubules necessary for centrosome reorientation in T cells

    PubMed Central

    Andrés-Delgado, Laura; Antón, Olga M.; Bartolini, Francesca; Ruiz-Sáenz, Ana; Correas, Isabel; Gundersen, Gregg G.

    2012-01-01

    T cell antigen receptor–proximal signaling components, Rho-family GTPases, and formin proteins DIA1 and FMNL1 have been implicated in centrosome reorientation to the immunological synapse of T lymphocytes. However, the role of these molecules in the reorientation process is not yet defined. Here we find that a subset of microtubules became rapidly stabilized and that their α-tubulin subunit posttranslationally detyrosinated after engagement of the T cell receptor. Formation of stabilized, detyrosinated microtubules required the formin INF2, which was also found to be essential for centrosome reorientation, but it occurred independently of T cell receptor–induced massive tyrosine phosphorylation. The FH2 domain, which was mapped as the INF2 region involved in centrosome repositioning, was able to mediate the formation of stable, detyrosinated microtubules and to restore centrosome translocation in DIA1-, FMNL1-, Rac1-, and Cdc42-deficient cells. Further experiments indicated that microtubule stabilization was required for centrosome polarization. Our work identifies INF2 and stable, detyrosinated microtubules as central players in centrosome reorientation in T cells. PMID:22986496

  4. Drosophila PTB promotes formation of high-order RNP particles and represses oskar translation

    PubMed Central

    Besse, Florence; López de Quinto, Sonia; Marchand, Virginie; Trucco, Alvar; Ephrussi, Anne

    2009-01-01

    Local translation of asymmetrically enriched mRNAs is a powerful mechanism for functional polarization of the cell. In Drosophila, exclusive accumulation of Oskar protein at the posterior pole of the oocyte is essential for development of the future embryo. This is achieved by the formation of a dynamic oskar ribonucleoprotein (RNP) complex regulating the transport of oskar mRNA, its translational repression while unlocalized, and its translational activation upon arrival at the posterior pole. We identified the nucleo–cytoplasmic shuttling protein PTB (polypyrimidine tract-binding protein)/hnRNP I as a new factor associating with the oskar RNP in vivo. While PTB function is largely dispensable for oskar mRNA transport, it is necessary for translational repression of the localizing mRNA. Unexpectedly, a cytoplasmic form of PTB can associate with oskar mRNA and repress its translation, suggesting that nuclear recruitment of PTB to oskar complexes is not required for its regulatory function. Furthermore, PTB binds directly to multiple sites along the oskar 3′ untranslated region and mediates assembly of high-order complexes containing multiple oskar RNA molecules in vivo. Thus, PTB is a key structural component of oskar RNP complexes that dually controls formation of high-order RNP particles and translational silencing. PMID:19131435

  5. Macrophagic CD146 promotes foam cell formation and retention during atherosclerosis.

    PubMed

    Luo, Yongting; Duan, Hongxia; Qian, Yining; Feng, Liqun; Wu, Zhenzhen; Wang, Fei; Feng, Jing; Yang, Dongling; Qin, Zhihai; Yan, Xiyun

    2017-03-01

    The persistence of cholesterol-engorged macrophages (foam cells) in the artery wall fuels the development of atherosclerosis. However, the mechanism that regulates the formation of macrophage foam cells and impedes their emigration out of inflamed plaques is still elusive. Here, we report that adhesion receptor CD146 controls the formation of macrophage foam cells and their retention within the plaque during atherosclerosis exacerbation. CD146 is expressed on the macrophages in human and mouse atheroma and can be upregulated by oxidized low-density lipoprotein (oxLDL). CD146 triggers macrophage activation by driving the internalization of scavenger receptor CD36 during lipid uptake. In response to oxLDL, macrophages show reduced migratory capacity toward chemokines CCL19 and CCL21; this capacity can be restored by blocking CD146. Genetic deletion of macrophagic CD146 or targeting of CD146 with an antibody result in much less complex plaques in high-fat diet-fed ApoE(-/-) mice by causing lipid-loaded macrophages to leave plaques. Collectively, our findings identify CD146 as a novel retention signal that traps macrophages within the artery wall, and a promising therapeutic target in atherosclerosis treatment.

  6. Loss of pRB and p107 disrupts cartilage development and promotes enchondroma formation

    PubMed Central

    Landman, Allison S.; Danielian, Paul S.; Lees, Jacqueline A.

    2013-01-01

    The pocket proteins pRB, p107, and p130 have established roles in regulating the cell cycle through the control of E2F activity. The pocket proteins regulate differentiation of a number of tissues in both cell cycle-dependent and –independent manners. Prior studies showed that mutation of p107 and p130 in the mouse leads to defects in cartilage development during endochondral ossification, the process by which long bones form. Despite evidence of a role for pRB in osteoblast differentiation, it is unknown whether it functions during cartilage development. Here, we show that mutation of Rb in the early mesenchyme of p107-mutant mice results in severe cartilage defects in the growth plates of long bones. This is attributable to inappropriate chondrocyte proliferation that persists after birth and leads to the formation of enchondromas in the growth plates as early as 8 weeks of age. Genetic crosses show that development of these tumorigenic lesions is E2f3 dependent. These results reveal an overlapping role for pRB and p107 in cartilage development, endochondral ossification and enchondroma formation that reflects their coordination of cell cycle exit at appropriate developmental stages. PMID:23146901

  7. Macrophagic CD146 promotes foam cell formation and retention during atherosclerosis

    PubMed Central

    Luo, Yongting; Duan, Hongxia; Qian, Yining; Feng, Liqun; Wu, Zhenzhen; Wang, Fei; Feng, Jing; Yang, Dongling; Qin, Zhihai; Yan, Xiyun

    2017-01-01

    The persistence of cholesterol-engorged macrophages (foam cells) in the artery wall fuels the development of atherosclerosis. However, the mechanism that regulates the formation of macrophage foam cells and impedes their emigration out of inflamed plaques is still elusive. Here, we report that adhesion receptor CD146 controls the formation of macrophage foam cells and their retention within the plaque during atherosclerosis exacerbation. CD146 is expressed on the macrophages in human and mouse atheroma and can be upregulated by oxidized low-density lipoprotein (oxLDL). CD146 triggers macrophage activation by driving the internalization of scavenger receptor CD36 during lipid uptake. In response to oxLDL, macrophages show reduced migratory capacity toward chemokines CCL19 and CCL21; this capacity can be restored by blocking CD146. Genetic deletion of macrophagic CD146 or targeting of CD146 with an antibody result in much less complex plaques in high-fat diet-fed ApoE−/− mice by causing lipid-loaded macrophages to leave plaques. Collectively, our findings identify CD146 as a novel retention signal that traps macrophages within the artery wall, and a promising therapeutic target in atherosclerosis treatment. PMID:28084332

  8. Syndapin promotes pseudocleavage furrow formation by actin organization in the syncytial Drosophila embryo

    PubMed Central

    Sherlekar, Aparna; Rikhy, Richa

    2016-01-01

    Coordinated membrane and cytoskeletal remodeling activities are required for membrane extension in processes such as cytokinesis and syncytial nuclear division cycles in Drosophila. Pseudocleavage furrow membranes in the syncytial Drosophila blastoderm embryo show rapid extension and retraction regulated by actin-remodeling proteins. The F-BAR domain protein Syndapin (Synd) is involved in membrane tubulation, endocytosis, and, uniquely, in F-actin stability. Here we report a role for Synd in actin-regulated pseudocleavage furrow formation. Synd localized to these furrows, and its loss resulted in short, disorganized furrows. Synd presence was important for the recruitment of the septin Peanut and distribution of Diaphanous and F-actin at furrows. Synd and Peanut were both absent in furrow-initiation mutants of RhoGEF2 and Diaphanous and in furrow-progression mutants of Anillin. Synd overexpression in rhogef2 mutants reversed its furrow-extension phenotypes, Peanut and Diaphanous recruitment, and F-actin organization. We conclude that Synd plays an important role in pseudocleavage furrow extension, and this role is also likely to be crucial in cleavage furrow formation during cell division. PMID:27146115

  9. Cholesterol gallstones and bile host diverse bacterial communities with potential to promote the formation of gallstones.

    PubMed

    Peng, Yuhong; Yang, Yang; Liu, Yongkang; Nie, Yuanyang; Xu, Peilun; Xia, Baixue; Tian, Fuzhou; Sun, Qun

    2015-01-01

    The prevalence of cholesterol gallstones has increased in recent years. Bacterial infection correlates with the formation of gallstones. We studied the composition and function of bacterial communities in cholesterol gallstones and bile from 22 cholesterol gallstone patients using culture-dependent and culture-independent methods. Altogether fourteen and eight bacterial genera were detected in cholesterol gallstones and bile, respectively. Pseudomonas spp. were the dominant bacteria in both cholesterol gallstones and bile. As judged by diversity indices, hierarchical clustering and principal component analysis, the bacterial communities in gallstones were different from those in bile. The gallstone microbiome was considered more stable than that of bile. The different microbial communities may be partially explained by differences in their habitats. We found that 30% of the culturable strains from cholesterol gallstones secreted β-glucuronidase and phospholipase A2. Pseudomonas aeruginosa strains showed the highest β-glucuronidase activity and produced the highest concentration of phospholipase A2, indicating that Ps. aeruginosa may be a major agent in the formation of cholesterol gallstones.

  10. microRNAs That Promote or Inhibit Memory Formation in Drosophila melanogaster

    PubMed Central

    Busto, Germain U.; Guven-Ozkan, Tugba; Fulga, Tudor A.; Van Vactor, David; Davis, Ronald L.

    2015-01-01

    microRNAs (miRNAs) are small noncoding RNAs that regulate gene expression post-transcriptionally. Prior studies have shown that they regulate numerous physiological processes critical for normal development, cellular growth control, and organismal behavior. Here, we systematically surveyed 134 different miRNAs for roles in olfactory learning and memory formation using “sponge” technology to titrate their activity broadly in the Drosophila melanogaster central nervous system. We identified at least five different miRNAs involved in memory formation or retention from this large screen, including miR-9c, miR-31a, miR-305, miR-974, and miR-980. Surprisingly, the titration of some miRNAs increased memory, while the titration of others decreased memory. We performed more detailed experiments on two miRNAs, miR-974 and miR-31a, by mapping their roles to subpopulations of brain neurons and testing the functional involvement in memory of potential mRNA targets through bioinformatics and a RNA interference knockdown approach. This screen offers an important first step toward the comprehensive identification of all miRNAs and their potential targets that serve in gene regulatory networks important for normal learning and memory. PMID:26088433

  11. Molecular cloning of the tomato Hairless gene implicates actin dynamics in trichome-mediated defense and mechanical properties of stem tissue

    PubMed Central

    Kang, Jin-Ho; Campos, Marcelo L.; Zemelis-Durfee, Starla; Al-Haddad, Jameel M.; Jones, A. Daniel; Telewski, Frank W.; Brandizzi, Federica; Howe, Gregg A.

    2016-01-01

    Trichomes are epidermal structures that provide a first line of defense against arthropod herbivores. The recessive hairless (hl) mutation in tomato (Solanum lycopersicum L.) causes severe distortion of trichomes on all aerial tissues, impairs the accumulation of sesquiterpene and polyphenolic compounds in glandular trichomes, and compromises resistance to the specialist herbivore Manduca sexta. Here, we demonstrate that the tomato Hl gene encodes a subunit (SRA1) of the highly conserved WAVE regulatory complex that controls nucleation of actin filaments in a wide range of eukaryotic cells. The tomato SRA1 gene spans a 42-kb region containing both Solyc11g013280 and Solyc11g013290. The hl mutation corresponds to a complex 3-kb deletion that removes the last exon of the gene. Expression of a wild-type SRA1 cDNA in the hl mutant background restored normal trichome development, accumulation of glandular trichome-derived metabolites, and resistance to insect herbivory. These findings establish a role for SRA1 in the development of tomato trichomes and also implicate the actin-cytoskeleton network in cytosolic control of specialized metabolism for plant defense. We also show that the brittleness of hl mutant stems is associated with altered mechanical and cell morphological properties of stem tissue, and demonstrate that this defect is directly linked to the mutation in SRA1. PMID:27481446

  12. Fine mapping and candidate gene analysis of Brtri1, a gene controlling trichome development in Chinese cabbage (Brassica rapa L. ssp pekinensis).

    PubMed

    Ye, X L; Hu, F Y; Ren, J; Huang, S N; Liu, W J; Feng, H; Liu, Z Y

    2016-12-02

    Trichomes are derived from the epidermis and constitute an ideal system for studying cell division in plants. Here, a Chinese cabbage doubled haploid (DH) line (FT) without trichomes was crossed with another DH line (PurDH-1) with trichomes to develop an F2 population for fine mapping of trichome control genes. Genetic analysis showed that the trichome phenotype was controlled by a single dominant gene, Brtri1. Using 1226 glabrous individuals in the F2 segregation population, Brtri1 was localized to a 16.84 kb region between markers Pur6-31 and Pur6-39 on chromosome A06. One of the four complete open reading frames within the mapping region, Bra025311, encodes a MYB transcription factor and is highly homologous to the trichome regulatory gene GL1 in Arabidopsis thaliana. It was thus regarded as a candidate gene for Brtri1. Comparative sequencing showed a 5-bp deletion in the third exon of Bra025311 in FT, resulting in a frame-shift mutation. No expression of Bra025311 was detected in FT. A co-dominant indel marker close to this mutation site was developed for marker-assisted selection in Chinese cabbage breeding.

  13. Molecular cloning of the tomato Hairless gene implicates actin dynamics in trichome-mediated defense and mechanical properties of stem tissue.

    PubMed

    Kang, Jin-Ho; Campos, Marcelo L; Zemelis-Durfee, Starla; Al-Haddad, Jameel M; Jones, A Daniel; Telewski, Frank W; Brandizzi, Federica; Howe, Gregg A

    2016-10-01

    Trichomes are epidermal structures that provide a first line of defense against arthropod herbivores. The recessive hairless (hl) mutation in tomato (Solanum lycopersicum L.) causes severe distortion of trichomes on all aerial tissues, impairs the accumulation of sesquiterpene and polyphenolic compounds in glandular trichomes, and compromises resistance to the specialist herbivore Manduca sexta Here, we demonstrate that the tomato Hl gene encodes a subunit (SRA1) of the highly conserved WAVE regulatory complex that controls nucleation of actin filaments in a wide range of eukaryotic cells. The tomato SRA1 gene spans a 42-kb region containing both Solyc11g013280 and Solyc11g013290 The hl mutation corresponds to a complex 3-kb deletion that removes the last exon of the gene. Expression of a wild-type SRA1 cDNA in the hl mutant background restored normal trichome development, accumulation of glandular trichome-derived metabolites, and resistance to insect herbivory. These findings establish a role for SRA1 in the development of tomato trichomes and also implicate the actin-cytoskeleton network in cytosolic control of specialized metabolism for plant defense. We also show that the brittleness of hl mutant stems is associated with altered mechanical and cell morphological properties of stem tissue, and demonstrate that this defect is directly linked to the mutation in SRA1.

  14. Does radiative feedback by the first stars promote or prevent second generation star formation?

    NASA Astrophysics Data System (ADS)

    Ahn, Kyungjin; Shapiro, Paul R.

    2007-03-01

    We study the effect of starlight from the first stars on the ability of other minihaloes in their neighbourhood to form additional stars. The first stars in the Λ cold dark matter (ΛCDM) universe are believed to have formed in minihaloes of total mass ~105-6 Msolar at redshifts z >~ 20, when molecular hydrogen (H2) formed and cooled the dense gas at their centres, leading to gravitational collapse. Simulations suggest that the Population III (Pop III) stars thus formed were massive (~100 Msolar) and luminous enough in ionizing radiation to cause an ionization front (I-front) to sweep outward, through their host minihalo and beyond, into the intergalactic medium. Our previous work suggested that this I-front was trapped when it encountered other, nearby minihaloes, and that it failed to penetrate the dense gas at their centres within the lifetime of the Pop III stars (<~3 Myr). The question of what the dynamical consequences were for these target minihaloes, of their exposure to the ionizing and dissociating starlight from the Pop III star requires further study, however. Towards this end, we have performed a series of detailed, one-dimensional (1D), radiation-hydrodynamical simulations to answer the question of whether star formation in these surrounding minihaloes was triggered or suppressed by radiation from the first stars. We have varied the distance to the source (and, hence, the flux) and the mass and evolutionary stage of the target haloes to quantify this effect. We find (1) trapping of the I-front and its transformation from R-type to D-type, preceded by a shock front; (2) photoevaporation of the ionized gas (i.e. all gas originally located outside the trapping radius); (3) formation of an H2 precursor shell which leads the I-front, stimulated by partial photoionization; and (4) the shock-induced formation of H2 in the minihalo neutral core when the shock speeds up and partially ionizes the gas. The fate of the neutral core is mostly determined by the

  15. The hot-spot p53R172H mutant promotes formation of giant spermatogonia triggered by DNA damage.

    PubMed

    Xue, Y; Raharja, A; Sim, W; Wong, E S M; Rahmat, S A B; Lane, D P

    2017-04-06

    Overexpression of mutant p53 is a common finding in most cancers but testicular tumours accumulate wild-type p53 (wtp53). In contrast to the accepted concept that p53 homozygous mutant mice do not accumulate mutant p53 in normal cells, our study on a mutant p53 mouse model of Li-Fraumeni syndrome harbouring the hot-spot p53R172H mutation described an elevated level of mutant p53 in non-cancerous mouse tissues. Here we use detailed immunohistochemical analysis to document the expression of p53R172H in mouse testis. In developing and adult testes, p53R172H was expressed in gonocytes, type A, Int, B spermatogonia as well as in pre-Sertoli cells and Leydig cells but was undetectable in spermatocytes and spermatids. A similar staining pattern was demonstrated for wtp53. However, the intensity of wtp53 staining was generally weaker than that of p53R172H, which indicates that the expression of p53R172H can be a surrogate marker of p53 gene transcription. Comparing the responses of wtp53 and p53R172H to irradiation, we found persistent DNA double-strand breaks in p53R172H testes and the formation of giant spermatogonia (GSG) following persistent DNA damage in p53R172H and p53-null mice. Strikingly, we found that p53R172H promotes spontaneous formation of GSG in non-stressed p53R172H ageing mice. Two types of GSG: Viable and Degenerative GSG were defined. We elucidate the factors involved in the formation of GSG: the loss of p53 function is a requirement for the formation of GSG whereas DNA damage acts as a promoting trigger. The formation of GSG does not translate to higher efficacy of testicular tumorigenesis arising from mutant p53 cells, which might be due to the presence of delayed-onset of p53-independent apoptosis.

  16. An Exploration of Teachers' Narratives: What Are the Facilitators and Constraints Which Promote or Inhibit "Good" Formative Assessment Practices in Schools?

    ERIC Educational Resources Information Center

    Sach, Elizabeth

    2015-01-01

    This paper set out to explore teachers' narratives in order to understand some of the facilitators and constraints which promote or inhibit good formative assessment practices in schools. A "responsive interview" approach was used to probe a small sample of lower and middle school teachers' perceptions of formative assessment.…

  17. Divergent Regulation of Terpenoid Metabolism in the Trichomes of Wild and Cultivated Tomato Species1[W][OA

    PubMed Central

    Besser, Katrin; Harper, Andrea; Welsby, Nicholas; Schauvinhold, Ines; Slocombe, Stephen; Li, Yi; Dixon, Richard A.; Broun, Pierre

    2009-01-01

    The diversification of chemical production in glandular trichomes is important in the development of resistance against pathogens and pests in two species of tomato. We have used genetic and genomic approaches to uncover some of the biochemical and molecular mechanisms that underlie the divergence in trichome metabolism between the wild species Solanum habrochaites LA1777 and its cultivated relative, Solanum lycopersicum. LA1777 produces high amounts of insecticidal sesquiterpene carboxylic acids (SCAs), whereas cultivated tomatoes lack SCAs and are more susceptible to pests. We show that trichomes of the two species have nearly opposite terpenoid profiles, consisting mainly of monoterpenes and low levels of sesquiterpenes in S. lycopersicum and mainly of SCAs and very low monoterpene levels in LA1777. The accumulation patterns of these terpenoids are different during development, in contrast to the developmental expression profiles of terpenoid pathway genes, which are similar in the two species, but they do not correlate in either case with terpenoid accumulation. However, our data suggest that the accumulation of monoterpenes in S. lycopersicum and major sesquiterpenes in LA1777 are linked both genetically and biochemically. Metabolite analyses after targeted gene silencing, inhibitor treatments, and precursor feeding all show that sesquiterpene biosynthesis relies mainly on products from the plastidic 2-C-methyl-d-erythritol-4-phosphate pathway in LA1777 but less so in the cultivated species. Furthermore, two classes of sesquiterpenes produced by the wild species may be synthesized from distinct pools of precursors via cytosolic and plastidial cyclases. However, highly trichome-expressed sesquiterpene cyclase-like enzymes were ruled out as being involved in the production of major LA1777 sesquiterpenes. PMID:18997116

  18. Mg(2+)/Ca(2+) promotes the adhesion of marine bacteria and algae and enhances following biofilm formation in artificial seawater.

    PubMed

    He, Xiaoyan; Wang, Jinpeng; Abdoli, Leila; Li, Hua

    2016-10-01

    Adhesion of microorganisms in the marine environment is essential for initiation and following development of biofouling. A variety of factors play roles in regulating the adhesion. Here we report the influence of Ca(2+) and Mg(2+) in artificial seawater on attachment and colonization of Bacillus sp., Chlorella and Phaeodactylum tricornutum on silicon wafer. Extra addition of the typical divalent cations in culturing solution gives rise to significantly enhanced adhesion of the microorganisms. Mg(2+) and Ca(2+) affect the adhesion of Bacillus sp. presumably by regulating aggregation and formation of extracellular polymeric substances (EPS). The ions alter quantity and types of the proteins in EPS, in turn affecting subsequent adhesion. However, it is noted that Mg(2+) promotes adhesion of Chlorella likely by regulating EPS formation and polysaccharide synthesis. Ca(2+) plays an important role in protein expression to enhance the adhesion of Chlorella. For Phaeodactylum tricornutum, Ca(2+) expedites protein synthesis for enhanced adhesion. The results shed some light on effective ways of utilizing divalent cations to mediate formation of biofilms on the marine structures for desired performances.

  19. The matrix protein CCN1 (CYR61) promotes proliferation, migration and tube formation of endothelial progenitor cells.

    PubMed

    Yu, Yang; Gao, Yu; Wang, Hong; Huang, Lan; Qin, Jun; Guo, Ruiwei; Song, Mingbao; Yu, Shiyong; Chen, Jianfei; Cui, Bin; Gao, Pan

    2008-10-15

    Neovascularization and re-endothelialization relies on circulating endothelial progenitor cells (EPCs), but their recruitment and angiogenic roles are subjected to regulation by the vascular microenvironment, which remains largely unknown. The present study was designed to investigate the effects of mature ECs and matrix protein CCN1 on the properties of EPCs. In a coculture system, effects of ECs on proliferation, migration and participation in tube-like formation of EPCs were evaluated, and functional assays were employed to identify the exact role of CCN1 in EPCs vitality and function. We demonstrated that ECs, as an indispensable part of the cellular milieu, significantly promoted the proliferation, migration and tube formation activities of EPCs, and more importantly, CCN1 was potentially involved in such effects of ECs. Expression of CCN1 in EPCs was significantly increased by serum, VEGF, ECs-cocultivation and ECs conditioned medium. Moreover, Ad-CCN1-mediated overexpression of CCN1 directly enhanced migration and tube formation of EPCs, whereas silencing of endogenous CCN1 in EPCs inhibits cell functions. Furthermore, CCN1 induced the expressions of chemokines and growth factors, such as MCP-1 and VEGF, suggesting a complex interaction between those proangiogenic factors. Our data suggest that matrix protein CCN1 may play an important role in microenvironment-mediated biological properties of EPCs.

  20. The matrix protein CCN1 (CYR61) promotes proliferation, migration and tube formation of endothelial progenitor cells

    SciTech Connect

    Yu Yang; Gao Yu; Wang, Hong; Huang Lan Qin Jun; Guo Ruiwei; Song Mingbao; Yu Shiyong; Chen Jianfei; Cui Bin; Gao Pan

    2008-10-15

    Neovascularization and re-endothelialization relies on circulating endothelial progenitor cells (EPCs), but their recruitment and angiogenic roles are subjected to regulation by the vascular microenvironment, which remains largely unknown. The present study was designed to investigate the effects of mature ECs and matrix protein CCN1 on the properties of EPCs. In a coculture system, effects of ECs on proliferation, migration and participation in tube-like formation of EPCs were evaluated, and functional assays were employed to identify the exact role of CCN1 in EPCs vitality and function. We demonstrated that ECs, as an indispensable part of the cellular milieu, significantly promoted the proliferation, migration and tube formation activities of EPCs, and more importantly, CCN1 was potentially involved in such effects of ECs. Expression of CCN1 in EPCs was significantly increased by serum, VEGF, ECs-cocultivation and ECs conditioned medium. Moreover, Ad-CCN1-mediated overexpression of CCN1 directly enhanced migration and tube formation of EPCs, whereas silencing of endogenous CCN1 in EPCs inhibits cell functions. Furthermore, CCN1 induced the expressions of chemokines and growth factors, such as MCP-1 and VEGF, suggesting a complex interaction between those proangiogenic factors. Our data suggest that matrix protein CCN1 may play an important role in microenvironment-mediated biological properties of EPCs.

  1. Wnt3a Promotes the Vasculogenic Mimicry Formation of Colon Cancer via Wnt/β-Catenin Signaling.

    PubMed

    Qi, Lisha; Song, Wangzhao; Liu, Zhiyong; Zhao, Xiulan; Cao, Wenfeng; Sun, Baocun

    2015-08-10

    Our previous study provided evidence that non-canonical Wnt signaling is involved in regulating vasculogenic mimicry (VM) formation. However, the functions of canonical Wnt signaling in VM formation have not yet been explored. In this study, we found the presence of VM was related to colon cancer histological differentiation (p < 0.001), the clinical stage (p < 0.001), and presence of metastasis and recurrence (p < 0.001). VM-positive colon cancer samples showed increased Wnt3a expression (p < 0.001) and β-catenin nuclear expression (p < 0.001) compared with the VM-negative samples. In vitro, over-regulated Wnt3a expression in HT29 colon cancer cells promoted the capacity to form tube-like structures in the three-dimensional (3-D) culture together with increased expression of endothelial phenotype-associated proteins such as VEGFR2 and VE-cadherin. The mouse xenograft model showed that Wnt3a-overexpressing cells grew into larger tumor masses and formed more VM than the control cells. In addition, the Wnt/β-catenin signaling antagonist Dickkopf-1(Dkk1) can reverse the capacity to form tube-like structures and can decrease the expressions of VEGFR2 and VE-cadherin in Wnt3a-overexpressing cells. Taken together, our results suggest that Wnt/β-catenin signaling is involved in VM formation in colon cancer and might contribute to the development of more accurate treatment modalities aimed at VM.

  2. Taxol-stabilized microtubules promote the formation of filaments from unmodified full-length Tau in vitro.

    PubMed

    Duan, Aranda R; Goodson, Holly V

    2012-12-01

    Tau is a neuronal protein that stabilizes the microtubule (MT) network, but it also forms filaments associated with Alzheimer's disease. Understanding Tau-MT and Tau-Tau interactions would help to establish Tau function in health and disease. For many years, literature reports on Tau-MT binding behavior and affinity have remained surprisingly contradictory (e.g., 10-fold variation in Tau-MT affinity). Tau-Tau interactions have also been investigated, but whether MTs might affect Tau filament formation is unknown. We have addressed these issues through binding assays and microscopy. We assessed Tau-MT interactions via cosedimentation and found that the measured affinity of Tau varies greatly, depending on the experimental design and the protein concentrations used. To investigate this dependence, we used fluorescence microscopy to examine Tau-MT binding. Strikingly, we found that Taxol-stabilized MTs promote Tau filament formation without characterized Tau-filament inducers. We propose that these novel Tau filaments account for the incongruence in Tau-MT affinity measurements. Moreover, electron microscopy reveals that these filaments appear similar to the heparin-induced Alzheimer's model. These observations suggest that the MT-induced Tau filaments provide a new model for Alzheimer's studies and that MTs might play a role in the formation of Alzheimer's-associated neurofibrillary tangles.

  3. Comparative glandular trichome transcriptome based gene characterization reveals reasons for differential (-)-menthol biosynthesis in Mentha species.

    PubMed

    Akhtar, Md Qussen; Qamar, Nida; Yadav, Pallavi; Kulkarni, Pallavi; Kumar, Ajay; Shasany, Ajit Kumar

    2017-02-11

    The genes involved in menthol biosynthesis are reported earlier in Mentha × piperita. But the information on these genes is not available in Mentha arvensis. To bridge the gap in knowledge on differential biosynthesis of monoterpenes leading to compositional variation in the essential oil of these species, a comparative transcriptome analysis of the glandular trichome was carried out. In addition to the MVA and MEP pathway genes, about 210 and 196 different terpene synthases (TPS) transcripts were identified from annotation in M. arvensis and M. × piperita, respectively, and correlated to several monoterpenes present in the essential oil. Six isoforms of (-)-menthol dehydrogenases (MD), the last enzyme of the menthol biosynthetic pathway, were identified, cloned and characterized from the transcriptome data (3 from each species). Varied expression levels and differential enzyme kinetics of these isoforms indicated the nature and composition of the product, as these isoforms generate both (-)-menthol and (+)-neomenthol from (-)-menthone and converts (-)-menthol to (-)-menthone in the reverse reaction, and hence together determine the quantity of (-)-menthol in the essential oil in these two species. Several genes for high value minor monoterpenes could also be identified from the transcriptome data. Abbreviations - AACT, acetyl-CoA C-acetyltransferase; CMK, 4-diphosphocytidyl-2-C-methyl-D-erythritol kinase; DGE, digital gene expression; DXR, 1-deoxy-D-xylulose-5-phosphate reductoisomerase; DXS, 1-deoxy-D-xylulose-5-phosphate synthase; FPPS, farnesyl pyrophosphate synthase; GC, gas chromatography; GPPS, geranyl pyrophosphate synthase; GT, glandular trichome; HDR, 4-hydroxy-3-methylbut-2-enyl diphosphate reductase; HDS, (E)-4-hydroxy-3-methylbut-2-enyl-diphosphate synthase; HMGR, 3-hydroxy-3-methylglutaryl-coenzyme A reductase; HMGS, hydroxymethylglutaryl-CoA synthase; IDI, isopentenyl-diphosphate delta-isomerase; IPD, isopiperitenol dehydrogenase; IPI

  4. Amelogenin Promotes the Formation of Elongated Apatite Microstructures in a Controlled Crystallization System

    PubMed Central

    Wang, Lijun; Guan, Xiangying; Du, Chang; Moradian-Oldak, Janet; Nancollas, George H.

    2009-01-01

    The organic matrix in forming enamel consists largely of the amelogenin protein self-assembled into nanospheres that play a pivotal role in controlling the oriented and elongated growth of highly ordered apatitic crystals during enamel biomineralization. However, the mechanisms of amelogenin-mediated mineralization have not yet been fully elucidated. Here we report that amelogenin dramatically accelerates the nucleation kinetics by decreasing the induction time in a dose-dependent manner in a controlled constant composition (CC) in vitro crystallization system. Remarkably, at very low protein concentrations, elongated microstructures which are similar in appearance to apatitic crystals in enamel were formed at relatively low supersaturations, through interfacial structural match/synergy between structured amelogenin assemblies and apatite nanocrystallites. This heterogeneous crystallization study provides experimental evidence to support the concept that templating by amelogenin very early in the crystallization process facilitates the formation of developing enamel crystals. PMID:20333260

  5. 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

  6. Zinc promotes clot stability by accelerating clot formation and modifying fibrin structure.

    PubMed

    Henderson, Sara J; Xia, Jing; Wu, Huayin; Stafford, Alan R; Leslie, Beverly A; Fredenburgh, James C; Weitz, David A; Weitz, Jeffrey I

    2016-03-01

    Zinc released from activated platelets binds fibrin(ogen) and attenuates fibrinolysis. Although zinc also affects clot formation, the mechanism and consequences are poorly understood. To address these gaps, the effect of zinc on clot formation and structure was examined in the absence or presence of factor (F) XIII. Zinc accelerated a) plasma clotting by 1.4-fold, b) fibrinogen clotting by 3.5- and 2.3-fold in the absence or presence of FXIII, respectively, c) fragment X clotting by 1.3-fold, and d) polymerisation of fibrin monomers generated with thrombin or batroxobin by 2.5- and 1.8-fold, respectively. Whereas absorbance increased up to 3.3-fold when fibrinogen was clotted in the presence of zinc, absorbance of fragment X clots was unaffected by zinc, consistent with reports that zinc binds to the αC-domain of fibrin(ogen). Scanning electron microscopic analysis revealed a two-fold increase in fibre diameter in the presence of zinc and in permeability studies, zinc increased clot porosity by 30-fold with or without FXIII. Whereas FXIII increased clot stiffness from 128 ± 19 Pa to 415 ± 27 Pa in rheological analyses, zinc reduced clot stiffness by 10- and 8.5-fold in the absence and presence of FXIII, respectively. Clots formed in the presence of zinc were more stable and resisted rupture with or without FXIII. Therefore, zinc accelerates clotting and reduces fibrin clot stiffness in a FXIII-independent manner, suggesting that zinc may work in concert with FXIII to modulate clot strength and stability.

  7. Inhibition of transforming growth factor β signaling promotes epiblast formation in mouse embryos.

    PubMed

    Ghimire, Sabitri; Heindryckx, Björn; Van der Jeught, Margot; Neupane, Jitesh; O'Leary, Thomas; Lierman, Sylvie; De Vos, Winnok H; Chuva de Sousa Lopes, Susana; Deroo, Tom; De Sutter, Petra

    2015-02-15

    Early lineage segregation in preimplantation embryos and maintenance of pluripotency in embryonic stem cells (ESCs) are both regulated by specific signaling pathways. Small molecules have been shown to modulate these signaling pathways. We examined the influence of several small molecules and growth factors on second-lineage segregation of the inner cell mass toward hypoblast and epiblast lineage during mouse embryonic preimplantation development. We found that the second-lineage segregation is influenced by activation or inhibition of the transforming growth factor (TGF)β pathway. Inhibition of the TGFβ pathway from the two-cell, four-cell, and morula stages onward up to the blastocyst stage significantly increased the epiblast cell proliferation. The epiblast formed in the embryos in which TGFβ signaling was inhibited was fully functional as demonstrated by the potential of these epiblast cells to give rise to pluripotent ESCs. Conversely, activating the TGFβ pathway reduced epiblast formation. Inhibition of the glycogen synthase kinase (GSK)3 pathway and activation of bone morphogenetic protein 4 signaling reduced the formation of both epiblast and hypoblast cells. Activation of the protein kinase A pathway and of the Janus kinase/signal transducer and activator of transcription 3 pathway did not influence the second-lineage segregation in mouse embryos. The simultaneous inhibition of three pathways--TGFβ, GSK3β, and the fibroblast growth factor (FGF)/extracellular signal-regulated kinases (Erk)--significantly enhanced the proliferation of epiblast cells than that caused by inhibition of either TGFβ pathway alone or by combined inhibition of the GSK3β and FGF/Erk pathways only.

  8. The Extract of Fructus Psoraleae Promotes Viability and Cartilaginous Formation of Rat Chondrocytes In Vitro

    PubMed Central

    Xu, Kang; Qiu, Xuefeng; Zhao, Wen; Wang, Dong

    2016-01-01

    This study aimed to investigate the extract components of FP on rat chondrocyte function and cartilaginous formation in vitro. Petroleum ether extract (P-e) of FP extract components was selected to treat Sprague-Dawley rat chondrocytes. Cell viability was tested with different concentrations (0.1, 1, 10, and 100 μg/mL) of P-e treatment. Concentrations of 0.1 and 1 μg/mL P-e conditioned culture mediums were used for treating chondrocytes in experiments. Cell proliferation was measured via DNA incorporation assay. Type II collagen, aggrecan, and Sox-9 genes expression levels were measured with RT-PCR. Additionally, cartilaginous formation was analyzed with type II collagen immunofluorescence, H&E, and alcian blue staining. Concentrations of 0.1 and 1 μg/mL P-e showed low cytotoxicity and demonstrated stimulatory effects on chondrocyte proliferation in early stages. Following 6 days of P-e culture, aggrecan and Sox-9 gene expression levels of the 1 μg/mL P-e group were upregulated by 1.82- (p < 0.05) and 2.06-fold (p < 0.05), respectively, versus controls. Moreover, 1 μg/mL P-e significantly stimulated cell aggregation and type II collagen deposits after 1 week of treatment. Noteworthy, tight cartilaginous structures formed in the 10-day 1 μg/mL P-e conditioned culture. These findings suggest that P-e has the potential to treat cartilage degeneration induced by chondrocyte failure. PMID:27994628

  9. Secretory trichomes, a substitutive floral nectar source in Lundia A. DC. (Bignoniaceae), a genus lacking a functional disc.

    PubMed

    Lopes, Ariadna Valentina; Vogel, Stefan; Machado, Isabel Cristina

    2002-08-01

    This is the first report of corolla-borne secretory trichomes that substitute in role for a non-functional disc in a species of the neotropical genus Lundia A. DC. (Bignoniaceae). The floral biology and flowering phenology of Lundia cordata were investigated at two remnants of tropical rainforest in northeastern Brazil. This species is a typically omithophilous liana, with reddish, tubular, scentless flowers. The flowers are resupinate, protandrous and last for 2 d. There is a vestigial non-functional perigynous disc and nectar is secreted by glandular trichomes distributed along the internal surface of the corolla. The nectar is stored at the base of the corolla tube, thus showing secondary nectar presentation. The nectariferous trichomes are multi-cellular, uniseriate, with a basal foot cell rooting in the epidermis, one neck cell, and a glandular head with 13 cells on average. Three species of hummingbirds (Amazilia fimbriata, Eupetomena macroura and Phaethornis pretrei) serve as pollinators. Phaethornis ruber, Xylocopa bees, wasps and diurnal moths are considered nectar thieves.

  10. EST analysis of hop glandular trichomes identifies an O-methyltransferase that catalyzes the biosynthesis of xanthohumol.

    PubMed

    Nagel, Jana; Culley, Lana K; Lu, Yuping; Liu, Enwu; Matthews, Paul D; Stevens, Jan F; Page, Jonathan E

    2008-01-01

    The glandular trichomes (lupulin glands) of hop (Humulus lupulus) synthesize essential oils and terpenophenolic resins, including the bioactive prenylflavonoid xanthohumol. To dissect the biosynthetic processes occurring in lupulin glands, we sequenced 10,581 ESTs from four trichome-derived cDNA libraries. ESTs representing enzymes of terpenoid biosynthesis, including all of the steps of the methyl 4-erythritol phosphate pathway, were abundant in the EST data set, as were ESTs for the known type III polyketide synthases of bitter acid and xanthohumol biosynthesis. The xanthohumol biosynthetic pathway involves a key O-methylation step. Four S-adenosyl-l-methionine-dependent O-methyltransferases (OMTs) with similarity to known flavonoid-methylating enzymes were present in the EST data set. OMT1, which was the most highly expressed OMT based on EST abundance and RT-PCR analysis, performs the final reaction in xanthohumol biosynthesis by methylating desmethylxanthohumol to form xanthohumol. OMT2 accepted a broad range of substrates, including desmethylxanthohumol, but did not form xanthohumol. Mass spectrometry and proton nuclear magnetic resonance analysis showed it methylated xanthohumol to 4-O-methylxanthohumol, which is not known from hop. OMT3 was inactive with all substrates tested. The lupulin gland-specific EST data set expands the genomic resources for H. lupulus and provides further insight into the metabolic specialization of glandular trichomes.

  11. EST Analysis of Hop Glandular Trichomes Identifies an O-Methyltransferase That Catalyzes the Biosynthesis of Xanthohumol[W][OA

    PubMed Central

    Nagel, Jana; Culley, Lana K.; Lu, Yuping; Liu, Enwu; Matthews, Paul D.; Stevens, Jan F.; Page, Jonathan E.

    2008-01-01

    The glandular trichomes (lupulin glands) of hop (Humulus lupulus) synthesize essential oils and terpenophenolic resins, including the bioactive prenylflavonoid xanthohumol. To dissect the biosynthetic processes occurring in lupulin glands, we sequenced 10,581 ESTs from four trichome-derived cDNA libraries. ESTs representing enzymes of terpenoid biosynthesis, including all of the steps of the methyl 4-erythritol phosphate pathway, were abundant in the EST data set, as were ESTs for the known type III polyketide synthases of bitter acid and xanthohumol biosynthesis. The xanthohumol biosynthetic pathway involves a key O-methylation step. Four S-adenosyl-l-methionine–dependent O-methyltransferases (OMTs) with similarity to known flavonoid-methylating enzymes were present in the EST data set. OMT1, which was the most highly expressed OMT based on EST abundance and RT-PCR analysis, performs the final reaction in xanthohumol biosynthesis by methylating desmethylxanthohumol to form xanthohumol. OMT2 accepted a broad range of substrates, including desmethylxanthohumol, but did not form xanthohumol. Mass spectrometry and proton nuclear magnetic resonance analysis showed it methylated xanthohumol to 4-O-methylxanthohumol, which is not known from hop. OMT3 was inactive with all substrates tested. The lupulin gland-specific EST data set expands the genomic resources for H. lupulus and provides further insight into the metabolic specialization of glandular trichomes. PMID:18223037

  12. Effect of prolonged water stress on specialized secondary metabolites, peltate glandular trichomes, and pathway gene expression in Artemisia annua L.

    PubMed

    Yadav, Ritesh K; Sangwan, Rajender S; Sabir, Farzana; Srivastava, Awadesh K; Sangwan, Neelam S

    2014-01-01

    Artemisia annua L. accumulates substantial quantities of unique sesquiternoid artemisinin and related phytomolecules and characteristic essential oil in glandular trichomes, present on its leaves and inflorescence. Water stress is a major concern in controlling plant growth and productivity. In this study, our aim was to find out the modulation of artemisinin and essential oil constituents in plants grown under prolonged water stress conditions. A. annua CIM-Arogya plants grown in pots were subjected to mild (60% ± 5) and moderate (40% ± 5) water stress treatment and continued during entire developmental period. Results revealed that artemisinin, arteannuin-B, artemisinic acid, dihydroartemisinic acid and essential oil content were positively controlled by the growth and development however negatively modulated by water deficit stress. Interestingly, some of minor monoterpenes, all sesquiterpenes and other low molecular weight volatiles of essential oil components were induced by water deficit treatment. Camphor which is the major essential oil constituents did not alter much while 1, 8 cineole was modulated during development of plant as well as under water stress conditions. Water deficit stress induces a decrease in glandular trichome density and size as well. The dynamics of various secondary metabolites is discussed in the light of growth responses, trichomes and pathway gene expression in plants grown under two levels of prolonged water stress conditions.

  13. Studies of a Biochemical Factory: Tomato Trichome Deep Expressed Sequence Tag Sequencing and Proteomics1[W][OA

    PubMed Central

    Schilmiller, Anthony L.; Miner, Dennis P.; Larson, Matthew; McDowell, Eric; Gang, David R.; Wilkerson, Curtis; Last, Robert L.

    2010-01-01

    Shotgun proteomics analysis allows hundreds of proteins to be identified and quantified from a single sample at relatively low cost. Extensive DNA sequence information is a prerequisite for shotgun proteomics, and it is ideal to have sequence for the organism being studied rather than from related species or accessions. While this requirement has limited the set of organisms that are candidates for this approach, next generation sequencing technologies make it feasible to obtain deep DNA sequence coverage from any organism. As part of our studies of specialized (secondary) metabolism in tomato (Solanum lycopersicum) trichomes, 454 sequencing of cDNA was combined with shotgun proteomics analyses to obtain in-depth profiles of genes and proteins expressed in leaf and stem glandular trichomes of 3-week-old plants. The expressed sequence tag and proteomics data sets combined with metabolite analysis led to the discovery and characterization of a sesquiterpene synthase that produces β-caryophyllene and α-humulene from E,E-farnesyl diphosphate in trichomes of leaf but not of stem. This analysis demonstrates the utility of combining high-throughput cDNA sequencing with proteomics experiments in a target tissue. These data can be used for dissection of other biochemical processes in these specialized epidermal cells. PMID:20431087

  14. Pyruvate kinase M2 activators promote tetramer formation and suppress tumorigenesis

    SciTech Connect

    Anastasiou, Dimitrios; Yu, Yimin; Israelsen, William J.; Jiang, Jian-Kang; Boxer, Matthew B.; Hong, Bum Soo; Tempel, Wolfram; Dimov, Svetoslav; Shen, Min; Jha, Abhishek; Yang, Hua; Mattaini, Katherine R.; Metallo, Christian M.; Fiske, Brian P.; Courtney, Kevin D.; Malstrom, Scott; Khan, Tahsin M.; Kung, Charles; Skoumbourdis, Amanda P.; Veith, Henrike; Southall, Noel; Walsh, Martin J.; Brimacombe, Kyle R.; Leister, William; Lunt, Sophia Y.; Johnson, Zachary R.; Yen, Katharine E.; Kunii, Kaiko; Davidson, Shawn M.; Christofk, Heather R.; Austin, Christopher P.; Inglese, James; Harris, Marian H.; Asara, John M.; Stephanopoulos, Gregory; Salituro, Francesco G.; Jin, Shengfang; Dang, Lenny; Auld, Douglas S.; Park, Hee-Won; Cantley, Lewis C.; Thomas, Craig J.; Vander Heiden, Matthew G.

    2012-08-26

    Cancer cells engage in a metabolic program to enhance biosynthesis and support cell proliferation. The regulatory properties of pyruvate kinase M2 (PKM2) influence altered glucose metabolism in cancer. The interaction of PKM2 with phosphotyrosine-containing proteins inhibits enzyme activity and increases the availability of glycolytic metabolites to support cell proliferation. This suggests that high pyruvate kinase activity may suppress tumor growth. We show that expression of PKM1, the pyruvate kinase isoform with high constitutive activity, or exposure to published small-molecule PKM2 activators inhibits the growth of xenograft tumors. Structural studies reveal that small-molecule activators bind PKM2 at the subunit interaction interface, a site that is distinct from that of the endogenous activator fructose-1,6-bisphosphate (FBP). However, unlike FBP, binding of activators to PKM2 promotes a constitutively active enzyme state that is resistant to inhibition by tyrosine-phosphorylated proteins. This data supports the notion that small-molecule activation of PKM2 can interfere with anabolic metabolism.

  15. Promotion of wound collagen formation in normal and diabetic mice by quadrol.

    PubMed

    Bhide, M V; Dunphy, M J; Mirkopulos, N; Smith, D J

    1988-01-01

    The rate of collagen deposition in implanted polytetrafluoroethylene (PTFE) tubing in non-diabetic and streptozotocin-induced (STZ) diabetic mice was measured during 14 days post-wounding. At the time of implantation, test groups received injections of either Quadrol [N,N,N',N'-tetrakis(2-hydroxypropyl)ethylenediamine], glucan, or buffer in an area adjacent to the wound site. The accumulation of collagen in the implants of Quadrol-treated non-diabetic animals was more than 200% above control on days 8 to 11 and was 50% above control on day 14. In Quadrol-treated STZ-diabetic mice, the collagen accumulation gradually increased from 50% above control on day 8 to 200% above control on day 14. Treatment with glucan increased the collagen accumulation in normal mice 200 to 300% above control from days 8 to 11 respectively and then 30% above control on day 14. Collagen accumulation in the implants of the glucan-treated STZ-diabetic mice was similar to the control group. These results indicate that Quadrol promotes in vivo collagen synthesis and that Quadrol may be effective as a stimulator of wound healing in diabetic and non-diabetic animals.

  16. Silicon-Doped Titanium Dioxide Nanotubes Promoted Bone Formation on Titanium Implants.

    PubMed

    Zhao, Xijiang; Wang, Tao; Qian, Shi; Liu, Xuanyong; Sun, Junying; Li, Bin

    2016-02-26

    While titanium (Ti) implants have been extensively used in orthopaedic and dental applications, the intrinsic bioinertness of untreated Ti surface usually results in insufficient osseointegration irrespective of the excellent biocompatibility and mechanical properties of it. In this study, we prepared surface modified Ti substrates in which silicon (Si) was doped into the titanium dioxide (TiO₂) nanotubes on Ti surface using plasma immersion ion implantation (PIII) technology. Compared to TiO₂ nanotubes and Ti alone, Si-doped TiO₂ nanotubes significantly enhanced the expression of genes related to osteogenic differentiation, including Col-I, ALP, Runx2, OCN, and OPN, in mouse pre-osteoblastic MC3T3-E1 cells and deposition of mineral matrix. In vivo, the pull-out mechanical tests after two weeks of implantation in rat femur showed that Si-doped TiO₂ nanotubes improved implant fixation strength by 18% and 54% compared to TiO₂-NT and Ti implants, respectively. Together, findings from this study indicate that Si-doped TiO₂ nanotubes promoted the osteogenic differentiation of osteoblastic cells and improved bone-Ti integration. Therefore, they may have considerable potential for the bioactive surface modification of Ti implants.

  17. Calcium carbonate hybrid coating promotes the formation of biomimetic hydroxyapatite on titanium surfaces

    NASA Astrophysics Data System (ADS)

    Cruz, Marcos Antônio E.; Ruiz, Gilia C. M.; Faria, Amanda N.; Zancanela, Daniela C.; Pereira, Lourivaldo S.; Ciancaglini, Pietro; Ramos, Ana P.

    2016-05-01

    CaCO3 particles dispersed in liquid media have proven to be good inductors of hydroxyapatite (HAp) growth. However, the use of CaCO3 deposited as thin films for this propose is unknown. Here, we report the growth of CaCO3 continuous films on Langmuir-Blodgett (LB) modified titanium surfaces and its use as HAp growth inductor. The Ti surfaces were modified with two, four, and six layers of dihexadecylphosphate (DHP)-LB films containing Ca2+, exposed to CO2 (g) for 12 h. The modified surfaces were immersed in simulated body fluid (SBF) at 37 °C for 36 h and submitted to bioactivity studies. This procedure originates bioactive coatings composed by non-stoichiometric HAp as evidenced by Fourier-Transform Infrared Spectroscopy (FTIR), X-ray Diffraction (XRD), and X-ray Photoelectron Spectroscopy (XPS). The presence of the CaCO3 film as pre-coating diminished the time necessary to growth continuous and homogeneous HAp films using a biomimetic approach. The surface properties of the films regarding their roughness, composition, charge, wettability, and surface free energy (γs) were accessed. The presence of HAp increased the wettability and γs of the surfaces. The coatings are not toxic for osteoblasts as observed for cell viability assays obtained after 7 and 14 days of culture. Moreover, the CaCO3 thin films promote the recovery of the osteoblasts viability more than the Ti surfaces themselves.

  18. Attenuation of AMPK signaling by ROQUIN promotes T follicular helper cell formation

    PubMed Central

    Ramiscal, Roybel R; Parish, Ian A; Lee-Young, Robert S; Babon, Jeffrey J; Blagih, Julianna; Pratama, Alvin; Martin, Jaime; Hawley, Naomi; Cappello, Jean Y; Nieto, Pablo F; Ellyard, Julia I; Kershaw, Nadia J; Sweet, Rebecca A; Goodnow, Christopher C; Jones, Russell G; Febbraio, Mark A; Vinuesa, Carola G; Athanasopoulos, Vicki

    2015-01-01

    T follicular helper cells (Tfh) are critical for the longevity and quality of antibody-mediated protection against infection. Yet few signaling pathways have been identified to be unique solely to Tfh development. ROQUIN is a post-transcriptional repressor of T cells, acting through its ROQ domain to destabilize mRNA targets important for Th1, Th17, and Tfh biology. Here, we report that ROQUIN has a paradoxical function on Tfh differentiation mediated by its RING domain: mice with a T cell-specific deletion of the ROQUIN RING domain have unchanged Th1, Th2, Th17, and Tregs during a T-dependent response but show a profoundly defective antigen-specific Tfh compartment. ROQUIN RING signaling directly antagonized the catalytic α1 subunit of adenosine monophosphate-activated protein kinase (AMPK), a central stress-responsive regulator of cellular metabolism and mTOR signaling, which is known to facilitate T-dependent humoral immunity. We therefore unexpectedly uncover a ROQUIN–AMPK metabolic signaling nexus essential for selectively promoting Tfh responses. DOI: http://dx.doi.org/10.7554/eLife.08698.001 PMID:26496200

  19. Silicon-Doped Titanium Dioxide Nanotubes Promoted Bone Formation on Titanium Implants

    PubMed Central

    Zhao, Xijiang; Wang, Tao; Qian, Shi; Liu, Xuanyong; Sun, Junying; Li, Bin

    2016-01-01

    While titanium (Ti) implants have been extensively used in orthopaedic and dental applications, the intrinsic bioinertness of untreated Ti surface usually results in insufficient osseointegration irrespective of the excellent biocompatibility and mechanical properties of it. In this study, we prepared surface modified Ti substrates in which silicon (Si) was doped into the titanium dioxide (TiO2) nanotubes on Ti surface using plasma immersion ion implantation (PIII) technology. Compared to TiO2 nanotubes and Ti alone, Si-doped TiO2 nanotubes significantly enhanced the expression of genes related to osteogenic differentiation, including Col-I, ALP, Runx2, OCN, and OPN, in mouse pre-osteoblastic MC3T3-E1 cells and deposition of mineral matrix. In vivo, the pull-out mechanical tests after two weeks of implantation in rat femur showed that Si-doped TiO2 nanotubes improved implant fixation strength by 18% and 54% compared to TiO2-NT and Ti implants, respectively. Together, findings from this study indicate that Si-doped TiO2 nanotubes promoted the osteogenic differentiation of osteoblastic cells and improved bone-Ti integration. Therefore, they may have considerable potential for the bioactive surface modification of Ti implants. PMID:26927080

  20. Pyruvate kinase M2 activators promote tetramer formation and suppress tumorigenesis

    PubMed Central

    Anastasiou, Dimitrios; Yu, Yimin; Israelsen, William J.; Jiang, Jian-kang; Boxer, Matthew B.; Hong, Bum Soo; Tempel, Wolfram; Dimov, Svetoslav; Shen, Min; Jha, Abhishek; Yang, Hua; Mattaini, Katherine R.; Metallo, Christian M.; Fiske, Brian P.; Courtney, Kevin D.; Malstrom, Scott; Khan, Tahsin M.; Kung, Charles; Skoumbourdis, Amanda P.; Veith, Henrike; Southall, Noel; Walsh, Martin J.; Brimacombe, Kyle R.; Leister, William; Lunt, Sophia Y.; Johnson, Zachary R.; Yen, Katharine E.; Kunii, Kaiko; Davidson, Shawn M.; Christofk, Heather R.; Austin, Christopher P.; Inglese, James; Harris, Marian H.; Asara, John M.; Stephanopoulos, Gregory; Salituro, Francesco G.; Jin, Shengfang; Dang, Lenny; Auld, Douglas S.; Park, Hee-Won; Cantley, Lewis C.; Thomas, Craig J.; Vander Heiden, Matthew G.

    2012-01-01

    Cancer cells engage in a metabolic program to enhance biosynthesis and support cell proliferation. The regulatory properties of pyruvate kinase M2 (PKM2) influence altered glucose metabolism in cancer. PKM2 interaction with phosphotyrosine-containing proteins inhibits enzyme activity and increases availability of glycolytic metabolites to support cell proliferation. This suggests that high pyruvate kinase activity may suppress tumor growth. We show that expression of PKM1, the pyruvate kinase isoform with high constitutive activity, or exposure to published small molecule PKM2 activators inhibit growth of xenograft tumors. Structural studies reveal that small molecule activators bind PKM2 at the subunit interaction interface, a site distinct from that of the endogenous activator fructose-1,6-bisphosphate (FBP). However, unlike FBP, binding of activators to PKM2 promotes a constitutively active enzyme state that is resistant to inhibition by tyrosine-phosphorylated proteins. These data support the notion that small molecule activation of PKM2 can interfere with anabolic metabolism. PMID:22922757

  1. Mesenchymal Wnt signaling promotes formation of sternum and thoracic body wall.

    PubMed

    Snowball, John; Ambalavanan, Manoj; Cornett, Bridget; Lang, Richard; Whitsett, Jeffrey; Sinner, Debora

    2015-05-15

    Midline defects account for approximately 5% of congenital abnormalities observed at birth. However, the molecular mechanisms underlying the formation of the ventral body wall are not well understood. Recent studies linked mutations in Porcupine-an O-acetyl transferase mediating Wnt ligand acylation-with defects in the thoracic body wall. We hypothesized that anomalous Wnt signaling is involved in the pathogenesis of defective closure of the thoracic body wall. We generated a mouse model wherein Wntless (Wls), which encodes a cargo receptor mediating secretion of Wnt ligands, was conditionally deleted from the developing mesenchyme using Dermo1Cre mice. Wls(f/f);Dermo1(Cre/+) embryos died during mid-gestation. At E13.5, skeletal defects were observed in the forelimbs, jaw, and rib cage. At E14.5, midline defects in the thoracic body wall began to emerge: the sternum failed to fuse and the heart protruded through the body wall at the midline (ectopia cordis). To determine the molecular mechanism underlying the phenotype observed in Wls(f/f);Dermo1(Cre/+) embryos, we tested whether Wnt/β-catenin signaling was operative in developing the embryonic ventral body wall using Axin2(LacZ) and BatGal reporter mice. While Wnt/β-catenin signaling activity was observed at the midline of the ventral body wall before sternal fusion, this pattern of activity was altered and scattered throughout the body wall after mesenchymal deletion of Wls. Mesenchymal cell migration was disrupted in Wls(f/f);Dermo1(Cre/+) thoracic body wall partially due to anomalous β-catenin independent Wnt signaling as determined by in vitro assays. Deletion of Lrp5 and Lrp6 receptors, which mediate Wnt/β-catenin signaling in the mesenchyme, partially recapitulated the phenotype observed in the chest midline of Wls(f/f);Dermo1(Cre/+) embryos supporting a role for Wnt/β-catenin signaling activity in the normal formation of the ventral body wall mesenchyme. We conclude that Wls-mediated secretion of Wnt

  2. SOXC Transcription Factors Induce Cartilage Growth Plate Formation in Mouse Embryos by Promoting Noncanonical WNT Signaling.

    PubMed

    Kato, Kenji; Bhattaram, Pallavi; Penzo-Méndez, Alfredo; Gadi, Abhilash; Lefebvre, Véronique

    2015-09-01

    Growth plates are specialized cartilage structures that ensure the elongation of most skeletal primordia during vertebrate development. They are made by chondrocytes that proliferate in longitudinal columns and then progress in a staggered manner towards prehypertrophic, hypertrophic and terminal maturation. Complex molecular networks control the formation and activity of growth plates, but remain incompletely understood. We investigated here the importance of the SoxC genes, which encode the SOX4, SOX11 and SOX12 transcription factors, in growth plates. We show that the three genes are expressed robustly in perichondrocytes and weakly in growth plate chondrocytes. SoxC(Prx1Cre) mice, which deleted SoxC genes in limb bud skeletogenic mesenchyme, were born with tiny appendicular cartilage primordia because of failure to form growth plates. In contrast, SoxC(Col2Cre) and SoxC(ATC) mice, which deleted SoxC genes primarily in chondrocytes, were born with mild dwarfism and fair growth plates. Chondrocytes in the latter mutants matured normally, but formed irregular columns, proliferated slowly and died ectopically. Asymmetric distribution of VANGL2 was defective in both SoxC(Prx1Cre) and SoxC(ATC) chondrocytes, indicating impairment of planar cell polarity, a noncanonical WNT signaling pathway that controls growth plate chondrocyte alignment, proliferation and survival. Accordingly, SoxC genes were necessary in perichondrocytes for expression of Wnt5a, which encodes a noncanonical WNT ligand required for growth plate formation, and in chondrocytes and perichondrocytes for expression of Fzd3 and Csnk1e, which encode a WNT receptor and casein kinase-1 subunit mediating planar cell polarity, respectively. Reflecting the differential strengths of the SOXC protein transactivation domains, SOX11 was more powerful than SOX4, and SOX12 interfered with the activity of SOX4 and SOX11. Altogether, these findings provide novel insights into the molecular regulation of skeletal

  3. Mesenchymal Wnt Signaling Promotes Formation of Sternum and Thoracic Body Wall

    PubMed Central

    Snowball, John; Ambalavanan, Manoj; Cornett, Bridget; Lang, Richard; Whitsett, Jeff; Sinner, Debora

    2015-01-01

    Midline defects account for approximately 5% of congenital abnormalities observed at birth. However, the molecular mechanisms underlying the formation of the ventral body wall are not well understood. Recent studies linked mutations in Porcupine—an O-acetyl transferase mediating Wnt ligand acylation—with defects in the thoracic body wall. We hypothesized that anomalous Wnt signaling is involved in the pathogenesis of defective closure of the thoracic body wall. We generated a mouse model wherein Wntless (Wls), which encodes a cargo receptor mediating secretion of Wnt ligands, was conditionally deleted from the developing mesenchyme using Dermo1Cre mice. Wlsf/f;Dermo1Cre/+ embryos died during mid-gestation. At E13.5, skeletal defects were observed in the forelimbs, jaw, and rib cage. At E14.5, midline defects in the thoracic body wall began to emerge: the sternum failed to fuse and the heart protruded through the body wall at the midline (ectopia cordis). To determine the molecular mechanism underlying the phenotype observed in Wlsf/f;Dermo1Cre/+ embryos, we tested whether Wnt/β-catenin signaling was operative in developing the embryonic ventral body wall using Axin2LacZ and BatGal reporter mice. While Wnt/β-catenin signaling activity was observed at the midline of the ventral body wall before sternal fusion, this pattern of activity was altered and scattered throughout the body wall after mesenchymal deletion of Wls. Mesenchymal cell migration was disrupted in Wlsf/f;Dermo1Cre/+ thoracic body wall partially due to anomalous non-canonical Wnt signaling as determined by in vitro assays. Deletion of Lrp5 and Lrp6 receptors, which mediate Wnt/β-catenin signaling in the mesenchyme, partially recapitulated the phenotype observed in the chest midline of Wlsf/f;Dermo1Cre/+ embryos supporting a role for Wnt/β-catenin signaling activity in the normal formation of the ventral body wall mesenchyme. We conclude that Wls-mediated secretion of Wnt ligands from the

  4. Pili of oral Streptococcus sanguinis bind to salivary amylase and promote the biofilm formation.

    PubMed

    Okahashi, Nobuo; Nakata, Masanobu; Terao, Yutaka; Isoda, Ryutaro; Sakurai, Atsuo; Sumitomo, Tomoko; Yamaguchi, Masaya; Kimura, Richard K; Oiki, Eiji; Kawabata, Shigetada; Ooshima, Takashi

    2011-01-01

    Streptococcus sanguinis is a member of oral streptococci and one of the most abundant species found in oral biofilm called dental plaque. Colonization of the oral streptococci on the tooth surface depends on the adhesion of bacteria to salivary components adsorbed to the tooth surface. Recently, we identified unique cell surface long filamentous structures named pili in this species. Herein, we investigated the role of S. sanguinis pili in biofilm formation. We found that pili-deficient mutant, in which the genes encoding the three pilus proteins PilA, PilB and PilC have been deleted, showed an impaired bacterial accumulation on saliva-coated surfaces. Confocal microscopic observations suggested that the mutant was incapable of producing typical three-dimensional layer of biofilm. Ligand blot analysis showed that the ancillary pilus proteins PilB and PilC bound to human whole saliva. Additional analysis demonstrated that PilC bound to multiple salivary components, and one of which was found to be salivary α-amylase. These results indicate that pilus proteins are members of saliva-binding proteins of oral S. sanguinis, and suggest the involvement of pili in its colonization on saliva-coated tooth surfaces and in the human oral cavity.

  5. Does Radiative Feedback by the First Stars Promote or Prevent Second Generation Star Formation?

    NASA Astrophysics Data System (ADS)

    Ahn, Kyungjin; Shapiro, Paul R.

    2008-03-01

    We present a self-consistent study of formation of Pop III stars in the early stage of cosmic reionization. We study the effect of starlight from the first stars on the ability of other minihalos in their neighborhood to form additional stars. We show that the ionization front (I-front) is trapped by the neighboring minihalos, after it is transformed from R-type to D-type and preceded by a shock front. The fate of the core of nearby minihalos is mostly determined by the response of the core to this shock front, which leads to molecular cooling and collapse that, when compared to the same halo without external radiation, is (a) expedited, (b) delayed, (c) unaltered, or (d) reversed and prevented, depending upon the flux and halo mass and evolutionary stage. Roughly speaking, most halos that were destined to cool, collapse and form stars in the absence of external radiation are found to do so even when exposed to the first Pop III star in their neighborhood, while those that would not have done so are still not able to.

  6. Type IV collagen aggregates promote keratinocyte proliferation and formation of epidermal layer in human skin equivalents.

    PubMed

    Matsuura-Hachiya, Yuko; Arai, Koji Y; Muraguchi, Taichi; Sasaki, Tasuku; Nishiyama, Toshio

    2017-03-07

    Type IV collagen isolated from lens capsule without enzymatic treatment is known to form a gel under physiological condition and influences cellular activities. In case of human keratinocytes, the suppression of proliferation on reconstituted type IV collagen gels was reported in monolayer culture. In this study, we examined effects of type IV collagen isolated from porcine lens capsule on epidermal formation in human skin equivalents. Type IV collagen aggregates were prepared under the culture condition and the aggregates suppressed keratinocyte proliferation in monolayer culture as well as the culture on the gels. In human skin equivalents type IV collagen aggregates were reconstituted on the surface of contracted collagen gels containing human dermal fibroblasts and the keratinocytes were then cultured on the aggregates for 14 days. Interestingly, in human skin equivalents with type IV collagen aggregates, the BrdU-positive keratinocytes were increased and the thickness of the epidermal layer was around twice than that of control culture. Epidermal differentiation markers were expressed in the upper layer of the epidermis and the defined deposition of human basement membrane components were increased at the dermal-epidermal junction. These results indicate that the type IV collagen aggregates stimulate the proliferation of basal keratinocytes and improve the stratification of epidermal layers in human skin equivalents. This article is protected by copyright. All rights reserved.

  7. Thromboxane A2 exacerbates acute lung injury via promoting edema formation

    PubMed Central

    Kobayashi, Koji; Horikami, Daiki; Omori, Keisuke; Nakamura, Tatsuro; Yamazaki, Arisa; Maeda, Shingo; Murata, Takahisa

    2016-01-01

    Thromboxane A2 (TXA2) is produced in the lungs of patients suffering from acute lung injury (ALI). We assessed its contribution in disease progression using three different ALI mouse models. The administration of hydrochloric acid (HCl) or oleic acid (OA)+ lipopolysaccharide (LPS) caused tissue edema and neutrophil infiltration with TXA2 production in the lungs of the experimental mice. The administration of LPS induced only neutrophil accumulation without TXA2 production. Pretreatment with T prostanoid receptor (TP) antagonist attenuated the tissue edema but not neutrophil infiltration in these models. Intravital imaging and immunostaining demonstrated that administration of TP agonist caused vascular hyper-permeability by disrupting the endothelial barrier formation in the mouse ear. In vitro experiments showed that TP-stimulation disrupted the endothelial adherens junction, and it was inhibited by Ca2+ channel blockade or Rho kinase inhibition. Thus endogenous TXA2 exacerbates ALI, and its blockade attenuates it by modulating the extent of lung edema. This can be explained by the endothelial hyper-permeability caused by the activation of TXA2-TP axis, via Ca2+- and Rho kinase-dependent signaling. PMID:27562142

  8. mTORC2 Signaling Promotes Skeletal Growth and Bone Formation in Mice

    PubMed Central

    Chen, Jianquan; Holguin, Nilsson; Shi, Yu; Silva, Matthew J.; Long, Fanxin

    2015-01-01

    Mammalian target of rapamycin (mTOR) is an evolutionarily conserved serine/threonine kinase controlling many physiological processes in mammals. mTOR functions in two distinct protein complexes, namely mTORC1 and mTORC2. Compared to mTORC1, the specific roles of mTORC2 are less well understood. To investigate the potential contribution of mTORC2 to skeletal development and homeostasis, we have genetically deleted Rictor, an essential component of mTORC2, in the limb skeletogenic mesenchyme of the mouse embryo. Loss of Rictor leads to shorter and narrower skeletal elements in both embryos and postnatal mice. In the embryo, Rictor deletion reduces the width but not the length of the initial cartilage anlage. Subsequently, the embryonic skeletal elements are shortened due to a delay in chondrocyte hypertrophy, with no change in proliferation, apoptosis, cell size, or matrix production. Postnatally, Rictor-deficient mice exhibit impaired bone formation, resulting in thinner cortical bone, but the trabecular bone mass is relatively normal thanks to a concurrent decrease in bone resorption. Moreover, Rictor-deficient bones exhibit a lesser anabolic response to mechanical loading. Thus, mTORC2 signaling is necessary for optimal skeletal growth and bone anabolism. PMID:25196701

  9. CD147 promotes the formation of functional osteoclasts through NFATc1 signalling.

    PubMed

    Nishioku, Tsuyoshi; Terasawa, Mariko; Baba, Misaki; Yamauchi, Atsushi; Kataoka, Yasufumi

    2016-04-29

    CD147, a membrane glycoprotein of the immunoglobulin superfamily, is highly upregulated during dynamic cellular events including tissue remodelling. Elevated CD147 expression is present in the joint of rheumatoid arthritis patients. However, the role of CD147 in bone destruction remains unclear. To determine whether CD147 is involved in osteoclastogenesis, we studied its expression in mouse osteoclasts and its role in osteoclast differentiation and function. CD147 expression was markedly upregulated during osteoclast differentiation. To investigate the role of CD147 in receptor activator of nuclear factor-kappa B ligand (RANKL)-induced osteoclastogenesis and bone resorption activity, osteoclast precursor cells were transfected with CD147 siRNA. Decreased CD147 expression inhibited osteoclast formation and bone resorption, inhibited RANKL-induced nuclear translocation of the nuclear factor of activated T cells (NFAT) c1 and decreased the expression of the d2 isoform of vacuolar ATPase Vo domain and cathepsin K. Therefore, CD147 plays a critical role in the differentiation and function of osteoclasts by upregulating NFATc1 through the autoamplification of its expression in osteoclastogenesis.

  10. The Gαo Activator Mastoparan-7 Promotes Dendritic Spine Formation in Hippocampal Neurons.

    PubMed

    Ramírez, Valerie T; Ramos-Fernández, Eva; Inestrosa, Nibaldo C

    2016-01-01

    Mastoparan-7 (Mas-7), an analogue of the peptide mastoparan, which is derived from wasp venom, is a direct activator of Pertussis toxin- (PTX-) sensitive G proteins. Mas-7 produces several biological effects in different cell types; however, little is known about how Mas-7 influences mature hippocampal neurons. We examined the specific role of Mas-7 in the development of dendritic spines, the sites of excitatory synaptic contact that are crucial for synaptic plasticity. We report here that exposure of hippocampal neurons to a low dose of Mas-7 increases dendritic spine density and spine head width in a time-dependent manner. Additionally, Mas-7 enhances postsynaptic density protein-95 (PSD-95) clustering in neurites and activates Gα(o) signaling, increasing the intracellular Ca(2+) concentration. To define the role of signaling intermediates, we measured the levels of phosphorylated protein kinase C (PKC), c-Jun N-terminal kinase (JNK), and calcium-calmodulin dependent protein kinase IIα (CaMKIIα) after Mas-7 treatment and determined that CaMKII activation is necessary for the Mas-7-dependent increase in dendritic spine density. Our results demonstrate a critical role for Gα(o) subunit signaling in the regulation of synapse formation.

  11. The Gαo Activator Mastoparan-7 Promotes Dendritic Spine Formation in Hippocampal Neurons

    PubMed Central

    Ramírez, Valerie T.; Ramos-Fernández, Eva; Inestrosa, Nibaldo C.

    2016-01-01

    Mastoparan-7 (Mas-7), an analogue of the peptide mastoparan, which is derived from wasp venom, is a direct activator of Pertussis toxin- (PTX-) sensitive G proteins. Mas-7 produces several biological effects in different cell types; however, little is known about how Mas-7 influences mature hippocampal neurons. We examined the specific role of Mas-7 in the development of dendritic spines, the sites of excitatory synaptic contact that are crucial for synaptic plasticity. We report here that exposure of hippocampal neurons to a low dose of Mas-7 increases dendritic spine density and spine head width in a time-dependent manner. Additionally, Mas-7 enhances postsynaptic density protein-95 (PSD-95) clustering in neurites and activates Gαo signaling, increasing the intracellular Ca2+ concentration. To define the role of signaling intermediates, we measured the levels of phosphorylated protein kinase C (PKC), c-Jun N-terminal kinase (JNK), and calcium-calmodulin dependent protein kinase IIα (CaMKIIα) after Mas-7 treatment and determined that CaMKII activation is necessary for the Mas-7-dependent increase in dendritic spine density. Our results demonstrate a critical role for Gαo subunit signaling in the regulation of synapse formation. PMID:26881110

  12. Tau and spectraplakins promote synapse formation and maintenance through Jun kinase and neuronal trafficking

    PubMed Central

    Voelzmann, Andre; Okenve-Ramos, Pilar; Qu, Yue; Chojnowska-Monga, Monika; del Caño-Espinel, Manuela; Prokop, Andreas; Sanchez-Soriano, Natalia

    2016-01-01

    The mechanisms regulating synapse numbers during development and ageing are essential for normal brain function and closely linked to brain disorders including dementias. Using Drosophila, we demonstrate roles of the microtubule-associated protein Tau in regulating synapse numbers, thus unravelling an important cellular requirement of normal Tau. In this context, we find that Tau displays a strong functional overlap with microtubule-binding spectraplakins, establishing new links between two different neurodegenerative factors. Tau and the spectraplakin Short Stop act upstream of a three-step regulatory cascade ensuring adequate delivery of synaptic proteins. This cascade involves microtubule stability as the initial trigger, JNK signalling as the central mediator, and kinesin-3 mediated axonal transport as the key effector. This cascade acts during development (synapse formation) and ageing (synapse maintenance) alike. Therefore, our findings suggest novel explanations for intellectual disability in Tau deficient individuals, as well as early synapse loss in dementias including Alzheimer’s disease. DOI: http://dx.doi.org/10.7554/eLife.14694.001 PMID:27501441

  13. Studies of the structure and function of Mms6, a bacterial protein that promotes the formation of magnetic nanoparticles

    SciTech Connect

    Wang, Lijun

    2011-01-01

    Here we report structural and functional studies of Mms6, a biomineralization protein that can promote the formation in vitro of magnetic nanoparticles with sizes and morphologies similar to the magnetites synthesized by magnetotactic bacteria. We found the binding pattern of Mms6 to ferric ion to be two-phase and multivalent. We quantatively determined that Mms6 binds one Fe3+ with a very high affinity (Kd = 1016 M). The second phase of iron binding is multivalent and cooperative with respect to iron with a Kd in the μM range and a stoichiometry of about 20 ferric ion per protein molecule. We found that Mms6 exists in large particles of two sizes, one consisting of 20-40 monomeric units and the other of 200 units. From proteolytic digestion, ultracentrifugation and liposome fusion studies, we found that Mms6 forms a large micellar quaternary structure with the N-terminal domain self-assembling into a uniformly sized micelle and the C-terminal domain on the surface. The two-phase iron-binding pattern may be relevant to iron crystal formation. We propose that the first high affinity phase may stabilize a new conformation of the C-terminal domain that allows interaction with other C-terminal domains leading to a structural change in the multimeric protein complex that enables the second low affinity iron binding phase to organize iron and initiate crystal formation. We also observed a dimeric apparent molecular mass of the Mms6 C-terminal peptide (C21Mms6). We speculate that the C-terminal domain may form higher order quaternary arrangements on the surface of the micelle or when anchored to a membrane by the N-terminal domain. The change in fluorescence quenching in the N-terminal domain with iron binding suggests a structural integrity between the C- and N-terminal domains. The slow change in trp fluorescence as a function of time after adding iron suggests a very slow conformational change in the protein that involves

  14. RacGAP1-driven focal adhesion formation promotes melanoma transendothelial migration through mediating adherens junction disassembly.

    PubMed

    Zhang, Pu; Bai, Huiyuan; Fu, Changliang; Chen, Feng; Zeng, Panying; Wu, Chengxiang; Ye, Qichao; Dong, Cheng; Song, Yang; Song, Erqun

    2015-03-27

    Melanoma cell migration across vascular endothelial cells is an essential step of tumor metastasis. Here, we provide evidence that RacGAP1, a cytokinesis-related Rho GTPase-activating protein, contributed to this process. Depletion of RacGAP1 with RacGAP1-targeting siRNA or overexpression of RacGAP1 mutant (T249A) attenuated melanoma cell transendothelial migration and concomitant changes of adherens junctions. In addition, RacGAP1 promoted the activations of RhoA, FAK, paxillin and triggered focal adhesion formation and cytoskeletal rearrangement. By overexpressing FAK-related non-kinase (FRNK) in endothelium, we showed that RacGAP1 mediated endothelial barrier function loss and melanoma transmigration in a focal adhesion-dependent manner. These results suggest that endothelial RacGAP1 may play critical roles in pathogenic processes of cancer by regulating endothelial permeability.

  15. DNA ligase III promotes alternative nonhomologous end-joining during chromosomal translocation formation.

    PubMed

    Simsek, Deniz; Brunet, Erika; Wong, Sunnie Yan-Wai; Katyal, Sachin; Gao, Yankun; McKinnon, Peter J; Lou, Jacqueline; Zhang, Lei; Li, James; Rebar, Edward J; Gregory, Philip D; Holmes, Michael C; Jasin, Maria

    2011-06-01

    Nonhomologous end-joining (NHEJ) is the primary DNA repair pathway thought to underlie chromosomal translocations and other genomic rearrangements in somatic cells. The canonical NHEJ pathway, including DNA ligase IV (Lig4), suppresses genomic instability and chromosomal translocations, leading to the notion that a poorly defined, alternative NHEJ (alt-NHEJ) pathway generates these rearrangements. Here, we investigate the DNA ligase requirement of chromosomal translocation formation in mouse cells. Mammals have two other DNA ligases, Lig1 and Lig3, in addition to Lig4. As deletion of Lig3 results in cellular lethality due to its requirement in mitochondria, we used recently developed cell lines deficient in nuclear Lig3 but rescued for mitochondrial DNA ligase activity. Further, zinc finger endonucleases were used to generate DNA breaks at endogenous loci to induce translocations. Unlike with Lig4 deficiency, which causes an increase in translocation frequency, translocations are reduced in frequency in the absence of Lig3. Residual translocations in Lig3-deficient cells do not show a bias toward use of pre-existing microhomology at the breakpoint junctions, unlike either wild-type or Lig4-deficient cells, consistent with the notion that alt-NHEJ is impaired with Lig3 loss. By contrast, Lig1 depletion in otherwise wild-type cells does not reduce translocations or affect microhomology use. However, translocations are further reduced in Lig3-deficient cells upon Lig1 knockdown, suggesting the existence of two alt-NHEJ pathways, one that is biased toward microhomology use and requires Lig3 and a back-up pathway which does not depend on microhomology and utilizes Lig1.

  16. Fibronectin promotes proplatelet formation in the human megakaryocytic cell line UT-7/TPO.

    PubMed

    Kawaguchi, Tatsuya; Hatano, Ryo; Yamaguchi, Kyoji; Nawa, Katsuhiko; Hashimoto, Ryuji; Yokota, Hiroshi

    2012-01-01

    We investigated PPF (proplatelet formation) in the human megakaryocytic cell line UT-7/TPO in vitro and signal transduction pathways responsible for PPF. The megakaryocytic cell lines are useful for studying megakaryocyte biology, although PPF is induced only in the presence of phorbol ester. TPO (thrombopoietin) stimulates megakaryocyte proliferation and differentiation; however, no PPF occurred in the megakaryocytic cell lines, even after the addition of TPO. Therefore, factors other than TPO may play an important role in the process of PPF. As PPF occurs in the bone marrow in vivo, we noted extracellular matrix proteins and found that soluble FN (fibronectin) induced potent PPF in UT-7/TPO without phorbol ester. A Western blot analysis showed that the expression of integrins was not increased by FN treatment. Anti-β1 antibody and the RGD (arginine-glycine-aspartate) peptide inhibited FN-induced PPF. This result indicates that the signal originated from integrin β1, which is essential to inducing PPF in UT-7/TPO. Results of the experiments using several inhibitors suggest that activation of the MEK [MAPK (mitogen-activated protein kinase)/ERK (extracellular-signal-regulated kinase) kinase]-ERK and PI3K (phosphoinositide 3-kinase) pathways are necessary for PPF. The phosphorylation of ERK gradually increased for 2 h after the addition of soluble FN, which suggests that activation of ERK is essential for the initial induction of FN-induced PPF in UT-7/TPO. UT-7/TPO is a useful cell line that enables us to study the signals of PPF without effects of chemical compounds.

  17. Effect of protein hydration on receptor conformation: decreased levels of bound water promote metarhodopsin II formation.

    PubMed

    Mitchell, D C; Litman, B J

    1999-06-15

    Neutral solutes were used to investigate the effects of osmotic stress both on the ability of rhodopsin to undergo its activating conformation change and on acyl chain packing in the rod outer segment (ROS) disk membrane. The equilibrium concentration of metarhodopsin II (MII), the conformation of photoactivated rhodopsin, which binds and activates transducin, was increased by glycerol, sucrose, and stachyose in a manner which was linear with osmolality. Analysis of this shift in equilibrium in terms of the dependence of ln(Keq) on osmolality revealed that 20 +/- 1 water molecules are released during the MI-to-MII transition at 20 degrees C, and at 35 degrees C 13 +/- 1 waters are released. At 35 degrees C the average time constant for MII formation was increased from 1.20 +/- 0.09 ms to 1.63 +/- 0.09 ms by addition of 1 osmolal sucrose or glycerol. The effect of the neutral solutes on acyl chain packing in the ROS disk membrane was assessed via measurements of the fluorescence lifetime and anisotropy decay of 1,6-diphenyl-1,3,5-hexatriene (DPH). Analysis of the anisotropy decay of DPH in terms of the rotational diffusion model showed that the angular width of the equilibrium orientational distribution of DPH about the membrane normal was progressively narrowed by increased osmolality. The parameter fv, which is proportional to the overlap between the DPH orientational probability distribution and a random orientational distribution, was reduced by the osmolytes in a manner which was linear with osmolality. This study highlights the potentially opposing interplay between the effect of membrane surface hydration on both the lipid bilayer and integral membrane protein structure. Our results further demonstrate that the binding and release of water molecules play an important role in modulating functional conformational changes for integral membrane proteins, as well as for soluble globular proteins.

  18. DNA Ligase III Promotes Alternative Nonhomologous End-Joining during Chromosomal Translocation Formation

    PubMed Central

    Wong, Sunnie Yan-Wai; Katyal, Sachin; Gao, Yankun; McKinnon, Peter J.; Lou, Jacqueline; Zhang, Lei; Li, James; Rebar, Edward J.; Gregory, Philip D.; Holmes, Michael C.; Jasin, Maria

    2011-01-01

    Nonhomologous end-joining (NHEJ) is the primary DNA repair pathway thought to underlie chromosomal translocations and other genomic rearrangements in somatic cells. The canonical NHEJ pathway, including DNA ligase IV (Lig4), suppresses genomic instability and chromosomal translocations, leading to the notion that a poorly defined, alternative NHEJ (alt-NHEJ) pathway generates these rearrangements. Here, we investigate the DNA ligase requirement of chromosomal translocation formation in mouse cells. Mammals have two other DNA ligases, Lig1 and Lig3, in addition to Lig4. As deletion of Lig3 results in cellular lethality due to its requirement in mitochondria, we used recently developed cell lines deficient in nuclear Lig3 but rescued for mitochondrial DNA ligase activity. Further, zinc finger endonucleases were used to generate DNA breaks at endogenous loci to induce translocations. Unlike with Lig4 deficiency, which causes an increase in translocation frequency, translocations are reduced in frequency in the absence of Lig3. Residual translocations in Lig3-deficient cells do not show a bias toward use of pre-existing microhomology at the breakpoint junctions, unlike either wild-type or Lig4-deficient cells, consistent with the notion that alt-NHEJ is impaired with Lig3 loss. By contrast, Lig1 depletion in otherwise wild-type cells does not reduce translocations or affect microhomology use. However, translocations are further reduced in Lig3-deficient cells upon Lig1 knockdown, suggesting the existence of two alt-NHEJ pathways, one that is biased toward microhomology use and requires Lig3 and a back-up pathway which does not depend on microhomology and utilizes Lig1. PMID:21655080

  19. Prenatal secondhand cigarette smoke promotes Th2 polarization and impairs goblet cell differentiation and airway mucus formation.

    PubMed

    Singh, Shashi P; Gundavarapu, Sravanthi; Peña-Philippides, Juan C; Rir-Sima-ah, Jules; Mishra, Neerad C; Wilder, Julie A; Langley, Raymond J; Smith, Kevin R; Sopori, Mohan L

    2011-11-01

    Parental, particularly maternal, smoking increases the risk for childhood allergic asthma and infection. Similarly, in a murine allergic asthma model, prenatal plus early postnatal exposure to secondhand cigarette smoke (SS) exacerbates airways hyperreactivity and Th2 responses in the lung. However, the mechanism and contribution of prenatal versus early postnatal SS exposure on allergic asthma remain unresolved. To identify the effects of prenatal and/or early postnatal SS on allergic asthma, BALB/c dams and their offspring were exposed gestationally and/or 8-10 wk postbirth to filtered air or SS. Prenatal, but not postnatal, SS strongly increased methacholine and allergen (Aspergillus)-induced airway resistance, Th2 cytokine levels, and atopy and activated the Th2-polarizing pathway GATA3/Lck/ERK1/2/STAT6. Either prenatal and/or early postnatal SS downregulated the Th1-specific transcription factor T-bet and, surprisingly, despite high levels of IL-4/IL-13, dramatically blocked the allergen-induced mucous cell metaplasia, airway mucus formation, and the expression of mucus-related genes/proteins: Muc5ac, γ-aminobutyric acid A receptors, and SAM pointed domain-containing Ets-like factor. Given that SS/nicotine exposure of normal adult mice promotes mucus formation, the results suggested that fetal and neonatal lung are highly sensitive to cigarette smoke. Thus, although the gestational SS promotes Th2 polarization/allergic asthma, it may also impair and/or delay the development of fetal and neonatal lung, affecting mucociliary clearance and Th1 responses. Together, this may explain the increased susceptibility of children from smoking parents to allergic asthma and childhood respiratory infections.

  20. Pyrrolidine and Piperidine Formation Via Copper(II) Carboxylate Promoted Intramolecular Carboamination of Unactivated Olefins: Diastereoselectivity and Mechanism

    PubMed Central

    Sherman, Eric S.; Fuller, Peter H.; Kasi, Dhanalakshmi; Chemler, Sherry R.

    2008-01-01

    An expanded substrate scope and in depth analysis of the reaction mechanism of the copper(II) carboxylate promoted intramolecular carboamination of unactivated alkenes is described. This method provides access to N-functionalized pyrrolidines and piperidines. Both aromatic and aliphatic γ- and δ-alkenyl N-arylsulfonamides undergo the oxidative cyclization reaction efficiently. N-Benzoyl-2-allylaniline also underwent the oxidative cyclization. The terminal olefin substrates examined were more reactive than those with internal olefins, and the latter terminated in elimination rather than carbon-carbon bond formation. The efficiency of the reaction was enhanced by the use of more organic soluble copper(II) carboxylate salts, copper(II) neodecanoate in particular. The reaction times were reduced by the use of microwave heating. High levels of diastereoselectivity were observed in the synthesis of 2,5-disubstituted pyrrolidines, wherein the cis substitution pattern predominates. The mechanism of the reaction is discussed in the context of the observed reactivity and in comparison to analogous reactions promoted by other reagents and conditions. Our evidence supports a mechanism wherein the N-C bond is formed via intramolecular syn aminocupration and the C-C bond is formed via intramolecular addition of a primary carbon radical to an aromatic ring. PMID:17428100

  1. Silicon Promotes Exodermal Casparian Band Formation in Si-Accumulating and Si-Excluding Species by Forming Phenol Complexes

    PubMed Central

    Hinrichs, Martin; Specht, André; Waßmann, Friedrich; Schreiber, Lukas; Schenk, Manfred K.

    2015-01-01

    We studied the effect of Silicon (Si) on Casparian band (CB) development, chemical composition of the exodermal CB and Si deposition across the root in the Si accumulators rice and maize and the Si non-accumulator onion. Plants were cultivated in nutrient solution with and without Si supply. The CB development was determined in stained root cross-sections. The outer part of the roots containing the exodermis was isolated after enzymatic treatment. The exodermal suberin was transesterified with MeOH/BF3 and the chemical composition was measured using gas chromatography-mass spectroscopy (GC-MS) and flame ionization detector (GC-FID). Laser ablation-inductively coupled plasma-mass spectroscopy (LA-ICP-MS) was used to determine the Si deposition across root cross sections. Si promoted CB formation in the roots of Si-accumulator and Si non-accumulator species. The exodermal suberin was decreased in rice and maize due to decreased amounts of aromatic suberin fractions. Si did not affect the concentration of lignin and lignin-like polymers in the outer part of rice, maize and onion roots. The highest Si depositions were found in the tissues containing CB. These data along with literature were used to suggest a mechanism how Si promotes the CB development by forming complexes with phenols. PMID:26383862

  2. Polygonatum sibiricum polysaccharide inhibits osteoporosis by promoting osteoblast formation and blocking osteoclastogenesis through Wnt/β-catenin signalling pathway

    PubMed Central

    Du, Li; Nong, Meng-Ni; Zhao, Jin-Min; Peng, Xiao-Ming; Zong, Shao-Hui; Zeng, Gao-Feng

    2016-01-01

    Bone homeostasis is maintained by a balance between bone formation by osteoblasts and bone resorption by osteoclasts. Osteoporosis occurs when osteoclast activity surpasses osteoblast activity. Our previous studies showed the plant-derived natural polysaccharide (Polygonatum sibiricum polysaccharide or PSP) had significant anti-ovariectomy (OVX)-induced osteoporosis effects in vivo, but the mechanisms of PSP’s anti-osteoporosis effect remains unclear. In this study, we assessed PSP’s effect on the generation of osteoblast and osteoclast in vitro. This study showed that PSP promoted the osteogenic differentiation of mouse bone marrow stromal cells (BMSCs) without affecting BMPs signaling pathway. This effect was due to the increased nuclear accumulation of β-catenin, resulting in a higher expression of osteoblast-related genes. Furthermore, the study showed PSP could inhibit the receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclastogenesis and exert prophylatic protection against LPS-induced osteolysis in vivo. This effect was also related to the increased nuclear accumulation of β-catenin, resulting in the decreased expression of osteoclast-related genes. In conclusion, our results showed that PSP effectively promoted the osteogenic differentiation of mouse BMSCs and suppressed osteoclastogenesis; therefore, it could be used to treat osteoporosis. PMID:27554324

  3. Thermomineral water promotes axonal sprouting but does not reduce glial scar formation in a mouse model of spinal cord injury

    PubMed Central

    Aleksić, Dubravka; Aksić, Milan; Divac, Nevena; Radonjić, Vidosava; Filipović, Branislav; Jakovčevski, Igor

    2014-01-01

    Thermomineral water from the Atomic Spa Gornja Trepča has been used for a century in the treatment of neurologic disease. The thermomineral water contains microelements, including lithium and magnesium, which show neural regeneration-promoting effects after central nervous system injury. In this study, we investigated the effects of oral intake of thermomineral water from the Atomic Spa Gornja Trepča on nerve regeneration in a 3-month-old mouse model of spinal cord injury. The mice receiving oral intake of thermomineral water showed better locomotor recovery than those without administration of thermomineral water at 8 and 12 weeks after lower thoracic spinal cord compression. At 12 weeks after injury, sprouting of catecholaminergic axons was better in mice that drank thermomineral water than in those without administration of thermomineral water, but there was no difference in glial reaction to injury between mice with and without administration of thermomineral water. These findings suggest that thermomineral water can promote the nerve regeneration but cannot reduce glial scar formation in a mouse model of spinal cord injury. PMID:25657739

  4. The glabra1 mutation affects cuticle formation and plant responses to microbes.

    PubMed

    Xia, Ye; Yu, Keshun; Navarre, Duroy; Seebold, Kenneth; Kachroo, Aardra; Kachroo, Pradeep

    2010-10-01

    Systemic acquired resistance (SAR) is a form of defense that provides resistance against a broad spectrum of pathogens in plants. Previous work indicates a role for plastidial glycerolipid biosynthesis in SAR. Specifically, mutations in FATTY ACID DESATURASE7 (FAD7), which lead to reduced trienoic fatty acid levels and compromised plastidial lipid biosynthesis, have been associated with defective SAR. We show that the defective SAR in Arabidopsis (Arabidopsis thaliana) fad7-1 plants is not associated with a mutation in FAD7 but rather with a second-site mutation in GLABRA1 (GL1), a gene well known for its role in trichome formation. The compromised SAR in gl1 plants is associated with impairment in their cuticles. Furthermore, mutations in two other components of trichome development, GL3 and TRANSPARENT TESTA GLABRA1, also impaired cuticle development and SAR. This suggests an overlap in the biochemical pathways leading to cuticle and trichome development. Interestingly, exogenous application of gibberellic acid (GA) not only enhanced SAR in wild-type plants but also restored SAR in gl1 plants. In contrast to GA, the defense phytohoromes salicylic acid and jasmonic acid were unable to restore SAR in gl1 plants. GA application increased levels of cuticular components but not trichome formation on gl1 plants, thus implicating cuticle, but not trichomes, as an important component of SAR. Our findings question the prudence of using mutant backgrounds for genetic screens and underscore a need to reevaluate phenotypes previously studied in the gl1 background.

  5. Lewis acid promoted titanium alkylidene formation: off-cycle intermediates relevant to olefin trimerization catalysis.

    PubMed

    Sattler, Aaron; VanderVelde, David G; Labinger, Jay A; Bercaw, John E

    2014-07-30

    Two new precatalysts for ethylene and α-olefin trimerization, (FI)Ti(CH2SiMe3)2Me and (FI)Ti(CH2CMe3)2Me (FI = phenoxy-imine), have been synthesized and structurally characterized by X-ray diffraction. (FI)Ti(CH2SiMe3)2Me can be activated with 1 equiv of B(C6F5)3 at room temperature to give the solvent-separated ion pair [(FI)Ti(CH2SiMe3)2][MeB(C6F5)3], which catalytically trimerizes ethylene or 1-pentene to produce 1-hexene or C15 olefins, respectively. The neopentyl analogue (FI)Ti(CH2CMe3)2Me is unstable toward activation with B(C6F5)3 at room temperature, giving no discernible diamagnetic titanium complexes, but at -30 °C the following can be observed by NMR spectroscopy: (i) formation of the bis-neopentyl cation [(FI)Ti(CH2CMe3)2](+), (ii) α-elimination of neopentane to give the neopentylidene complex [(FI)Ti(═CHCMe3)](+), and (iii) subsequent conversion to the imido-olefin complex [(MeOAr2N═)Ti(OArHC═CHCMe3)](+) via an intramolecular metathesis reaction with the imine fragment of the (FI) ligand. If the reaction is carried out at low temperature in the presence of ethylene, catalytic production of 1-hexene is observed, in addition to the titanacyclobutane complex [(FI)Ti(CH(CMe3)CH2CH2)](+), resulting from addition of ethylene to the neopentylidene [(FI)Ti(═CHCMe3)](+). None of the complexes observed spectroscopically subsequent to [(FI)Ti(CH2CMe3)2](+) is an intermediate or precursor for ethylene trimerization, but notwithstanding these off-cycle pathways, [(FI)Ti(CH2CMe3)2](+) is a precatalyst that undergoes rapid initiation to generate a catalyst for trimerizing ethylene or 1-pentene.

  6. aPKC phosphorylates JAM-A at Ser285 to promote cell contact maturation and tight junction formation.

    PubMed

    Iden, Sandra; Misselwitz, Steve; Peddibhotla, Swetha S D; Tuncay, Hüseyin; Rehder, Daniela; Gerke, Volker; Robenek, Horst; Suzuki, Atsushi; Ebnet, Klaus

    2012-03-05

    The PAR-3-atypical protein kinase C (aPKC)-PAR-6 complex has been implicated in the development of apicobasal polarity and the formation of tight junctions (TJs) in vertebrate epithelial cells. It is recruited by junctional adhesion molecule A (JAM-A) to primordial junctions where aPKC is activated by Rho family small guanosine triphosphatases. In this paper, we show that aPKC can interact directly with JAM-A in a PAR-3-independent manner. Upon recruitment to primordial junctions, aPKC phosphorylates JAM-A at S285 to promote the maturation of immature cell-cell contacts. In fully polarized cells, S285-phosphorylated JAM-A is localized exclusively at the TJs, and S285 phosphorylation of JAM-A is required for the development of a functional epithelial barrier. Protein phosphatase 2A dephosphorylates JAM-A at S285, suggesting that it antagonizes the activity of aPKC. Expression of nonphosphorylatable JAM-A/S285A interferes with single lumen specification during cyst development in three-dimensional culture. Our data suggest that aPKC phosphorylates JAM-A at S285 to regulate cell-cell contact maturation, TJ formation, and single lumen specification.

  7. Collagen-nanofiber hydrogel composites promote contact guidance of human lymphatic microvascular endothelial cells and directed capillary tube formation.

    PubMed

    Laco, Filip; Grant, M Helen; Black, Richard A

    2013-06-01

    Collagen and fibronectin matrices are known to stimulate migration of microvascular endothelial cells and the process of tubulogenesis, but the physical, chemical, and topographical cues for directed vessel formation have yet to be determined. In this study, growth, migration, elongation, and tube formation of human lymphatic microvascular endothelial cells (LECs) were investigated on electrospun poly(D,L-lactic-co-glycolic acid) (PLGA) and poly(L-lactic-co-D-lactic acid) (PLDL) nanofiber-coated substrates, and correlated with fiber density and diameter. Directed migration of LECs was observed in the presence of aligned nanofibers, whereas random fiber alignment slowed down migration and growth of LECs. Cell guidance was significantly enhanced in the presence of more hydrophobic PLDL polymer nanofibers compared to PLGA (10:90). Subsequent experiments with tube-forming assays reveal the ability of resorbable hydrophobic nanofibers >300 nm in diameter to promote cell guidance in collagen gels without direct cell-fiber contact, in contrast to the previously reported contact-guidance phenomena. Our results show that endothelial cell guidance is possible within nanofiber/collagen-gel constructs that mimic the native extracellular matrix in terms of size and orientation of fibrillar components.

  8. Formation of 100S ribosomes in Staphylococcus aureus by the hibernation promoting factor homolog SaHPF.

    PubMed

    Ueta, Masami; Wada, Chieko; Wada, Akira

    2010-01-01

    In the stationary growth phase of Escherichia coli, the 70S ribosomes are dimerized by the ribosome modulation factor (RMF) and hibernation promoting factor (HPF) proteins to form 100S ribosomes, which lose translational activity. In this study we found 100S ribosomes in the gram-positive bacterium Staphylococcus aureus, which has an HPF homolog (named SaHPF) but no RMF homolog. Unlike in E. coli, 100S ribosomes exist in all growth phases of S. aureus, with the highest levels at the transition from the exponential phase to the stationary phase. To find the key factors involved in 100S formation, we analyzed proteins associated with crude ribosomes using radical-free and highly reducing 2-D PAGE and MALDI TOF/MS. Only the SaHPF levels changed in parallel with the changes in 100S levels. SaHPF bound preferentially to 70S components in 100S ribosomes, with a molar ratio of 1 : 1 relative to the 70S, but some SaHPF was also detected in free 70S ribosomes. High-salt washing of the crude ribosomes released SaHPF and dissociated the 100S ribosomes to their 70S components. When these 70S components were incubated with purified SaHPF in vitro, they re-associated to form 100S. These results suggest that SaHPF is a key protein involved in 100S ribosome formation in S. aureus.

  9. Pulsed electromagnetic fields partially preserve bone mass, microarchitecture, and strength by promoting bone formation in hindlimb-suspended rats.

    PubMed

    Jing, Da; Cai, Jing; Wu, Yan; Shen, Guanghao; Li, Feijiang; Xu, Qiaoling; Xie, Kangning; Tang, Chi; Liu, Juan; Guo, Wei; Wu, Xiaoming; Jiang, Maogang; Luo, Erping

    2014-10-01

    A large body of evidence indicates that pulsed electromagnetic fields (PEMF), as a safe and noninvasive method, could promote in vivo and in vitro osteogenesis. Thus far, the effects and underlying mechanisms of PEMF on disuse osteopenia and/or osteoporosis remain poorly understood. Herein, the efficiency of PEMF on osteoporotic bone microarchitecture, bone strength, and bone metabolism, together with its associated signaling pathway mechanism, was systematically investigated in hindlimb-unloaded (HU) rats. Thirty young mature (3-month-old), male Sprague-Dawley rats were equally assigned to control, HU, and HU + PEMF groups. The HU + PEMF group was subjected to daily 2-hour PEMF exposure at 15 Hz, 2.4 mT. After 4 weeks, micro-computed tomography (µCT) results showed that PEMF ameliorated the deterioration of trabecular and cortical bone microarchitecture. Three-point bending test showed that PEMF mitigated HU-induced reduction in femoral mechanical properties, including maximum load, stiffness, and elastic modulus. Moreover, PEMF increased serum bone formation markers, including osteocalcin (OC) and N-terminal propeptide of type 1 procollagen (P1NP); nevertheless, PEMF exerted minor inhibitory effects on bone resorption markers, including C-terminal crosslinked telopeptides of type I collagen (CTX-I) and tartrate-resistant acid phosphatase 5b (TRAcP5b). Bone histomorphometric analysis demonstrated that PEMF increased mineral apposition rate, bone formation rate, and osteoblast numbers in cancellous bone, but PEMF caused no obvious changes on osteoclast numbers. Real-time PCR showed that PEMF promoted tibial gene expressions of Wnt1, LRP5, β-catenin, OPG, and OC, but did not alter RANKL, RANK, or Sost mRNA levels. Moreover, the inhibitory effects of PEMF on disuse-induced osteopenia were further confirmed in 8-month-old mature adult HU rats. Together, these results demonstrate that PEMF alleviated disuse-induced bone loss by promoting skeletal anabolic activities

  10. Hydathode trichomes actively secreting water from leaves play a key role in the physiology and evolution of root-parasitic rhinanthoid Orobanchaceae

    PubMed Central

    Světlíková, Petra; Hájek, Tomáš; Těšitel, Jakub

    2015-01-01

    Background and Aims Root hemiparasites from the rhinanthoid clade of Orobanchaceae possess metabolically active glandular trichomes that have been suggested to function as hydathode trichomes actively secreting water, a process that may facilitate resource acquisition from the host plant’s root xylem. However, no direct evidence relating the trichomes to water secretion exists, and carbon budgets associated with this energy-demanding process have not been determined. Methods Macro- and microscopic observations of the leaves of hemiparasitic Rhinanthus alectorolophus were conducted and night-time gas exchange was measured. Correlations were examined among the intensity of guttation, respiration and transpiration, and analysis of these correlations allowed the carbon budget of the trichome activity to be quantified. We examined the intensity of guttation, respiration and transpiration, correlations among which indicate active water secretion. Key Results Guttation was observed on the leaves of 50 % of the young, non-flowering plants that were examined, and microscopic observations revealed water secretion from the glandular trichomes present on the abaxial leaf side. Night-time rates of respiration and transpiration and the presence of guttation drops were positively correlated, which is a clear indicator of hydathode trichome activity. Subsequent physiological measurements on older, flowering plants indicated neither intense guttation nor the presence of correlations, which suggests that the peak activity of hydathodes is in the juvenile stage. Conclusions This study provides the first unequivocal evidence for the physiological role of the hydathode trichomes in active water secretion in the rhinanthoid Orobanchaceae. Depending on the concentration of organic elements calculated to be in the host xylem sap, the direct effect of water secretion on carbon balance ranges from close to neutral to positive. However, it is likely to be positive in the xylem-only feeding

  11. The Listeria monocytogenes Hibernation-Promoting Factor Is Required for the Formation of 100S Ribosomes, Optimal Fitness, and Pathogenesis

    PubMed Central

    Kline, Benjamin C.; McKay, Susannah L.; Tang, William W.

    2014-01-01

    During exposure to certain stresses, bacteria dimerize pairs of 70S ribosomes into translationally silent 100S particles in a process called ribosome hibernation. Although the biological roles of ribosome hibernation are not completely understood, this process appears to represent a conserved and adaptive response that contributes to optimal survival during stress and post-exponential-phase growth. Hibernating ribosomes are formed by the activity of one or more highly conserved proteins; gammaproteobacteria produce two relevant proteins, ribosome modulation factor (RMF) and hibernation promoting factor (HPF), while most Gram-positive bacteria produce a single, longer HPF protein. Here, we report the formation of 100S ribosomes by an HPF homolog in Listeria monocytogenes. L. monocytogenes 100S ribosomes were observed by sucrose density gradient centrifugation of bacterial extracts during mid-logarithmic phase, peaked at the transition to stationary phase, and persisted at lower levels during post-exponential-phase growth. 100S ribosomes were undetectable in bacteria carrying an hpf::Himar1 transposon insertion, indicating that HPF is required for ribosome hibernation in L. monocytogenes. Additionally, epitope-tagged HPF cosedimented with 100S ribosomes, supporting its previously described direct role in 100S formation. We examined hpf mRNA by quantitative PCR (qPCR) and identified several conditions that upregulated its expression, including carbon starvation, heat shock, and exposure to high concentrations of salt or ethanol. Survival of HPF-deficient bacteria was impaired under certain conditions both in vitro and during animal infection, providing evidence for the biological relevance of 100S ribosome formation. PMID:25422304

  12. The Listeria monocytogenes hibernation-promoting factor is required for the formation of 100S ribosomes, optimal fitness, and pathogenesis.

    PubMed

    Kline, Benjamin C; McKay, Susannah L; Tang, William W; Portnoy, Daniel A

    2015-02-01

    During exposure to certain stresses, bacteria dimerize pairs of 70S ribosomes into translationally silent 100S particles in a process called ribosome hibernation. Although the biological roles of ribosome hibernation are not completely understood, this process appears to represent a conserved and adaptive response that contributes to optimal survival during stress and post-exponential-phase growth. Hibernating ribosomes are formed by the activity of one or more highly conserved proteins; gammaproteobacteria produce two relevant proteins, ribosome modulation factor (RMF) and hibernation promoting factor (HPF), while most Gram-positive bacteria produce a single, longer HPF protein. Here, we report the formation of 100S ribosomes by an HPF homolog in Listeria monocytogenes. L. monocytogenes 100S ribosomes were observed by sucrose density gradient centrifugation of bacterial extracts during mid-logarithmic phase, peaked at the transition to stationary phase, and persisted at lower levels during post-exponential-phase growth. 100S ribosomes were undetectable in bacteria carrying an hpf::Himar1 transposon insertion, indicating that HPF is required for ribosome hibernation in L. monocytogenes. Additionally, epitope-tagged HPF cosedimented with 100S ribosomes, supporting its previously described direct role in 100S formation. We examined hpf mRNA by quantitative PCR (qPCR) and identified several conditions that upregulated its expression, including carbon starvation, heat shock, and exposure to high concentrations of salt or ethanol. Survival of HPF-deficient bacteria was impaired under certain conditions both in vitro and during animal infection, providing evidence for the biological relevance of 100S ribosome formation.

  13. Solcoseryl, a tissue respiration stimulating agent, significantly enhances the effect of capacitively coupled electric field on the promotion of bone formation around dental implants.

    PubMed

    Ochi, Morio; Wang, Pao-Li; Ohura, Kiyoshi; Takashima, Shigenori; Kagami, Hiroyuki; Hirose, Yukito; Kaku, Tohru; Sakaguchi, Kunihiko

    2003-06-01

    In the present study we examined the combined effect of application of a capacitively coupled electric field (CCEF) and the tissue respiration stimulating agent, Solcoseryl, on the promotion of bone formation around dental implants histologically and mechanically. After a dental implant was inserted into each femur of Japanese white rabbits, Solcoseryl (2 ml/kg) was administered intravenously in the ear vein and a CCEF was applied for 4 h per day for 14 days. The degree of bone formation on microscopic observation, bone contact ratio, bone surface area ratio, and the level of removal torque of the implant in the Solcoseryl- and CCEF-treated group were significantly higher than the respective value in the control group, which had not been treated with Solcoseryl nor CCEF. Thus, the combination of CCEF stimulation and Solcoseryl effectively promoted the formation of new bone. It is suggested that the clinical use of a combination of CCEF stimulation and Solcoseryl for dental implants promotes osseointegration.

  14. Human Choline Kinase-α Promotes Hepatitis C Virus RNA Replication through Modulation of Membranous Viral Replication Complex Formation

    PubMed Central

    Wong, Mun-Teng

    2016-01-01

    ABSTRACT Hepatitis C virus (HCV) infection reorganizes cellular membranes to create an active viral replication site named the membranous web (MW). The role that human choline kinase-α (hCKα) plays in HCV replication remains elusive. Here, we first showed that hCKα activity, not the CDP-choline pathway, promoted viral RNA replication. Confocal microscopy and subcellular fractionation of HCV-infected cells revealed that a small fraction of hCKα colocalized with the viral replication complex (RC) on the endoplasmic reticulum (ER) and that HCV infection increased hCKα localization to the ER. In the pTM-NS3-NS5B model, NS3-NS5B expression increased the localization of the wild-type, not the inactive D288A mutant, hCKα on the ER, and hCKα activity was required for effective trafficking of hCKα and NS5A to the ER. Coimmunoprecipitation showed that hCKα was recruited onto the viral RC presumably through its binding to NS5A domain 1 (D1). hCKα silencing or treatment with CK37, an hCKα activity inhibitor, abolished HCV-induced MW formation. In addition, hCKα depletion hindered NS5A localization on the ER, interfered with NS5A and NS5B colocalization, and mitigated NS5A-NS5B interactions but had no apparent effect on NS5A-NS4B and NS4B-NS5B interactions. Nevertheless, hCKα activity was not essential for the binding of NS5A to hCKα or NS5B. These findings demonstrate that hCKα forms a complex with NS5A and that hCKα activity enhances the targeting of the complex to the ER, where hCKα protein, not activity, mediates NS5A binding to NS5B, thereby promoting functional membranous viral RC assembly and viral RNA replication. IMPORTANCE HCV infection reorganizes the cellular membrane to create an active viral replication site named the membranous web (MW). Here, we report that human choline kinase-α (hCKα) acts as an essential host factor for HCV RNA replication. A fraction of hCKα colocalizes with the viral replication complex (RC) on the endoplasmic reticulum

  15. Ultrastructure of hydathode trichomes of hemiparasitic Rhinanthus alectorolophus and Odontites vernus: how important is their role in physiology and evolution of parasitism in Orobanchaceae?

    PubMed

    Těšitel, J; Tesařová, M

    2013-01-01

    The Rhinanthoid clade of the family Orobanchaceae comprises plants displaying a hemiparasitic or holoparasitic strategy of resource acquisition. Some of its species (mainly Rhinanthus spp.) are often used as models for studies of hemiparasite physiology. Although there is a well-developed concept covering their physiological processes, most recent studies have neglected the existence of hydathode trichomes present on leaves of these hemiparasitic plants. As a first step for the proposed integration of these structures in the theory of physiological processes of the hemiparasites, we described the outer micromorphology and ultrastructure of the hydathode trichomes on leaves of hemiparasitic Rhinanthus alectorolophus and Odontites vernus with scanning and transmission electron microscopy (SEM and TEM, respectively). The TEM inspections of both types of trichome revealed typical ultrastructural features: labyrinthine cell wall, high content of cytoplasm in cells with numerous mitochondria and presence of plasmodesmata. All these features indicate high metabolic activity complying with their function as glandular trichomes actively secreting water. The active secretion of water by the hydathode trichomes (evidence for which is summarised here) also presents a possible mechanism explaining results of previous gas exchange measurements detecting high dark respiration and transpiration rates and a tight inter-correlation between them in hemiparasitic Orobanchaceae. In addition, this process is hypothesised to have allowed multiple evolutionary transitions from facultative to obligate hemiparasitism and unique xylem-feeding holoparasitism of Lathraea with a long-lived underground stage featuring a rhizome covered by scales of leaf origin.

  16. Promoted reduction of tellurite and formation of extracellular tellurium nanorods by concerted reaction between iron and Shewanella oneidensis MR-1.

    PubMed

    Kim, Dong-Hun; Kim, Min-Gyu; Jiang, Shenghua; Lee, Ji-Hoon; Hur, Hor-Gil

    2013-08-06

    The reduction of tellurite (Te(IV)) by dissimilatory metal reducing bacterium, Shewanella oneidensis MR-1, was promoted in the presence of Fe(III) in comparison with Te(IV) bioreduction in the absence of Fe(III). Electron microscopic analyses revealed that iron promoted Te(IV) reduction led to form exclusively extracellular crystalline Te(0) nanorods, as compared to the mostly intracellular formation of Te(0) nanorods in the absence of Fe(III). The Te K-edge X-ray absorption spectrometric analyses demonstrated that S. oneidensis MR-1 in the presence of Fe(III) reduced Te(IV) to less harmful metallic Te(0) nanorods through the precipitation of tellurite (Te(IV)Ox) complex by the bacterial respiration of Fe(III) to Fe(II) under anaerobic conditions. However, Fe(II) ion itself was only able to precipitate the solid tellurite (Te(IV)Ox) complex from the Te(IV) solution, which was not further reduced to Te(0). The results clearly indicated that bacterial S. oneidensis MR-1 plays important roles in the reduction and crystallization of Te(0) nanorods by as yet undetermined biochemical mechanisms. As compared to the slow bacterial Te(IV) reduction in the absence of Fe(III), the rapid reduction of Te(IV) to Te(0) by the concerted biogeochemical reaction between Fe(II) and S. oneidensis MR-1 could be applied for the sequestration and detoxification of Te(IV) in the environments as well as for the preparation of extracellular Te(0) nanorod structures.

  17. Vascular mimicry formation is promoted by paracrine TGF-β and SDF1 of cancer-associated fibroblasts and inhibited by miR-101 in hepatocellular carcinoma.

    PubMed

    Yang, Jine; Lu, Yang; Lin, Ying-Ying; Zheng, Zhi-Yuan; Fang, Jian-Hong; He, Shuai; Zhuang, Shi-Mei

    2016-12-01

    Vascular mimicry (VM) describes the phenomenon that tumor cells but not endothelial cells form vascular-like channels, which provide blood perfusion for tumor tissues. VM is associated with tumor growth, metastasis and worse survival of different cancers. The mechanisms of VM formation remain largely unknown. We showed that the conditioned medium of cancer-associated fibroblast (CM-CAF) promoted tumor cells to form capillary-like structure in vitro. Consistently, co-implantation of CAFs with tumor cells significantly enhanced VM formation in mouse xenografts, and higher amount of CAFs was found in VM(+) human HCC tissues compared to VM(-) ones. However, the CM-CAF-promoted VM formation was attenuated when TGF-β or SDF1 signaling was abrogated. Similar to CM-CAF, recombinant TGF-β1 and SDF1 induced VM formation. We further disclosed that the CAF-secreted TGF-β and SDF1 enhanced the expression of VE-cadherin, MMP2 and laminin5γ2 via TGF-βR1 and CXCR4 in tumor cells, thereby promoted VM formation. Moreover, tumor cells with high activity of self-sustaining TGF-β signaling displayed strong capability of VM formation. Subsequent investigations showed that miR-101, which was down-regulated in both tumor cells and CAFs, suppressed the CAF-promoted VM formation in vitro and in vivo. Gain- and loss-of-function analyses revealed that miR-101 attenuated TGF-β signaling transduction by targeting TGF-βR1 and Smad2 in tumor cells, and simultaneously abrogated SDF1 signaling by suppressing SDF1 expression in CAFs and inhibiting VE-cadherin expression in tumor cells. Our findings suggest that the miR-101-TGF-β/SDF1-VE-cadherin/MMP2/LAMC2 networks regulate VM formation and represent the potential targets for cancer therapy.

  18. Shoot bending promotes flower bud formation by miRNA-mediated regulation in apple (Malus domestica Borkh.).

    PubMed

    Xing, Libo; Zhang, Dong; Zhao, Caiping; Li, Youmei; Ma, Juanjuan; An, Na; Han, Mingyu

    2016-02-01

    Flower induction in apple (Malus domestica Borkh.) trees plays an important life cycle role, but young trees produce fewer and inferior quality flower buds. Therefore, shoot bending has become an important cultural practice, significantly promoting the capacity to develop more flower buds during the growing seasons. Additionally, microRNAs (miRNAs) play essential roles in plant growth, flower induction and stress responses. In this study, we identified miRNAs potentially involved in the regulation of bud growth, and flower induction and development, as well as in the response to shoot bending. Of the 195 miRNAs identified, 137 were novel miRNAs. The miRNA expression profiles revealed that the expression levels of 68 and 27 known miRNAs were down-regulated and up-regulated, respectively, in response to shoot bending, and that the 31 differentially expressed novel miRNAs between them formed five major clusters. Additionally, a complex regulatory network associated with auxin, cytokinin, abscisic acid (ABA) and gibberellic acid (GA) plays important roles in cell division, bud growth and flower induction, in which related miRNAs and targets mediated regulation. Among them, miR396, 160, 393, and their targets associated with AUX, miR159, 319, 164, and their targets associated with ABA and GA, and flowering-related miRNAs and genes, regulate bud growth and flower bud formation in response to shoot bending. Meanwhile, the flowering genes had significantly higher expression levels during shoot bending, suggesting that they are involved in this regulatory process. This study provides a framework for the future analysis of miRNAs associated with multiple hormones and their roles in the regulation of bud growth, and flower induction and formation in response to shoot bending in apple trees.

  19. GhAGL15s, preferentially expressed during somatic embryogenesis, promote embryogenic callus formation in cotton (Gossypium hirsutum L.).

    PubMed

    Yang, Zuoren; Li, Changfeng; Wang, Ye; Zhang, Chaojun; Wu, Zhixia; Zhang, Xueyan; Liu, Chuanliang; Li, Fuguang

    2014-10-01

    Somatic embryogenesis is a useful tool for gene transfer and propagation of plants. AGAMOUS-LIKE15 (AGL15) promotes somatic embryogenesis in many plant species. In this study, three homologous AGL15 genes were isolated from Gossypium hirsutum L., namely GhAGL15-1, GhAGL15-3, and GhAGL15-4. Their putative proteins contained a highly conserved MADS-box DNA-binding domain and a less conserved K domain. Phylogenetic analysis suggested that the three GhAGL15s clustered most closely with AGL15 proteins in other plants. Subcellular location analyses revealed that three GhAGL15s were localized in the nucleus. Furthermore, their expression levels increased following embryogenic callus induction, but sharply decreased during the embryoid stage. GhAGL15-1 and GhAGL15-3 were significantly induced by 2,4-D and kinetin, whereas GhAGL15-4 was only responsive to 2,4-D treatment. Over-expression of the three GhAGL15s in cotton callus improved callus quality and significantly increased the embryogenic callus formation rate, while GhAGL15-4 had the highest positive effect on the embryogenic callus formation rate (an increase from 38.1 to 65.2%). These results suggest that over-expression of GhAGL15s enhances embryogenic potential of transgenic calli. Therefore, spatiotemporal manipulation of GhAGL15s expression may prove valuable in improving cotton transformation efficiency.

  20. Foliar trichome- and aquaporin-aided water uptake in a drought-resistant epiphyte Tillandsia ionantha Planchon.

    PubMed

    Ohrui, T; Nobira, H; Sakata, Y; Taji, T; Yamamoto, C; Nishida, K; Yamakawa, T; Sasuga, Y; Yaguchi, Y; Takenaga, H; Tanaka, Shigeo

    2007-12-01

    The atmospheric epiphyte Tillandsia ionantha is capable of surviving drought stress for 6 months or more without any exogenous water supply via an as of yet to be determined mechanism. When plants were soaked in water for 3 h, leaves absorbed a remarkably large amount of water (30-40% on the basis of fresh weight), exhibiting a bimodal absorption pattern. Radiolabeled water was taken up by the leaves by capillary action of the epidermal trichomes within 1 min (phase 1) and then transported intracellularly to leaf tissues over 3 h (phase 2). The removal of epidermal trichome wings from leaves as well as rinsing leaves with water significantly lowered the extracellular accumulation of water on leaf surfaces. The intracellular transport of water was inhibited by mercuric chloride, implicating the involvement of a water channel aquaporin in second-phase water absorption. Four cDNA clones (TiPIP1a, TiPIP1b, TiPIP1c, and TiPIP2a) homologous to PIP family aquaporins were isolated from the leaves, and RT-PCR showed that soaking plants in water stimulated the expression of TiPIP2a mRNA, suggesting the reinforcement in ability to rapidly absorb a large amount of water. The expression of TiPIP2a complementary RNA in Xenopus oocytes enhanced permeability, and treatment with inhibitors suggested that the water channel activity of TiPIP2a protein was regulated by phosphorylation. Thus, the high water uptake capability of T. ionantha leaves surviving drought is attributable to a bimodal trichome- and aquaporin-aided water uptake system based on rapid physical collection of water and subsequent, sustained chemical absorption.

  1. The Function of the Early Trichomes Gene in Arabidopsis and Maize.

    SciTech Connect

    Scott Poethig

    2011-12-05

    Lateral organ polarity in Arabidopsis is regulated by antagonistic interactions between genes that promote either adaxial or abaxial identity, but the molecular basis of this interaction is largely unknown. We show that the adaxial regulator ASYMMETRIC LEAVES2 (AS2) is a direct target of the abaxial regulator KANADI1 (KAN1), and that KAN1 represses the transcription of AS2 in abaxial cells. Mutation of a single nucleotide in a KAN1 binding site in the AS2 promoter causes AS2 to be ectopically expressed in abaxial cells, resulting in a dominant, adaxialized phenotype. We also show that the abaxial expression of KAN1 is mediated directly or indirectly by AS2. These results demonstrate that KAN1 acts as a transcriptional repressor and that mutually repressive interactions between KAN1 and AS2 contribute to the establishment of adaxial-abaxial polarity in plants. A screen for mutations that affect the expression of a GFP reporter for KANADI2 produced mutations in two genes, CENTER CITY (CCT) and GRAND CENTRAL (GCT). Mutations in GCT and CCT delay the specification of central and peripheral identity and the globular-to-heart transition, but have little or no effect on the initial growth rate of the embryo. Mutant embryos eventually recover and undergo relatively normal patterning, albeit at an inappropriate size. GCT and CCT were identified as the Arabidopsis orthologs of MED12 and MED13--evolutionarily conserved proteins that act in association with the Mediator complex to negatively regulate transcription. The predicted function of these proteins combined with the effect of gct and cct on embryo development suggests that MED12 and MED13 regulate pattern formation during Arabidopsis embryogenesis by transiently repressing a transcriptional program that interferes with this process. Their mutant phenotype reveals the existence of a previously unknown temporal regulatory mechanism in plant embryogenesis.

  2. Glycoprotein130 (Gp130)/interleukin-6 (IL-6) signalling in osteoclasts promotes bone formation in periosteal and trabecular bone.

    PubMed

    Johnson, Rachelle W; McGregor, Narelle E; Brennan, Holly J; Crimeen-Irwin, Blessing; Poulton, Ingrid J; Martin, T John; Sims, Natalie A

    2015-12-01

    Interleukin-6 (IL-6) and interleukin-11 (IL-11) receptors (IL-6R and IL-11R, respectively) are both expressed in osteoclasts and transduce signal via the glycoprotein130 (gp130) co-receptor, but the physiological role of this pathway is unclear. To determine the critical roles of gp130 signalling in the osteoclast, we generated mice using cathepsin K Cre (CtskCre) to disrupt gp130 signalling in osteoclasts. Bone marrow macrophages from CtskCre.gp130(f/f) mice generated more osteoclasts in vitro than cells from CtskCre.gp130(w/w) mice; these osteoclasts were also larger and had more nuclei than controls. While no increase in osteoclast numbers was observed in vivo, osteoclasts on trabecular bone surfaces of CtskCre.gp130(f/f) mice were more spread out than in control mice, but had no functional defect detectable by serum CTX1 levels or trabecular bone cartilage remnants. However, trabecular osteoblast number and mineralising surfaces were significantly lower in male CtskCre.gp130(f/f) mice compared to controls, and this was associated with a significantly lower trabecular bone volume at 12 weeks of age. Furthermore, CtskCre.gp130(f/f) mice exhibited greatly suppressed periosteal bone formation at this age, indicated by significant reductions in both double-labelled surface and mineral apposition rate. By 26 weeks of age, CtskCre.gp130(f/f) mice exhibited narrower femora, with lower periosteal and endocortical perimeters than CtskCre.gp130(w/w) controls. Since IL-6 and IL-11R global knockout mice exhibited a similar reduction in femoral width, we also assessed periosteal bone formation in those strains, and found bone forming surfaces were also reduced in male IL-6 null mice. These data suggest that IL-6/gp130 signalling in the osteoclast is not essential for normal bone resorption in vivo, but maintains both trabecular and periosteal bone formation in male mice by promoting osteoblast activity through the stimulation of osteoclast-derived "coupling factors" and

  3. An auxin-mediated shift toward growth isotropy promotes organ formation at the shoot meristem in Arabidopsis.

    PubMed

    Sassi, Massimiliano; Ali, Olivier; Boudon, Frédéric; Cloarec, Gladys; Abad, Ursula; Cellier, Coralie; Chen, Xu; Gilles, Benjamin; Milani, Pascale; Friml, Jiří; Vernoux, Teva; Godin, Christophe; Hamant, Olivier; Traas, Jan

    2014-10-06

    To control morphogenesis, molecular regulatory networks have to interfere with the mechanical properties of the individual cells of developing organs and tissues, but how this is achieved is not well known. We study this issue here in the shoot meristem of higher plants, a group of undifferentiated cells where complex changes in growth rates and directions lead to the continuous formation of new organs. Here, we show that the plant hormone auxin plays an important role in this process via a dual, local effect on the extracellular matrix, the cell wall, which determines cell shape. Our study reveals that auxin not only causes a limited reduction in wall stiffness but also directly interferes with wall anisotropy via the regulation of cortical microtubule dynamics. We further show that to induce growth isotropy and organ outgrowth, auxin somehow interferes with the cortical microtubule-ordering activity of a network of proteins, including AUXIN BINDING PROTEIN 1 and KATANIN 1. Numerical simulations further indicate that the induced isotropy is sufficient to amplify the effects of the relatively minor changes in wall stiffness to promote organogenesis and the establishment of new growth axes in a robust manner.

  4. Laminin-521 Promotes Rat Bone Marrow Mesenchymal Stem Cell Sheet Formation on Light-Induced Cell Sheet Technology

    PubMed Central

    2017-01-01

    Rat bone marrow mesenchymal stem cell sheets (rBMSC sheets) are attractive for cell-based tissue engineering. However, methods of culturing rBMSC sheets are critically limited. In order to obtain intact rBMSC sheets, a light-induced cell sheet method was used in this study. TiO2 nanodot films were coated with (TL) or without (TN) laminin-521. We investigated the effects of laminin-521 on rBMSCs during cell sheet culturing. The fabricated rBMSC sheets were subsequently assessed to study cell sheet viability, reattachment ability, cell sheet thickness, collagen type I deposition, and multilineage potential. The results showed that laminin-521 could promote the formation of rBMSC sheets with good viability under hyperconfluent conditions. Cell sheet thickness increased from an initial 26.7 ± 1.5 μm (day 5) up to 47.7 ± 3.0 μm (day 10). Moreover, rBMSC sheets maintained their potential of osteogenic, adipogenic, and chondrogenic differentiation. This study provides a new strategy to obtain rBMSC sheets using light-induced cell sheet technology. PMID:28164129

  5. Substrate-promoted formation of a catalytically competent binuclear center and regulation of reactivity in a glycerophosphodiesterase from Enterobacter aerogenes.

    PubMed

    Hadler, Kieran S; Tanifum, Eric A; Yip, Sylvia Hsu-Chen; Mitić, Natasa; Guddat, Luke W; Jackson, Colin J; Gahan, Lawrence R; Nguyen, Kelly; Carr, Paul D; Ollis, David L; Hengge, Alvan C; Larrabee, James A; Schenk, Gerhard

    2008-10-29

    The glycerophosphodiesterase (GpdQ) from Enterobacter aerogenes is a promiscuous binuclear metallohydrolase that catalyzes the hydrolysis of mono-, di-, and triester substrates, including some organophosphate pesticides and products of the degradation of nerve agents. GpdQ has attracted recent attention as a promising enzymatic bioremediator. Here, we have investigated the catalytic mechanism of this versatile enzyme using a range of techniques. An improved crystal structure (1.9 A resolution) illustrates the presence of (i) an extended hydrogen bond network in the active site, and (ii) two possible nucleophiles, i.e., water/hydroxide ligands, coordinated to one or both metal ions. While it is at present not possible to unambiguously distinguish between these two possibilities, a reaction mechanism is proposed whereby the terminally bound H2O/OH(-) acts as the nucleophile, activated via hydrogen bonding by the bridging water molecule. Furthermore, the presence of substrate promotes the formation of a catalytically competent binuclear center by significantly enhancing the binding affinity of one of the metal ions in the active site. Asn80 appears to display coordination flexibility that may modulate enzyme activity. Kinetic data suggest that the rate-limiting step occurs after hydrolysis, i.e., the release of the phosphate moiety and the concomitant dissociation of one of the metal ions and/or associated conformational changes. Thus, it is proposed that GpdQ employs an intricate regulatory mechanism for catalysis, where coordination flexibility in one of the two metal binding sites is essential for optimal activity.

  6. BRCA2 and RAD51 promote double-strand break formation and cell death in response to gemcitabine.

    PubMed

    Jones, Rebecca M; Kotsantis, Panagiotis; Stewart, Grant S; Groth, Petra; Petermann, Eva

    2014-10-01

    Replication inhibitors cause replication fork stalling and double-strand breaks (DSB) that result from processing of stalled forks. During recovery from replication blocks, the homologous recombination (HR) factor RAD51 mediates fork restart and DSB repair. HR defects therefore sensitize cells to replication inhibitors, with clear implications for cancer therapy. Gemcitabine is a potent replication inhibitor used to treat cancers with mutations in HR genes such as BRCA2. Here, we investigate why, paradoxically, mutations in HR genes protect cells from killing by gemcitabine. Using DNA replication and DNA damage assays in mammalian cells, we show that even short gemcitabine treatments cause persistent replication inhibition. BRCA2 and RAD51 are recruited to chromatin early after removal of the drug, actively inhibit replication fork progression, and promote the formation of MUS81- and XPF-dependent DSBs that remain unrepaired. Our data suggest that HR intermediates formed at gemcitabine-stalled forks are converted into DSBs and thus contribute to gemcitabine-induced cell death, which could have implications for the treatment response of HR-deficient tumors.

  7. Top3β is an RNA topoisomerase that works with Fragile X syndrome protein to promote synapse formation

    PubMed Central

    Xu, Dongyi; Shen, Weiping; Guo, Rong; Xue, Yutong; Peng, Wei; Sima, Jian; Yang, Jay; Sharov, Alexei; Srikantan, Subramanya; Yang, Jiandong; Fox, David; Qian, Yong; Martindale, Jennifer L.; Piao, Yulan; Machamer, James; Joshi, Samit R.; Mohanty, Subhasis; Shaw, Albert C.; Lloyd, Thomas E.; Brown, Grant W.; Ko, Minoru S.H.; Gorospe, Myriam; Zou, Sige; Wang, Weidong

    2013-01-01

    Topoisomerases are crucial to solve DNA topological problems, but they have not been linked to RNA metabolism. Here we show that human topoisomerase 3β (Top3β) is an RNA topoisomerase that biochemically and genetically interacts with FMRP, a protein deficient in Fragile X syndrome and known to regulate translation of mRNAs important for neuronal function and autism. Notably, the FMRP-Top3β interaction is abolished by a disease-associated FMRP mutation, suggesting that Top3β may contribute to pathogenesis of mental disorders. Top3β binds multiple mRNAs encoded by genes with neuronal functions related to schizophrenia and autism. Expression of one such gene, ptk2/FAK, is reduced in neuromuscular junctions of Top3β mutant flies. Synapse formation is defective in Top3β mutant flies and mice, as observed in FMRP mutant animals. Our findings suggest that Top3β acts as an RNA topoisomerase and works with FMRP to promote expression of mRNAs critical for neurodevelopment and mental health. PMID:23912945

  8. Multiple Ligands of von Willebrand Factor-binding Protein (vWbp) Promote Staphylococcus aureus Clot Formation in Human Plasma*

    PubMed Central

    Thomer, Lena; Schneewind, Olaf; Missiakas, Dominique

    2013-01-01

    Staphylococcus aureus secretes coagulase (Coa) and von Willebrand factor-binding protein (vWbp) to activate host prothrombin and form fibrin cables, thereby promoting the establishment of infectious lesions. The D1-D2 domains of Coa and vWbp associate with, and non-proteolytically activate prothrombin. Moreover, Coa encompasses C-terminal tandem repeats for binding to fibrinogen, whereas vWbp has been reported to associate with von Willebrand factor and fibrinogen. Here we used affinity chromatography with non-catalytic Coa and vWbp to identify the ligands for these virulence factors in human plasma. vWbp bound to prothrombin, fibrinogen, fibronectin, and factor XIII, whereas Coa co-purified with prothrombin and fibrinogen. vWbp association with fibrinogen and factor XIII, but not fibronectin, required prothrombin and triggered the non-proteolytic activation of FXIII in vitro. Staphylococcus aureus coagulation of human plasma was associated with the recruitment of prothrombin, FXIII, and fibronectin as well as the formation of cross-linked fibrin. FXIII activity in staphylococcal clots could be attributed to thrombin-dependent proteolytic activation as well as vWbp-mediated non-proteolytic activation of FXIII zymogen. PMID:23960083

  9. Smooth muscle cell-specific Tgfbr1 deficiency promotes aortic aneurysm formation by stimulating multiple signaling events

    PubMed Central

    Yang, Pu; Schmit, Bradley M.; Fu, Chunhua; DeSart, Kenneth; Oh, S. Paul; Berceli, Scott A.; Jiang, Zhihua

    2016-01-01

    Transforming growth factor (TGF)-β signaling disorder has emerged as a common molecular signature for aortic aneurysm development. The timing of postnatal maturation plays a key role in dictating the biological outcome of TGF-β signaling disorders in the aortic wall. In this study, we investigated the impact of deficiency of TGFβ receptors on the structural homeostasis of mature aortas. We used an inducible Cre-loxP system driven by a Myh11 promoter to delete Tgfbr1, Tgfbr2, or both in smooth muscle cells (SMCs) of adult mice. TGFBR1 deficiency resulted in rapid and severe aneurysmal degeneration, with 100% penetrance of ascending thoracic aortas, whereas TGFBR2 deletion only caused mild aortic pathology with low (26%) lesion prevalence. Removal of TGFBR2 attenuated the aortic pathology caused by TGFBR1 deletion and correlated with a reduction of early ERK phosphorylation. In addition, the production of angiotensin (Ang)-converting enzyme was upregulated in TGFBR1 deficient aortas at the early stage of aneurysmal degeneration. Inhibition of ERK phosphorylation or blockade of AngII type I receptor AT1R prevented aneurysmal degeneration of TGFBR1 deficient aortas. In conclusion, loss of SMC-Tgfbr1 triggers multiple deleterious pathways, including abnormal TGFBR2, ERK, and AngII/AT1R signals that disrupt aortic wall homeostasis to cause aortic aneurysm formation. PMID:27739498

  10. Leaf trichome density may explain herbivory patterns of Actinote sp. (Lepidoptera: Acraeidae) on Liabum mandonii (Asteraceae) in a montane humid forest (Nor Yungas, Bolivia)

    NASA Astrophysics Data System (ADS)

    Molina-Montenegro, Marco A.; Ávila, Pamela; Hurtado, Rosember; Valdivia, Alejandra I.; Gianoli, Ernesto

    2006-09-01

    Increasing evidence shows that most insect herbivores of tropical forests show specific associations with their hosts. Environmental factors as well as foliar characters can modify insect preference. In the present work, we evaluated in a montane humid forest the preference and herbivory rate of Actinote sp. caterpillars (Lepidoptera: Acraeidae) on mature and young leaves of their specific host plant Liabum mandonii (Asteraceae) in two contrasting sites. Additionally, the density of non-glandular trichomes in young and mature leaves of plant at each site was evaluated. Analysis of variance of herbivory showed significant effects of site, leaf age, and the interaction of these factors. Higher herbivory levels were found on leaves from the site with lower levels of radiation and on mature leaves. On the other hand, trichome density was significantly higher in leaves from the site with higher levels of radiation and in young leaves. This suggests trichomes may explain the observed pattern of herbivory.

  11. Functionally Divergent Alleles and Duplicated Loci Encoding an Acyltransferase Contribute to Acylsugar Metabolite Diversity in Solanum Trichomes[OPEN

    PubMed Central

    Schilmiller, Anthony L.; Moghe, Gaurav D.; Fan, Pengxiang; Ghosh, Banibrata; Ning, Jing; Jones, A. Daniel; Last, Robert L.

    2015-01-01

    Glandular trichomes from tomato (Solanum lycopersicum) and other species in the Solanaceae produce and secrete a mixture of O-acylsugars (aliphatic esters of sucrose and glucose) that contribute to insect defense. Despite their phylogenetic distribution and diversity, relatively little is known about how these specialized metabolites are synthesized. Mass spectrometric profiling of acylsugars in the S. lycopersicum x Solanum pennellii introgression lines identified a chromosome 11 locus containing a cluster of BAHD acyltransferases with one gene (named Sl-ASAT3) expressed in tip cells of type I trichomes where acylsugars are made. Sl-ASAT3 was shown to encode an acyl-CoA-dependent acyltransferase that catalyzes the transfer of short (four to five carbons) branched acyl chains to the furanose ring of di-acylsucrose acceptors to produce tri-acylsucroses, which can be further acetylated by Sl-ASAT4 (previously Sl-AT2). Among the wild tomatoes, diversity in furanose ring acyl chains on acylsucroses was most striking in Solanum habrochaites. S. habrochaites accessions from Ecuador and northern Peru produced acylsucroses with short (≤C5) or no acyl chains on the furanose ring. Accessions from central and southern Peru had the ability to add short or long (up to C12) acyl chains to the furanose ring. Multiple ASAT3-like sequences were found in most accessions, and their in vitro activities correlated with observed geographical diversity in acylsugar profiles. PMID:25862303

  12. The Active Component of Aspirin, Salicylic Acid, Promotes Staphylococcus aureus Biofilm Formation in a PIA-dependent Manner.

    PubMed

    Dotto, Cristian; Lombarte Serrat, Andrea; Cattelan, Natalia; Barbagelata, María S; Yantorno, Osvaldo M; Sordelli, Daniel O; Ehling-Schulz, Monika; Grunert, Tom; Buzzola, Fernanda R

    2017-01-01

    Aspirin has provided clear benefits to human health. But salicylic acid (SAL) -the main aspirin biometabolite- exerts several effects on eukaryote and prokaryote cells. SAL can affect, for instance, the expression of Staphylococcus aureus virulence factors. SAL can also form complexes with iron cations and it has been shown that different iron chelating molecules diminished the formation of S. aureus biofilm. The aim of this study was to elucidate whether the iron content limitation caused by SAL can modify the S. aureus metabolism and/or metabolic regulators thus changing the expression of the main polysaccharides involved in biofilm formation. The exposure of biofilm to 2 mM SAL induced a 27% reduction in the intracellular free Fe(2+) concentration compared with the controls. In addition, SAL depleted 23% of the available free Fe(2+) cation in culture media. These moderate iron-limited conditions promoted an intensification of biofilms formed by strain Newman and by S. aureus clinical isolates related to the USA300 and USA100 clones. The slight decrease in iron bioavailability generated by SAL was enough to induce the increase of PIA expression in biofilms formed by methicillin-resistant as well as methicillin-sensitive S. aureus strains. S. aureus did not produce capsular polysaccharide (CP) when it was forming biofilms under any of the experimental conditions tested. Furthermore, SAL diminished aconitase activity and stimulated the lactic fermentation pathway in bacteria forming biofilms. The polysaccharide composition of S. aureus biofilms was examined and FTIR spectroscopic analysis revealed a clear impact of SAL in a codY-dependent manner. Moreover, SAL negatively affected codY transcription in mature biofilms thus relieving the CodY repression of the ica operon. Treatment of mice with SAL induced a significant increase of S aureus colonization. It is suggested that the elevated PIA expression induced by SAL might be responsible for the high nasal

  13. The Active Component of Aspirin, Salicylic Acid, Promotes Staphylococcus aureus Biofilm Formation in a PIA-dependent Manner

    PubMed Central

    Dotto, Cristian; Lombarte Serrat, Andrea; Cattelan, Natalia; Barbagelata, María S.; Yantorno, Osvaldo M.; Sordelli, Daniel O.; Ehling-Schulz, Monika; Grunert, Tom; Buzzola, Fernanda R.

    2017-01-01

    Aspirin has provided clear benefits to human health. But salicylic acid (SAL) -the main aspirin biometabolite- exerts several effects on eukaryote and prokaryote cells. SAL can affect, for instance, the expression of Staphylococcus aureus virulence factors. SAL can also form complexes with iron cations and it has been shown that different iron chelating molecules diminished the formation of S. aureus biofilm. The aim of this study was to elucidate whether the iron content limitation caused by SAL can modify the S. aureus metabolism and/or metabolic regulators thus changing the expression of the main polysaccharides involved in biofilm formation. The exposure of biofilm to 2 mM SAL induced a 27% reduction in the intracellular free Fe2+ concentration compared with the controls. In addition, SAL depleted 23% of the available free Fe2+ cation in culture media. These moderate iron-limited conditions promoted an intensification of biofilms formed by strain Newman and by S. aureus clinical isolates related to the USA300 and USA100 clones. The slight decrease in iron bioavailability generated by SAL was enough to induce the increase of PIA expression in biofilms formed by methicillin-resistant as well as methicillin-sensitive S. aureus strains. S. aureus did not produce capsular polysaccharide (CP) when it was forming biofilms under any of the experimental conditions tested. Furthermore, SAL diminished aconitase activity and stimulated the lactic fermentation pathway in bacteria forming biofilms. The polysaccharide composition of S. aureus biofilms was examined and FTIR spectroscopic analysis revealed a clear impact of SAL in a codY-dependent manner. Moreover, SAL negatively affected codY transcription in mature biofilms thus relieving the CodY repression of the ica operon. Treatment of mice with SAL induced a significant increase of S aureus colonization. It is suggested that the elevated PIA expression induced by SAL might be responsible for the high nasal colonization

  14. Dipeptidyl peptidase IV inhibition prevents the formation and promotes the healing of indomethacin-induced intestinal ulcers in rats

    PubMed Central

    Inoue, Takuya; Higashiyama, Masaaki; Kaji, Izumi; Rudenkyy, Sergiy; Higuchi, Kazuhide; Guth, Paul H.; Engel, Eli; Kaunitz, Jonathan D; Akiba, Yasutada

    2014-01-01

    Backgrounds & Aims We studied the intestinotrophic hormone glucagon-like peptide-2 (GLP-2) as a possible therapy for non-steroidal anti-inflammatory drug (NSAID)-induced intestinal ulcers. Luminal nutrients release endogenous GLP-2 from enteroendocrine L cells. Since GLP-2 is degraded by dipeptidyl peptidase IV (DPPIV), we hypothesized that DPPIV inhibition combined with luminal administration of nutrients potentiates the effects of endogenous GLP-2 on intestinal injury. Methods Intestinal injury was induced by indomethacin (10 mg/kg, sc) in fed rats. The long-acting DPPIV inhibitor K579 was intragastrically (ig) or intraperitoneally (ip) given before or after indomethacin treatment. L-alanine (L-Ala) and 5′-inosine monophosphate (IMP) were co-administered ig after the treatment. Results Indomethacin treatment induced intestinal ulcers which gradually healed after treatment. Pretreatment with ig or ip K579 given either at 1 mg/kg reduced total ulcer length, whereas K579 at 3 mg/kg had no effect. Exogenous GLP-2 also reduced intestinal ulcers. The preventive effect of K579 was dose-dependently inhibited by a GLP-2 receptor antagonist. Daily treatment with K579 (1 mg/kg), GLP-2, or L-Ala + IMP after indomethacin treatment reduced total ulcer length. Co-administration (ig) of K579 and L-Ala + IMP further accelerated intestinal ulcer healing. Conclusion DPPIV inhibition and exogenous GLP-2 prevented the formation and promoted the healing of indomethacin-induced intestinal ulcers, although high-dose DPPIV inhibition reversed the preventive effect. Umami receptor agonists also enhanced the healing effects of the DPPIV inhibitor. The combination of DPPIV inhibition and luminal nutrient-induced GLP-2 release may be a useful therapeutic tool for the treatment of NSAIDs-induced intestinal ulcers. PMID:24379150

  15. Allotropic control: How certain fluorinated carbonate electrolytes protect aluminum current collectors by promoting the formation of insoluble coordination polymers

    SciTech Connect

    Shkrob, Ilya A.; Pupek, Krzysztof Z.; Abraham, Daniel P.

    2016-07-28

    Here, there is a strong incentive for increasing the operation voltage of Li-ion cells above 4.5 V in order to increase the density of stored energy. Aluminum is an inexpensive, lightweight metal that is commonly used as a positive electrode current collector in these cells. Imide LiX salts, such as lithium bis(trifluoromethylsulfonyl)imide (X = TFSI), and lithium bis(fluorosulfonyl)imide (X = FSI), are chemically stable on the energized lithiated transition metal oxide electrodes, but their presence in the electrolyte causes rapid anodic dissolution and pitting of Al current collectors at potentials exceeding 4.0 V versus Li/Li+. For LiBF4 and LiPF6, the release of HF near the energized surfaces passivates the exposed Al metal, inhibiting this pitting corrosion, but it also causes the gradual degradation of the cathode active material, negating this important advantage. Here we report that in certain electrolytes containing fluorinated carbonate solvents and LiX salts, the threshold voltage for safe operation of Al current collectors can be increased to 5.5 V versus Li/Li+. Interestingly, the most efficient solvent also facilitates the formation of an insoluble gel when AlX3 is introduced into this solvent. We suggest that this solvent promotes the aggregation of coordination polymers of AlX3 at the exposed Al surface that isolate this surface from the electrolyte, thereby preventing further Al dissolution and corrosion. Other examples of Al collector protection may also involve this mechanism. Our study suggests that such “allotropic control” could be a way of widening the operation window of Li-ion cells without electrode deterioration, Al current collector corrosion, and electrolyte breakdown.

  16. Androgens inhibit tumor necrosis factor-α-induced cell adhesion and promote tube formation of human coronary artery endothelial cells.

    PubMed

    Liao, Chun-Hou; Lin, Feng-Yen; Wu, Yi-No; Chiang, Han-Sun

    2012-06-01

    Endothelial cells contribute to the function and integrity of the vascular wall, and a functional aberration may lead to atherogenesis. There is increasing evidence on the atheroprotective role of androgens. Therefore, we studied the effect of the androgens-testosterone and dihydrotestosterone-and estradiol on human coronary artery endothelial cell (HCAEC) function. We found by MTT assay that testosterone is not cytotoxic and enhances HCAEC proliferation. The effect of testosterone (10-50 nM), dihydrotestosterone (5-50 nM), and estradiol (0.1-0.4 nM) on the adhesion of tumor necrosis factor-α (TNF-α)-stimulated HCAECs was determined at different time points (12-96 h) by assessing their binding with human monocytic THP-1 cells. In addition, the expression of adhesion molecules, vascular cell adhesion molecule-1 (VCAM-1) and intracellular adhesion molecule-1 (ICAM-1), was determined by ELISA and Western blot analysis. Both testosterone and dihydrotestosterone attenuated cell adhesion and the expression of VCAM-1 and ICAM-1 in a dose- and time-dependent manner. Furthermore, androgen treatment for a longer duration inhibited cell migration, as demonstrated by wound-healing assay, and promoted tube formation on a Matrigel. Western blot analysis demonstrated that the expression of phosphorylated endothelial nitric oxide synthase (eNOS) increased, whereas that of inducible nitric oxide synthase (iNOS) decreased following the 96-h steroid treatment of TNF-α-stimulated HCAECs. Our findings suggest that androgens modulate endothelial cell functions by suppressing the inflammatory process and enhancing wound-healing and regenerative angiogenesis, possibly through an androgen receptor (AR)-dependent mechanism.

  17. Steviol Reduces MDCK Cyst Formation and Growth by Inhibiting CFTR Channel Activity and Promoting Proteasome-Mediated CFTR Degradation

    PubMed Central

    Yuajit, Chaowalit; Homvisasevongsa, Sureeporn; Chatsudthipong, Lisa; Soodvilai, Sunhapas; Muanprasat, Chatchai; Chatsudthipong, Varanuj

    2013-01-01

    Cyst enlargement in polycystic kidney disease (PKD) involves cAMP-activated proliferation of cyst-lining epithelial cells and transepithelial fluid secretion into the cyst lumen via cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel. This study aimed to investigate an inhibitory effect and detailed mechanisms of steviol and its derivatives on cyst growth using a cyst model in Madin-Darby canine kidney (MDCK) cells. Among 4 steviol-related compounds tested, steviol was found to be the most potent at inhibiting MDCK cyst growth. Steviol inhibition of cyst growth was dose-dependent; steviol (100 microM) reversibly inhibited cyst formation and cyst growth by 72.53.6% and 38.2±8.5%, respectively. Steviol at doses up to 200 microM had no effect on MDCK cell viability, proliferation and apoptosis. However, steviol acutely inhibited forskolin-stimulated apical chloride current in MDCK epithelia, measured with the Ussing chamber technique, in a dose-dependent manner. Prolonged treatment (24 h) with steviol (100 microM) also strongly inhibited forskolin-stimulated apical chloride current, in part by reducing CFTR protein expression in MDCK cells. Interestingly, proteasome inhibitor, MG-132, abolished the effect of steviol on CFTR protein expression. Immunofluorescence studies demonstrated that prolonged treatment (24 h) with steviol (100 microM) markedly reduced CFTR expression at the plasma membrane. Taken together, the data suggest that steviol retards MDCK cyst progression in two ways: first by directly inhibiting CFTR chloride channel activity and second by reducing CFTR expression, in part, by promoting proteasomal degradation of CFTR. Steviol and related compounds therefore represent drug candidates for treatment of polycystic kidney disease. PMID:23536832

  18. Allotropic control: How certain fluorinated carbonate electrolytes protect aluminum current collectors by promoting the formation of insoluble coordination polymers

    DOE PAGES

    Shkrob, Ilya A.; Pupek, Krzysztof Z.; Abraham, Daniel P.

    2016-07-28

    Here, there is a strong incentive for increasing the operation voltage of Li-ion cells above 4.5 V in order to increase the density of stored energy. Aluminum is an inexpensive, lightweight metal that is commonly used as a positive electrode current collector in these cells. Imide LiX salts, such as lithium bis(trifluoromethylsulfonyl)imide (X = TFSI), and lithium bis(fluorosulfonyl)imide (X = FSI), are chemically stable on the energized lithiated transition metal oxide electrodes, but their presence in the electrolyte causes rapid anodic dissolution and pitting of Al current collectors at potentials exceeding 4.0 V versus Li/Li+. For LiBF4 and LiPF6, themore » release of HF near the energized surfaces passivates the exposed Al metal, inhibiting this pitting corrosion, but it also causes the gradual degradation of the cathode active material, negating this important advantage. Here we report that in certain electrolytes containing fluorinated carbonate solvents and LiX salts, the threshold voltage for safe operation of Al current collectors can be increased to 5.5 V versus Li/Li+. Interestingly, the most efficient solvent also facilitates the formation of an insoluble gel when AlX3 is introduced into this solvent. We suggest that this solvent promotes the aggregation of coordination polymers of AlX3 at the exposed Al surface that isolate this surface from the electrolyte, thereby preventing further Al dissolution and corrosion. Other examples of Al collector protection may also involve this mechanism. Our study suggests that such “allotropic control” could be a way of widening the operation window of Li-ion cells without electrode deterioration, Al current collector corrosion, and electrolyte breakdown.« less

  19. Identification of Fic-1 as an enzyme that inhibits bacterial DNA replication by AMPylating GyrB, promoting filament formation.

    PubMed

    Lu, Canhua; Nakayasu, Ernesto S; Zhang, Li-Qun; Luo, Zhao-Qing

    2016-01-26

    The morphology of bacterial cells is important for virulence, evasion of the host immune system, and coping with environmental stresses. The widely distributed Fic proteins (filamentation induced by cAMP) are annotated as proteins involved in cell division because of the presence of the HPFx[D/E]GN[G/K]R motif. We showed that the presence of Fic-1 from Pseudomonas fluorescens significantly reduced the yield of plasmid DNA when expressed in Escherichia coli or P. fluorescens. Fic-1 interacted with GyrB, a subunit of DNA gyrase, which is essential for bacterial DNA replication. Fic-1 catalyzed the AMPylation of GyrB at Tyr(109), a residue critical for binding ATP, and exhibited auto-AMPylation activity. Mutation of the Fic-1 auto-AMPylated site greatly reduced AMPylation activity toward itself and toward GyrB. Fic-1-dependent AMPylation of GyrB triggered the SOS response, indicative of DNA replication stress or DNA damage. Fic-1 also promoted the formation of elongated cells when the SOS response was blocked. We identified an α-inhibitor protein that we named anti-Fic-1 (AntF), encoded by a gene immediately upstream of Fic-1. AntF interacted with Fic-1, inhibited the AMPylation activity of Fic-1 for GyrB in vitro, and blocked Fic-1-mediated inhibition of DNA replication in bacteria, suggesting that Fic-1 and AntF comprise a toxin-antitoxin module. Our work establishes Fic-1 as an AMPylating enzyme that targets GyrB to inhibit DNA replication and may target other proteins to regulate bacterial morphology.

  20. Platelet rich plasma promotes skeletal muscle cell migration in association with up-regulation of FAK, paxillin, and F-Actin formation.

    PubMed

    Tsai, Wen-Chung; Yu, Tung-Yang; Lin, Li-Ping; Lin, Mioa-Sui; Tsai, Ting-Ta; Pang, Jong-Hwei S

    2017-02-24

    Platelet rich plasma (PRP) contains various cytokines and growth factors which may be beneficial to the healing process of injured muscle. The aim of this study was to investigate the effect and molecular mechanism of PRP on migration of skeletal muscle cells. Skeletal muscle cells intrinsic to Sprague-Dawley rats were treated with PRP. The cell migration was evaluated by transwell filter migration assay and electric cell-substrate impedance sensing. The spreading of cells was evaluated microscopically. The formation of filamentous actin (F-actin) cytoskeleton was assessed by immunofluorescence staining. The protein expressions of paxillin and focal adhesion kinase (FAK) were assessed by Western blot analysis. Transfection of paxillin small-interfering RNA (siRNAs) to muscle cells was performed to validate the role of paxillin in PRP-mediated promotion of cell migration. Dose-dependently PRP promotes migration of and spreading and muscle cells. Protein expressions of paxillin and FAK were up-regulated dose-dependently. F-actin formation was also enhanced by PRP treatment. Furthermore, the knockdown of paxillin expression impaired the effect of PRP to promote cell migration. It was concluded that PRP promoting migration of muscle cells is associated with up-regulation of proteins expression of paxillin and FAK as well as increasing F-actin formation. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res.

  1. The Ras-related protein Cdc42Hs and bradykinin promote formation of peripheral actin microspikes and filopodia in Swiss 3T3 fibroblasts.

    PubMed Central

    Kozma, R; Ahmed, S; Best, A; Lim, L

    1995-01-01

    The Ras-related protein Cdc42 plays a role in yeast cell budding and polarity. Two related proteins, Rac1 and RhoA, promote formation in mammalian cells of membrane ruffles and stress fibers, respectively, which contain actin microfilaments. We now show that microinjection of the related human Cdc42Hs into Swiss 3T3 fibroblasts induced the formation of peripheral actin microspikes, determined by staining with phalloidin. A proportion of these microspikes was found to be components of filopodia, as analyzed by time-lapse phase-contrast microscopy. The formation of filopodia was also found to be promoted by Cdc42Hs microinjection. This was followed by activation of Rac1-mediated membrane ruffling. Treatment with bradykinin also promoted formation of microspikes and filopodia as well as subsequent effects similar to that seen upon Cdc42Hs microinjection. These effects of bradykinin were specifically inhibited by prior microinjection of dominant negative Cdc42HsT17N, suggesting that bradykinin acts by activating cellular Cdc42Hs. Since filopodia have been ascribed an important sensory function in fibroblasts and are required for guidance of neuronal growth cones, these results indicate that Cdc42Hs plays an important role in determining mammalian cell morphology. PMID:7891688

  2. Bromodomain protein Brd3 promotes Ifnb1 transcription via enhancing IRF3/p300 complex formation and recruitment to Ifnb1 promoter in macrophages

    PubMed Central

    Ren, Wenhui; Wang, Chunmei; Wang, Qinlan; Zhao, Dezhi; Zhao, Kai; Sun, Donghao; Liu, Xingguang; Han, Chaofeng; Hou, Jin; Li, Xia; Zhang, Qian; Cao, Xuetao; Li, Nan

    2017-01-01

    As members of bromodomain and extra-terminal motif protein family, bromodomain-containing proteins regulate a wide range of biological processes including protein scaffolding, mitosis, cell cycle progression and transcriptional regulation. The function of these bromodomain proteins (Brds) in innate immune response has been reported but the role of Brd3 remains unclear. Here we find that virus infection significantly downregulate Brd3 expression in macrophages and Brd3 knockout inhibits virus-triggered IFN-β production. Brd3 interacts with both IRF3 and p300, increases p300-mediated acetylation of IRF3, and enhances the association of IRF3 with p300 upon virus infection. Importantly, Brd3 promotes the recruitment of IRF3/p300 complex to the promoter of Ifnb1, and increases the acetylation of histone3/histone4 within the Ifnb1 promoter, leading to the enhancement of type I interferon production. Therefore, our work indicated that Brd3 may act as a coactivator in IRF3/p300 transcriptional activation of Ifnb1 and provided new epigenetic mechanistic insight into the efficient activation of the innate immune response. PMID:28045112

  3. The C. elegans DSB-2 protein reveals a regulatory network that controls competence for meiotic DSB formation and promotes crossover assurance.

    PubMed

    Rosu, Simona; Zawadzki, Karl A; Stamper, Ericca L; Libuda, Diana E; Reese, Angela L; Dernburg, Abby F; Villeneuve, Anne M

    2013-01-01

    For most organisms, chromosome segregation during meiosis relies on deliberate induction of DNA double-strand breaks (DSBs) and repair of a subset of these DSBs as inter-homolog crossovers (COs). However, timing and levels of DSB formation must be tightly controlled to avoid jeopardizing genome integrity. Here we identify the DSB-2 protein, which is required for efficient DSB formation during C. elegans meiosis but is dispensable for later steps of meiotic recombination. DSB-2 localizes to chromatin during the time of DSB formation, and its disappearance coincides with a decline in RAD-51 foci marking early recombination intermediates and precedes appearance of COSA-1 foci marking CO-designated sites. These and other data suggest that DSB-2 and its paralog DSB-1 promote competence for DSB formation. Further, immunofluorescence analyses of wild-type gonads and various meiotic mutants reveal that association of DSB-2 with chromatin is coordinated with multiple distinct aspects of the meiotic program, including the phosphorylation state of nuclear envelope protein SUN-1 and dependence on RAD-50 to load the RAD-51 recombinase at DSB sites. Moreover, association of DSB-2 with chromatin is prolonged in mutants impaired for either DSB formation or formation of downstream CO intermediates. These and other data suggest that association of DSB-2 with chromatin is an indicator of competence for DSB formation, and that cells respond to a deficit of CO-competent recombination intermediates by prolonging the DSB-competent state. In the context of this model, we propose that formation of sufficient CO-competent intermediates engages a negative feedback response that leads to cessation of DSB formation as part of a major coordinated transition in meiotic prophase progression. The proposed negative feedback regulation of DSB formation simultaneously (1) ensures that sufficient DSBs are made to guarantee CO formation and (2) prevents excessive DSB levels that could have deleterious

  4. The Tomato odorless-2 Mutant Is Defective in Trichome-Based Production of Diverse Specialized Metabolites and Broad-Spectrum Resistance to Insect Herbivores1[W][OA

    PubMed Central

    Kang, Jin-Ho; Liu, Guanghui; Shi, Feng; Jones, A. Daniel; Beaudry, Randolph M.; Howe, Gregg A.

    2010-01-01

    Glandular secreting trichomes of cultivated tomato (Solanum lycopersicum) produce a wide array of volatile and nonvolatile specialized metabolites. Many of these compounds contribute to the characteristic aroma of tomato foliage and constitute a key part of the language by which plants communicate with other organisms in natural environments. Here, we describe a novel recessive mutation called odorless-2 (od-2) that was identified on the basis of an altered leaf-aroma phenotype. od-2 plants exhibit pleiotrophic phenotypes, including alterations in the morphology, density, and chemical composition of glandular trichomes. Type VI glandular trichomes isolated from od-2 leaves accumulate only trace levels of monoterpenes, sesquiterpenes, and flavonoids. Other foliar defensive compounds, including acyl sugars, glycoalkaloids, and jasmonate-regulated proteinase inhibitors, are produced in od-2 leaves. Growth of od-2 plants under natural field conditions showed that the mutant is highly susceptible to attack by an indigenous flea beetle, Epitrix cucumeris, and the Colorado potato beetle, Leptinotarsa decemlineata. The increased susceptibility of od-2 plants to Colorado potato beetle larvae and to the solanaceous specialist Manduca sexta was verified in no-choice bioassays. These findings indicate that Od-2 is essential for the synthesis of diverse trichome-borne compounds and further suggest that these compounds influence host plant selection and herbivore community composition under natural conditions. PMID:20668059

  5. The tomato odorless-2 mutant is defective in trichome-based production of diverse specialized metabolites and broad-spectrum resistance to insect herbivores.

    PubMed

    Kang, Jin-Ho; Liu, Guanghui; Shi, Feng; Jones, A Daniel; Beaudry, Randolph M; Howe, Gregg A

    2010-09-01

    Glandular secreting trichomes of cultivated tomato (Solanum lycopersicum) produce a wide array of volatile and nonvolatile specialized metabolites. Many of these compounds contribute to the characteristic aroma of tomato foliage and constitute a key part of the language by which plants communicate with other organisms in natural environments. Here, we describe a novel recessive mutation called odorless-2 (od-2) that was identified on the basis of an altered leaf-aroma phenotype. od-2 plants exhibit pleiotrophic phenotypes, including alterations in the morphology, density, and chemical composition of glandular trichomes. Type VI glandular trichomes isolated from od-2 leaves accumulate only trace levels of monoterpenes, sesquiterpenes, and flavonoids. Other foliar defensive compounds, including acyl sugars, glycoalkaloids, and jasmonate-regulated proteinase inhibitors, are produced in od-2 leaves. Growth of od-2 plants under natural field conditions showed that the mutant is highly susceptible to attack by an indigenous flea beetle, Epitrix cucumeris, and the Colorado potato beetle, Leptinotarsa decemlineata. The increased susceptibility of od-2 plants to Colorado potato beetle larvae and to the solanaceous specialist Manduca sexta was verified in no-choice bioassays. These findings indicate that Od-2 is essential for the synthesis of diverse trichome-borne compounds and further suggest that these compounds influence host plant selection and herbivore community composition under natural conditions.

  6. RNA polymerase I transcription factors in active yeast rRNA gene promoters enhance UV damage formation and inhibit repair.

    PubMed

    Meier, Andreas; Thoma, Fritz

    2005-03-01

    UV photofootprinting and repair of pyrimidine dimers by photolyase was used to investigate chromatin structure, protein-DNA interactions, and DNA repair in the spacer and promoter of Saccharomyces cerevisiae rRNA genes. Saccharomyces cerevisiae contains about 150 copies of rRNA genes separated by nontranscribed spacers. Under exponential growth conditions about half of the genes are transcribed by RNA polymerase I (RNAP-I). Initiation of transcription requires the assembly of the upstream activating factor (UAF), the core factor (CF), TATA binding protein, and RNAP-I with Rrn3p on the upstream element and core promoter. We show that UV irradiation of wild-type cells and transcription factor mutants generates photofootprints in the promoter elements. The core footprint depends on UAF, while the UAF footprint was also detected in absence of the CFs. Fractionation of active and inactive promoters showed the core footprint mainly in the active fraction and similar UAF footprints in both fractions. DNA repair by photolyase was strongly inhibited in active promoters but efficient in inactive promoters. The data suggest that UAF is present in vivo in active and inactive promoters and that recruitment of CF and RNAP-I to active promoters generates a stable complex which inhibits repair.

  7. Parent and Clinician Preferences for an Asthma App to Promote Adolescent Self-Management: A Formative Study

    PubMed Central

    Roberts, Courtney A; Sage, Adam J; Coyne-Beasley, Tamera; Sleath, Betsy L; Carpenter, Delesha M

    2016-01-01

    Background Most youth asthma apps are not designed with parent and clinician use in mind, and rarely is the app development process informed by parent or clinician input. Objective This study was conducted to generate formative data on the use, attitudes, and preferences for asthma mHealth app features among parents and clinicians, the important stakeholders who support adolescents with asthma and promote adolescent self-management skills. Methods We conducted a mixed-methods study from 2013 to 2014 employing a user-centered design philosophy to acquire feedback from a convenience sample of 20 parents and 6 clinicians. Participants were given an iPod Touch and asked to evaluate 10 features on 2 existing asthma apps. Participant experiences using the apps were collected from questionnaires and a thematic analysis of audio-recorded and transcribed (verbatim) interviews using MAXQDA. Descriptive statistics were calculated to characterize the study sample and app feature feedback. Independent samples t tests were performed to compare parent and clinician ratings of app feature usefulness (ratings: 1=not at all useful to 5=very useful). Results All parents were female (n=20), 45% were black, 20% had an income ≥US $50,000, and 45% had a bachelor’s degree or higher education. The clinician sample included 2 nurses and 4 physicians with a mean practice time of 13 years. Three main themes provided an understanding of how participants perceived their roles and use of asthma app features to support adolescent asthma self-management: monitoring and supervision, education, and communication/information sharing. Parents rated the doctor report feature highest, and clinicians rated the doctor appointment reminder highest of all evaluated app features on usefulness. The peak flow monitoring feature was the lowest ranked feature by both parents and clinicians. Parents reported higher usefulness for the doctor report (t(10)=2.7, P<.02), diary (t(10)=2.7, P<.03), and self

  8. EZH2 and BCL6 Cooperate to Assemble CBX8-BCOR Complex to Repress Bivalent Promoters, Mediate Germinal Center Formation and Lymphomagenesis.

    PubMed

    Béguelin, Wendy; Teater, Matt; Gearhart, Micah D; Calvo Fernández, María Teresa; Goldstein, Rebecca L; Cárdenas, Mariano G; Hatzi, Katerina; Rosen, Monica; Shen, Hao; Corcoran, Connie M; Hamline, Michelle Y; Gascoyne, Randy D; Levine, Ross L; Abdel-Wahab, Omar; Licht, Jonathan D; Shaknovich, Rita; Elemento, Olivier; Bardwell, Vivian J; Melnick, Ari M

    2016-08-08

    The EZH2 histone methyltransferase mediates the humoral immune response and drives lymphomagenesis through formation of bivalent chromatin domains at critical germinal center (GC) B cell promoters. Herein we show that the actions of EZH2 in driving GC formation and lymphoma precursor lesions require site-specific binding by the BCL6 transcriptional repressor and the presence of a non-canonical PRC1-BCOR-CBX8 complex. The chromodomain protein CBX8 is induced in GC B cells, binds to H3K27me3 at bivalent promoters, and is required for stable association of the complex and the resulting histone modifications. Moreover, oncogenic BCL6 and EZH2 cooperate to accelerate diffuse large B cell lymphoma (DLBCL) development and combinatorial targeting of these repressors results in enhanced anti-lymphoma activity in DLBCLs.

  9. Support vector regression-guided unravelling: antioxidant capacity and quantitative structure-activity relationship predict reduction and promotion effects of flavonoids on acrylamide formation

    PubMed Central

    Huang, Mengmeng; Wei, Yan; Wang, Jun; Zhang, Yu

    2016-01-01

    We used the support vector regression (SVR) approach to predict and unravel reduction/promotion effect of characteristic flavonoids on the acrylamide formation under a low-moisture Maillard reaction system. Results demonstrated the reduction/promotion effects by flavonoids at addition levels of 1–10000 μmol/L. The maximal inhibition rates (51.7%, 68.8% and 26.1%) and promote rates (57.7%, 178.8% and 27.5%) caused by flavones, flavonols and isoflavones were observed at addition levels of 100 μmol/L and 10000 μmol/L, respectively. The reduction/promotion effects were closely related to the change of trolox equivalent antioxidant capacity (ΔTEAC) and well predicted by triple ΔTEAC measurements via SVR models (R: 0.633–0.900). Flavonols exhibit stronger effects on the acrylamide formation than flavones and isoflavones as well as their O-glycosides derivatives, which may be attributed to the number and position of phenolic and 3-enolic hydroxyls. The reduction/promotion effects were well predicted by using optimized quantitative structure-activity relationship (QSAR) descriptors and SVR models (R: 0.926–0.994). Compared to artificial neural network and multi-linear regression models, SVR models exhibited better fitting performance for both TEAC-dependent and QSAR descriptor-dependent predicting work. These observations demonstrated that the SVR models are competent for predicting our understanding on the future use of natural antioxidants for decreasing the acrylamide formation. PMID:27586851

  10. Support vector regression-guided unravelling: antioxidant capacity and quantitative structure-activity relationship predict reduction and promotion effects of flavonoids on acrylamide formation

    NASA Astrophysics Data System (ADS)

    Huang, Mengmeng; Wei, Yan; Wang, Jun; Zhang, Yu

    2016-09-01

    We used the support vector regression (SVR) approach to predict and unravel reduction/promotion effect of characteristic flavonoids on the acrylamide formation under a low-moisture Maillard reaction system. Results demonstrated the reduction/promotion effects by flavonoids at addition levels of 1–10000 μmol/L. The maximal inhibition rates (51.7%, 68.8% and 26.1%) and promote rates (57.7%, 178.8% and 27.5%) caused by flavones, flavonols and isoflavones were observed at addition levels of 100 μmol/L and 10000 μmol/L, respectively. The reduction/promotion effects were closely related to the change of trolox equivalent antioxidant capacity (ΔTEAC) and well predicted by triple ΔTEAC measurements via SVR models (R: 0.633–0.900). Flavonols exhibit stronger effects on the acrylamide formation than flavones and isoflavones as well as their O-glycosides derivatives, which may be attributed to the number and position of phenolic and 3-enolic hydroxyls. The reduction/promotion effects were well predicted by using optimized quantitative structure-activity relationship (QSAR) descriptors and SVR models (R: 0.926–0.994). Compared to artificial neural network and multi-linear regression models, SVR models exhibited better fitting performance for both TEAC-dependent and QSAR descriptor-dependent predicting work. These observations demonstrated that the SVR models are competent for predicting our understanding on the future use of natural antioxidants for decreasing the acrylamide formation.

  11. The hexanoyl-CoA precursor for cannabinoid biosynthesis is formed by an acyl-activating enzyme in Cannabis sativa trichomes.

    PubMed

    Stout, Jake M; Boubakir, Zakia; Ambrose, Stephen J; Purves, Randy W; Page, Jonathan E

    2012-08-01

    The psychoactive and analgesic cannabinoids (e.g. Δ(9) -tetrahydrocannabinol (THC)) in Cannabis sativa are formed from the short-chain fatty acyl-coenzyme A (CoA) precursor hexanoyl-CoA. Cannabinoids are synthesized in glandular trichomes present mainly on female flowers. We quantified hexanoyl-CoA using LC-MS/MS and found levels of 15.5 pmol g(-1) fresh weight in female hemp flowers with lower amounts in leaves, stems and roots. This pattern parallels the accumulation of the end-product cannabinoid, cannabidiolic acid (CBDA). To search for the acyl-activating enzyme (AAE) that synthesizes hexanoyl-CoA from hexanoate, we analyzed the transcriptome of isolated glandular trichomes. We identified 11 unigenes that encoded putative AAEs including CsAAE1, which shows high transcript abundance in glandular trichomes. In vitro assays showed that recombinant CsAAE1 activates hexanoate and other short- and medium-chained fatty acids. This activity and the trichome-specific expression of CsAAE1 suggest that it is the hexanoyl-CoA synthetase that supplies the cannabinoid pathway. CsAAE3 encodes a peroxisomal enzyme that activates a variety of fatty acid substrates including hexanoate. Although phylogenetic analysis showed that CsAAE1 groups with peroxisomal AAEs, it lacked a peroxisome targeting sequence 1 (PTS1) and localized to the cytoplasm. We suggest that CsAAE1 may have been recruited to the cannabinoid pathway through the loss of its PTS1, thereby redirecting it to the cytoplasm. To probe the origin of hexanoate, we analyzed the trichome expressed sequence tag (EST) dataset for enzymes of fatty acid metabolism. The high abundance of transcripts that encode desaturases and a lipoxygenase suggests that hexanoate may be formed through a pathway that involves the oxygenation and breakdown of unsaturated fatty acids.

  12. The genetics of extreme microgeographic adaptation: an integrated approach identifies a major gene underlying leaf trichome divergence in Yellowstone Mimulus guttatus.

    PubMed

    Hendrick, Margaret F; Finseth, Findley R; Mathiasson, Minna E; Palmer, Kristen A; Broder, Emma M; Breigenzer, Peter; Fishman, Lila

    2016-11-01

    Microgeographic adaptation provides a particularly interesting context for understanding the genetic basis of phenotypic divergence and may also present unique empirical challenges. In particular, plant adaptation to extreme soil mosaics may generate barriers to gene flow or shifts in mating system that confound simple genomic scans for adaptive loci. Here, we combine three approaches - quantitative trait locus (QTL) mapping of candidate intervals in controlled crosses, population resequencing (PoolSeq) and analyses of wild recombinant individuals - to investigate one trait associated with Mimulus guttatus (yellow monkeyflower) adaptation to geothermal soils in Yellowstone National Park. We mapped a major QTL causing dense leaf trichomes in thermally adapted plants to a <50-kb region of linkage Group 14 (Tr14) previously implicated in trichome divergence between independent M. guttatus populations. A PoolSeq scan of Tr14 region revealed a cluster of six genes, coincident with the inferred QTL peak, with high allele frequency differences sufficient to explain observed phenotypic differentiation. One of these, the R2R3 MYB transcription factor Migut.N02661, is a plausible functional candidate and was also strongly associated (r(2)  = 0.27) with trichome phenotype in analyses of wild-collected admixed individuals. Although functional analyses will be necessary to definitively link molecular variants in Tr14 with trichome divergence, our analyses are a major step in that direction. They point to a simple, and parallel, genetic basis for one axis of Mimulus guttatus adaptation to an extreme habitat, suggest a broadly conserved genetic basis for trichome variation across flowering plants and pave the way for further investigations of this challenging case of microgeographic incipient speciation.

  13. Epitope-tagged yeast strains reveal promoter driven changes to 3'-end formation and convergent antisense-transcription from common 3' UTRs.

    PubMed

    Swaminathan, Angavai; Beilharz, Traude H

    2016-01-08

    Epitope-tagging by homologous recombination is ubiquitously used to study gene expression, protein localization and function in yeast. This is generally thought to insulate the regulation of gene expression to that mediated by the promoter and coding regions because native 3' UTR are replaced. Here we show that the 3' UTRs, CYC1 and ADH1, contain cryptic promoters that generate abundant convergent antisense-transcription in Saccharomyces cerevisiae. Moreover we show that aberrant, truncating 3' -end formation is often associated with regulated transcription in TAP-tagged strains. Importantly, the steady-state level of both 3' -truncated and antisense transcription products is locus dependent. Using TAP and GFP-tagged strains we show that the transcriptional state of the gene-of-interest induces changes to 3' -end formation by alternative polyadenylation and antisense transcription from a universal 3' UTR. This means that these 3' UTRs contains plastic features that can be molded to reflect the regulatory architecture of the locus rather than bringing their own regulatory paradigm to the gene-fusions as would be expected. Our work holds a cautionary note for studies utilizing tagged strains for quantitative biology, but also provides a new model for the study of promoter driven rewiring of 3' -end formation and regulatory non-coding transcription.

  14. Injectable hydrogel promotes early survival of induced pluripotent stem cell-derived oligodendrocytes and attenuates longterm teratoma formation in a spinal cord injury model.

    PubMed

    Führmann, T; Tam, R Y; Ballarin, B; Coles, B; Elliott Donaghue, I; van der Kooy, D; Nagy, A; Tator, C H; Morshead, C M; Shoichet, M S

    2016-03-01

    Transplantation of pluripotent stem cells and their differentiated progeny has the potential to preserve or regenerate functional pathways and improve function after central nervous system injury. However, their utility has been hampered by poor survival and the potential to form tumors. Peptide-modified biomaterials influence cell adhesion, survival and differentiation in vitro, but their effectiveness in vivo remains uncertain. We synthesized a peptide-modified, minimally invasive, injectable hydrogel comprised of hyaluronan and methylcellulose to enhance the survival and differentiation of human induced pluripotent stem cell-derived oligodendrocyte progenitor cells. Cells were transplanted subacutely after a moderate clip compression rat spinal cord injury. The hydrogel, modified with the RGD peptide and platelet-derived growth factor (PDGF-A), promoted early survival and integration of grafted cells. However, prolific teratoma formation was evident when cells were transplanted in media at longer survival times, indicating that either this cell line or the way in which it was cultured is unsuitable for human use. Interestingly, teratoma formation was attenuated when cells were transplanted in the hydrogel, where most cells differentiated to a glial phenotype. Thus, this hydrogel promoted cell survival and integration, and attenuated teratoma formation by promoting cell differentiation.

  15. Effects of Plant Density on the Number of Glandular Trichomes and on Yield and Quality of Essential Oils from Oregano.

    PubMed

    Tuttolomondo, Teresa; La Bella, Salvatore; Leto, Claudio; Bonsangue, Giuseppe; Leone, Raffaele; Gennaro, Maria Cristina; Virga, Giuseppe; Inguanta, Rosalinda; Licata, Mario

    2016-06-01

    Plants yields are influenced by agronomic techniques. Plant density is a complex issue and extremely important when maximizing both crop quality, and biomass and essential oil yields. Plants belonging to the Origanum vulgare subspecies hirtum (Link) Ietswaart were grown adopting four types of plant density and were characterized in biometric and chemical terms. The samples were analyzed using the ANOVA (Principal Component Analysis) statistical method regarding biometric aspects, EO yield and peltate hair density. Essential oil (EO) was extracted by hydrodistillation and analyzed using GC-FID and GC-MS. GC-FID and GC-MS analysis led to the identification of 45 compounds from the EO. Plant density affected production both in terms of biomass and EO. However, it was not found to have affected peltate glandular trichome density or EO quality.

  16. A Model Formative Assessment Strategy to Promote Student-Centered Self-Regulated Learning in Higher Education

    ERIC Educational Resources Information Center

    Bose, Jayakumar; Rengel, Zed

    2009-01-01

    Adult learners are already involved in the process of self-regulation; hence, higher education institutions should focus on strengthening students' self-regulatory skills. Self-regulation can be facilitated through formative assessment. This paper proposes a model formative assessment strategy that would complement existing university teaching,…

  17. Synaptopodin-2 induces assembly of peripheral actin bundles and immature focal adhesions to promote lamellipodia formation and prostate cancer cell migration

    PubMed Central

    Kai, FuiBoon; Fawcett, James P.; Duncan, Roy

    2015-01-01

    Synaptopodin-2 (Synpo2), an actin-binding protein and invasive cancer biomarker, induces formation of complex stress fiber networks in the cell body and promotes PC3 prostate cancer cell migration in response to serum stimulation. The role of these actin networks in enhanced cancer cell migration is unknown. Using time-course analysis and live cell imaging of mock- and Synpo2-transduced PC3 cells, we now show that Synpo2 induces assembly of actin fibers near the cell periphery and Arp2/3-dependent lamellipodia formation. Lamellipodia formed in a non-directional manner or repeatedly changed direction, explaining the enhanced chemokinetic activity of PC3 cells in response to serum stimulation. Myosin contraction promotes retrograde flow of the Synpo2-associated actin filaments at the leading edge and their merger with actin networks in the cell body. Enhanced PC3 cell migration correlates with Synpo2-induced formation of lamellipodia and immature focal adhesions (FAs), but is not dependent on myosin contraction or FA maturation. The previously reported correlation between Synpo2-induced stress fiber assembly and enhanced PC3 cell migration therefore reflects the role of Synpo2 as a newly identified regulator of actin bundle formation and nascent FA assembly near the leading cell edge. PMID:25883213

  18. Myosin 1b promotes the formation of post-Golgi carriers by regulating actin assembly and membrane remodelling at the trans-Golgi network.

    PubMed

    Almeida, Claudia G; Yamada, Ayako; Tenza, Danièle; Louvard, Daniel; Raposo, Graça; Coudrier, Evelyne

    2011-06-12

    The function of organelles is intimately associated with rapid changes in membrane shape. By exerting force on membranes, the cytoskeleton and its associated motors have an important role in membrane remodelling. Actin and myosin 1 have been implicated in the invagination of the plasma membrane during endocytosis. However, whether myosin 1 and actin contribute to the membrane deformation that gives rise to the formation of post-Golgi carriers is unknown. Here we report that myosin 1b regulates the actin-dependent post-Golgi traffic of cargo, generates force that controls the assembly of F-actin foci and, together with the actin cytoskeleton, promotes the formation of tubules at the TGN. Our results provide evidence that actin and myosin 1 regulate organelle shape and uncover an important function for myosin 1b in the initiation of post-Golgi carrier formation by regulating actin assembly and remodelling TGN membranes.

  19. TolC promotes ExPEC biofilm formation and curli production in response to medium osmolarity.

    PubMed

    Hou, Bo; Meng, Xian-Rong; Zhang, Li-Yuan; Tan, Chen; Jin, Hui; Zhou, Rui; Gao, Jian-Feng; Wu, Bin; Li, Zi-Li; Liu, Mei; Chen, Huan-Chun; Bi, Ding-Ren; Li, Shao-Wen

    2014-01-01

    While a high osmolarity medium activates Cpx signaling and causes CpxR to repress csgD expression, and efflux protein TolC protein plays an important role in biofilm formation in Escherichia coli, whether TolC also responds to an osmolarity change to regulate biofilm formation in extraintestinal pathogenic E. coli (ExPEC) remains unknown. In this study, we constructed ΔtolC mutant and complement ExPEC strains to investigate the role of TolC in the retention of biofilm formation and curli production capability under different osmotic conditions. The ΔtolC mutant showed significantly decreased biofilm formation and lost the ability to produce curli fimbriae compared to its parent ExPEC strain PPECC42 when cultured in M9 medium or 1/2 M9 medium of increased osmolarity with NaCl or sucrose at 28°C. However, biofilm formation and curli production levels were restored to wild-type levels in the ΔtolC mutant in 1/2 M9 medium. We propose for the first time that TolC protein is able to form biofilm even under high osmotic stress. Our findings reveal an interplay between the role of TolC in ExPEC biofilm formation and the osmolarity of the surrounding environment, thus providing guidance for the development of a treatment for ExPEC biofilm formation.

  20. The primary function of gp130 signaling in osteoblasts is to maintain bone formation and strength, rather than promote osteoclast formation.

    PubMed

    Johnson, Rachelle W; Brennan, Holly J; Vrahnas, Christina; Poulton, Ingrid J; McGregor, Narelle E; Standal, Therese; Walker, Emma C; Koh, Thuan-Tzen; Nguyen, Huynh; Walsh, Nicole C; Forwood, Mark R; Martin, T John; Sims, Natalie A

    2014-06-01

    Interleukin-6 (IL-6) family cytokines act via gp130 in the osteoblast lineage to stimulate the formation of osteoclasts (bone resorbing cells) and the activity of osteoblasts (bone forming cells), and to inhibit expression of the osteocyte protein, sclerostin. We report here that a profound reduction in trabecular bone mass occurs both when gp130 is deleted in the entire osteoblast lineage (Osx1Cre gp130 f/f) and when this deletion is restricted to osteocytes (DMP1Cre gp130 f/f). This was caused not by an alteration in osteoclastogenesis, but by a low level of bone formation specific to the trabecular compartment. In contrast, cortical diameter increased to maintain ultimate bone strength, despite a reduction in collagen type 1 production. We conclude that osteocytic gp130 signaling is required for normal trabecular bone mass and proper cortical bone composition.

  1. The BR domain of PsrP interacts with extracellular DNA to promote bacterial aggregation; structural insights into pneumococcal biofilm formation

    PubMed Central

    Schulte, Tim; Mikaelsson, Cecilia; Beaussart, Audrey; Kikhney, Alexey; Deshmukh, Maya; Wolniak, Sebastian; Pathak, Anuj; Ebel, Christine; Löfling, Jonas; Fogolari, Federico; Henriques-Normark, Birgitta; Dufrêne, Yves F.; Svergun, Dmitri; Nygren, Per-Åke; Achour, Adnane

    2016-01-01

    The major human pathogen Streptococcus pneumoniae is a leading cause of disease and death worldwide. Pneumococcal biofilm formation within the nasopharynx leads to long-term colonization and persistence within the host. We have previously demonstrated that the capsular surface-associated pneumococcal serine rich repeat protein (PsrP), key factor for biofilm formation, binds to keratin-10 (KRT10) through its microbial surface component recognizing adhesive matrix molecule (MSCRAMM)-related globular binding region domain (BR187–385). Here, we show that BR187–385 also binds to DNA, as demonstrated by electrophoretic mobility shift assays and size exclusion chromatography. Further, heterologous expression of BR187–378 or the longer BR120–378 construct on the surface of a Gram-positive model host bacterium resulted in the formation of cellular aggregates that was significantly enhanced in the presence of DNA. Crystal structure analyses revealed the formation of BR187–385 homo-dimers via an intermolecular β-sheet, resulting in a positively charged concave surface, shaped to accommodate the acidic helical DNA structure. Furthermore, small angle X-ray scattering and circular dichroism studies indicate that the aggregate-enhancing N-terminal region of BR120–166 adopts an extended, non-globular structure. Altogether, our results suggest that PsrP adheres to extracellular DNA in the biofilm matrix and thus promotes pneumococcal biofilm formation. PMID:27582320

  2. Promotion of peripheral nerve regeneration and prevention of neuroma formation by PRGD/PDLLA/β-TCP conduit: report of two cases.

    PubMed

    Yin, Yixia; Li, Binbin; Yan, Qiongjiao; Dai, Honglian; Wang, Xinyu; Huang, Jifeng; Li, Shipu

    2015-06-01

    In the field of nerve repair, one major challenge is the formation of neuroma. However, reports on both the promotion of nerve regeneration and prevention of traumatic neuroma in the clinical settings are rare in the field of nerve repair. One of the reasons could be the insufficiency in the follow-up system. We have conducted 33 cases of nerve repair using PRGD/PDLLA/β-TCP conduit without any sign of adverse reaction, especially no neuroma formation. Among them, we have selected two cases as representatives to report in this article. The first case was a patient with an upper limb nerve wound was bridged by PRGD/PDLLA/β-TCP conduit and a plate fixation was given. After nearly 3-years' follow-up, the examination results demonstrated that nerve regeneration effect was very good. When the reoperation was performed to remove the steel plate we observed a uniform structure of the regenerated nerve without the formation of neuroma, and to our delight, the implanted conduit was completely degraded 23 months after the implantation. The second case had an obsolete nerve injury with neuroma formation. After removal of the neuroma, the nerve was bridged by PRGD/PDLLA/β-TCP conduit. Follow-up examinations showed that the structure and functional recovery were improved gradually in the 10-month follow-up; no end-enlargement and any other abnormal reaction associated with the characteristic of neuroma were found. Based on our 33-case studies, we have concluded that PRGD/PDLLA/β-TCP nerve conduit could both promote nerve regeneration and prevent neuroma formation; therefore, it is a good alternative for peripheral nerve repair.

  3. Tumor Necrosis Factor Receptor-associated Factor 6 (TRAF6) Associates with Huntingtin Protein and Promotes Its Atypical Ubiquitination to Enhance Aggregate Formation*

    PubMed Central

    Zucchelli, Silvia; Marcuzzi, Federica; Codrich, Marta; Agostoni, Elena; Vilotti, Sandra; Biagioli, Marta; Pinto, Milena; Carnemolla, Alisia; Santoro, Claudio; Gustincich, Stefano; Persichetti, Francesca

    2011-01-01

    Huntington disease (HD) is a neurodegenerative disorder caused by an expansion of polyglutamines in the first exon of huntingtin (HTT), which confers aggregation-promoting properties to amino-terminal fragments of the protein (N-HTT). Mutant N-HTT aggregates are enriched for ubiquitin and contain ubiquitin E3 ligases, thus suggesting a role for ubiquitination in aggregate formation. Here, we report that tumor necrosis factor receptor-associated factor 6 (TRAF6) binds to WT and polyQ-expanded N-HTT in vitro as well as to endogenous full-length proteins in mouse and human brain in vivo. Endogenous TRAF6 is recruited to cellular inclusions formed by mutant N-HTT. Transient overexpression of TRAF6 promotes WT and mutant N-HTT atypical ubiquitination with Lys6, Lys27, and Lys29 linkage formation. Both interaction and ubiquitination seem to be independent from polyQ length. In cultured cells, TRAF6 enhances mutant N-HTT aggregate formation, whereas it has no effect on WT N-HTT protein localization. Mutant N-HTT inclusions are enriched for ubiquitin staining only when TRAF6 and Lys6, Lys27, and Lys29 ubiquitin mutants are expressed. Finally, we show that TRAF6 is up-regulated in post-mortem brains from HD patients where it is found in the insoluble fraction. These results suggest that TRAF6 atypical ubiquitination warrants investigation in HD pathogenesis. PMID:21454471

  4. Tumor necrosis factor receptor-associated factor 6 (TRAF6) associates with huntingtin protein and promotes its atypical ubiquitination to enhance aggregate formation.

    PubMed

    Zucchelli, Silvia; Marcuzzi, Federica; Codrich, Marta; Agostoni, Elena; Vilotti, Sandra; Biagioli, Marta; Pinto, Milena; Carnemolla, Alisia; Santoro, Claudio; Gustincich, Stefano; Persichetti, Francesca

    2011-07-15

    Huntington disease (HD) is a neurodegenerative disorder caused by an expansion of polyglutamines in the first exon of huntingtin (HTT), which confers aggregation-promoting properties to amino-terminal fragments of the protein (N-HTT). Mutant N-HTT aggregates are enriched for ubiquitin and contain ubiquitin E3 ligases, thus suggesting a role for ubiquitination in aggregate formation. Here, we report that tumor necrosis factor receptor-associated factor 6 (TRAF6) binds to WT and polyQ-expanded N-HTT in vitro as well as to endogenous full-length proteins in mouse and human brain in vivo. Endogenous TRAF6 is recruited to cellular inclusions formed by mutant N-HTT. Transient overexpression of TRAF6 promotes WT and mutant N-HTT atypical ubiquitination with Lys(6), Lys(27), and Lys(29) linkage formation. Both interaction and ubiquitination seem to be independent from polyQ length. In cultured cells, TRAF6 enhances mutant N-HTT aggregate formation, whereas it has no effect on WT N-HTT protein localization. Mutant N-HTT inclusions are enriched for ubiquitin staining only when TRAF6 and Lys(6), Lys(27), and Lys(29) ubiquitin mutants are expressed. Finally, we show that TRAF6 is up-regulated in post-mortem brains from HD patients where it is found in the insoluble fraction. These results suggest that TRAF6 atypical ubiquitination warrants investigation in HD pathogenesis.

  5. Met promotes the formation of double minute chromosomes induced by Sei-1 in NIH-3T3 murine fibroblasts

    PubMed Central

    You, Jia; Wu, Di; Yu, Yang; Liu, Chang; Wang, Lei; Wang, Fei; Xu, Lu; Wang, Liqun; Wang, Nan; Tian, Xing; Wang, Falin; Liang, Hongbin; Gao, Yating; Cui, Xiaobo; Ji, Guohua; Bai, Jing; Yu, Jingcui; Meng, Xiangning; Jin, Yan; Sun, Wenjing; Guan, Xin-yuan; Zhang, Chunyu; Fu, Songbin

    2016-01-01

    Background Sei-1 is an oncogene capable of inducing double minute chromosomes (DMs) formation. DMs are hallmarks of amplification and contribute to oncogenesis. However, the mechanism of Sei-1 inducing DMs formation remains unelucidated. Results DMs formation significantly increased during serial passage in vivo and gradually decreased following culture in vitro. micro nuclei (MN) was found to be responsible for the reduction. Of the DMs-carrying genes, Met was found to be markedly amplified, overexpressed and highly correlated with DMs formation. Inhibition of Met signaling decreased the number of DMs and reduced the amplification of the DMs-carrying genes. We identified a 3.57Mb DMs representing the majority population, which consists of the 1.21 Mb AMP1 from locus 6qA2 and the 2.36 Mb AMP2 from locus 6qA2-3. Materials and Methods We employed NIH-3T3 cell line with Sei-1 overexpression to monitor and characterize DMs in vivo and in vitro. Array comparative genome hybridization (aCGH) and fluorescence in situ hybridization (FISH) were performed to reveal amplification regions and DMs-carrying genes. Metaphase spread was prepared to count the DMs. Western blot and Met inhibition rescue experiments were performed to examine for involvement of altered Met signaling in Sei-1 induced DMs. Genomic walking and PCR were adopted to reveal DMs structure. Conclusions Met is an important promotor of DMs formation. PMID:27494853

  6. Cooperativity between Al Sites Promotes Hydrogen Transfer and Carbon–Carbon Bond Formation upon Dimethyl Ether Activation on Alumina

    PubMed Central

    2015-01-01

    The methanol-to-olefin (MTO) process allows the conversion of methanol/dimethyl ether into olefins on acidic zeolites via the so-called hydrocarbon pool mechanism. However, the site and mechanism of formation of the first carbon–carbon bond are still a matter of debate. Here, we show that the Lewis acidic Al sites on the 110 facet of γ-Al2O3 can readily activate dimethyl ether to yield CH4, alkenes, and surface formate species according to spectroscopic studies combined with a computational approach. The carbon–carbon forming step as well as the formation of methane and surface formate involves a transient oxonium ion intermediate, generated by a hydrogen transfer between surface methoxy species and coordinated methanol on adjacent Al sites. These results indicate that extra framework Al centers in acidic zeolites, which are associated with alumina, can play a key role in the formation of the first carbon–carbon bond, the initiation step of the industrial MTO process. PMID:27162986

  7. Fibrinogen triggers astrocyte scar formation by promoting the availability of active TGF-β after vascular damage

    PubMed Central

    Schachtrup, Christian; Ryu, Jae K.; Helmrick, Matthew; Vagena, Eirini; Galanakis, Dennis K.; Degen, Jay L.; Margolis, Richard U.; Akassoglou, Katerina

    2010-01-01

    Scar formation in the nervous system begins within hours after traumatic injury and is characterized primarily by reactive astrocytes depositing proteoglycans that inhibit regeneration. A fundamental question in CNS repair has been the identity of the initial molecular mediator that triggers glial scar formation. Here we show that the blood protein fibrinogen, which leaks into the CNS immediately after blood-brain barrier (BBB) disruption or vascular damage, serves as an early signal for the induction of glial scar formation via the TGF-β/Smad signaling pathway. Our studies revealed that fibrinogen is a carrier of latent TGF-β and induces phosphorylation of Smad2 in astrocytes that leads to inhibition of neurite outgrowth. Consistent with these findings, genetic or pharmacologic depletion of fibrinogen in mice reduces active TGF-β, Smad2 phosphorylation, glial cell activation and neurocan deposition following cortical injury. Furthermore, stereotactic injection of fibrinogen into the mouse cortex is sufficient to induce astrogliosis. Inhibition of the TGF-β receptor pathway abolishes the fibrinogen-induced effects on glial scar formation in vivo and in vitro. These results identify fibrinogen as a primary astrocyte activation signal, provide evidence that deposition of inhibitory proteoglycans is induced by a blood protein that leaks in the CNS after vasculature rupture, and point to TGF-β as a molecular link between vascular permeability and scar formation. PMID:20427645

  8. In Vitro Oxidation of Collagen Promotes the Formation of Advanced Oxidation Protein Products and the Activation of Human Neutrophils.

    PubMed

    Bochi, Guilherme Vargas; Torbitz, Vanessa Dorneles; de Campos, Luízi Prestes; Sangoi, Manuela Borges; Fernandes, Natieli Flores; Gomes, Patrícia; Moretto, Maria Beatriz; Barbisan, Fernanda; da Cruz, Ivana Beatrice Mânica; Moresco, Rafael Noal

    2016-04-01

    The accumulation of advanced oxidation protein products (AOPPs) has been linked to several pathological conditions. Here, we investigated collagen as a potential source for AOPP formation and determined the effects of hypochlorous acid (HOCl)-treated collagen (collagen-AOPPs) on human neutrophil activity. We also assessed whether alpha-tocopherol could counteract these effects. Exposure to HOCl increased the levels of collagen-AOPPs. Collagen-AOPPs also stimulated the production of AOPPs, nitric oxide (NO), superoxide radicals (O2(-)), and HOCl by neutrophils. Collagen-AOPPs induced apoptosis and decreased the number of viable cells. Alpha-tocopherol prevented the formation of collagen-AOPPs, strongly inhibited the collagen-AOPP-induced production of O2(-) and HOCl, and increased the viability of neutrophils. Our results suggest that collagen is an important protein that interacts with HOCl to form AOPPs, and consequently, collagen-AOPP formation is related to human neutrophil activation and cell death.

  9. Protein kinase D promotes plasticity-induced F-actin stabilization in dendritic spines and regulates memory formation

    PubMed Central

    Bencsik, Norbert; Szíber, Zsófia; Liliom, Hanna; Tárnok, Krisztián; Borbély, Sándor; Gulyás, Márton; Rátkai, Anikó; Szűcs, Attila; Hazai-Novák, Diána; Ellwanger, Kornelia; Rácz, Bence; Pfizenmaier, Klaus; Hausser, Angelika

    2015-01-01

    Actin turnover in dendritic spines influences spine development, morphology, and plasticity, with functional consequences on learning and memory formation. In nonneuronal cells, protein kinase D (PKD) has an important role in stabilizing F-actin via multiple molecular pathways. Using in vitro models of neuronal plasticity, such as glycine-induced chemical long-term potentiation (LTP), known to evoke synaptic plasticity, or long-term depolarization block by KCl, leading to homeostatic morphological changes, we show that actin stabilization needed for the enlargement of dendritic spines is dependent on PKD activity. Consequently, impaired PKD functions attenuate activity-dependent changes in hippocampal dendritic spines, including LTP formation, cause morphological alterations in vivo, and have deleterious consequences on spatial memory formation. We thus provide compelling evidence that PKD controls synaptic plasticity and learning by regulating actin stability in dendritic spines. PMID:26304723

  10. Acetylcholine Protects against Candida albicans Infection by Inhibiting Biofilm Formation and Promoting Hemocyte Function in a Galleria mellonella Infection Model

    PubMed Central

    Rajendran, Ranjith; Borghi, Elisa; Falleni, Monica; Perdoni, Federica; Tosi, Delfina; Lappin, David F.; O'Donnell, Lindsay; Greetham, Darren; Ramage, Gordon

    2015-01-01

    Both neuronal acetylcholine and nonneuronal acetylcholine have been demonstrated to modulate inflammatory responses. Studies investigating the role of acetylcholine in the pathogenesis of bacterial infections have revealed contradictory findings with regard to disease outcome. At present, the role of acetylcholine in the pathogenesis of fungal infections is unknown. Therefore, the aim of this study was to determine whether acetylcholine plays a role in fungal biofilm formation and the pathogenesis of Candida albicans infection. The effect of acetylcholine on C. albicans biofilm formation and metabolism in vitro was assessed using a crystal violet assay and phenotypic microarray analysis. Its effect on the outcome of a C. albicans infection, fungal burden, and biofilm formation were investigated in vivo using a Galleria mellonella infection model. In addition, its effect on modulation of host immunity to C. albicans infection was also determined in vivo using hemocyte counts, cytospin analysis, larval histology, lysozyme assays, hemolytic assays, and real-time PCR. Acetylcholine was shown to have the ability to inhibit C. albicans biofilm formation in vitro and in vivo. In addition, acetylcholine protected G. mellonella larvae from C. albicans infection mortality. The in vivo protection occurred through acetylcholine enhancing the function of hemocytes while at the same time inhibiting C. albicans biofilm formation. Furthermore, acetylcholine also inhibited inflammation-induced damage to internal organs. This is the first demonstration of a role for acetylcholine in protection against fungal infections, in addition to being the first report that this molecule can inhibit C. albicans biofilm formation. Therefore, acetylcholine has the capacity to modulate complex host-fungal interactions and plays a role in dictating the pathogenesis of fungal infections. PMID:26092919

  11. Sterol and triterpene derivatives from plants inhibit the effects of a tumor promoter, and sitosterol and betulinic acid inhibit tumor formation in mouse skin two-stage carcinogenesis.

    PubMed

    Yasukawa, K; Takido, M; Matsumoto, T; Takeuchi, M; Nakagawa, S

    1991-01-01

    A single topical application of 1 microgram of 12-O-tetradecanoylphorbol- 13-acetate (TPA) to the ears of mice was shown to induce edema, and this TPA-induced inflammation was inhibited by 4-methylsterol and triterpene derivatives. The ED50 of these compounds against TPA-induced inflammation was 0.1-3 mumol. Phytosterols had only slight inhibitory effects. Furthermore, application of 5 micrograms TPA to mouse skin rapidly caused accumulation of ornithine decarboxylase (ODC). Similarly, sitosterol and lupane-type triterpene derivatives markedly inhibited this TPA-induced ODC accumulation. In addition, 5 mumol betulinic acid markedly inhibited the promoting effect of 2.5 micrograms TPA applied twice weekly on skin tumor formation in mice initiated with 50 micrograms of 7,12-dimethylbenz[a]anthracene, and 5 mumol of sitosterol caused slight suppression. Thus, the inhibitory effects of sterol and triterpene derivatives on TPA-induced inflammation roughly parallelled their inhibitory activities against tumor promotion.

  12. Mechanistic insights on iodine(III) promoted metal-free dual C-H activation involved in the formation of a spirocyclic bis-oxindole.

    PubMed

    Sreenithya, A; Sunoj, Raghavan B

    2014-12-05

    The mechanism of a metal-free, phenyliodine(III) bis(trifluoroacetate) promoted, dual aryl C-H activation of an anilide to a spirocyclic bis-oxindole is examined using density functional theory (M06-2X). The most preferred pathway proceeds through the involvement of a novel iodonium ion intermediate and a pivotal trifluoroacetate counterion. The two sequential aryl C-H activations, assisted by trifluoroacetate as well as the superior leaving group ability of PhI, facilitate the formation of spirocyclic bis-oxindole.

  13. Task-Appropriate Visualizations: Can the Very Same Visualization Format Either Promote or Hinder Learning Depending on the Task Requirements?

    ERIC Educational Resources Information Center

    Soemer, Alexander; Schwan, Stephan

    2016-01-01

    In a series of experiments, we tested a recently proposed hypothesis stating that the degree of alignment between the form of a mental representation resulting from learning with a particular visualization format and the specific requirements of a learning task determines learning performance (task-appropriateness). Groups of participants were…

  14. An Exploratory Study of a Novel Online Formative Assessment and Instructional Tool to Promote Students' Circuit Problem Solving

    ERIC Educational Resources Information Center

    Chung, Gregory K. W. K.; Shel, Tammy; Kaiser, William J.

    2006-01-01

    We examined a novel formative assessment and instructional approach with 89 students in three electrical engineering classes in special computer-based discussion sections. The technique involved students individually solving circuit problems online, with their real-time responses observed by the instructor. While exploratory, survey and…

  15. A Formative Evaluation of Customized Pamphlets to Promote Physical Activity and Symptom Self-Management in Women with Multiple Sclerosis

    ERIC Educational Resources Information Center

    Plow, Matthew; Bethoux, Francois; Mai, Kimloan; Marcus, Bess

    2014-01-01

    Inactivity is a prevalent problem in the population affected with multiple sclerosis (MS). Thus, there is a need to develop and test physical activity (PA) interventions that can be widely disseminated. We conducted a formative evaluation as part of a randomized controlled trial of a pamphlet-based PA intervention among 30 women with MS. Pamphlets…

  16. Ammonia Catalyzed Formation of Sulfuric Acid in Troposphere: The Curious Case of A Base Promoting Acid Rain.

    PubMed

    Bandyopadhyay, Biman; Kumar, Pradeep; Biswas, Partha

    2017-04-03

    Electronic structure calculations have been performed to investigate the role of ammonia in catalyzing the formation of sulfuric acid through hydrolysis of SO3 in Earth's atmosphere. The uncatalyzed process involves a high activation barrier and, till date, is mainly known to occur in Earth's atmosphere only when catalyzed by water and acids. Here we show that hydrolysis of SO3 can be very efficiently catalyzed by ammonia, the most abundant basic component in Earth's atmosphere. It was found, based on magnitude of relative potential energies as well as rate coefficients, that ammonia is the best among all the catalysts studied until now (water and acids) and could be a considerable factor in formation of sulfuric acid in troposphere. The calculated rate coefficient (at 298 K) of ammonia catalyzed reaction has been found to be ~10^5 - 10^7 times greater than that for water catalyzed ones. It was found, based on relative rates of ammonia and water catalyzed processes that in troposphere ammonia, together with water, could be the key factor in determining the rate of formation of sulfuric acid. In fact ammonia could surpass water in catalyzing formation of sulfuric acid via hydrolysis of SO3 at various altitudes in troposphere depending upon their relative concentrations.

  17. Formative evaluation for promoting adoption of the DGA, 2005 among African American parents and children in the Lower Mississippi Delta

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Formative research was conducted to increase adherence to the healthful food and physical activity patterns set forth in the Dietary Guidelines for Americans, 2005 (DGA, 2005) and thereby reduce weight gain and risk factors for obesity-related chronic diseases in African American parents and their c...

  18. Differential microRNA Analysis of Glandular Trichomes and Young Leaves in Xanthium strumarium L. Reveals Their Putative Roles in Regulating Terpenoid Biosynthesis

    PubMed Central

    Fan, Rongyan; Li, Yuanjun; Li, Changfu; Zhang, Yansheng

    2015-01-01

    The medicinal plant Xanthium strumarium L. (X. strumarium) is covered with glandular trichomes, which are the sites for synthesizing pharmacologically active terpenoids such as xanthatin. MicroRNAs (miRNAs) are a class of 21–24 nucleotide (nt) non-coding RNAs, most of which are identified as regulators of plant growth development. Identification of miRNAs involved in the biosynthesis of plant secondary metabolites remains limited. In this study, high-throughput Illumina sequencing, combined with target gene prediction, was performed to discover novel and conserved miRNAs with potential roles in regulating terpenoid biosynthesis in X. strumarium glandular trichomes. Two small RNA libraries from leaves and glandular trichomes of X. strumarium were established. In total, 1,185 conserved miRNAs and 37 novel miRNAs were identified, with 494 conserved miRNAs and 18 novel miRNAs being differentially expressed between the two tissue sources. Based on the X. strumarium transcriptome data that we recently constructed, 3,307 annotated mRNA transcripts were identified as putative targets of the differentially expressed miRNAs. KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analysis suggested that some of the differentially expressed miRNAs, including miR6435, miR5021 and miR1134, might be involved in terpenoid biosynthesis in the X. strumarium glandular trichomes. This study provides the first comprehensive analysis of miRNAs in X. strumarium, which forms the basis for further understanding of miRNA-based regulation on terpenoid biosynthesis. PMID:26406988

  19. TGF-β1 promotes linear invadosome formation in hepatocellular carcinoma cells, through DDR1 up-regulation and collagen I cross-linking.

    PubMed

    Ezzoukhry, Zakaria; Henriet, Elodie; Piquet, Léo; Boyé, Kevin; Bioulac-Sage, Paulette; Balabaud, Charles; Couchy, Gabrielle; Zucman-Rossi, Jessica; Moreau, Violaine; Saltel, Frédéric

    2016-11-01

    Transforming growth factor-β1 (TGF-β1) is an important player in chronic liver diseases inducing fibrogenesis and hepatocellular carcinoma (HCC) development. TGF-β1 promotes pleiotropic modifications at the cellular and matrix microenvironment levels. TGF-β1 was described to enhance production of type I collagen and its associated cross-linking enzyme, the lysyl oxidase-like2 (LOXL2). In addition, TGF-β1 and type I collagen are potent inducers of invadosomes. Indeed, type I collagen fibers induce the formation of active linear invadosomes through the discoidin domain receptor 1 (DDR1). The goal of our study was to address the role of TGF-β1 in collagen cross-linking and its impact on the formation of linear invadosomes in liver cancer cells. We first report a significant correlation between expressions of TGF-β1, and type I collagen, LOXL2, DDR1 and MT1-MMP in human HCCs. We demonstrate that TGF-β1 promotes a Smad4-dependent up-regulation of DDR1, together with LOXL2, in cultured HCC cells. Moreover, we show that LOXL2-induced collagen cross-linking enhances linear invadosome formation. Altogether, our data demonstrate that TGF-β1 favors linear invadosome formation through the expressions of both the inducers, such as collagen and LOXL2, and the components such as DDR1 and MT1-MMP of linear invadosomes in cancer cells. Meanwhile, our data uncover a new TGF-β1-dependent regulation of DDR1 expression.

  20. A Sensitized Screen for Genes Promoting Invadopodia Function In Vivo: CDC-42 and Rab GDI-1 Direct Distinct Aspects of Invadopodia Formation.

    PubMed

    Lohmer, Lauren L; Clay, Matthew R; Naegeli, Kaleb M; Chi, Qiuyi; Ziel, Joshua W; Hagedorn, Elliott J; Park, Jieun E; Jayadev, Ranjay; Sherwood, David R

    2016-01-01

    Invadopodia are specialized membrane protrusions composed of F-actin, actin regulators, signaling proteins, and a dynamically trafficked invadopodial membrane that drive cell invasion through basement membrane (BM) barriers in development and cancer. Due to the challenges of studying invasion in vivo, mechanisms controlling invadopodia formation in their native environments remain poorly understood. We performed a sensitized genome-wide RNAi screen and identified 13 potential regulators of invadopodia during anchor cell (AC) invasion into the vulval epithelium in C. elegans. Confirming the specificity of this screen, we identified the Rho GTPase cdc-42, which mediates invadopodia formation in many cancer cell lines. Using live-cell imaging, we show that CDC-42 localizes to the AC-BM interface and is activated by an unidentified vulval signal(s) that induces invasion. CDC-42 is required for the invasive membrane localization of WSP-1 (N-WASP), a CDC-42 effector that promotes polymerization of F-actin. Loss of CDC-42 or WSP-1 resulted in fewer invadopodia and delayed BM breaching. We also characterized a novel invadopodia regulator, gdi-1 (Rab GDP dissociation inhibitor), which mediates membrane trafficking. We show that GDI-1 functions in the AC to promote invadopodia formation. In the absence of GDI-1, the specialized invadopodial membrane was no longer trafficked normally to the invasive membrane, and instead was distributed to plasma membrane throughout the cell. Surprisingly, the pro-invasive signal(s) from the vulval cells also controls GDI-1 activity and invadopodial membrane trafficking. These studies represent the first in vivo screen for genes regulating invadopodia and demonstrate that invadopodia formation requires the integration of distinct cellular processes that are coordinated by an extracellular cue.

  1. ‘And then there were three’: highly efficient uptake of potassium by foliar trichomes of epiphytic bromeliads

    PubMed Central

    Winkler, Uwe; Zotz, Gerhard

    2010-01-01

    Background and Aims Vascular epiphytes have to acquire nutrients from atmospheric wash out, stem-flow, canopy soils and trapped litter. Physiological studies on the adaptations to nutrient acquisition and plant utilization of nutrients have focused on phosphorus and nitrogen; potassium, as a third highly abundant nutrient element, has received minor attention. In the present study, potassium uptake kinetics by leaves, within-plant distribution and nutrient accumulation were analysed to gain an improved understanding of physiological adaptations to non-terrestrial nutrient supply of plants. Methods Radioactively labelled 86RbCl was used as an analogue to study uptake kinetics of potassium absorbed from tanks of epiphytes, its plant distribution and the correlation between uptake efficiency and abundance of trichomes, functioning as uptake organs of leaves. Potassium in leaves was additionally analysed by atomic absorption spectroscopy to assess plant responses to potassium deficiency. Key Results Labelled rubidium was taken up from tanks over a wide range of concentrations, 0·01–90 mm, which was achieved by two uptake systems. In four tank epiphytes, the high-affinity transporters had average Km values of 41·2 µm, and the low-affinity transporters average Km values of 44·8 mm. Further analysis in Vriesea splenriet showed that high-affinity uptake of rubidium was an ATP-dependent process, while low-affinity uptake was mediated by a K+-channel. The kinetic properties of both types of transporters are comparable with those of potassium transporters in roots of terrestrial plants. Specific differences in uptake velocities of epiphytes are correlated with the abundance of trichomes on their leaf surfaces. The main sinks for potassium were fully grown leaves. These leaves thus function as internal potassium sources, which allow growth to be maintained during periods of low external potassium availability. Conclusions Vascular epiphytes possess effective mechanisms

  2. Effect of carbodiimide-derivatized hyaluronic acid gelatin on preventing postsurgical intra-abdominal adhesion formation and promoting healing in a rat model.

    PubMed

    Yuan, Fang; Lin, Long-Xiang; Zhang, Hui-Hui; Huang, Dan; Sun, Yu-Long

    2016-05-01

    Adhesions often occur after abdominal surgery. It could cause chronic pelvic pain, intestinal obstruction, and infertility. A hydrogel biomaterial, carbodiimide-derivatized hyaluronic acid gelatin (cd-HA gelatin), has been successfully used to reduce adhesion formation after flexor tendon grafting. This study investigated the efficacy of cd-HA gelatin in preventing postsurgical peritoneal adhesions in a rat model. The surgical traumas were created on the underlying muscle of the abdominal wall and the serosal layer of the cecum. The wounds were covered with or without cd-HA gelatin. Animals were euthanized at day 14 after surgery. Adhesion formation was assessed with adhesion degree and adhesion breaking strength. The healing of abdominal wall was evaluated with biomechanical testing and histological analysis. The adhesions occurred in all rats (n = 12) without cd-HA gelatin treatment. The application of cd-HA gelatin significantly reduced the adhesion rate from 100% to 58%. The decrease of adhesion breaking strength also manifested that cd-HA gelatin could reduce postsurgical intra-abdominal adhesion formation. Moreover, it was found that cd-HA gelatin was a safe material and could promote tissue healing. The cd-HA gelatin hydrogel could reduce the formation of intra-abdominal adhesions without adversely effects on wound healing.

  3. miR-29b promotes skin wound healing and reduces excessive scar formation by inhibition of the TGF-β1/Smad/CTGF signaling pathway.

    PubMed

    Guo, Jingdong; Lin, Quan; Shao, Ying; Rong, Li; Zhang, Duo

    2017-04-01

    The hypertrophic scar is a medical difficulty of humans, which has caused great pain to patients. Here, we investigated the inhibitory effect of miR-29b on scar formation. The scalded model was established in mice and miR-29b mimics or a negative control was subcutaneously injected into the injury skin. Then various molecular biological experiments were performed to assess the effect of miR-29b on scar formation. According to our present study, first, the results demonstrated that miR-29b was down-regulated in thermal injury tissue and miR-29b treatment could promote wound healing, inhibit scar formation, and alleviate histopathological morphologic alteration in scald tissues. Additionally, miR-29b treatment suppressed collagen deposition and fibrotic gene expression in scar tissues. Finally, we found that miR-29b treatment inhibited the TGF-β1/Smad/CTGF signaling pathway. Taken together, our data suggest that miR-29b treatment has an inhibitory effect against scar formation via inhibition of the TGF-β1/Smad/CTGF signaling pathway and may provide a potential molecular basis for future treatments for hypertrophic scars.

  4. Methylerythritol and mevalonate pathway contributions to biosynthesis of mono-, sesqui-, and diterpenes in glandular trichomes and leaves of Stevia rebaudiana Bertoni.

    PubMed

    Wölwer-Rieck, Ursula; May, Bianca; Lankes, Christa; Wüst, Matthias

    2014-03-19

    The biosynthesis of the diterpenoid steviol glycosides rebaudioside A and stevioside in nonrooted cuttings of Stevia rebaudiana was investigated by feeding experiments using the labeled key precursors [5,5-(2)H2]-mevalonic acid lactone (d2-MVL) and [5,5-(2)H2]-1-deoxy-d-xylulose (d2-DOX). Labeled glycosides were extracted from the leaves and stems and were directly analyzed by LC-(-ESI)-MS/MS and by GC-MS after hydrolysis and derivatization of the resulting isosteviol to the corresponding TMS-ester. Additionally, the incorporation of the proffered d2-MVL and d2-DOX into volatile monoterpenes, sesquiterpenes, and diterpenes in glandular trichomes on leaves and stems was investigated by headspace-solid phase microextraction-GC-MS (HS-SPME-GC-MS). Incorporation of the labeled precursors indicated that diterpenes in leaves and monoterpenes and diterpenes in glandular trichomes are predominately biosynthesized via the methylerythritol phosphate (MEP) pathway, whereas both the MEP and mevalonate (MVA) pathways contribute to the biosynthesis of sesquiterpenes at equal rates in glandular trichomes. These findings give evidence for a transport of MEP pathway derived farnesyl diphosphate precursors from plastids to the cytosol. Contrarily, the transport of MVA pathway derived geranyl diphosphate and geranylgeranyl diphosphate precursors from the cytosol to the plastid is limited.

  5. TRICHOME BIREFRINGENCE and Its Homolog AT5G01360 Encode Plant-Specific DUF231 Proteins Required for Cellulose Biosynthesis in Arabidopsis1[W][OA

    PubMed Central

    Bischoff, Volker; Nita, Silvia; Neumetzler, Lutz; Schindelasch, Dana; Urbain, Aurélie; Eshed, Ravit; Persson, Staffan; Delmer, Deborah; Scheible, Wolf-Rüdiger

    2010-01-01

    The Arabidopsis (Arabidopsis thaliana) trichome birefringence (tbr) mutant has severely reduced crystalline cellulose in trichomes, but the molecular nature of TBR was unknown. We determined TBR to belong to the plant-specific DUF231 domain gene family comprising 46 members of unknown function in Arabidopsis. The genes harbor another plant-specific domain, called the TBL domain, which contains a conserved GDSL motif known from some esterases/lipases. TBR and TBR-like3 (TBL3) are transcriptionally coordinated with primary and secondary CELLULOSE SYNTHASE (CESA) genes, respectively. The tbr and tbl3 mutants hold lower levels of crystalline cellulose and have altered pectin composition in trichomes and stems, respectively, tissues generally thought to contain mainly secondary wall crystalline cellulose. In contrast, primary wall cellulose levels remain unchanged in both mutants as measured in etiolated tbr and tbl3 hypocotyls, while the amount of esterified pectins is reduced and pectin methylesterase activity is increased in this tissue. Furthermore, etiolated tbr hypocotyls have reduced length with swollen epidermal cells, a phenotype characteristic for primary cesa mutants or the wild type treated with cellulose synthesis inhibitors. Taken together, we show that two TBL genes contribute to the synthesis and deposition of secondary wall cellulose, presumably by influencing the esterification state of pectic polymers. PMID:20388664

  6. A ribozyme selected from variants of U6 snRNA promotes 2',5'-branch formation.

    PubMed Central

    Tuschl, T; Sharp, P A; Bartel, D P

    2001-01-01

    In vitro selection was used to sample SnRNA-related sequences for ribozyme activities, and several 2',5'-branch-forming ribozymes were isolated. One such ribozyme is highly dependent upon an 11-nt motif that contains a conserved U6 snRNA sequence (ACAGAGA-box) known to be important for pre-mRNA splicing. The ribozyme reaction is similar to the first step of splicing in that an internal 2'-hydroxyl of an unpaired adenosine attacks at the 5'-phosphate of a guanosine. It differs in that the leaving group is diphosphate rather than a 5' exon. The finding that lariat formation can be accomplished by a small RNA with sequences related to U6 snRNA indicates that the RNA available in the spliceosome may be involved in RNA-catalyzed branch formation. PMID:11214178

  7. Ficolin-1-PTX3 complex formation promotes clearance of altered self-cells and modulates IL-8 production.

    PubMed

    Ma, Ying Jie; Doni, Andrea; Romani, Luigina; Jürgensen, Henrik Jessen; Behrendt, Niels; Mantovani, Alberto; Garred, Peter

    2013-08-01

    The long pentraxin 3 (PTX3) has been shown to be important in maintaining internal tissue homeostasis and in protecting against fungal Aspergillus fumigatus infection. However, the molecular mechanisms of how these functions are elicited are poorly delineated. Ficolin-1 is a soluble pattern recognition molecule that interacts with PTX3. We hypothesized that heterocomplexes between ficolin-1 and PTX3 might mediate the signals necessary for sequestration of altered self-cells and A. fumigatus. We were able to show that ficolin-1 interacts with PTX3 via its fibrinogen-like domain. The interaction was affected in a pH- and divalent cation-sensitive manner. The primary binding site for ficolin-1 on PTX3 was located in the N-terminal domain portion of PTX3. Ficolin-1 and PTX3 heterocomplex formation occurred on dying host cells, but not on A. fumigatus. The heterocomplex formation was a prerequisite for enhancement of phagocytosis by human monocyte-derived macrophages and downregulation of IL-8 production during phagocytosis. On A. fumigatus, PTX3 exposed the C-terminal portion of the molecule, probably resulting in steric hindrance of ficolin-1 interaction with PTX3. These results demonstrate that ficolin-1 and PTX3 heterocomplex formation acts as a noninflammatory "find me and eat me" signal to sequester altered-host cells. The fact that the ficolin-1-PTX3 complex formation did not occur on A. fumigatus shows that PTX3 uses different molecular effector mechanisms, depending on which domains it exposes during ligand interaction.

  8. Phosphoinositide 3-kinase p110δ promotes lumen formation through enhancement of apico-basal polarity and basal membrane organization

    PubMed Central

    Sar, Sokhavuth; Komaiha, Ola Hamze; Moyano, Romina; Rayal, Amel; Samuel, Didier; Shewan, Annette; Vanhaesebroeck, Bart; Mostov, Keith; Gassama-Diagne, Ama

    2016-01-01

    Signaling triggered by adhesion to the extracellular matrix plays a key role in the spatial orientation of epithelial polarity and formation of lumens in glandular tissues. Phosphoinositide 3-kinase signaling in particular is known to influence the polarization process during epithelial cell morphogenesis. Here, using Madin-Darby canine kidney epithelial cells grown in 3D culture, we show that the p110δ isoform of phosphoinositide 3-kinase colocalizes with focal adhesion proteins at the basal surface of polarized cells. Pharmacological, siRNA- or kinase-dead mediated inhibition of p110δ impair the early stages of lumen formation, resulting in inverted polarized cysts, with no laminin or type IV collagen assembly at cell/extracellular matrix contacts. p110δ also regulates the organization of focal adhesions and membrane localization of dystroglycan. Thus, we uncover a previously unrecognized role for p110δ in epithelial cells in the orientation of the apico-basal axis and lumen formation. PMID:25583025

  9. Capsicum annuum S (CaS) promotes reproductive transition and is required for flower formation in pepper (Capsicum annuum).

    PubMed

    Cohen, Oded; Borovsky, Yelena; David-Schwartz, Rakefet; Paran, Ilan

    2014-05-01

    The genetic control of the transition to flowering has mainly been studied in model species, while few data are available in crop species such as pepper (Capsicum spp.). To elucidate the genetic control of the transition to flowering in pepper, mutants that lack flowers were isolated and characterized. Genetic mapping and sequencing allowed the identification of the gene disrupted in the mutants. Double mutants and expression analyses were used to characterize the relationships between the mutated gene and other genes controlling the transition to flowering and flower differentiation. The mutants were characterized by a delay in the initiation of sympodial growth, a delay in the termination of sympodial meristems and complete inhibition of flower formation. Capsicum annuum S (CaS), the pepper (Capsicum annuum) ortholog of tomato (Solanum lycopersicum) COMPOUND INFLORESCENCE and petunia (Petunia hybrida) EVERGREEN, was found to govern the mutant phenotype. CaS is required for the activity of the flower meristem identity gene Ca-ANANTHA and does not affect the expression of CaLEAFY. CaS is epistatic over other genes controlling the transition to flowering with respect to flower formation. Comparative homologous mutants in the Solanaceae indicate that CaS has uniquely evolved to have a critical role in flower formation, while its role in meristem maturation is conserved in pepper, tomato and petunia.

  10. Chromosomal gain promotes formation of a steep RanGTP gradient that drives mitosis in aneuploid cells

    PubMed Central

    Hasegawa, Keisuke; Ryu, Sung Jin

    2013-01-01

    Many mitotic factors were shown to be activated by Ran guanosine triphosphatase. Previous studies in Xenopus laevis egg extracts and in highly proliferative cells showed that mitotic chromosomes were surrounded by steep Ran guanosine triphosphate (GTP) concentration gradients, indicating that RanGTP-activated factors promote spindle assembly around chromosomes. However, the mitotic role of Ran in normal differentiated cells is not known. In this paper, we show that although the steep mitotic RanGTP gradients were present in rapidly growing cell lines and were required for chromosome congression in mitotic HeLa cells, the gradients were strongly reduced in slow-growing primary cells, such as HFF-1 fibroblasts. The overexpression of RCC1, the guanine nucleotide exchange factor for Ran, induced steeper mitotic RanGTP gradients in HFF-1 cells, showing the critical role of RCC1 levels in the regulation of mitosis by Ran. Remarkably, in vitro fusion of HFF-1 cells produced cells with steep mitotic RanGTP gradients comparable to HeLa cells, indicating that chromosomal gain can promote mitosis in aneuploid cancer cells via Ran. PMID:23319601

  11. The Arabidopsis Transcription Factor MYB112 Promotes Anthocyanin Formation during Salinity and under High Light Stress1[OPEN

    PubMed Central

    Lotkowska, Magda E.; Tohge, Takayuki; Fernie, Alisdair R.; Xue, Gang-Ping; Balazadeh, Salma; Mueller-Roeber, Bernd

    2015-01-01

    MYB transcription factors (TFs) are important regulators of flavonoid biosynthesis in plants. Here, we report MYB112 as a formerly unknown regulator of anthocyanin accumulation in Arabidopsis (Arabidopsis thaliana). Expression profiling after chemically induced overexpression of MYB112 identified 28 up- and 28 down-regulated genes 5 h after inducer treatment, including MYB7 and MYB32, which are both induced. In addition, upon extended induction, MYB112 also positively affects the expression of PRODUCTION OF ANTHOCYANIN PIGMENT1, a key TF of anthocyanin biosynthesis, but acts negatively toward MYB12 and MYB111, which both control flavonol biosynthesis. MYB112 binds to an 8-bp DNA fragment containing the core sequence (A/T/G)(A/C)CC(A/T)(A/G/T)(A/C)(T/C). By electrophoretic mobility shift assay and chromatin immunoprecipitation coupled to quantitative polymerase chain reaction, we show that MYB112 binds in vitro and in vivo to MYB7 and MYB32 promoters, revealing them as direct downstream target genes. We further show that MYB112 expression is up-regulated by salinity and high light stress, environmental parameters that both require the MYB112 TF for anthocyanin accumulation under these stresses. In contrast to several other MYB TFs affecting anthocyanin biosynthesis, MYB112 expression is not controlled by nitrogen limitation or an excess of carbon. Thus, MYB112 constitutes a regulator that promotes anthocyanin accumulation under abiotic stress conditions. PMID:26378103

  12. Translocations of Chromosome End-Segments and Facultative Heterochromatin Promote Meiotic Ring Formation in Evening Primroses[W][OPEN

    PubMed Central

    Golczyk, Hieronim; Massouh, Amid; Greiner, Stephan

    2014-01-01

    Due to reciprocal chromosomal translocations, many species of Oenothera (evening primrose) form permanent multichromosomal meiotic rings. However, regular bivalent pairing is also observed. Chiasmata are restricted to chromosomal ends, which makes homologous recombination virtually undetectable. Genetic diversity is achieved by changing linkage relations of chromosomes in rings and bivalents via hybridization and reciprocal translocations. Although the structural prerequisite for this system is enigmatic, whole-arm translocations are widely assumed to be the mechanistic driving force. We demonstrate that this prerequisite is genome compartmentation into two epigenetically defined chromatin fractions. The first one facultatively condenses in cycling cells into chromocenters negative both for histone H3 dimethylated at lysine 4 and for C-banding, and forms huge condensed middle chromosome regions on prophase chromosomes. Remarkably, it decondenses in differentiating cells. The second fraction is euchromatin confined to distal chromosome segments, positive for histone H3 lysine 4 dimethylation and for histone H3 lysine 27 trimethylation. The end-segments are deprived of canonical telomeres but capped with constitutive heterochromatin. This genomic organization promotes translocation breakpoints between the two chromatin fractions, thus facilitating exchanges of end-segments. We challenge the whole-arm translocation hypothesis by demonstrating why reciprocal translocations of chromosomal end-segments should strongly promote meiotic rings and evolution toward permanent translocation heterozygosity. Reshuffled end-segments, each possessing a major crossover hot spot, can furthermore explain meiotic compatibility between genomes with different translocation histories. PMID:24681616

  13. Transplantation of hUC-MSCs seeded collagen scaffolds reduces scar formation and promotes functional recovery in canines with chronic spinal cord injury

    PubMed Central

    Li, Xing; Tan, Jun; Xiao, Zhifeng; Zhao, Yannan; Han, Sufang; Liu, Dingyang; Yin, Wen; Li, Jing; Li, Juan; Wanggou, Siyi; Chen, Bing; Ren, Caiping; Jiang, Xingjun; Dai, Jianwu

    2017-01-01

    Spinal cord injury (SCI) can lead to locomotor deficits, and the repair of chronic SCI is considered one of the most challenging clinical problems. Although extensive studies have evaluated treatments for acute SCI in small animals, comparatively fewer studies have been conducted on large-animal SCI in the chronic phase, which is more clinically relevant. Here, we used a collagen-based biomaterial, named the NeuroRegen scaffold, loaded with human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) in a canine chronic SCI model. To generate chronic SCI, the T8 spinal cord segment was removed by complete transection of the spinal cord. Two months later, glial scar tissue was removed and a NeuroRegen scaffold was transplanted into the lesion area. Functionalized NeuroRegen scaffold implantation promoted both locomotor recovery and endogenous neurogenesis in the lesion area. Moreover, some newly generated neurons successfully matured into 5-HT-positive neurons at 1 year post-injury. In addition, many regenerated axon fibers in the lesion area exhibited remyelination and synapse formation at 1 year post-injury in the functionalized NeuroRegen scaffold group. In conclusion, the NeuroRegen scaffold functionalized with hUC-MSCs is a promising potential therapeutic approach to chronic SCI that promotes neuronal regeneration, reduces glial scar formation, and ultimately improves locomotor recovery. PMID:28262732

  14. Transgenic analysis of sugar beet xyloglucan endo-transglucosylase/hydrolase Bv-XTH1 and Bv-XTH2 promoters reveals overlapping tissue-specific and wound-inducible expression profiles.

    PubMed

    Dimmer, Emily; Roden, Laura; Cai, Daguang; Kingsnorth, Crawford; Mutasa-Göttgens, Effie

    2004-03-01

    The identification and analysis of tissue-specific gene regulatory elements will improve our knowledge of the molecular mechanisms that control the growth and development of different plant tissues and offer potentially useful tools for the genetic engineering of plants. A polymerase chain reaction (PCR)-based 5'-genome walk from sequences of an isolated sugar beet xyloglucan endo-transglucosylase hydrolase (XTH) gene led to the isolation of two independent upstream fragments. They were 1332 and 2163 base pairs upstream of the XTH ATG start site, respectively. In vivo transgenic assays in sugar beet hairy roots and Arabidopsis thaliana revealed that both fragments had promoter function and, in A. thaliana, directed expression in vascular tissues within the root, leaves and petals. Promoter activity was also observed in the leaf trichomes and within rapidly expanding stem internodes. Expression driven by both promoters was found to be wound inducible. Overall, the spatial and temporal expression pattern of these promoters suggested that the corresponding Bv-XTH genes (designated Bv-XTH1 and Bv-XTH2) may be involved in secondary cell wall formation. This work provides new insights on molecular mechanisms that could be exploited for the genetic engineering of sugar beet crops.

  15. The sex-determining factors SRY and SOX9 regulate similar target genes and promote testis cord formation during testicular differentiation.

    PubMed

    Li, Yunmin; Zheng, Ming; Lau, Yun-Fai Chris

    2014-08-07

    Male sex determination is mediated sequentially by sex-determining region Y (SRY) and related SRY-box 9 (SOX9) transcription factors. To understand the gene regulatory hierarchy for SRY and SOX9, a series of chromatin immunoprecipitation and whole-genome promoter tiling microarray (ChIP-Chip) experiments were conducted with mouse gonadal cells at the time of sex determination. SRY and SOX9 bind to the promoters of many common targets involved in testis differentiation and regulate their expression in Sertoli cells. SRY binds to various ovarian differentiation genes and represses their activation through WNT/β-catenin signaling. Sertoli cell-Sertoli cell junction signaling, important for testis cord formation, is the top canonical pathway among the SRY and SOX9 targets. Hence, SRY determines Sertoli cell fate by repressing ovarian and activating testicular differentiation genes, promotes early Sertoli cells to form testis cord, and then passes on its functions to SOX9, which regulates common targets and activates its own gene regulatory program, beyond SRY actions, in sex determination.

  16. A novel activity of HMG domains: promotion of the triple-stranded complex formation between DNA containing (GGA/TCC)11 and d(GGA)11 oligonucleotides.

    PubMed Central

    Suda, T; Mishima, Y; Takayanagi, K; Asakura, H; Odani, S; Kominami, R

    1996-01-01

    The high mobility group protein (HMG)-box is a DNA-binding domain found in many proteins that bind preferentially to DNA of irregular structures in a sequence-independent manner and can bend the DNA. We show here that GST-fusion proteins of HMG domains from HMG1 and HMG2 promote a triple-stranded complex formation between DNA containing the (GGA/TCC)11 repeat and oligonucleotides of d(GGA)11 probably due to G:G base pairing. The activity is to reduce association time and requirements of Mg2+ and oligonucleotide concentrations. The HMG box of SRY, the protein determining male-sex differentiation, also has the activity, suggesting that it is not restricted to the HMG-box domains derived from HMG1/2 but is common to those from other members of the HMG-box family of proteins. Interestingly, the box-AB and box-B of HMG1 bend DNA containing the repeat, but SRY fails to bend in a circularization assay. The difference suggests that the two activities of association-promotion and DNA bending are distinct. These results suggest that the HMG-box domain has a novel activity of promoting the association between GGA repeats which might be involved in higher-order architecture of chromatin. PMID:8972860

  17. Trichomes as dangerous lollipops: do lizards also use caterpillar body and frass odor to optimize their foraging?

    PubMed

    Stork, William F J; Weinhold, Alexander; Baldwin, Ian T

    2011-12-01

    When attacked by herbivores, plants produce toxic secondary metabolites that function as direct defenses, as well as indirect defenses that attract and reward predators of the offending herbivores. These indirect defenses include both nutritive rewards such as extra floral nectar, as well as informational rewards, such as the production and release of volatile compounds that betray the location of feeding herbivores to predators. Herbivory of Nicotiana attenuata by the tobacco hornworm (Manduca larvae) alters the volatile profiles of both the plant and larval headspace. Herbivory-elicited specific changes in the volatile profiles are detected by arthropod predators of Manduca larvae. The known predators that perceive volatile cues induced by Manduca herbivory of N. attenuata are insects that target Manduca at early developmental stages, when the larvae are still small; large, late-instar larvae may have outgrown these predation risks. However, here we offer evidence that branched chain aliphatic acids derived from the digestion of plant O-acyl sugars from trichomes may betray Manduca larvae to lizard predators during late developmental stages as well.

  18. Glandular trichome density and essential oil composition in leaves and inflorescences of Lippia origanoides Kunth (Verbenaceae) in the Brazilian Cerrado.

    PubMed

    Tozin, Luiz R S; Marques, Marcia O M; Rodrigues, Tatiane M

    2015-01-01

    The essential oils from leaves and inflorescences of Lippia origanoides Kunth present aromatic and medicinal potential and have been used to treat several diseases, including melanoma. In Brazil, L. origanoides is commonly found in campo cerrado and cerrado stricto sensu, physiognomies featured mainly by the differential light conditions to which short and medium-sized plants are subjected. Our aim was to investigate the glandular trichome density and the yield and chemical composition of the essential oils in leaves and inflorescences of L. origanoides from campo cerrado and cerrado stricto sensu. For glandular density analysis, leaves and inflorescences were processed according to conventional techniques for scanning electron microscopy. The essential oils of leaves and inflorescences were obtained by hydrodistillation and identified with gas chromatography. Bracts and sepals showed the highest glandular density, followed by petals and leaves. The glandular density in the abaxial leaf surface was higher in individuals from the campo cerrado. In both populations the essential oil yield was higher in inflorescences than in leaves. The chemical composition of the essential oils varied among individuals from different areas and inside a same population. Our results demonstrated the chemical plasticity of L. origanoides suggesting the importance of monitoring its popular use.

  19. Purification and characterization of an NADPH-cytochrome P450 (cytochrome c) reductase from spearmint (Mentha spicata) glandular trichomes.

    PubMed

    Ponnamperuma, K; Croteau, R

    1996-05-01

    Solubilized NADPH-cytochrome c (P450) reductase was purified to homogeneity from an extract of spearmint (Mentha spicata) glandular trichomes by dye-ligand interaction chromatography on Matrex-Gel Red A and affinity chromatography on 2', 5'-adenosine diphosphate agarose. SDS-PAGE of the purified enzyme preparation revealed the presence of two similar proteins with masses of 82 kDa (major) and 77 kDa (minor) that crossreacted on immunoblot analysis with polyclonal antibodies directed against NADPH-cytochrome P450 reductase from Jerusalem artichoke and from mung bean. Complete immunoinhibition of reductase activity was observed with both types of polyclonal antibodies, while only partial inhibition of activity resulted using a family of monoclonal antibodies directed against the Jerusalem artichoke cytochrome P450 reductase. Inhibition of the spearmint oil gland cytochrome c reductase was also observed with the diphenyliodonium ion. The K(m) values for the cosubstrates NADPH and cytochrome c were 6.2 and 3.7 microM, respectively, and the pH optimum for activity was at 8.5. The NADPH-cytochrome c reductase reconstituted NADPH-dependent (-)-4S-limonene-6-hydroxylase activity in the presence of cytochrome P450, purified from the microsomal fraction of spearmint oil gland cells and dilauroyl phosphatidyl choline. These characteristics establish the identity of the purified enzyme as a NADPH-cytochrome P450 reductase.

  20. The characterization of SaPIN2b, a plant trichome-localized proteinase inhibitor from Solanum americanum.

    PubMed

    Luo, Ming; Ding, Ling-Wen; Ge, Zhi-Juan; Wang, Zhen-Yu; Hu, Bo-Lun; Yang, Xiao-Bei; Sun, Qiao-Yang; Xu, Zeng-Fu

    2012-11-16

    Proteinase inhibitors play an important role in plant resistance of insects and pathogens. In this study, we characterized the serine proteinase inhibitor SaPIN2b, which is constitutively expressed in Solanum americanum trichomes and contains two conserved motifs of the proteinase inhibitor II (PIN2) family. The recombinant SaPIN2b (rSaPIN2b), which was expressed in Escherichia coli, was demonstrated to be a potent proteinase inhibitor against a panel of serine proteinases, including subtilisin A, chymotrypsin and trypsin. Moreover, rSaPIN2b also effectively inhibited the proteinase activities of midgut trypsin-like proteinases that were extracted from the devastating pest Helicoverpa armigera. Furthermore, the overexpression of SaPIN2b in transgenic tobacco plants resulted in enhanced resistance against H. armigera. Taken together, our results demonstrated that SaPIN2b is a potent serine proteinase inhibitor that may act as a protective protein in plant defense against insect attacks.

  1. Dense white trichome production by plants as possible mimicry of arthropod silk or fungal hyphae that deter herbivory.

    PubMed

    Yamazaki, Kazuo; Lev-Yadun, Simcha

    2015-01-07

    Some spiders are well-known to mimic flowers or other plant surfaces in order to be cryptic to both their prey and their predators. We propose that dense, thread-like white trichomes of some plants from Estonia, Greece, Israel and Japan visually mimic spider webs, lepidopteran and spider-mite web nests and plant-pathogenic fungi, and that it may result in reduced herbivory, since various herbivores avoid spider- or other arthropod webs to circumvent predation or toxic attacks, or refrain from colonizing plants that have already been occupied by other herbivores and pathogens. Spiders and other web-forming arthropods are also the prey of certain vertebrate predators and wasps, and therefore such predators may be attracted to these web-like plant structures and prey on the invertebrate herbivores occupying them. We do not dismiss the possibility that these web-like structures may also have other defensive or physiological functions or that they are not classic mimics but rather exploit the herbivore׳s perceptual state concerning the avoidance of potentially risky objects.

  2. Insights on co-catalyst-promoted enamine formation between dimethylamine and propanal through ab initio and density functional theory study.

    PubMed

    Patil, Mahendra P; Sunoj, Raghavan B

    2007-10-26

    The mechanistic details on enamine formation between dimethylamine and propanal are unraveled using the ab initio and density functional theory methods. The addition of secondary amine to the electrophile and simultaneous proton transfer results in a carbinolamine intermediate, which subsequently undergoes dehydration to form enamine. The direct addition of amine as well as the dehydration of the resulting carbinolamine intermediate is predicted to possess fairly high activation barrier implying that a unimolecular process is unlikely to be responsible for enamine formation. Different models are therefore proposed which could explain the relative ease of enamine formation under neat condition as well as under the influence of methanol as the co-catalyst. The explicit inclusion of either the reagent or the co-catalyst is considered in the transition states as stabilizing agents. The participation of the reagent or the co-catalyst as a monofunctional ancillary species is found to stabilize the transition states relative to the unassisted or the direct addition/dehydration pathways. The reduction in enthalpy of activation is found to be much more dramatic when two co-catalysts participate in an active bifunctional mode in the rate-determining dehydration step. The transition structures exhibited characteristic features of a relay proton transfer mechanism. The free energy of activation associated with the two methanol-assisted pathway is found to be 16.7 kcal/mol lower than that of the unassisted pathway. The results are found to be in concurrence with the available reports on the rate acceleration by co-catalysts in the Michael reaction between enamine and methyl vinyl ketone under neat conditions.

  3. Loss of nuclear receptor RXRα in epidermal keratinocytes promotes the formation of Cdk4-activated invasive melanomas.

    PubMed

    Hyter, Stephen; Bajaj, Gaurav; Liang, Xiaobo; Barbacid, Mariano; Ganguli-Indra, Gitali; Indra, Arup Kumar

    2010-10-01

    Keratinocytes contribute to melanocyte transformation by affecting their microenvironment, in part through the secretion of paracrine factors. Here we report a loss of expression of nuclear receptor RXRα in epidermal keratinocytes during human melanoma progression. In the absence of keratinocytic RXRα, in combination with mutant Cdk4, cutaneous melanoma was generated that metastasized to lymph nodes in a bigenic mouse model. Expression of several keratinocyte-derived mitogenic growth factors (Et-1, Hgf, Scf, α-MSH and Fgf 2 ) was elevated in skin of bigenic mice, whereas Fas, E-cadherin and Pten, implicated in apoptosis, cellular invasion and melanomagenesis, respectively, were downregulated within the microdissected melanocytic tumors. We demonstrated that RXRα is recruited on the proximal promoter of both Et-1 and Hgf, possibly directly regulating their transcription in keratinocytes. These studies demonstrate the contribution of keratinocytic paracrine signaling during the cellular transformation and malignant conversion of melanocytes.

  4. Precise Formation of a Hollow Carbon Nitride Structure with a Janus Surface To Promote Water Splitting by Photoredox Catalysis

    PubMed Central

    Zheng, Dandan; Cao, Xu‐Ning

    2016-01-01

    Abstract The precise modification of redox species on the inner and outer surfaces of hollow nanostructures is relevant in catalysis, surface science, and nanotechnology, but has proven difficult to achieve. Herein, we develop a facile approach to specifically fabricate Pt and Co3O4 nanoparticles (NPs) onto the interior and exterior surface of hollow carbon nitride spheres (HCNS), respectively, to promote the surface redox functions of the polymer semiconductors. The photocatalytic water splitting activities of HCNS with spatially separated oxidation and reduction centers at their nanodomains were enhanced. The origin of the enhanced activity was attributed to the spatially separated reactive sites for the evolution of H2 and O2 and also to the unidirectional migration of the electron and hole on the Janus surfaces, thereby preventing the unwanted reverse reaction of water splitting and decreasing charge recombination. PMID:27533739

  5. FGFR3 promotes angiogenesis-dependent metastasis of hepatocellular carcinoma via facilitating MCP-1-mediated vascular formation.

    PubMed

    Liu, Xinyu; Jing, Xiaoqian; Cheng, Xi; Ma, Ding; Jin, Zhijian; Yang, Weiping; Qiu, Weihua

    2016-05-01

    The biological role of fibroblast growth factor receptor 3 (FGFR3) in tumor angiogenesis of hepatocellular carcinoma (HCC) has not been discussed before. Our previous work had indicated FGFR3 was overexpressed in HCC, and silencing FGFR3 in Hu7 cells could regulate tumorigenesis via down-regulating the phosphorylation level of key members of classic signaling pathways including ERK and AKT. In the present work, we explored the role of FGFR3 in angiogenesis-dependent metastasis by using SMMC-7721 and QGY-7703 stable cell lines. Our results indicated FGFR3 could regulate in vitro cell migration ability and in vivo lung metastasis ability of HCC, which was in accordance with increased angiogenesis ability in vitro and in vivo. Using the supernatant from SMMC-7721/FGFR3 cells, we conducted a human angiogenesis protein microarray including 43 angiogenesis factors and found that FGFR3 modulated angiogenesis and metastasis of HCC mainly by promoting the protein level of monocyte chemotactic protein 1 (MCP-1). Silencing FGFR3 by short hairpin RNA (shRNA) could reduce MCP-1 level in lysates and supernatant of QGY-7703 cells and SMMC-7721 cells. Silencing MCP-1 in QGY-7703 or SMMC-7721 cells could induce similar phenotypes compared with silencing FGFR3. Our results suggested FGFR3 promoted metastasis potential of HCC, at least partially if not all, via facilitating MCP-1-mediated angiogenesis, in addition to previously found cell growth and metastasis. MCP-1, a key medium between HCC cells and HUVECs, might be a novel anti-vascular target in HCC.

  6. KISS1R signaling promotes invadopodia formation in human breast cancer cell via β-arrestin2/ERK.

    PubMed

    Goertzen, Cameron G; Dragan, Magdalena; Turley, Eva; Babwah, Andy V; Bhattacharya, Moshmi

    2016-03-01

    Kisspeptins (KPs), peptide products of the KISS1 gene are endogenous ligands for the kisspeptin receptor (KISS1R), a G protein-coupled receptor. In numerous cancers, KISS1R signaling plays anti-metastatic roles. However, we have previously shown that in breast cancer cells lacking the estrogen receptor (ERα), kisspeptin-10 stimulates cell migration and invasion by cross-talking with the epidermal growth factor receptor (EGFR), via a β-arrestin-2-dependent mechanism. To further define the mechanisms by which KISS1R stimulates invasion, we determined the effect of down-regulating KISS1R expression in triple negative breast cancer cells. We found that depletion of KISS1R reduced their mesenchymal phenotype and invasiveness. We show for the first time that KISS1R signaling induces invadopodia formation and activation of key invadopodia proteins, cortactin, cofilin and membrane type I matrix metalloproteases (MT1-MMP). Moreover, KISS1R stimulated invadopodia formation occurs via a new pathway involving a β-arrestin2 and ERK1/2-dependent mechanism, independent of Src. Taken together, our findings suggest that targeting the KISS1R signaling axis might be a promising strategy to inhibit invasiveness and metastasis.

  7. Developmental Control of a parAB Promoter Leads to Formation of Sporulation-Associated ParB Complexes in Streptomyces coelicolor

    PubMed Central

    Jakimowicz, Dagmara; Mouz, Sebastien; Zakrzewska-Czerwińska, Jolanta; Chater, Keith F.

    2006-01-01

    The Streptomyces coelicolor partitioning protein ParB binds to numerous parS sites in the oriC-proximal part of the linear chromosome. ParB binding results in the formation of large complexes, which behave differentially during the complex life cycle (D. Jakimowicz, B. Gust, J. Zakrzewska-Czerwinska, and K. F. Chater, J. Bacteriol. 187:3572-3580, 2005). Here we have analyzed the transcriptional regulation that underpins this developmentally specific behavior. Analysis of promoter mutations showed that the irregularly spaced complexes present in vegetative hyphae are dependent on the constitutive parABp1 promoter, while sporulation-specific induction of the promoter parABp2 is required for the assembly of arrays of ParB complexes in aerial hyphae and thus is necessary for efficient chromosome segregation. Expression from parABp2 depended absolutely on two sporulation regulatory genes, whiA and whiB, and partially on two others, whiH and whiI, all four of which are needed for sporulation septation. Because of this pattern of dependence, we investigated the transcription of these four whi genes in whiA and whiB mutants, revealing significant regulatory interplay between whiA and whiB. A strain in which sporulation septation (but not vegetative septation) was blocked by mutation of a sporulation-specific promoter of ftsZ showed close to wild-type induction of parABp2 and formed fairly regular ParB-enhanced green fluorescent protein foci in aerial hyphae, ruling out strong morphological coupling or checkpoint regulation between septation and DNA partitioning during sporulation. A model for developmental regulation of parABp2 expression is presented. PMID:16484182

  8. Using Formal Embedded Formative Assessments Aligned with a Short-Term Learning Progression to Promote Conceptual Change and Achievement in Science

    NASA Astrophysics Data System (ADS)

    Yin, Yue; Tomita, Miki K.; Shavelson, Richard J.

    2014-03-01

    This study examined the effect of learning progression-aligned formal embedded formative assessment on conceptual change and achievement in middle-school science. Fifty-two sixth graders were randomly assigned to either an experimental group or a control group. Both groups were taught about sinking and floating by the same teacher with identical curriculum materials and activities. The experimental group received, in addition, three sets of formal embedded formative assessments with qualitative feedback as to how to improve their understandings aligned with an expected learning progression during instruction. The control group spent the corresponding time between new curriculum activities conducting curriculum-specific extension activities. Overall, the experimental group experienced on average greater conceptual change than the control group. The experimental group also scored higher on average than the control group on general achievement tests, especially the performance assessment. This study supported the contention, then, that embedding formal formative assessments within a curricular sequence built around an expected learning progression is a useful way to promote conceptual change along that learning progression in science classrooms.

  9. Promoting oxygen vacancy formation and p-type conductivity in SrTiO3via alkali metal doping: a first principles study.

    PubMed

    Triggiani, Leonardo; Muñoz-García, Ana B; Agostiano, Angela; Pavone, Michele

    2016-10-19

    Strontium titanate (SrTiO3, STO) is a prototypical perovskite oxide, widely exploited in many technological applications, from catalysis to energy conversion devices. In the context of solid-oxide fuel cells, STO has been recently applied as an epitaxial substrate for nano-sized layers of mixed ion-electron conductive catalysts with enhanced electrochemical performances. To extend the applications of such heterogeneous nano-cathodes in real devices, also the STO support should be active for both electron transport and oxide diffusion. To this end, we explored using first-principles calculations the strategy of doping of STO at the Sr site with sodium and potassium. These two ions fit in the perovskite structure and induce holes in the STO valence band, so as to obtain the desired p-type electronic conduction. At the same time, the doping with alkali ions also promotes the formation of oxygen vacancies in STO, a prerequisite for effective oxide diffusion. Analysis of electron density rearrangements upon defect formation allows relating the favorable vacancy formation energies to an improved electronic delocalization over the oxide sub-lattice, as observed in closely related materials (e.g. Sr2Fe1.5Mo0.5O6). Overall, our results suggest the alkali-doped STO as a new potential substrate material in nanoscale heterogeneous electrodes for solid oxide electrochemical cells.

  10. Akt1-Inhibitor of DNA binding2 is essential for growth cone formation and axon growth and promotes central nervous system axon regeneration

    PubMed Central

    Ko, Hyo Rim; Kwon, Il-Sun; Hwang, Inwoo; Jin, Eun-Ju; Shin, Joo-Ho; Brennan-Minnella, Angela M; Swanson, Raymond; Cho, Sung-Woo; Lee, Kyung-Hoon; Ahn, Jee-Yin

    2016-01-01

    Mechanistic studies of axon growth during development are beneficial to the search for neuron-intrinsic regulators of axon regeneration. Here, we discovered that, in the developing neuron from rat, Akt signaling regulates axon growth and growth cone formation through phosphorylation of serine 14 (S14) on Inhibitor of DNA binding 2 (Id2). This enhances Id2 protein stability by means of escape from proteasomal degradation, and steers its localization to the growth cone, where Id2 interacts with radixin that is critical for growth cone formation. Knockdown of Id2, or abrogation of Id2 phosphorylation at S14, greatly impairs axon growth and the architecture of growth cone. Intriguingly, reinstatement of Akt/Id2 signaling after injury in mouse hippocampal slices redeemed growth promoting ability, leading to obvious axon regeneration. Our results suggest that Akt/Id2 signaling is a key module for growth cone formation and axon growth, and its augmentation plays a potential role in CNS axonal regeneration. DOI: http://dx.doi.org/10.7554/eLife.20799.001 PMID:27938661

  11. Extracorporeal shock waves alone or combined with raloxifene promote bone formation and suppress resorption in ovariectomized rats

    PubMed Central

    Corrado, Bruno; Pirozzi, Claudio; Paciello, Orlando; Pagano, Teresa Bruna; Russo, Sergio; Calignano, Antonio; Mattace Raso, Giuseppina; Meli, Rosaria

    2017-01-01

    Osteoporosis is a metabolic skeletal disease characterized by an imbalance between osteoclast-mediated bone resorption and osteoblast-mediated bone formation. We examined the beneficial effect of shock waves (SW) alone or in combination with raloxifene (RAL) on bone loss in ovariectomized rats (OVX). Sixteen weeks after surgery, OVX were treated for five weeks with SW at the antero-lateral side of the right hind leg, one session weekly, at 3 Hz (EFD of 0.33 mJ/mm2), or with RAL (5 mg/kg/die, per os) or with SW+RAL. Sera, femurs, tibiae and vertebrae were sampled for following biochemical and histological analysis. SW, alone or combined with RAL, prevented femur weight reduction and the deterioration of trabecular microarchitecture both in femur and vertebrae. All treatments increased Speed of Sound (SoS) values, improving bone mineral density, altered by OVX. Serum parameters involved in bone remodeling (alkaline phosphatase, receptor activator of nuclear factor kappa-B ligand, osteoprotegerin) and osteoblast proliferation (PTH), altered by ovariectomy, were restored by SW and RAL alone or in combination. In tibiae, SW+RAL significantly reduced cathepsin k and TNF-α levels, indicating the inhibition of osteoclast activity, while all treatments significantly increased runt-related transcription factor 2 and bone morphogenetic-2 expression, suggesting an increase in osteoblastogenic activity. Finally, in bone marrow from tibiae, SW or RAL reduced PPARγ and adiponectin transcription, indicating a shift of mesenchymal cells toward osteoblastogenesis, without showing a synergistic effect. Our data indicate SW therapy, alone and in combination with raloxifene, as an innovative strategy to limit the hypoestrogenic bone loss, restoring the balance between bone formation and resorption. PMID:28158228

  12. Geochemical factors promoting die-back gap formation in colonizing patches of Spartina densiflora in an irregularly flooded marsh

    NASA Astrophysics Data System (ADS)

    Mirlean, Nicolai; Costa, Cesar S. B.

    2017-04-01

    Circular (RP) and ring-shape (RP) patches of vegetation in intertidal flats have been associated with the radial expansion of tussock growth forms and die-back gap in older central stands, respectively. RP formation has not yet been sufficiently explained. We accomplished a comparative geochemical study of CP and RP structures of Spartina densiflora within a single saltmarsh in a microtidal estuary (<0.5 m). The pore water under these structures demonstrated distinctive physical-chemical properties by marked seasonal changing in water level and salinity. During high-water period dissolved H2S was frequently low in pore waters of S. densiflora structures due to reactive-Fe, which scavenge the sulfide from solution and form solid sulfides. During less flooded-brackish water period, pore water pH goes down below 4 inside the vegetated bordering areas of RP. In these locations the concentration of soluble sulfides dramatically increases up to 140 μM L-1. The high concentration of protons in pore water is the result of solid sulfides atmospheric oxidation to sulfuric acid. High dissolution of H2S, along with the low pH, creates a toxic environment for S. densiflora and die-back central gap formation in RP. CP structure was 5 cm higher in the intertidal than RP but shows frequent presence of a water layer, less severe oxidation of sulfides and limited building-up of toxic condition to plants. Development of S. densiflora RP probably indicates the uplift of sediment by this bioengineer grass and/or periodic lowering of the water surface below a certain critical level.

  13. Is Active Management the Key to the Conservation of Saproxylic Biodiversity? Pollarding Promotes the Formation of Tree Hollows

    PubMed Central

    Sebek, Pavel; Altman, Jan; Platek, Michal; Cizek, Lukas

    2013-01-01

    Trees with hollows are key features sustaining biodiversity in wooded landscapes. They host rich assemblages of often highly specialised organisms. Hollow trees, however, have become rare and localised in Europe. Many of the associated biota is thus declining or endangered. The challenge of its conservation, therefore, is to safeguard the presence of hollow trees in sufficient numbers. Populations of numerous species associated with tree hollows and dead wood are often found in habitats that were formed by formerly common traditional silvicultural practices such as coppicing, pollarding or pasture. Although it has been occasionally mentioned that such practices increase the formation of hollows and the availability of often sun-exposed dead wood, their effect has never been quantified. Our study examined the hollow incidence in pollard and non-pollard (unmanaged) willows and the effect of pollarding on incremental growth rate by tree ring analysis. The probability of hollow occurrence was substantially higher in pollard than in non-pollard trees. Young pollards, especially, form hollows much more often than non-pollards; for instance, in trees of 50 cm DBH, the probability of hollow ocurrence was ∼0.75 in pollards, but only ∼0.3 in non-pollards. No difference in growth rate was found. Pollarding thus leads to the rapid formation of tree hollows, a habitat usually associated with old trees. It is therefore potentially a very important tool in the restoration of saproxylic habitats and conservation of hollow-dependent fauna. If applied along e.g. roads and watercourses, pollarding could also be used to increase landscape connectivity for saproxylic organisms. In reserves where pollarding was formerly practiced, its restoration would be necessary to prevent loss of saproxylic biodiversity. Our results point to the importance of active management measures for maintaining availability, and spatial and temporal continuity of deadwood microhabitats. PMID:23544142

  14. Progesterone promotes focal adhesion formation and migration in breast cancer cells through induction of protease-activated receptor-1.

    PubMed

    Diaz, Jorge; Aranda, Evelyn; Henriquez, Soledad; Quezada, Marisol; Espinoza, Estefanía; Bravo, Maria Loreto; Oliva, Bárbara; Lange, Soledad; Villalon, Manuel; Jones, Marius; Brosens, Jan J; Kato, Sumie; Cuello, Mauricio A; Knutson, Todd P; Lange, Carol A; Leyton, Lisette; Owen, Gareth I

    2012-08-01

    Progesterone and progestins have been demonstrated to enhance breast cancer cell migration, although the mechanisms are still not fully understood. The protease-activated receptors (PARs) are a family of membrane receptors that are activated by serine proteases in the blood coagulation cascade. PAR1 (F2R) has been reported to be involved in cancer cell migration and overexpressed in breast cancer. We herein demonstrate that PAR1 mRNA and protein are upregulated by progesterone treatment of the breast cancer cell lines ZR-75 and T47D. This regulation is dependent on the progesterone receptor (PR) but does not require PR phosphorylation at serine 294 or the PR proline-rich region mPRO. The increase in PAR1 mRNA was transient, being present at 3  h and returning to basal levels at 18  h. The addition of a PAR1-activating peptide (aPAR1) to cells treated with progesterone resulted in an increase in focal adhesion (FA) formation as measured by the cellular levels of phosphorylated FA kinase. The combined but not individual treatment of progesterone and aPAR1 also markedly increased stress fiber formation and the migratory capacity of breast cancer cells. In agreement with in vitro findings, data mining from the Oncomine platform revealed that PAR1 expression was significantly upregulated in PR-positive breast tumors. Our observation that PAR1 expression and signal transduction are modulated by progesterone provides new insight into how the progestin component in hormone therapies increases the risk of breast cancer in postmenopausal women.

  15. SerpinB2 (PAI-2) Modulates Proteostasis via Binding Misfolded Proteins and Promotion of Cytoprotective Inclusion Formation

    PubMed Central

    Farrawell, Natalie; Shearer, Robert F.; Constantinescu, Patrick; Hatters, Danny M.; Schroder, Wayne A.; Suhrbier, Andreas; Wilson, Mark R.; Saunders, Darren N.; Ranson, Marie

    2015-01-01

    SerpinB2 (PAI-2), a member of the clade B family of serine protease inhibitors, is one of the most upregulated proteins following cellular stress. Originally described as an inhibitor of urokinase plasminogen activator, its predominant cytoplasmic localisation suggests an intracellular function. SerpinB2 has been reported to display cytoprotective properties in neurons and to interact with intracellular proteins including components of the ubiquitin-proteasome system (UPS). In the current study we explored the potential role of SerpinB2 as a modulator of proteotoxic stress. Initially, we transiently transfected wild-type SerpinB2 and SerpinB2-/- murine embryonic fibroblasts (MEFs) with Huntingtin exon1-polyglutamine (fused C-terminally to mCherry). Inclusion body formation as result of Huntingtin aggregation was evident in the SerpinB2 expressing cells but significantly impaired in the SerpinB2-/- cells, the latter concomitant with loss in cell viability. Importantly, recovery of the wild-type phenotype and cell viability was rescued by retroviral transduction of SerpinB2 expression. SerpinB2 modestly attenuated Huntingtin and amyloid beta fibril formation in vitro and was able to bind preferentially to misfolded proteins. Given the modest chaperone-like activity of SerpinB2 we tested the ability of SerpinB2 to modulate UPS and autophagy activity using a GFP reporter system and autophagy reporter, respectively. Activity of the UPS was reduced and autophagy was dysregulated in SerpinB2-/- compared to wild-type MEFs. Moreover, we observed a non-covalent interaction between ubiquitin and SerpinB2 in cells using GFP-pulldown assays and bimolecular fluorescence complementation. We conclude that SerpinB2 plays an important role in proteostasis as its loss leads to a proteotoxic phenotype associated with an inability to compartmentalize aggregating proteins and a reduced capacity of the UPS. PMID:26083412

  16. Strain differences in urinary factors that promote calcium oxalate crystal formation in the kidneys of ethylene glycol-treated rats.

    PubMed

    Li, Yan; McMartin, Kenneth E

    2009-05-01

    Ethylene glycol (EG)-induced hyperoxaluria is the most commonly employed experimental regimen as an animal model of calcium oxalate (CaOx) stone formation. The variant sensitivity to CaOx among different rat strains has not been fully explored, although the Wistar rat is known to accumulate more CaOx in kidney tissue after low-dose EG exposure than in the Fischer 344 (F344) rats. Supersaturation of CaOx in tubular fluid contributes to the amount of CaOx crystal formation in the kidney. We hypothesized that the urinary supersaturation of CaOx in Wistar rats is higher than that of F344 rats, thereby allowing for greater CaOx crystal deposition in the Wistar rat. Age-matched male Wistar and F344 rats were treated with 0.75% EG or drinking water for 8 wk. Twenty-four-hour urine was collected at 0, 2, 4, 6, and 8 wk for analysis of key electrolytes to calculate the CaOx supersaturation. Plasma oxalate level was also measured. Our data confirmed the different sensitivity to renal toxicity from EG between the two rat strains (Wistar > F344). After EG treatment, the plasma oxalate level and urine oxalate excretion were markedly greater in the Wistar rats than in the F344 rats, while urine calcium was slightly decreased in Wistars. Thus, the CaOx supersaturation in urine of Wistar rats was higher, which led to a greater crystal deposition in kidney in Wistar rats. These studies suggest that during EG treatment, changes in urine electrolytes and in CaOx supersaturation occur to a greater extent in the Wistar rat, in agreement with its greater sensitivity to EG toxicity.

  17. Respiratory Syncytial Virus Fusion Protein Promotes TLR-4–Dependent Neutrophil Extracellular Trap Formation by Human Neutrophils

    PubMed Central

    Funchal, Giselle A.; Jaeger, Natália; Czepielewski, Rafael S.; Machado, Mileni S.; Muraro, Stéfanie P.; Stein, Renato T.; Bonorino, Cristina B. C.; Porto, Bárbara N.

    2015-01-01

    Acute viral bronchiolitis by Respiratory Syncytial Virus (RSV) is the most common respiratory illness in children in the first year of life. RSV bronchiolitis generates large numbers of hospitalizations and an important burden to health systems. Neutrophils and their products are present in the airways of RSV-infected patients who developed increased lung disease. Neutrophil Extracellular Traps (NETs) are formed by the release of granular and nuclear contents of neutrophils in the extracellular space in response to different stimuli and recent studies have proposed a role for NETs in viral infections. In this study, we show that RSV particles and RSV Fusion protein were both capable of inducing NET formation by human neutrophils. Moreover, we analyzed the mechanisms involved in RSV Fusion protein-induced NET formation. RSV F protein was able to induce NET release in a concentration-dependent fashion with both neutrophil elastase and myeloperoxidase expressed on DNA fibers and F protein-induced NETs was dismantled by DNase treatment, confirming that their backbone is chromatin. This viral protein caused the release of extracellular DNA dependent on TLR-4 activation, NADPH Oxidase-derived ROS production and ERK and p38 MAPK phosphorylation. Together, these results demonstrate a coordinated signaling pathway activated by F protein that led to NET production. The massive production of NETs in RSV infection could aggravate the inflammatory symptoms of the infection in young children and babies. We propose that targeting the binding of TLR-4 by F protein could potentially lead to novel therapeutic approaches to help control RSV-induced inflammatory consequences and pathology of viral bronchiolitis. PMID:25856628

  18. Bacterial genotoxins promote inside-out integrin β1 activation, formation of focal adhesion complexes and cell spreading.

    PubMed

    Levi, Laura; Toyooka, Tatsushi; Patarroyo, Manuel; Frisan, Teresa

    2015-01-01

    Integrins are membrane bound receptors that regulate several cellular processes, such as cell adhesion, migration, survival and proliferation, and may contribute to tumor initiation/progression in cells exposed to genotoxic stress. The extent of integrin activation and its role in cell survival upon intoxication with bacterial genotoxins are still poorly characterized. These toxins induce DNA strand breaks in the target cells and activate the DNA damage response (DDR), coordinated by the Ataxia Telangectasia Mutated (ATM) kinase. In the present study, we demonstrate that induction of DNA damage by two bacterial genotoxins promotes activation of integrin β1, leading to enhanced assembly of focal adhesions and cell spreading on fibronectin, but not on vitronectin. This phenotype is mediated by an ATM-dependent inside-out integrin signaling, and requires the actin cytoskeleton remodeler NET1. The toxin-mediated cell spreading and anchorage-independent survival further relies on ALIX and TSG101, two components of the endosomal sorting complex required for transport (ESCRT), known to regulate integrin intracellular trafficking. These data reveal a novel aspect of the cellular response to bacterial genotoxins, and provide new tools to understand the carcinogenic potential of these effectors in the context of chronic intoxication and infection.

  19. α-Synuclein assembles into higher-order multimers upon membrane binding to promote SNARE complex formation

    PubMed Central

    Burré, Jacqueline; Sharma, Manu; Südhof, Thomas C.

    2014-01-01

    Physiologically, α-synuclein chaperones soluble NSF attachment protein receptor (SNARE) complex assembly and may also perform other functions; pathologically, in contrast, α-synuclein misfolds into neurotoxic aggregates that mediate neurodegeneration and propagate between neurons. In neurons, α-synuclein exists in an equilibrium between cytosolic and membrane-bound states. Cytosolic α-synuclein appears to be natively unfolded, whereas membrane-bound α-synuclein adopts an α-helical conformation. Although the majority of studies showed that cytosolic α-synuclein is monomeric, it is unknown whether membrane-bound α-synuclein is also monomeric, and whether chaperoning of SNARE complex assembly by α-synuclei