Role and regulation of autophagy in heat stress responses of tomato plants

PubMed Central

Zhou, Jie; Wang, Jian; Yu, Jing-Quan; Chen, Zhixiang

2014-01-01

As sessile organisms, plants are constantly exposed to a wide spectrum of stress conditions such as high temperature, which causes protein misfolding. Misfolded proteins are highly toxic and must be efficiently removed to reduce cellular proteotoxic stress if restoration of native conformations is unsuccessful. Although selective autophagy is known to function in protein quality control by targeting degradation of misfolded and potentially toxic proteins, its role and regulation in heat stress responses have not been analyzed in crop plants. In the present study, we found that heat stress induced expression of autophagy-related (ATG) genes and accumulation of autophagosomes in tomato plants. Virus-induced gene silencing (VIGS) of tomato ATG5 and ATG7 genes resulted in increased sensitivity of tomato plants to heat stress based on both increased development of heat stress symptoms and compromised photosynthetic parameters of heat-stressed leaf tissues. Silencing of tomato homologs for the selective autophagy receptor NBR1, which targets ubiquitinated protein aggregates, also compromised tomato heat tolerance. To better understand the regulation of heat-induced autophagy, we found that silencing of tomato ATG5, ATG7, or NBR1 compromised heat-induced expression of not only the targeted genes but also other autophagy-related genes. Furthermore, we identified two tomato genes encoding proteins highly homologous to Arabidopsis WRKY33 transcription factor, which has been previously shown to interact physically with an autophagy protein. Silencing of tomato WRKY33 genes compromised tomato heat tolerance and reduced heat-induced ATG gene expression and autophagosome accumulation. Based on these results, we propose that heat-induced autophagy in tomato is subject to cooperative regulation by both WRKY33 and ATG proteins and plays a critical role in tomato heat tolerance, mostly likely through selective removal of heat-induced protein aggregates. PMID:24817875

Protein Kinase A Regulates Molecular Chaperone Transcription and Protein Aggregation

PubMed Central

Prince, Thomas; Calderwood, Stuart K.

2011-01-01

Heat shock factor 1 (HSF1) regulates one of the major pathways of protein quality control and is essential for deterrence of protein-folding disorders, particularly in neuronal cells. However, HSF1 activity declines with age, a change that may open the door to progression of neurodegenerative disorders such as Huntington's disease. We have investigated mechanisms of HSF1 regulation that may become compromised with age. HSF1 binds stably to the catalytic domain of protein kinase A (PKAcα) and becomes phosphorylated on at least one regulatory serine residue (S320). We show here that PKA is essential for effective transcription of HSP genes by HSF1. PKA triggers a cascade involving HSF1 binding to the histone acetylase p300 and positive translation elongation factor 1 (p-TEFb) and phosphorylation of the c-terminal domain of RNA polymerase II, a key mechanism in the downstream steps of HSF1-mediated transcription. This cascade appears to play a key role in protein quality control in neuronal cells expressing aggregation-prone proteins with long poly-glutamine (poly-Q) tracts. Such proteins formed inclusion bodies that could be resolved by HSF1 activation during heat shock. Resolution of the inclusions was inhibited by knockdown of HSF1, PKAcα, or the pTEFb component CDK9, indicating a key role for the HSF1-PKA cascade in protein quality control. PMID:22216146

Rilmenidine promotes MTOR-independent autophagy in the mutant SOD1 mouse model of amyotrophic lateral sclerosis without slowing disease progression

PubMed Central

Perera, Nirma D.; Sheean, Rebecca K.; Lau, Chew L.; Shin, Yea Seul; Beart, Philip M.; Horne, Malcolm K.; Turner, Bradley J.

2018-01-01

ABSTRACT Macroautophagy/autophagy is the main intracellular catabolic pathway in neurons that eliminates misfolded proteins, aggregates and damaged organelles associated with ageing and neurodegeneration. Autophagy is regulated by both MTOR-dependent and -independent pathways. There is increasing evidence that autophagy is compromised in neurodegenerative disorders, which may contribute to cytoplasmic sequestration of aggregation-prone and toxic proteins in neurons. Genetic or pharmacological modulation of autophagy to promote clearance of misfolded proteins may be a promising therapeutic avenue for these disorders. Here, we demonstrate robust autophagy induction in motor neuronal cells expressing SOD1 or TARDBP/TDP-43 mutants linked to amyotrophic lateral sclerosis (ALS). Treatment of these cells with rilmenidine, an anti-hypertensive agent and imidazoline-1 receptor agonist that induces autophagy, promoted autophagic clearance of mutant SOD1 and efficient mitophagy. Rilmenidine administration to mutant SOD1G93A mice upregulated autophagy and mitophagy in spinal cord, leading to reduced soluble mutant SOD1 levels. Importantly, rilmenidine increased autophagosome abundance in motor neurons of SOD1G93A mice, suggesting a direct action on target cells. Despite robust induction of autophagy in vivo, rilmenidine worsened motor neuron degeneration and symptom progression in SOD1G93A mice. These effects were associated with increased accumulation and aggregation of insoluble and misfolded SOD1 species outside the autophagy pathway, and severe mitochondrial depletion in motor neurons of rilmenidine-treated mice. These findings suggest that rilmenidine treatment may drive disease progression and neurodegeneration in this mouse model due to excessive mitophagy, implying that alternative strategies to beneficially stimulate autophagy are warranted in ALS. PMID:28980850

Sigma-1 receptor agonist PRE084 is protective against mutant huntingtin-induced cell degeneration: involvement of calpastatin and the NF-κB pathway

PubMed Central

Hyrskyluoto, A; Pulli, I; Törnqvist, K; Huu Ho, T; Korhonen, L; Lindholm, D

2013-01-01

Alterations in mitochondria and increased oxidative stress are associated with the disease progression in Huntington's disease (HD). Endoplasmic reticulum (ER) stress and oxidative damage are linked through the close communication between the ER and mitochondria. Sigma-1 receptor (Sig-1R) is a chaperone protein in the ER that is involved in ER stress regulation, but little is known about its role in HD or the mechanisms for cell protection. Here we show that the Sig-1R agonist, PRE084 increases cell survival and counteracts the deleterious effects caused by N-terminal mutant huntingtin proteins in neuronal PC6.3 cells. Particularly, PRE084 increased the levels of cellular antioxidants by activating the NF-κB pathway that is compromised by the expression of mutant huntingtin proteins. These results show that the Sig-1R agonist has beneficial effects in models of HD and that compounds affecting the Sig-1R may be promising targets for future drug development in HD. PMID:23703391

Ki-67 Contributes to Normal Cell Cycle Progression and Inactive X Heterochromatin in p21 Checkpoint-Proficient Human Cells

PubMed Central

Sun, Xiaoming; Bizhanova, Aizhan; Matheson, Timothy D.; Yu, Jun; Zhu, Lihua Julie

2017-01-01

ABSTRACT The Ki-67 protein is widely used as a tumor proliferation marker. However, whether Ki-67 affects cell cycle progression has been controversial. Here we demonstrate that depletion of Ki-67 in human hTERT-RPE1, WI-38, IMR90, and hTERT-BJ cell lines and primary fibroblast cells slowed entry into S phase and coordinately downregulated genes related to DNA replication. Some gene expression changes were partially relieved in Ki-67-depleted hTERT-RPE1 cells by codepletion of the Rb checkpoint protein, but more thorough suppression of the transcriptional and cell cycle defects was observed upon depletion of the cell cycle inhibitor p21. Notably, induction of p21 upon depletion of Ki-67 was a consistent hallmark of cell types in which transcription and cell cycle distribution were sensitive to Ki-67; these responses were absent in cells that did not induce p21. Furthermore, upon Ki-67 depletion, a subset of inactive X (Xi) chromosomes in female hTERT-RPE1 cells displayed several features of compromised heterochromatin maintenance, including decreased H3K27me3 and H4K20me1 labeling. These chromatin alterations were limited to Xi chromosomes localized away from the nuclear lamina and were not observed in checkpoint-deficient 293T cells. Altogether, our results indicate that Ki-67 integrates normal S-phase progression and Xi heterochromatin maintenance in p21 checkpoint-proficient human cells. PMID:28630280

Sera from remitting and secondary progressive multiple sclerosis patients disrupt the blood-brain barrier.

PubMed

Shimizu, Fumitaka; Tasaki, Ayako; Sano, Yasuteru; Ju, Mihua; Nishihara, Hideaki; Oishi, Mariko; Koga, Michiaki; Kawai, Motoharu; Kanda, Takashi

2014-01-01

Pathological destruction of blood-brain barrier (BBB) has been thought to be the initial key event in the process of developing multiple sclerosis (MS). The purpose of the present study was to clarify the possible molecular mechanisms responsible for the malfunction of BBB by sera from relapse-remitting MS (RRMS) and secondary progressive MS (SPMS) patients. We evaluated the effects of sera from the patients in the relapse phase of RRMS (RRMS-R), stable phase of RRMS (RRMS-S) and SPMS on the expression of tight junction proteins and vascular cell adhesion protein-1 (VCAM-1), and on the transendothelial electrical resistance (TEER) in human brain microvascular endothelial cells (BMECs). Sera from the RRMS-R or SPMS patients decreased the claudin-5 protein expression and the TEER in BMECs. In RRMS-R, this effect was restored after adding an MMP inhibitor, and the MMP-2/9 secretion by BMECs was significantly increased after the application of patients' sera. In SPMS, the immunoglobulin G (IgG) purified from patients' sera also decreased the claudin-5 protein expression and the TEER in BMECs. The sera and purified IgG from all MS patients increased the VCAM-1 protein expression in BMECs. The up-regulation of autocrine MMP-2/9 by BMECs after exposure to sera from RRMS-R patients or the autoantibodies against BMECs from SPMS patients can compromise the BBB. Both RRMS-S and SPMS sera increased the VCAM-1 expression in the BBB, thus indicating that targeting the VCAM-1 in the BBB could represent a possible therapeutic strategy for even the stable phase of MS and SPMS.

Political Compromise Makes the World Go 'Round

ERIC Educational Resources Information Center

Everett, Diana

2007-01-01

Compromise in any context is often hard to accept. It feels like a person is giving up on his or her ideals. This is especially true in dealing with politics. Legislative and congressional bills can be written with the highest of ideals in mind. By the time the bill progresses through committees and the floor debate process, it can look like a…

Mutations Allow JC Polyomaviruses to Elude Antibody Recognition | Center for Cancer Research

Cancer.gov

JC polyomavirus (JCV) infects the urinary tract of most adults. In healthy individuals, JCV infection does not cause noticeable symptoms. However, in those with compromised immune systems, JCV can cause a lethal brain disease called progressive multifocal leukoencephalopathy (PML). Data from a recently approved assay to detect serum antibodies specific for the JCV protein VP1 revealed that patients with antibodies are at increased risk of developing PML. At the same time, sequencing studies of JCV in cerebrospinal fluid (CSF) identified a number of mutations in VP1. Christopher Buck, Ph.D., and Diana Pastrana, Ph.D., of CCR’s Laboratory of Cellular Oncology, and their colleagues hypothesized that the VP1 mutations could allow the virus to evade antibody-mediated elimination.

Positive and negative early life experiences differentially modulate long term survival and amyloid protein levels in a mouse model of Alzheimer's disease.

PubMed

Lesuis, Sylvie L; Maurin, Herve; Borghgraef, Peter; Lucassen, Paul J; Van Leuven, Fred; Krugers, Harm J

2016-06-28

Stress has been implicated as a risk factor for the severity and progression of sporadic Alzheimer's disease (AD). Early life experiences determine stress responsivity in later life, and modulate age-dependent cognitive decline. Therefore, we examined whether early life experiences influence AD outcome in a bigenic mouse model which progressively develops combined tau and amyloid pathology (biAT mice).Mice were subjected to either early life stress (ELS) or to 'positive' early handling (EH) postnatally (from day 2 to 9). In biAT mice, ELS significantly compromised long term survival, in contrast to EH which increased life expectancy. In 4 month old mice, ELS-reared biAT mice displayed increased hippocampal Aβ levels, while these levels were reduced in EH-reared biAT mice. No effects of ELS or EH were observed on the brain levels of APP, protein tau, or PSD-95. Dendritic morphology was moderately affected after ELS and EH in the amygdala and medial prefrontal cortex, while object recognition memory and open field performance were not affected. We conclude that despite the strong transgenic background, early life experiences significantly modulate the life expectancy of biAT mice. Parallel changes in hippocampal Aβ levels were evident, without affecting cognition of young adult biAT mice.

Re-examining the phosphorus-protein dilemma: Does phosphorus restriction compromise protein status?

PubMed Central

St-Jules, David E; Woolf, Kathleen; Pompeii, Mary-Lou; Kalantar-Zadeh, Kamyar; Sevick, Mary Ann

2015-01-01

Dietary phosphorus restriction is recommended to help control hyperphosphatemia in hemodialysis (HD) patients, but many high-phosphorus foods are important sources of protein. In this review, we examine whether restricting dietary phosphorus compromises protein status in HD patients. Although dietary phosphorus and protein are highly correlated, phosphorus intakes can range up to 600 mg/day for a given energy and protein intake level. Further, the collinearity of phosphorus and protein may be biased because the phosphorus burden of food depends on: (1) the presence of phosphate additives; (2) food preparation method; and (3) bioavailability of phosphorus; which are often unaccounted for in nutrition assessments. Ultimately, we argue that clinically relevant reductions in phosphorus intake can be made without limiting protein intake by avoiding phosphate additives in processed foods, using wet cooking methods such as boiling, and if needed, substituting high-phosphorus foods for nutritionally-equivalent foods that are lower in bioavailable phosphorus. PMID:26873260

Ionizing radiation induces heritable disruption of epithelial cell interactions

NASA Technical Reports Server (NTRS)

Park, Catherine C.; Henshall-Powell, Rhonda L.; Erickson, Anna C.; Talhouk, Rabih; Parvin, Bahram; Bissell, Mina J.; Barcellos-Hoff, Mary Helen; Chatterjee, A. (Principal Investigator)

2003-01-01

Ionizing radiation (IR) is a known human breast carcinogen. Although the mutagenic capacity of IR is widely acknowledged as the basis for its action as a carcinogen, we and others have shown that IR can also induce growth factors and extracellular matrix remodeling. As a consequence, we have proposed that an additional factor contributing to IR carcinogenesis is the potential disruption of critical constraints that are imposed by normal cell interactions. To test this hypothesis, we asked whether IR affected the ability of nonmalignant human mammary epithelial cells (HMEC) to undergo tissue-specific morphogenesis in culture by using confocal microscopy and imaging bioinformatics. We found that irradiated single HMEC gave rise to colonies exhibiting decreased localization of E-cadherin, beta-catenin, and connexin-43, proteins necessary for the establishment of polarity and communication. Severely compromised acinar organization was manifested by the majority of irradiated HMEC progeny as quantified by image analysis. Disrupted cell-cell communication, aberrant cell-extracellular matrix interactions, and loss of tissue-specific architecture observed in the daughters of irradiated HMEC are characteristic of neoplastic progression. These data point to a heritable, nonmutational mechanism whereby IR compromises cell polarity and multicellular organization.

Air Duster abuse causing rapid airway compromise

PubMed Central

Winston, Amanda; Kanzy, Abed; Bachuwa, Ghassan

2015-01-01

Inhalant abuse is potentially life-threatening and has resulted in many complications such as central nervous system depression, cardiac dysrhythmia and hypoxia. Inhalant abuse causing angioedema is rarely reported in the medical literature. In this report we present a case of rapidly progressive airway compromise following recreational huffing. Our patient required intubation and intensive care unit admission with complete recovery after 5 days. The aetiology of airway compromise is postulated to be due to commonly reported frost bite injury and rarely reported angioedema. To the best of our knowledge this the second case reporting angioedema secondary to huffing Air Duster. PMID:25568278

Separate RNA-binding surfaces on the multifunctional La protein mediate distinguishable activities in tRNA maturation.

PubMed

Huang, Ying; Bayfield, Mark A; Intine, Robert V; Maraia, Richard J

2006-07-01

By sequence-specific binding to 3' UUU-OH, the La protein shields precursor (pre)-RNAs from 3' end digestion and is required to protect defective pre-transfer RNAs from decay. Although La is comprised of a La motif and an RNA-recognition motif (RRM), a recent structure indicates that the RRM beta-sheet surface is not involved in UUU-OH recognition, raising questions as to its function. Progressively defective suppressor tRNAs in Schizosaccharomyces pombe reveal differential sensitivities to La and Rrp6p, a 3' exonuclease component of pre-tRNA decay. 3' end protection is compromised by mutations to the La motif but not the RRM surface. The most defective pre-tRNAs require a second activity of La, in addition to 3' protection, that requires an intact RRM surface. The two activities of La in tRNA maturation map to its two conserved RNA-binding surfaces and suggest a modular model that has implications for its other ligands.

Reexamining the Phosphorus-Protein Dilemma: Does Phosphorus Restriction Compromise Protein Status?

PubMed

St-Jules, David E; Woolf, Kathleen; Pompeii, Mary Lou; Kalantar-Zadeh, Kamyar; Sevick, Mary Ann

2016-05-01

Dietary phosphorus restriction is recommended to help control hyperphosphatemia in hemodialysis patients, but many high-phosphorus foods are important sources of protein. In this review, we examine whether restricting dietary phosphorus compromises protein status in hemodialysis patients. Although dietary phosphorus and protein are highly correlated, phosphorus intakes can range up to 600 mg/day for a given energy and protein intake level. Furthermore, the collinearity of phosphorus and protein may be biased because the phosphorus burden of food depends on: (1) the presence of phosphate additives, (2) food preparation method, and (3) bioavailability of phosphorus, which are often unaccounted for in nutrition assessments. Ultimately, we argue that clinically relevant reductions in phosphorus intake can be made without limiting protein intake by avoiding phosphate additives in processed foods, using wet cooking methods such as boiling, and if needed, substituting high-phosphorus foods for nutritionally equivalent foods that are lower in bioavailable phosphorus. Copyright © 2016 National Kidney Foundation, Inc. Published by Elsevier Inc. All rights reserved.

Air Duster abuse causing rapid airway compromise.

PubMed

Winston, Amanda; Kanzy, Abed; Bachuwa, Ghassan

2015-01-07

Inhalant abuse is potentially life-threatening and has resulted in many complications such as central nervous system depression, cardiac dysrhythmia and hypoxia. Inhalant abuse causing angioedema is rarely reported in the medical literature. In this report we present a case of rapidly progressive airway compromise following recreational huffing. Our patient required intubation and intensive care unit admission with complete recovery after 5 days. The aetiology of airway compromise is postulated to be due to commonly reported frost bite injury and rarely reported angioedema. To the best of our knowledge this the second case reporting angioedema secondary to huffing Air Duster. 2015 BMJ Publishing Group Ltd.

Inadequate satellite cell replication compromises muscle regrowth following postnatal nutrient restriction

USDA-ARS?s Scientific Manuscript database

Perinatal growth impairment permanently compromises skeletal muscle mass. The present study assessed the contribution of muscle satellite cell replicative capacity to this deficit. Mouse dams were fed either a low protein (LP, n=7) or control (C, n=6) diet during lactation. Pups were weaned at 21 d ...

CNGA3 deficiency affects cone synaptic terminal structure and function and leads to secondary rod dysfunction and degeneration.

PubMed

Xu, Jianhua; Morris, Lynsie M; Michalakis, Stylianos; Biel, Martin; Fliesler, Steven J; Sherry, David M; Ding, Xi-Qin

2012-03-01

To investigate rod function and survival after cone dysfunction and degeneration in a mouse model of cone cyclic nucleotide-gated (CNG) channel deficiency. Rod function and survival in mice with cone CNG channel subunit CNGA3 deficiency (CNGA3-/- mice) were evaluated by electroretinographic (ERG), morphometric, and Western blot analyses. The arrangement, integrity, and ultrastructure of photoreceptor terminals were investigated by immunohistochemistry and electron microscopy. The authors found loss of cone function and cone death accompanied by impairment of rods and rod-driven signaling in CNGA3-/- mice. Scotopic ERG b-wave amplitudes were reduced by 15% at 1 month, 30% at 6 months, and 40% at 9 months and older, while scotopic a-wave amplitudes were decreased by 20% at 9 months, compared with ERGs of age-matched wild-type mice. Outer nuclear layer thickness in CNGA3-/- retina was reduced by 15% at 12 months compared with age-matched wild-type controls. This was accompanied by a 30%-40% reduction in expression of rod-specific proteins, including rhodopsin, rod transducin α-subunit, and glutamic acid-rich protein (GARP). Cone terminals in the CNGA3-/- retina showed a progressive loss of neurochemical and ultrastructural integrity. Abnormalities were observed as early as 1 month. Disorganized rod terminal ultrastructure was noted by 12 months. These findings demonstrate secondary rod impairment and degeneration after cone degeneration in mice with cone CNG channel deficiency. Loss of cone phototransduction accompanies the compromised integrity of cone terminals. With time, rod synaptic structure, function, and viability also become compromised.

Straight A's: Public Education Policy and Progress. Volume 6, Number 11

ERIC Educational Resources Information Center

Amos, Jason, Ed.

2006-01-01

"Straight A's: Public Education Policy and Progress" is a biweekly newsletter that focuses on education news and events both in Washington, DC and around the country. The following articles are included in this issue: (1) House Passes Budget Resolution: Prospects of a Compromise with the Senate Appear Dim; (2) Older Students' Science…

A proangiogenic signaling axis in myeloid cells promotes malignant progression of glioma.

PubMed

Huang, Yujie; Rajappa, Prajwal; Hu, Wenhuo; Hoffman, Caitlin; Cisse, Babacar; Kim, Joon-Hyung; Gorge, Emilie; Yanowitch, Rachel; Cope, William; Vartanian, Emma; Xu, Raymond; Zhang, Tuo; Pisapia, David; Xiang, Jenny; Huse, Jason; Matei, Irina; Peinado, Hector; Bromberg, Jacqueline; Holland, Eric; Ding, Bi-Sen; Rafii, Shahin; Lyden, David; Greenfield, Jeffrey

2017-05-01

Tumors are capable of coopting hematopoietic cells to create a suitable microenvironment to support malignant growth. Here, we have demonstrated that upregulation of kinase insert domain receptor (KDR), also known as VEGFR2, in a myeloid cell sublineage is necessary for malignant progression of gliomas in transgenic murine models and is associated with high-grade tumors in patients. KDR expression increased in myeloid cells as myeloid-derived suppressor cells (MDSCs) accumulated, which was associated with the transformation and progression of low-grade fibrillary astrocytoma to high-grade anaplastic gliomas. KDR deficiency in murine BM-derived cells (BMDCs) suppressed the differentiation of myeloid lineages and reduced granulocytic/monocytic populations. The depletion of myeloid-derived KDR compromised its proangiogenic function, which inhibited the angiogenic switch necessary for malignant progression of low-grade to high-grade tumors. We also identified inhibitor of DNA binding protein 2 (ID2) as a key upstream regulator of KDR activation during myeloid differentiation. Deficiency of ID2 in BMDCs led to downregulation of KDR, suppression of proangiogenic myeloid cells, and prevention of low-grade to high-grade transition. Tumor-secreted TGF-β and granulocyte-macrophage CSF (GM-CSF) enhanced the KDR/ID2 signaling axis in BMDCs. Our results suggest that modulation of KDR/ID2 signaling may restrict tumor-associated myeloid cells and could potentially be a therapeutic strategy for preventing transformation of premalignant gliomas.

A proangiogenic signaling axis in myeloid cells promotes malignant progression of glioma

PubMed Central

Huang, Yujie; Rajappa, Prajwal; Hu, Wenhuo; Hoffman, Caitlin; Cisse, Babacar; Kim, Joon-Hyung; Gorge, Emilie; Yanowitch, Rachel; Cope, William; Vartanian, Emma; Xu, Raymond; Pisapia, David; Xiang, Jenny; Huse, Jason; Matei, Irina; Peinado, Hector; Bromberg, Jacqueline; Holland, Eric; Ding, Bi-sen; Rafii, Shahin; Lyden, David; Greenfield, Jeffrey

2017-01-01

Tumors are capable of coopting hematopoietic cells to create a suitable microenvironment to support malignant growth. Here, we have demonstrated that upregulation of kinase insert domain receptor (KDR), also known as VEGFR2, in a myeloid cell sublineage is necessary for malignant progression of gliomas in transgenic murine models and is associated with high-grade tumors in patients. KDR expression increased in myeloid cells as myeloid-derived suppressor cells (MDSCs) accumulated, which was associated with the transformation and progression of low-grade fibrillary astrocytoma to high-grade anaplastic gliomas. KDR deficiency in murine BM-derived cells (BMDCs) suppressed the differentiation of myeloid lineages and reduced granulocytic/monocytic populations. The depletion of myeloid-derived KDR compromised its proangiogenic function, which inhibited the angiogenic switch necessary for malignant progression of low-grade to high-grade tumors. We also identified inhibitor of DNA binding protein 2 (ID2) as a key upstream regulator of KDR activation during myeloid differentiation. Deficiency of ID2 in BMDCs led to downregulation of KDR, suppression of proangiogenic myeloid cells, and prevention of low-grade to high-grade transition. Tumor-secreted TGF-β and granulocyte-macrophage CSF (GM-CSF) enhanced the KDR/ID2 signaling axis in BMDCs. Our results suggest that modulation of KDR/ID2 signaling may restrict tumor-associated myeloid cells and could potentially be a therapeutic strategy for preventing transformation of premalignant gliomas. PMID:28394259

Mutant Ataxin-1 Inhibits Neural Progenitor Cell Proliferation in SCA1

PubMed Central

Cvetanovic, Marija; Hu, Yuan-Shih; Opal, Puneet

2017-01-01

Spinocerebellar ataxia type 1 (SCA1) is a dominantly inherited neurodegenerative disease caused by the expansion of a polyglutamine (Q) repeat tract in the protein ataxin-1 (ATXN1). Beginning as a cerebellar ataxic disorder, SCA1 progresses to involve the cerebral cortex, hippocampus, and brainstem. Using SCA1 knock-in mice that mirror the complexity of the human disease, we report a significant decrease in the capacity of adult neuronal progenitor cells (NPCs) to proliferate. Remarkably, a decrease in NPCs proliferation can be observed in vitro, outside the degenerative milieu of surrounding neurons or glia, demonstrating that mutant ATXN1 acting cell autonomously within progenitor cells interferes with their ability to proliferate. Our findings suggest that compromised adult neurogenesis contributes to the progressive pathology of the disease particularly in areas such as the hippocampus and cerebral cortex where stem cells provide neurotropic factors and participate in adult neurogenesis. These findings not only shed light on the biology of the disease but also have therapeutic implications in any future stem cell- based clinical trials. PMID:27306906

Mutant Ataxin-1 Inhibits Neural Progenitor Cell Proliferation in SCA1.

PubMed

Cvetanovic, Marija; Hu, Yuan-Shih; Opal, Puneet

2017-04-01

Spinocerebellar ataxia type 1 (SCA1) is a dominantly inherited neurodegenerative disease caused by the expansion of a polyglutamine (Q) repeat tract in the protein ataxin-1 (ATXN1). Beginning as a cerebellar ataxic disorder, SCA1 progresses to involve the cerebral cortex, hippocampus, and brainstem. Using SCA1 knock-in mice that mirror the complexity of the human disease, we report a significant decrease in the capacity of adult neuronal progenitor cells (NPCs) to proliferate. Remarkably, a decrease in NPCs proliferation can be observed in vitro, outside the degenerative milieu of surrounding neurons or glia, demonstrating that mutant ATXN1 acting cell autonomously within progenitor cells interferes with their ability to proliferate. Our findings suggest that compromised adult neurogenesis contributes to the progressive pathology of the disease particularly in areas such as the hippocampus and cerebral cortex where stem cells provide neurotropic factors and participate in adult neurogenesis. These findings not only shed light on the biology of the disease but also have therapeutic implications in any future stem cell-based clinical trials.

Fatal breathing dysfunction in a mouse model of Leigh syndrome.

PubMed

Quintana, Albert; Zanella, Sebastien; Koch, Henner; Kruse, Shane E; Lee, Donghoon; Ramirez, Jan M; Palmiter, Richard D

2012-07-01

Leigh syndrome (LS) is a subacute necrotizing encephalomyelopathy with gliosis in several brain regions that usually results in infantile death. Loss of murine Ndufs4, which encodes NADH dehydrogenase (ubiquinone) iron-sulfur protein 4, results in compromised activity of mitochondrial complex I as well as progressive neurodegenerative and behavioral changes that resemble LS. Here, we report the development of breathing abnormalities in a murine model of LS. Magnetic resonance imaging revealed hyperintense bilateral lesions in the dorsal brain stem vestibular nucleus (VN) and cerebellum of severely affected mice. The mutant mice manifested a progressive increase in apnea and had aberrant responses to hypoxia. Electrophysiological recordings within the ventral brain stem pre-Bötzinger respiratory complex were also abnormal. Selective inactivation of Ndufs4 in the VN, one of the principle sites of gliosis, also led to breathing abnormalities and premature death. Conversely, Ndufs4 restoration in the VN corrected breathing deficits and prolonged the life span of knockout mice. These data demonstrate that mitochondrial dysfunction within the VN results in aberrant regulation of respiration and contributes to the lethality of Ndufs4-knockout mice.

Negative feedback via RSK modulates Erk-dependent progression from naïve pluripotency.

PubMed

Nett, Isabelle Re; Mulas, Carla; Gatto, Laurent; Lilley, Kathryn S; Smith, Austin

2018-06-12

Mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) signalling is implicated in initiation of embryonic stem (ES) cell differentiation. The pathway is subject to complex feedback regulation. Here, we examined the ERK-responsive phosphoproteome in ES cells and identified the negative regulator RSK1 as a prominent target. We used CRISPR/Cas9 to create combinatorial mutations in RSK family genes. Genotypes that included homozygous null mutations in Rps6ka1, encoding RSK1, resulted in elevated ERK phosphorylation. These RSK-depleted ES cells exhibit altered kinetics of transition into differentiation, with accelerated downregulation of naïve pluripotency factors, precocious expression of transitional epiblast markers and early onset of lineage specification. We further show that chemical inhibition of RSK increases ERK phosphorylation and expedites ES cell transition without compromising multilineage potential. These findings demonstrate that the ERK activation profile influences the dynamics of pluripotency progression and highlight the role of signalling feedback in temporal control of cell state transitions. © 2018 The Authors. Published under the terms of the CC BY 4.0 license.

An orthologue of the host-defense protein psoriasin (S100A7) is expressed in frog skin.

PubMed

Matthijs, Severine; Hernalsteens, Jean-Pierre; Roelants, Kim

2017-02-01

Host-defense peptides and proteins are vital for first line protection against bacteria. Most host-defense peptides and proteins common in vertebrates have been studied primarily in mammals, while their orthologues in non-mammalian vertebrates received less attention. We found that the European Common Frog Rana temporaria expresses a protein in its skin that is evolutionarily related to the host-defense protein S100A7. This prompted us to test if the encoded protein, which is an important microbicidal protein in human skin, shows similar activity in frogs. The R. temporaria protein lacks the zinc-binding sites that are key to the antimicrobial activity of human S100A7 at neutral pH. However, despite being less potent, the R. temporaria protein does compromise bacterial membranes at low pH, similar to its human counterpart. We postulate that, while amphibian S100A7 likely serves other functions, the capacity to compromise bacterial cell membranes evolved early in tetrapod evolution. Copyright © 2016 Elsevier Ltd. All rights reserved.

[Clinical and therapeutic differences in neuro-ophthalmological involvement secondary to syphilis].

PubMed

Crespo-Burillo, J A; Gil-Perez, D; Alarcia-Alejos, R; Hernando-Quintana, N; Garcia-Rubio, S; Martin-Martinez, J

2014-09-16

INTRODUCTION. There are many forms of neuro-ophthalmological involvement secondary to syphilis, and not all of them are well known. Our aim is to determine the clinical and therapeutic differences in these patients. CASE REPORTS. Our sample included eight patients diagnosed with an ocular and neuro-ophthalmological disorder due to syphilis over the years 2012 and 2013. Five of them presented uveitis, pan-eveitis being the most frequent, with three cases. Two cases presented papilloedema and another displayed retrobulbar optic neuropathy. A total of 62.5% were diagnosed with neurosyphilis, the presence of which was related with compromise of the optic nerve (p = 0.035). None of them gave positive for VDRL in cerebrospinal fluid and they were diagnosed by the presence of FTA antibodies together with high protein levels in cerebrospinal fluid, lymphocytic pleocytosis or intrathecal synthesis of antibodies. In the absence of uveitis, diagnosis was delayed by a mean time of 2.6 months (p = 0.047). All the patients, except one who required a vitrectomy, progressed favourably with intravenous antibiotic therapy. CONCLUSIONS. In cases of neuro-ophthalmological compromise, whether inflammatory or non-inflammatory, the physician must bear syphilis in mind as a potential causation in order to avoid delays in the diagnosis, since early well-tailored treatment can prevent permanent loss of sight.

Adaptive changes in renal mitochondrial redox status in diabetic nephropathy

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

Putt, David A.; Zhong, Qing; Lash, Lawrence H., E-mail: l.h.lash@wayne.edu

2012-01-15

Nephropathy is a serious and common complication of diabetes. In the streptozotocin (STZ)-treated rat model of diabetes, nephropathy does not typically develop until 30 to 45 days post-injection, although hyperglycemia occurs within 24 h. We tested the hypothesis that chronic hyperglycemia results in a modest degree of oxidative stress that is accompanied by compensatory changes in certain antioxidants and mitochondrial redox status. We propose that as kidneys progress to a state of diabetic nephropathy, further adaptations occur in mitochondrial redox status. Basic parameters of renal function in vivo and several parameters of mitochondrial function and glutathione (GSH) and redox statusmore » in isolated renal cortical mitochondria from STZ-treated and age-matched control rats were examined at 30 days and 90 days post-injection. While there was no effect of diabetes on blood urea nitrogen, measurement of other, more sensitive parameters, such as urinary albumin and protein, and histopathology showed significant and progressive worsening in diabetic rats. Thus, renal function is compromised even prior to the onset of frank nephropathy. Changes in mitochondrial respiration and enzyme activities indicated existence of a hypermetabolic state. Higher mitochondrial GSH content and rates of GSH transport into mitochondria in kidneys from diabetic rats were only partially due to changes in expression of mitochondrial GSH carriers and were mostly due to higher substrate supply. Although there are few clear indicators of oxidative stress, there are several redox changes that occur early and change further as nephropathy progresses, highlighting the complexity of the disease. Highlights: ►Adaptive changes in renal mitochondrial and redox status in diabetic rats. ►Modest renal dysfunction even prior to onset of nephropathy. ►Elevated concentrations of mitochondrial GSH in diabetic kidneys. ►Change in GSH due partly to increased protein expression of transporter. ►Oxidatively modified proteins in renal mitochondria from diabetic rats.« less

Respiratory Compromise as a New Paradigm for the Care of Vulnerable Hospitalized Patients.

PubMed

Morris, Timothy A; Gay, Peter C; MacIntyre, Neil R; Hess, Dean R; Hanneman, Sandra K; Lamberti, James P; Doherty, Dennis E; Chang, Lydia; Seckel, Maureen A

2017-04-01

Acute respiratory compromise describes a deterioration in respiratory function with a high likelihood of rapid progression to respiratory failure and death. Identifying patients at risk for respiratory compromise coupled with monitoring of patients who have developed respiratory compromise might allow earlier interventions to prevent or mitigate further decompensation. The National Association for the Medical Direction of Respiratory Care (NAMDRC) organized a workshop meeting with representation from many national societies to address the unmet needs of respiratory compromise from a clinical practice perspective. Respiratory compromise may arise de novo or may complicate preexisting lung disease. The group identified distinct subsets of respiratory compromise that present similar opportunities for early detection and useful intervention to prevent respiratory failure. The subtypes were characterized by the pathophysiological mechanisms they had in common: impaired control of breathing, impaired airway protection, parenchymal lung disease, increased airway resistance, hydrostatic pulmonary edema, and right-ventricular failure. Classification of acutely ill respiratory patients into one or more of these categories may help in selecting the screening and monitoring strategies that are most appropriate for the patient's particular pathophysiology. Standardized screening and monitoring practices for patients with similar mechanisms of deterioration may enhance the ability to predict respiratory failure early and prevent its occurrence. Copyright © 2017 by Daedalus Enterprises.

Safeguards of Neurotransmission: Endocytic Adaptors as Regulators of Synaptic Vesicle Composition and Function

PubMed Central

Kaempf, Natalie; Maritzen, Tanja

2017-01-01

Communication between neurons relies on neurotransmitters which are released from synaptic vesicles (SVs) upon Ca2+ stimuli. To efficiently load neurotransmitters, sense the rise in intracellular Ca2+ and fuse with the presynaptic membrane, SVs need to be equipped with a stringently controlled set of transmembrane proteins. In fact, changes in SV protein composition quickly compromise neurotransmission and most prominently give rise to epileptic seizures. During exocytosis SVs fully collapse into the presynaptic membrane and consequently have to be replenished to sustain neurotransmission. Therefore, surface-stranded SV proteins have to be efficiently retrieved post-fusion to be used for the generation of a new set of fully functional SVs, a process in which dedicated endocytic sorting adaptors play a crucial role. The question of how the precise reformation of SVs is achieved is intimately linked to how SV membranes are retrieved. For a long time both processes were believed to be two sides of the same coin since Clathrin-mediated endocytosis (CME), the proposed predominant SV recycling mode, will jointly retrieve SV membranes and proteins. However, with the recent proposal of Clathrin-independent SV recycling pathways SV membrane retrieval and SV reformation turn into separable events. This review highlights the progress made in unraveling the molecular mechanisms mediating the high-fidelity retrieval of SV proteins and discusses how the gathered knowledge about SV protein recycling fits in with the new notions of SV membrane endocytosis. PMID:29085282

ROS-activated calcium signaling mechanisms regulating endothelial barrier function.

PubMed

Di, Anke; Mehta, Dolly; Malik, Asrar B

2016-09-01

Increased vascular permeability is a common pathogenic feature in many inflammatory diseases. For example in acute lung injury (ALI) and its most severe form, the acute respiratory distress syndrome (ARDS), lung microvessel endothelia lose their junctional integrity resulting in leakiness of the endothelial barrier and accumulation of protein rich edema. Increased reactive oxygen species (ROS) generated by neutrophils (PMNs) and other inflammatory cells play an important role in increasing endothelial permeability. In essence, multiple inflammatory syndromes are caused by dysfunction and compromise of the barrier properties of the endothelium as a consequence of unregulated acute inflammatory response. This review focuses on the role of ROS signaling in controlling endothelial permeability with particular focus on ALI. We summarize below recent progress in defining signaling events leading to increased endothelial permeability and ALI. Copyright © 2016 Elsevier Ltd. All rights reserved.

Flotillins control zebrafish epiboly through their role in cadherin-mediated cell-cell adhesion.

PubMed

Morris, Eduardo A Rios; Bodin, Stéphane; Delaval, Bénédicte; Comunale, Franck; Georget, Virginie; Costa, Manoel L; Lutfalla, Georges; Gauthier-Rouvière, Cécile

2017-05-01

Zebrafish gastrulation and particularly epiboly that involves coordinated movements of several cell layers is a dynamic process for which regulators remain to be identified. We show here that Flotillin 1 and 2, ubiquitous and highly conserved proteins, are required for epiboly. Flotillins knockdown compromised embryo survival, strongly delayed epiboly and impaired deep cell radial intercalation and directed collective migration without affecting enveloping layer cell movement. At the molecular level, we identified that Flotillins are required for the formation of E-cadherin-mediated cell-cell junctions. These results provide the first in vivo evidence that Flotillins regulate E-cadherin-mediated cell-cell junctions to allow epiboly progression. © 2017 Société Française des Microscopies and Société de Biologie Cellulaire de France. Published by John Wiley & Sons Ltd.

High fat diet-induced modifications in membrane lipid and mitochondrial-membrane protein signatures precede the development of hepatic insulin resistance in mice

PubMed Central

Kahle, M.; Schäfer, A.; Seelig, A.; Schultheiß, J.; Wu, M.; Aichler, M.; Leonhardt, J.; Rathkolb, B.; Rozman, J.; Sarioglu, H.; Hauck, S.M.; Ueffing, M.; Wolf, E.; Kastenmueller, G.; Adamski, J.; Walch, A.; Hrabé de Angelis, M.; Neschen, S.

2014-01-01

Objective Excess lipid intake has been implicated in the pathophysiology of hepatosteatosis and hepatic insulin resistance. Lipids constitute approximately 50% of the cell membrane mass, define membrane properties, and create microenvironments for membrane-proteins. In this study we aimed to resolve temporal alterations in membrane metabolite and protein signatures during high-fat diet (HF)-mediated development of hepatic insulin resistance. Methods We induced hepatosteatosis by feeding C3HeB/FeJ male mice an HF enriched with long-chain polyunsaturated C18:2n6 fatty acids for 7, 14, or 21 days. Longitudinal changes in hepatic insulin sensitivity were assessed via the euglycemic-hyperinsulinemic clamp, in membrane lipids via t-metabolomics- and membrane proteins via quantitative proteomics-analyses, and in hepatocyte morphology via electron microscopy. Data were compared to those of age- and litter-matched controls maintained on a low-fat diet. Results Excess long-chain polyunsaturated C18:2n6 intake for 7 days did not compromise hepatic insulin sensitivity, however, induced hepatosteatosis and modified major membrane lipid constituent signatures in liver, e.g. increased total unsaturated, long-chain fatty acid-containing acyl-carnitine or membrane-associated diacylglycerol moieties and decreased total short-chain acyl-carnitines, glycerophosphocholines, lysophosphatidylcholines, or sphingolipids. Hepatic insulin sensitivity tended to decrease within 14 days HF-exposure. Overt hepatic insulin resistance developed until day 21 of HF-intervention and was accompanied by morphological mitochondrial abnormalities and indications for oxidative stress in liver. HF-feeding progressively decreased the abundance of protein-components of all mitochondrial respiratory chain complexes, inner and outer mitochondrial membrane substrate transporters independent from the hepatocellular mitochondrial volume in liver. Conclusions We assume HF-induced modifications in membrane lipid- and protein-signatures prior to and during changes in hepatic insulin action in liver alter membrane properties – in particular those of mitochondria which are highly abundant in hepatocytes. In turn, a progressive decrease in the abundance of mitochondrial membrane proteins throughout HF-exposure likely impacts on mitochondrial energy metabolism, substrate exchange across mitochondrial membranes, contributes to oxidative stress, mitochondrial damage, and the development of insulin resistance in liver. PMID:25685688

Effect of simvastatin on the antihypertensive activity of losartan in hypertensive hypercholesterolemic animals and patients: role of nitric oxide, oxidative stress, and high-sensitivity C-reactive protein.

PubMed

Abdel-Zaher, Ahmed O; Elkoussi, Alaa Eldin A; Abudahab, Lotfy H; Elbakry, Mohammed H; Elsayed, Elsayed Abu-Elwafa

2014-06-01

This study investigated whether simvastatin has antihypertensive activity and can enhance the antihypertensive effect of losartan in hypertensive hypercholesterolemic animals and patients. Hypertension and hypercholesterolemia were induced in rats by L-NAME and cholesterol-enriched diet, respectively. In these animals, repeated administration of simvastatin decreased the systolic blood pressure, enhanced its progressive reductions induced by repeated administration of losartan, and corrected the compromised lipid profile. Concomitantly, repeated administration of simvastatin, losartan, or simvastatin in combination with losartan to these animals increased nitric oxide (NO) production and decreased the elevated serum malondialdehyde (MDA) and high-sensitivity C-reactive protein (hs-CRP) levels. Effects of combined treatment were greater than those of simvastatin or losartan alone. In hypertensive hypercholesterolemic patients, repeated administration of losartan decreased systolic and diastolic blood pressure, increased NO production, and decreased the elevated serum MDA and hs-CRP levels. Addition of simvastatin to losartan therapy enhanced these effects and corrected the compromised lipid profile. Simvastatin inhibited the contractile responses of isolated aortic rings induced by angiotensin II and enhanced the inhibitory effect of losartan on this preparation. l-arginine and acetylcholine enhanced, while L-NAME inhibited the effects of simvastatin, losartan, and their combination on these contractile responses. Thus, simvastatin exerts antihypertensive effect in hypertensive hypercholesterolemic animals and enhances the antihypertensive effect of losartan in hypertensive hypercholesterolemic animals and patients. Besides, its cholesterol-lowering effect, the ability of simvastatin to ameliorate endothelial dysfunction through increasing NO bioavailability and through suppression of oxidative stress and vascular inflammation may play an important role in these effects. © 2013 The Authors Fundamental and Clinical Pharmacology © 2013 Société Française de Pharmacologie et de Thérapeutique.

CNGA3 Deficiency Affects Cone Synaptic Terminal Structure and Function and Leads to Secondary Rod Dysfunction and Degeneration

PubMed Central

Xu, Jianhua; Morris, Lynsie M.; Michalakis, Stylianos; Biel, Martin; Fliesler, Steven J.; Sherry, David M.

2012-01-01

Purpose. To investigate rod function and survival after cone dysfunction and degeneration in a mouse model of cone cyclic nucleotide-gated (CNG) channel deficiency. Methods. Rod function and survival in mice with cone CNG channel subunit CNGA3 deficiency (CNGA3−/− mice) were evaluated by electroretinographic (ERG), morphometric, and Western blot analyses. The arrangement, integrity, and ultrastructure of photoreceptor terminals were investigated by immunohistochemistry and electron microscopy. Results. The authors found loss of cone function and cone death accompanied by impairment of rods and rod-driven signaling in CNGA3−/− mice. Scotopic ERG b-wave amplitudes were reduced by 15% at 1 month, 30% at 6 months, and 40% at 9 months and older, while scotopic a-wave amplitudes were decreased by 20% at 9 months, compared with ERGs of age-matched wild-type mice. Outer nuclear layer thickness in CNGA3−/− retina was reduced by 15% at 12 months compared with age-matched wild-type controls. This was accompanied by a 30%–40% reduction in expression of rod-specific proteins, including rhodopsin, rod transducin α-subunit, and glutamic acid-rich protein (GARP). Cone terminals in the CNGA3−/− retina showed a progressive loss of neurochemical and ultrastructural integrity. Abnormalities were observed as early as 1 month. Disorganized rod terminal ultrastructure was noted by 12 months. Conclusions. These findings demonstrate secondary rod impairment and degeneration after cone degeneration in mice with cone CNG channel deficiency. Loss of cone phototransduction accompanies the compromised integrity of cone terminals. With time, rod synaptic structure, function, and viability also become compromised. PMID:22247469

The Role of Heparanase and Sulfatases in the Modification of Heparan Sulfate Proteoglycans within the Tumor Microenvironment and Opportunities for Novel Cancer Therapeutics

PubMed Central

Hammond, Edward; Khurana, Ashwani; Shridhar, Viji; Dredge, Keith

2014-01-01

Heparan sulfate proteoglycans (HSPGs) are an integral and dynamic part of normal tissue architecture at the cell surface and within the extracellular matrix. The modification of HSPGs in the tumor microenvironment is known to result not just in structural but also functional consequences, which significantly impact cancer progression. As substrates for the key enzymes sulfatases and heparanase, the modification of HSPGs is typically characterized by the degradation of heparan sulfate (HS) chains/sulfation patterns via the endo-6-O-sulfatases (Sulf1 and Sulf2) or by heparanase, an endo-glycosidase that cleaves the HS polymers releasing smaller fragments from HSPG complexes. Numerous studies have demonstrated how these enzymes actively influence cancer cell proliferation, signaling, invasion, and metastasis. The activity or expression of these enzymes has been reported to be modified in a variety of cancers. Such observations are consistent with the degradation of normal architecture and basement membranes, which are typically compromised in metastatic disease. Moreover, recent studies elucidating the requirements for these proteins in tumor initiation and progression exemplify their importance in the development and progression of cancer. Thus, as the influence of the tumor microenvironment in cancer progression becomes more apparent, the focus on targeting enzymes that degrade HSPGs highlights one approach to maintain normal tissue architecture, inhibit tumor progression, and block metastasis. This review discusses the role of these enzymes in the context of the tumor microenvironment and their promise as therapeutic targets for the treatment of cancer. PMID:25105093

Temporal and Spatial Effects of Blast Overpressure on Blood-Brain Barrier Permeability in Traumatic Brain Injury.

PubMed

Kuriakose, Matthew; Rama Rao, Kakulavarapu V; Younger, Daniel; Chandra, Namas

2018-06-06

Blast-induced traumatic brain injury (bTBI) is a "signature wound" in soldiers during training and in combat and has also become a major cause of morbidity in civilians due to increased insurgency. This work examines the role of blood-brain barrier (BBB) disruption as a result of both primary biomechanical and secondary biochemical injury mechanisms in bTBI. Extravasation of sodium fluorescein (NaF) and Evans blue (EB) tracers were used to demonstrate that compromise of the BBB occurs immediately following shock loading, increases in intensity up to 4 hours and returns back to normal in 24 hours. This BBB compromise occurs in multiple regions of the brain in the anterior-posterior direction of the shock wave, with maximum extravasation seen in the frontal cortex. Compromise of the BBB is confirmed by (a) extravasation of tracers into the brain, (b) quantification of tight-junction proteins (TJPs) in the brain and the blood, and (c) tracking specific blood-borne molecules into the brain and brain-specific proteins into the blood. Taken together, this work demonstrates that the BBB compromise occurs as a part of initial biomechanical loading and is a function of increasing blast overpressures.

Petri Net-Based Model of Helicobacter pylori Mediated Disruption of Tight Junction Proteins in Stomach Lining during Gastric Carcinoma

PubMed Central

Naz, Anam; Obaid, Ayesha; Awan, Faryal M.; Ikram, Aqsa; Ahmad, Jamil; Ali, Amjad

2017-01-01

Tight junctions help prevent the passage of digestive enzymes and microorganisms through the space between adjacent epithelial cells lining. However, Helicobacter pylori encoded virulence factors negatively regulate these tight junctions and contribute to dysfunction of gastric mucosa. Here, we have predicted the regulation of important tight junction proteins, such as Zonula occludens-1, Claudin-2 and Connexin32 in the presence of pathogenic proteins. Molecular events such as post translational modifications and crosstalk between phosphorylation, O-glycosylation, palmitoylation and methylation are explored which may compromise the integrity of these tight junction proteins. Furthermore, the signaling pathways disrupted by dysregulated kinases, proteins and post-translational modifications are reviewed to design an abstracted computational model showing the situation-dependent dynamic behaviors of these biological processes and entities. A qualitative hybrid Petri Net model is therefore constructed showing the altered host pathways in the presence of virulence factor cytotoxin-associated gene A, leading to the disruption of tight junction proteins. The model is qualitative logic-based, which does not depend on any kinetic parameter and quantitative data and depends on knowledge derived from experiments. The designed model provides insights into the tight junction disruption and disease progression. Model is then verified by the available experimental data, nevertheless formal in vitro experimentation is a promising way to ensure its validation. The major findings propose that H. pylori activated kinases are responsible to trigger specific post translational modifications within tight junction proteins, at specific sites. These modifications may favor alterations in gastric barrier and provide a route to bacterial invasion into host cells. PMID:28932213

Petri Net-Based Model of Helicobacter pylori Mediated Disruption of Tight Junction Proteins in Stomach Lining during Gastric Carcinoma.

PubMed

Naz, Anam; Obaid, Ayesha; Awan, Faryal M; Ikram, Aqsa; Ahmad, Jamil; Ali, Amjad

2017-01-01

Tight junctions help prevent the passage of digestive enzymes and microorganisms through the space between adjacent epithelial cells lining. However, Helicobacter pylori encoded virulence factors negatively regulate these tight junctions and contribute to dysfunction of gastric mucosa. Here, we have predicted the regulation of important tight junction proteins, such as Zonula occludens-1, Claudin-2 and Connexin32 in the presence of pathogenic proteins. Molecular events such as post translational modifications and crosstalk between phosphorylation, O-glycosylation, palmitoylation and methylation are explored which may compromise the integrity of these tight junction proteins. Furthermore, the signaling pathways disrupted by dysregulated kinases, proteins and post-translational modifications are reviewed to design an abstracted computational model showing the situation-dependent dynamic behaviors of these biological processes and entities. A qualitative hybrid Petri Net model is therefore constructed showing the altered host pathways in the presence of virulence factor cytotoxin-associated gene A, leading to the disruption of tight junction proteins. The model is qualitative logic-based, which does not depend on any kinetic parameter and quantitative data and depends on knowledge derived from experiments. The designed model provides insights into the tight junction disruption and disease progression. Model is then verified by the available experimental data, nevertheless formal in vitro experimentation is a promising way to ensure its validation. The major findings propose that H. pylori activated kinases are responsible to trigger specific post translational modifications within tight junction proteins, at specific sites. These modifications may favor alterations in gastric barrier and provide a route to bacterial invasion into host cells.

Compromised fidelity of endocytic synaptic vesicle protein sorting in the absence of stonin 2

PubMed Central

Kononenko, Natalia L.; Diril, M. Kasim; Puchkov, Dmytro; Kintscher, Michael; Koo, Seong Joo; Pfuhl, Gerit; Winter, York; Wienisch, Martin; Klingauf, Jürgen; Breustedt, Jörg; Schmitz, Dietmar; Maritzen, Tanja; Haucke, Volker

2013-01-01

Neurotransmission depends on the exocytic fusion of synaptic vesicles (SVs) and their subsequent reformation either by clathrin-mediated endocytosis or budding from bulk endosomes. How synapses are able to rapidly recycle SVs to maintain SV pool size, yet preserve their compositional identity, is poorly understood. We demonstrate that deletion of the endocytic adaptor stonin 2 (Stn2) in mice compromises the fidelity of SV protein sorting, whereas the apparent speed of SV retrieval is increased. Loss of Stn2 leads to selective missorting of synaptotagmin 1 to the neuronal surface, an elevated SV pool size, and accelerated SV protein endocytosis. The latter phenotype is mimicked by overexpression of endocytosis-defective variants of synaptotagmin 1. Increased speed of SV protein retrieval in the absence of Stn2 correlates with an up-regulation of SV reformation from bulk endosomes. Our results are consistent with a model whereby Stn2 is required to preserve SV protein composition but is dispensable for maintaining the speed of SV recycling. PMID:23345427

Protein catabolism in pregnant snakes (Epicrates cenchria maurus Boidae) compromises musculature and performance after reproduction.

PubMed

Lourdais, O; Brischoux, F; DeNardo, D; Shine, R

2004-07-01

In many species the high energetic demands of reproduction induce a negative energy balance, and thus females must rely on tissue catabolism to complete the reproductive process. Previous works have shown that both fat and protein are energy resources during prolonged fasting in vertebrates. While many ecological studies on energy costs of reproduction have focused on variations in fat stores, the impact of protein investment on the female has not been thoroughly investigated. Notably, as there is no specialized storage form for proteins, intense catabolism is likely to entail structural (musculature) loss that may compromise maternal physical performance after reproduction. Measurements on captive rainbow boas ( Epicrates cenchria maurus) confirm that reproducing females undergo significant protein catabolism (as indicated by elevated plasma uric acid levels) and show considerable musculature loss during gestation (as detected by reduced width of the epaxial muscles). Protein mobilization entailed a significant functional loss that was illustrated by decrements in tests of strength and constriction after parturition. In wild situations, such effects are likely to decrease the snakes' ability to forage and apprehend prey. Hence, the time period needed to recover from reproduction can be extended not only because the female must compensate losses of both fat stores and functional muscle, but also because the ability to do so may be compromised. Performance alteration is likely to be of equal or greater importance than reduced energy stores in the physiological mediation of elevated post-reproduction mortality rates and infrequent reproductive bouts (e.g. biannual or triannual), two common ecological traits of female snakes.

The endoplasmic reticulum stress response in aging and age-related diseases

PubMed Central

Brown, Marishka K.; Naidoo, Nirinjini

2012-01-01

The endoplasmic reticulum(ER) is a multifunctional organelle within which protein folding, lipid biosynthesis, and calcium storage occurs. Perturbations such as energy or nutrient depletion, disturbances in calcium or redox status that disrupt ER homeostasis lead to the misfolding of proteins, ER stress and up-regulation of several signaling pathways coordinately called the unfolded protein response (UPR). The UPR is characterized by the induction of chaperones, degradation of misfolded proteins and attenuation of protein translation. The UPR plays a fundamental role in the maintenance of cellular homeostasis and thus is central to normal physiology. However, sustained unresolved ER stress leads to apoptosis. Aging linked declines in expression and activity of key ER molecular chaperones and folding enzymes compromise proper protein folding and the adaptive response of the UPR. One mechanism to explain age associated declines in cellular functions and age-related diseases is a progressive failure of chaperoning systems. In many of these diseases, proteins or fragments of proteins convert from their normally soluble forms to insoluble fibrils or plaques that accumulate in a variety of organs including the liver, brain or spleen. This group of diseases, which typically occur late in life includes Alzheimer's, Parkinson's, type II diabetes and a host of less well known but often equally serious conditions such as fatal familial insomnia. The UPR is implicated in many of these neurodegenerative and familial protein folding diseases as well as several cancers and a host of inflammatory diseases including diabetes, atherosclerosis, inflammatory bowel disease and arthritis. This review will discuss age-related changes in the ER stress response and the role of the UPR in age-related diseases. PMID:22934019

Evolutionary Tradeoffs between Economy and Effectiveness in Biological Homeostasis Systems

PubMed Central

Szekely, Pablo; Sheftel, Hila; Mayo, Avi; Alon, Uri

2013-01-01

Biological regulatory systems face a fundamental tradeoff: they must be effective but at the same time also economical. For example, regulatory systems that are designed to repair damage must be effective in reducing damage, but economical in not making too many repair proteins because making excessive proteins carries a fitness cost to the cell, called protein burden. In order to see how biological systems compromise between the two tasks of effectiveness and economy, we applied an approach from economics and engineering called Pareto optimality. This approach allows calculating the best-compromise systems that optimally combine the two tasks. We used a simple and general model for regulation, known as integral feedback, and showed that best-compromise systems have particular combinations of biochemical parameters that control the response rate and basal level. We find that the optimal systems fall on a curve in parameter space. Due to this feature, even if one is able to measure only a small fraction of the system's parameters, one can infer the rest. We applied this approach to estimate parameters in three biological systems: response to heat shock and response to DNA damage in bacteria, and calcium homeostasis in mammals. PMID:23950698

Evolutionary tradeoffs between economy and effectiveness in biological homeostasis systems.

PubMed

Szekely, Pablo; Sheftel, Hila; Mayo, Avi; Alon, Uri

2013-01-01

Biological regulatory systems face a fundamental tradeoff: they must be effective but at the same time also economical. For example, regulatory systems that are designed to repair damage must be effective in reducing damage, but economical in not making too many repair proteins because making excessive proteins carries a fitness cost to the cell, called protein burden. In order to see how biological systems compromise between the two tasks of effectiveness and economy, we applied an approach from economics and engineering called Pareto optimality. This approach allows calculating the best-compromise systems that optimally combine the two tasks. We used a simple and general model for regulation, known as integral feedback, and showed that best-compromise systems have particular combinations of biochemical parameters that control the response rate and basal level. We find that the optimal systems fall on a curve in parameter space. Due to this feature, even if one is able to measure only a small fraction of the system's parameters, one can infer the rest. We applied this approach to estimate parameters in three biological systems: response to heat shock and response to DNA damage in bacteria, and calcium homeostasis in mammals.

Pharmacological modulation of dietary lipid-induced cerebral capillary dysfunction: Considerations for reducing risk for Alzheimer's disease.

PubMed

Pallebage-Gamarallage, Menuka; Takechi, Ryusuke; Lam, Virginie; Elahy, Mina; Mamo, John

2016-01-01

An increasing body of evidence suggests that cerebrovascular dysfunction and microvessel disease precede the evolution of hallmark pathological features that characterise Alzheimer's disease (AD), consistent with a causal association for onset or progression. Recent studies, principally in genetically unmanipulated animal models, suggest that chronic ingestion of diets enriched in saturated fats and cholesterol may compromise blood-brain barrier (BBB) integrity resulting in inappropriate blood-to-brain extravasation of plasma proteins, including lipid macromolecules that may be enriched in amyloid-β (Aβ). Brain parenchymal retention of blood proteins and lipoprotein bound Aβ is associated with heightened neurovascular inflammation, altered redox homeostasis and nitric oxide (NO) metabolism. Therefore, it is a reasonable proposition that lipid-lowering agents may positively modulate BBB integrity and by extension attenuate risk or progression of AD. In addition to their robust lipid lowering properties, reported beneficial effects of lipid-lowering agents were attributed to their pleiotropic properties via modulation of inflammation, oxidative stress, NO and Aβ metabolism. The review is a contemporary consideration of a complex body of literature intended to synthesise focussed consideration of mechanisms central to regulation of BBB function and integrity. Emphasis is given to dietary fat driven significant epidemiological evidence consistent with heightened risk amongst populations consuming greater amounts of saturated fats and cholesterol. In addition, potential neurovascular benefits associated with the use of hypolipidemic statins, probucol and fenofibrate are also presented in the context of lipid-lowering and pleiotropic properties.

Rationally engineered Troponin C modulates in vivo cardiac function and performance in health and disease.

PubMed

Shettigar, Vikram; Zhang, Bo; Little, Sean C; Salhi, Hussam E; Hansen, Brian J; Li, Ning; Zhang, Jianchao; Roof, Steve R; Ho, Hsiang-Ting; Brunello, Lucia; Lerch, Jessica K; Weisleder, Noah; Fedorov, Vadim V; Accornero, Federica; Rafael-Fortney, Jill A; Gyorke, Sandor; Janssen, Paul M L; Biesiadecki, Brandon J; Ziolo, Mark T; Davis, Jonathan P

2016-02-24

Treatment for heart disease, the leading cause of death in the world, has progressed little for several decades. Here we develop a protein engineering approach to directly tune in vivo cardiac contractility by tailoring the ability of the heart to respond to the Ca(2+) signal. Promisingly, our smartly formulated Ca(2+)-sensitizing TnC (L48Q) enhances heart function without any adverse effects that are commonly observed with positive inotropes. In a myocardial infarction (MI) model of heart failure, expression of TnC L48Q before the MI preserves cardiac function and performance. Moreover, expression of TnC L48Q after the MI therapeutically enhances cardiac function and performance, without compromising survival. We demonstrate engineering TnC can specifically and precisely modulate cardiac contractility that when combined with gene therapy can be employed as a therapeutic strategy for heart disease.

Rationally engineered Troponin C modulates in vivo cardiac function and performance in health and disease

PubMed Central

Shettigar, Vikram; Zhang, Bo; Little, Sean C.; Salhi, Hussam E.; Hansen, Brian J.; Li, Ning; Zhang, Jianchao; Roof, Steve R.; Ho, Hsiang-Ting; Brunello, Lucia; Lerch, Jessica K.; Weisleder, Noah; Fedorov, Vadim V.; Accornero, Federica; Rafael-Fortney, Jill A.; Gyorke, Sandor; Janssen, Paul M. L.; Biesiadecki, Brandon J.; Ziolo, Mark T.; Davis, Jonathan P.

2016-01-01

Treatment for heart disease, the leading cause of death in the world, has progressed little for several decades. Here we develop a protein engineering approach to directly tune in vivo cardiac contractility by tailoring the ability of the heart to respond to the Ca2+ signal. Promisingly, our smartly formulated Ca2+-sensitizing TnC (L48Q) enhances heart function without any adverse effects that are commonly observed with positive inotropes. In a myocardial infarction (MI) model of heart failure, expression of TnC L48Q before the MI preserves cardiac function and performance. Moreover, expression of TnC L48Q after the MI therapeutically enhances cardiac function and performance, without compromising survival. We demonstrate engineering TnC can specifically and precisely modulate cardiac contractility that when combined with gene therapy can be employed as a therapeutic strategy for heart disease. PMID:26908229

Medical Management of Endometriosis: Emerging Evidence Linking Inflammation to Disease Pathophysiology

PubMed Central

Bruner-Tran, Kaylon L.; Herington, Jennifer L.; Duleba, Antoni J.; Taylor, Hugh S.; Osteen, Kevin G.

2013-01-01

Progesterone action normally mediates the balance between anti-inflammatory and pro-inflammatory processes throughout the female reproductive tract. However, in women with endometriosis, endometrial progesterone resistance, characterized by alterations in progesterone responsive gene and protein expression, is now considered a central element in disease pathophysiology. Recent studies additionally suggest that the peritoneal microenvironment of endometriosis patients exhibits altered physiological characteristics that may further promote inflammation-driven disease development and progression. Within this review, we summarize our current understanding of the pathogenesis of endometriosis with an emphasis on the role that inflammation plays in generating not only the progesterone-resistant eutopic endometrium but also a peritoneal microenvironment that may contribute significantly to disease establishment. Viewing endometriosis from the emerging perspective that a progesterone resistant endometrium and an immunologically compromised peritoneal microenvironment are biologically linked risk factors for disease development provides a novel mechanistic framework to identify new therapeutic targets for appropriate medical management. PMID:23598784

A novel NDUFS4 frameshift mutation causes Leigh disease in the Hutterite population.

PubMed

Lamont, Ryan E; Beaulieu, Chandree L; Bernier, Francois P; Sparkes, Rebecca; Innes, A Micheil; Jackel-Cram, Candice; Ober, Carole; Parboosingh, Jillian S; Lemire, Edmond G

2017-03-01

Leigh disease is a progressive, infantile-onset, neurodegenerative disorder characterized by feeding difficulties, failure to thrive, hypotonia, seizures, and central respiratory compromise. Metabolic and neuroimaging investigations typically identify abnormalities consistent with a disorder of mitochondrial energy metabolism. Mutations in more than 35 genes affecting the mitochondrial respiratory chain encoded from both the nuclear and mitochondrial genomes have been associated with Leigh disease. The clinical presentations of five individuals of Hutterite descent with Leigh disease are described herein. An identity-by-descent mapping and candidate gene approach was used to identify a novel homozygous c.393dupA frameshift mutation in the NADH dehydrogenase (ubiquinone) Fe-S protein 4 (NDUFS4) gene. The carrier frequency of this mutation was estimated in >1,300 Hutterite individuals to be 1 in 27. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Rett syndrome and MeCP2.

PubMed

Liyanage, Vichithra R B; Rastegar, Mojgan

2014-06-01

Rett syndrome (RTT) is a severe and progressive neurological disorder, which mainly affects young females. Mutations of the methyl-CpG binding protein 2 (MECP2) gene are the most prevalent cause of classical RTT cases. MECP2 mutations or altered expression are also associated with a spectrum of neurodevelopmental disorders such as autism spectrum disorders with recent links to fetal alcohol spectrum disorders. Collectively, MeCP2 relation to these neurodevelopmental disorders highlights the importance of understanding the molecular mechanisms by which MeCP2 impacts brain development, mental conditions, and compromised brain function. Since MECP2 mutations were discovered to be the primary cause of RTT, a significant progress has been made in the MeCP2 research, with respect to the expression, function and regulation of MeCP2 in the brain and its contribution in RTT pathogenesis. To date, there have been intensive efforts in designing effective therapeutic strategies for RTT benefiting from mouse models and cells collected from RTT patients. Despite significant progress in MeCP2 research over the last few decades, there is still a knowledge gap between the in vitro and in vivo research findings and translating these findings into effective therapeutic interventions in human RTT patients. In this review, we will provide a synopsis of Rett syndrome as a severe neurological disorder and will discuss the role of MeCP2 in RTT pathophysiology.

Morphine induces albuminuria by compromising podocyte integrity.

PubMed

Lan, Xiqian; Rai, Partab; Chandel, Nirupama; Cheng, Kang; Lederman, Rivka; Saleem, Moin A; Mathieson, Peter W; Husain, Mohammad; Crosson, John T; Gupta, Kalpna; Malhotra, Ashwani; Singhal, Pravin C

2013-01-01

Morphine has been reported to accelerate the progression of chronic kidney disease. However, whether morphine affects slit diaphragm (SD), the major constituent of glomerular filtration barrier, is still unclear. In the present study, we examined the effect of morphine on glomerular filtration barrier in general and podocyte integrity in particular. Mice were administered either normal saline or morphine for 72 h, then urine samples were collected and kidneys were subsequently isolated for immunohistochemical studies and Western blot. For in vitro studies, human podocytes were treated with morphine and then probed for the molecular markers of slit diaphragm. Morphine-receiving mice displayed a significant increase in albuminuria and showed effacement of podocyte foot processes. In both in vivo and in vitro studies, the expression of synaptopodin, a molecular marker for podocyte integrity, and the slit diaphragm constituting molecules (SDCM), such as nephrin, podocin, and CD2-associated protein (CD2AP), were decreased in morphine-treated podocytes. In vitro studies indicated that morphine modulated podocyte expression of SDCM through opiate mu (MOR) and kappa (KOR) receptors. Since morphine also enhanced podocyte oxidative stress, the latter seems to contribute to decreased SDCM expression. In addition, AKT, p38, and JNK pathways were involved in morphine-induced down regulation of SDCM in human podocytes. These findings demonstrate that morphine has the potential to alter the glomerular filtration barrier by compromising the integrity of podocytes.

Nutrition support of the pediatric patient with AIDS.

PubMed

Bentler, M; Stanish, M

1987-04-01

Maintaining optimal nutrition in the pediatric patient with Acquired Immune Deficiency Syndrome (AIDS) is challenging, but it may be one of the most effective therapies. Patients experience numerous complications that compromise nutritional status. Infection, fever, diarrhea, feeding problems, and decreased intake all contribute to malnutrition, which in turn predisposes the patient even more to infection and malabsorption. Nutrition assessment should be done routinely so that new problems may be identified and treated. High-calorie, high-protein feedings, vitamin supplementation, and, when necessary, gavage feedings or parenteral nutrition are recommended to improve nutritional status and prevent further deficits. Maintaining optimal nutrition in the pediatric patient with Acquired Immune Deficiency Syndrome (AIDS) poses a significant challenge to the health care team. Patients may experience numerous complications that compromise nutritional status. The patient is at high risk for opportunistic infections, especially of the lungs, central nervous system, gastrointestinal (GI) tract, and skin. Such infections are common causes of morbidity and mortality. Impaired nutritional status may further impair the patient's immunocompetence. A study by Kotler and Gaety demonstrated severe progressive malnutrition in adult AIDS patients, with the lowest measures of lean body mass occurring in those patients close to death at the time of the study. While no studies of children with AIDS have been done to date, we have subjectively observed feeding problems, weight loss, and malnutrition in most of the patients we have seen.

Nrf2 Deficiency Exacerbates Obesity-Induced Oxidative Stress, Neurovascular Dysfunction, Blood-Brain Barrier Disruption, Neuroinflammation, Amyloidogenic Gene Expression, and Cognitive Decline in Mice, Mimicking the Aging Phenotype.

PubMed

Tarantini, Stefano; Valcarcel-Ares, M Noa; Yabluchanskiy, Andriy; Tucsek, Zsuzsanna; Hertelendy, Peter; Kiss, Tamas; Gautam, Tripti; Zhang, Xin A; Sonntag, William E; de Cabo, Rafael; Farkas, Eszter; Elliott, Michael H; Kinter, Michael T; Deak, Ferenc; Ungvari, Zoltan; Csiszar, Anna

2018-06-14

Obesity has deleterious effects on cognitive function in the elderly adults. In mice, aging exacerbates obesity-induced oxidative stress, microvascular dysfunction, blood-brain barrier (BBB) disruption, and neuroinflammation, which compromise cognitive health. However, the specific mechanisms through which aging and obesity interact to remain elusive. Previously, we have shown that Nrf2 signaling plays a critical role in microvascular resilience to obesity and that aging is associated with progressive Nrf2 dysfunction, promoting microvascular impairment. To test the hypothesis that Nrf2 deficiency exacerbates cerebromicrovascular dysfunction induced by obesity Nrf2+/+ and Nrf2-/-, mice were fed an adipogenic high-fat diet (HFD). Nrf2 deficiency significantly exacerbated HFD-induced oxidative stress and cellular senescence, impairment of neurovascular coupling responses, BBB disruption, and microglia activation, mimicking the aging phenotype. Obesity in Nrf2-/- mice elicited complex alterations in the amyloidogenic gene expression profile, including upregulation of amyloid precursor protein. Nrf2 deficiency and obesity additively reduced long-term potentiation in the CA1 area of the hippocampus. Collectively, Nrf2 dysfunction exacerbates the deleterious effects of obesity, compromising cerebromicrovascular and brain health by impairing neurovascular coupling mechanisms, BBB integrity and synaptic function and promoting neuroinflammation. These results support a possible role for age-related Nrf2 dysfunction in the pathogenesis of vascular cognitive impairment and Alzheimer's disease.

I-DIRT, a general method for distinguishing between specific and nonspecific protein interactions.

PubMed

Tackett, Alan J; DeGrasse, Jeffrey A; Sekedat, Matthew D; Oeffinger, Marlene; Rout, Michael P; Chait, Brian T

2005-01-01

Isolation of protein complexes via affinity-tagged proteins provides a powerful tool for studying biological systems, but the technique is often compromised by co-enrichment of nonspecifically interacting proteins. We describe a new technique (I-DIRT) that distinguishes contaminants from bona fide interactors in immunopurifications, overcoming this most challenging problem in defining protein complexes. I-DIRT will be of broad value for studying protein complexes in biological systems that can be metabolically labeled.

Canada's experience in applying C&I to measure progress towards SFM - perspectives from the National, Regional and local levels

Treesearch

John E. Hall; S. R. J. Bridge; Brian D. Haddon

2006-01-01

This paper will provide perspectives of Canada’s experiences in applying Criteria and Indicators (C&I) to measure progress towards Sustainable Forest Management (SFM) at the National, Regional (Provincial) and local levels. SFM is rooted in Bruntland’s concept of Sustainable Development and is about providing for present forest-based needs without compromising...

Efficacy of whey protein supplementation on resistance exercise-induced changes in muscle strength, lean mass, and function in mobility-limited older adults

USDA-ARS?s Scientific Manuscript database

Whey protein supplementation may augment resistance exercise-induced increases in muscle strength and mass. Further studies are required to determine whether this effect extends to functionally compromised older adults. The objectives of the study were to compare the effects of whey protein concent...

The Childhood Obesity Epidemic As a Result of Non-Genetic Evolution: the Maternal Resources Hypothesis

PubMed Central

Archer, Edward

2014-01-01

Over the past century, socio-environmental evolution (e.g., reduced pathogenic load, decreased physical activity [PA], improved nutrition) led to cumulative increments in maternal energy resources (i.e., body mass, adiposity) and decrements in energy expenditure and metabolic control. These decrements reduced the competition between maternal and fetal energy demands and increased the availability of energy substrates to the intrauterine milieu. This perturbation of mother-conceptus energy partitioning stimulated fetal pancreatic beta-cell and adipocyte hyperplasia, thereby inducing an enduring competitive advantage of adipocytes over other tissues in the acquisition and sequestering of nutrient-energy via intensified insulin secretion and hyperplastic adiposity. At menarche, the competitive dominance of adipocytes was further amplified via hormone-induced adipocyte hyperplasia and weight-induced decrements in PA. These metabolic and behavioral effects were propagated progressively when obese, inactive, metabolically compromised women produced progressively larger, more inactive and metabolically compromised children. Consequently, the evolution of human energy metabolism was significantly altered. This phenotypic evolution was exacerbated by increments in the use of Caesarian sections that allowed both the larger fetuses and the metabolically compromised mothers who produced them to survive and reproduce. Thus, natural selection was iatrogenically rendered artificial selection, and the frequency of obese, inactive, metabolically compromised phenotypes increased in the global population. By the late 20th century, a metabolic tipping point was reached in which the post-prandial insulin response was so intense, the relative number of adipocytes so magnified, and inactivity so pervasive that the competitive dominance of adipocytes in the sequestering of nutrient-energy was inevitable, and obesity was unavoidable. PMID:25440888

An Amphipathic Helix Directs Cellular Membrane Curvature Sensing and Function of the BAR Domain Protein PICK1.

PubMed

Herlo, Rasmus; Lund, Viktor K; Lycas, Matthew D; Jansen, Anna M; Khelashvili, George; Andersen, Rita C; Bhatia, Vikram; Pedersen, Thomas S; Albornoz, Pedro B C; Johner, Niklaus; Ammendrup-Johnsen, Ina; Christensen, Nikolaj R; Erlendsson, Simon; Stoklund, Mikkel; Larsen, Jannik B; Weinstein, Harel; Kjærulff, Ole; Stamou, Dimitrios; Gether, Ulrik; Madsen, Kenneth L

2018-05-15

BAR domains are dimeric protein modules that sense, induce, and stabilize lipid membrane curvature. Here, we show that membrane curvature sensing (MCS) directs cellular localization and function of the BAR domain protein PICK1. In PICK1, and the homologous proteins ICA69 and arfaptin2, we identify an amphipathic helix N-terminal to the BAR domain that mediates MCS. Mutational disruption of the helix in PICK1 impaired MCS without affecting membrane binding per se. In insulin-producing INS-1E cells, super-resolution microscopy revealed that disruption of the helix selectively compromised PICK1 density on insulin granules of high curvature during their maturation. This was accompanied by reduced hormone storage in the INS-1E cells. In Drosophila, disruption of the helix compromised growth regulation. By demonstrating size-dependent binding on insulin granules, our finding highlights the function of MCS for BAR domain proteins in a biological context distinct from their function, e.g., at the plasma membrane during endocytosis. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

Suboptimal T-cell receptor signaling compromises protein translation, ribosome biogenesis, and proliferation of mouse CD8 T cells.

PubMed

Tan, Thomas C J; Knight, John; Sbarrato, Thomas; Dudek, Kate; Willis, Anne E; Zamoyska, Rose

2017-07-25

Global transcriptomic and proteomic analyses of T cells have been rich sources of unbiased data for understanding T-cell activation. Lack of full concordance of these datasets has illustrated that important facets of T-cell activation are controlled at the level of translation. We undertook translatome analysis of CD8 T-cell activation, combining polysome profiling and microarray analysis. We revealed that altering T-cell receptor stimulation influenced recruitment of mRNAs to heavy polysomes and translation of subsets of genes. A major pathway that was compromised, when TCR signaling was suboptimal, was linked to ribosome biogenesis, a rate-limiting factor in both cell growth and proliferation. Defective TCR signaling affected transcription and processing of ribosomal RNA precursors, as well as the translation of specific ribosomal proteins and translation factors. Mechanistically, IL-2 production was compromised in weakly stimulated T cells, affecting the abundance of Myc protein, a known regulator of ribosome biogenesis. Consequently, weakly activated T cells showed impaired production of ribosomes and a failure to maintain proliferative capacity after stimulation. We demonstrate that primary T cells respond to various environmental cues by regulating ribosome biogenesis and mRNA translation at multiple levels to sustain proliferation and differentiation.

Axonal degeneration in Alzheimer’s disease: When signaling abnormalities meet the axonal transport system

PubMed Central

Kanaan, Nicholas M.; Pigino, Gustavo F.; Brady, Scott T.; Lazarov, Orly; Binder, Lester I.; Morfini, Gerardo A.

2012-01-01

Alzheimer’s disease (AD) is characterized by progressive, age-dependent degeneration of neurons in the central nervous system. A large body of evidence indicates that neurons affected in AD follow a dying-back pattern of degeneration, where abnormalities in synaptic function and axonal connectivity long precede somatic cell death. Mechanisms underlying dying-back degeneration of neurons in AD remain elusive but several have been proposed, including deficits in fast axonal transport (FAT). Accordingly, genetic evidence linked alterations in FAT to dying-back degeneration of neurons, and FAT defects have been widely documented in various AD models. In light of these findings, we discuss experimental evidence linking several AD-related pathogenic polypeptides to aberrant activation of signaling pathways involved in the phosphoregulation of microtubule-based motor proteins. While each pathway appears to affect FAT in a unique manner, in the context of AD, many of these pathways might work synergistically to compromise the delivery of molecular components critical for the maintenance and function of synapses and axons. Therapeutic approaches aimed at preventing FAT deficits by normalizing the activity of specific protein kinases may help prevent degeneration of vulnerable neurons in AD. PMID:22721767

New Perspectives on Osteogenesis Imperfecta

PubMed Central

Forlino, Antonella; Cabral, Wayne A.; Barnes, Aileen M.; Marini, Joan C.

2012-01-01

A new paradigm has emerged for osteogenesis imperfecta (OI) as a collagen-related disorder. The more prevalent autosomal dominant forms of OI are caused by primary defects in type I collagen, while autosomal recessive forms are caused by deficiency of proteins which interact with type I procollagen for post-translational modification and/or folding. Factors contributing to the mechanism of dominant OI include intracellular stress, disruption of interactions between collagen and non-collagenous proteins, compromised matrix structure, abnormal cell-cell and cell-matrix interactions and tissue mineralization. Recessive OI is caused by deficiency of any of the three components of the collagen prolyl 3-hydroxylation complex; absence of 3-hydroxylation is associated with increased modification of the collagen helix, supporting delayed collagen folding. Other causes of recessive OI include deficiency of collagen chaperones, FKBP65 or HSP47. Murine models are crucial to uncovering the common pathways in dominant and recessive OI bone dysplasia. Clinical management of OI is multidiscipinary, encompassing substantial progress in physical rehabilitation and surgical procedures, managment of hearing, dental and pulmonary abnormalities, as well as drugs such as bisphosphonates and rGH. Novel treatments using cell therapy or new drug regimens hold promise for the future. PMID:21670757

A mutation in MT-TW causes a tRNA processing defect and reduced mitochondrial function in a family with Leigh syndrome.

PubMed

Duff, Rachael M; Shearwood, Anne-Marie J; Ermer, Judith; Rossetti, Giulia; Gooding, Rebecca; Richman, Tara R; Balasubramaniam, Shanti; Thorburn, David R; Rackham, Oliver; Lamont, Phillipa J; Filipovska, Aleksandra

2015-11-01

Leigh syndrome (LS) is a progressive mitochondrial neurodegenerative disorder, whose symptoms most commonly include psychomotor delay with regression, lactic acidosis and a failure to thrive. Here we describe three siblings with LS, but with additional manifestations including hypertrophic cardiomyopathy, hepatosplenomegaly, cholestatic hepatitis, and seizures. All three affected siblings were found to be homoplasmic for an m. 5559A>G mutation in the T stem of the mitochondrial DNA-encoded MT-TW by next generation sequencing. The m.5559A>G mutation causes a reduction in the steady state levels of tRNA(Trp) and this decrease likely affects the stability of other mitochondrial RNAs in the patient fibroblasts. We observe accumulation of an unprocessed transcript containing tRNA(Trp), decreased de novo protein synthesis and consequently lowered steady state levels of mitochondrial DNA-encoded proteins that compromise mitochondrial respiration. Our results show that the m.5559A>G mutation at homoplasmic levels causes LS in association with severe multi-organ disease (LS-plus) as a consequence of dysfunctional mitochondrial RNA metabolism. Copyright © 2015 Elsevier B.V. and Mitochondria Research Society. All rights reserved.

The greenhouse stakes of globalization

EPA Science Inventory

As 2012 approaches, twenty-five years have passed since the Brundtland report defining sustainable development as a progress that meets the needs of the present without compromising the ability of future generations to meet their own needs (United Nations). Currently, the fight a...

Coping with uncertainty: nutrient deficiencies motivate insect migration at a cost to immunity.

PubMed

Srygley, Robert B; Lorch, Patrick D

2013-12-01

Migration often is associated with movement away from areas with depleted nutrients or other resources, and yet migration itself is energetically demanding. Migrating Mormon crickets Anabrus simplex (Orthoptera: Tettigoniidae) lack nutrients, and supplementation of deficient nutrients slows migratory movements and enhances specific aspects of their immune systems. Migrants deficient in proteins have less spontaneous phenoloxidase (PO) activity, whereas those deficient in carbohydrates have lower lysozyme-like anti-bacterial titers with a proposed compromise between migratory and anti-bacterial activities. To investigate the relationship between diet, movement, and immunity further, we removed Mormon crickets from a migratory band and offered each cricket one of five diets: high protein, high carbohydrate, equal weight of proteins and carbohydrates (P + C), vitamins only, or water only for 1 h. We then attached a radio, returned each to the migratory band, and recaptured them 18-24 h later. Mormon crickets fed protein moved the furthest, those with only water or only vitamins moved less, and those fed carbohydrates or P + C moved the least. Standard intake trials also indicated that the Mormon crickets were deficient in carbohydrates. Consistent with a previous study, lysozyme-like anti-bacterial activity was greatest in those fed carbohydrates, and there was no difference between those fed water, protein, or P + C. Crickets were removed from the same migratory band and fed one of four diets: high P, high C, P + C, or vitamins only, for 1 h. Then the crickets were held in captivity with water only for 4 or 24 h before blood was drawn. Immunity measures did not differ between times of draw. Diet treatments had no effect on anti-bacterial activity of captive Mormon crickets, whereas total PO was greater in those fed protein. These results support the hypothesis of a direct compromise between migratory and anti-bacterial activities, whereas PO is compromised by low protein independent of migratory activities. We discuss the potential effects of climate on nutritional deficits and susceptibility to different pathogens.

Meiosis evolves: adaptation to external and internal environments.

PubMed

Bomblies, Kirsten; Higgins, James D; Yant, Levi

2015-10-01

306 I. 306 II. 307 III. 312 IV. 317 V. 318 319 References 319 SUMMARY: Meiosis is essential for the fertility of most eukaryotes and its structures and progression are conserved across kingdoms. Yet many of its core proteins show evidence of rapid or adaptive evolution. What drives the evolution of meiosis proteins? How can constrained meiotic processes be modified in response to challenges without compromising their essential functions? In surveying the literature, we found evidence of two especially potent challenges to meiotic chromosome segregation that probably necessitate adaptive evolutionary responses: whole-genome duplication and abiotic environment, especially temperature. Evolutionary solutions to both kinds of challenge are likely to involve modification of homologous recombination and synapsis, probably via adjustments of core structural components important in meiosis I. Synthesizing these findings with broader patterns of meiosis gene evolution suggests that the structural components of meiosis coevolve as adaptive modules that may change in primary sequence and function while maintaining three-dimensional structures and protein interactions. The often sharp divergence of these genes among species probably reflects periodic modification of entire multiprotein complexes driven by genomic or environmental changes. We suggest that the pressures that cause meiosis to evolve to maintain fertility may cause pleiotropic alterations of global crossover rates. We highlight several important areas for future research. © 2015 The Authors. New Phytologist © 2015 New Phytologist Trust.

Low-density lipoprotein receptor-related protein 1: a physiological Aβ homeostatic mechanism with multiple therapeutic opportunities

PubMed Central

Sagare, Abhay P.; Deane, Rashid; Zlokovic, Berislav V.

2012-01-01

Low-density lipoprotein receptor-related protein-1 (LRP1) is the main cell surface receptor involved in brain and systemic clearance of the Alzheimer's disease (AD) toxin amyloid-beta (Aβ). In plasma, a soluble form of LRP1 (sLRP1) is the major transport protein for peripheral Aβ. LRP1 in brain endothelium and mural cells mediates Aβ efflux from brain by providing a transport mechanism for A across the blood-brain barrier (BBB). sLRP1 maintains a plasma ‘sink’ activity for Aβ through binding of peripheral Aβ which in turn inhibits re-entry of free plasma Aβ into the brain. LRP1 in the liver mediates systemic clearance of Aβ. In AD, LRP1 expression at the BBB is reduced and Aβ binding to circulating sLRP1 is compromised by oxidation. Cell surface LRP1 and circulating sLRP1 represent druggable targets which can be therapeutically modified to restore the physiological mechanisms of brain Aβ homeostasis. In this review, we discuss how increasing LRP1 expression at the BBB and liver with lifestyle changes, statins, plant-based active principles and/or gene therapy on one hand, and how replacing dysfunctional plasma sLRP1 on the other regulate Aβ clearance from brain ultimately controlling the onset and/or progression of AD. PMID:22820095

A Founder Mutation in VPS11 Causes an Autosomal Recessive Leukoencephalopathy Linked to Autophagic Defects.

PubMed

Zhang, Jinglan; Lachance, Véronik; Schaffner, Adam; Li, Xianting; Fedick, Anastasia; Kaye, Lauren E; Liao, Jun; Rosenfeld, Jill; Yachelevich, Naomi; Chu, Mary-Lynn; Mitchell, Wendy G; Boles, Richard G; Moran, Ellen; Tokita, Mari; Gorman, Elizabeth; Bagley, Kaytee; Zhang, Wei; Xia, Fan; Leduc, Magalie; Yang, Yaping; Eng, Christine; Wong, Lee-Jun; Schiffmann, Raphael; Diaz, George A; Kornreich, Ruth; Thummel, Ryan; Wasserstein, Melissa; Yue, Zhenyu; Edelmann, Lisa

2016-04-01

Genetic leukoencephalopathies (gLEs) are a group of heterogeneous disorders with white matter abnormalities affecting the central nervous system (CNS). The causative mutation in ~50% of gLEs is unknown. Using whole exome sequencing (WES), we identified homozygosity for a missense variant, VPS11: c.2536T>G (p.C846G), as the genetic cause of a leukoencephalopathy syndrome in five individuals from three unrelated Ashkenazi Jewish (AJ) families. All five patients exhibited highly concordant disease progression characterized by infantile onset leukoencephalopathy with brain white matter abnormalities, severe motor impairment, cortical blindness, intellectual disability, and seizures. The carrier frequency of the VPS11: c.2536T>G variant is 1:250 in the AJ population (n = 2,026). VPS11 protein is a core component of HOPS (homotypic fusion and protein sorting) and CORVET (class C core vacuole/endosome tethering) protein complexes involved in membrane trafficking and fusion of the lysosomes and endosomes. The cysteine 846 resides in an evolutionarily conserved cysteine-rich RING-H2 domain in carboxyl terminal regions of VPS11 proteins. Our data shows that the C846G mutation causes aberrant ubiquitination and accelerated turnover of VPS11 protein as well as compromised VPS11-VPS18 complex assembly, suggesting a loss of function in the mutant protein. Reduced VPS11 expression leads to an impaired autophagic activity in human cells. Importantly, zebrafish harboring a vps11 mutation with truncated RING-H2 domain demonstrated a significant reduction in CNS myelination following extensive neuronal death in the hindbrain and midbrain. Thus, our study reveals a defect in VPS11 as the underlying etiology for an autosomal recessive leukoencephalopathy disorder associated with a dysfunctional autophagy-lysosome trafficking pathway.

The childhood obesity epidemic as a result of nongenetic evolution: the maternal resources hypothesis.

PubMed

Archer, Edward

2015-01-01

Over the past century, socioenvironmental evolution (eg, reduced pathogenic load, decreased physical activity, and improved nutrition) led to cumulative increments in maternal energy resources (ie, body mass and adiposity) and decrements in energy expenditure and metabolic control. These decrements reduced the competition between maternal and fetal energy demands and increased the availability of energy substrates to the intrauterine milieu. This perturbation of mother-conceptus energy partitioning stimulated fetal pancreatic β-cell and adipocyte hyperplasia, thereby inducing an enduring competitive dominance of adipocytes over other tissues in the acquisition and sequestering of nutrient energy via intensified insulin secretion and hyperplastic adiposity. At menarche, the competitive dominance of adipocytes was further amplified via hormone-induced adipocyte hyperplasia and weight-induced decrements in physical activity. These metabolic and behavioral effects were propagated progressively when obese, inactive, metabolically compromised women produced progressively larger, more inactive, metabolically compromised children. Consequently, the evolution of human energy metabolism was markedly altered. This phenotypic evolution was exacerbated by increments in the use of cesarean sections, which allowed both the larger fetuses and the metabolically compromised mothers who produced them to survive and reproduce. Thus, natural selection was iatrogenically rendered artificial selection, and the frequency of obese, inactive, metabolically compromised phenotypes increased in the global population. By the late 20th century, a metabolic tipping point was reached at which the postprandial insulin response was so intense, the relative number of adipocytes so large, and inactivity so pervasive that the competitive dominance of adipocytes in the sequestering of nutrient energy was inevitable and obesity was unavoidable. Copyright © 2015 Mayo Foundation for Medical Education and Research. Published by Elsevier Inc. All rights reserved.

Advances in genetic therapeutic strategies for Duchenne muscular dystrophy.

PubMed

Guiraud, Simon; Chen, Huijia; Burns, David T; Davies, Kay E

2015-12-01

What is the topic of this review? This review highlights recent progress in genetically based therapies targeting the primary defect of Duchenne muscular dystrophy. What advances does it highlight? Over the last two decades, considerable progress has been made in understanding the mechanisms underlying Duchenne muscular dystrophy, leading to the development of genetic therapies. These include manipulation of the expression of the gene or related genes, the splicing of the gene and its translation, and replacement of the gene using viral approaches. Duchenne muscular dystrophy is a lethal X-linked disorder caused by mutations in the dystrophin gene. In the absence of the dystrophin protein, the link between the cytoskeleton and extracellular matrix is destroyed, and this severely compromises the strength, flexibility and stability of muscle fibres. The devastating consequence is progressive muscle wasting and premature death in Duchenne muscular dystrophy patients. There is currently no cure, and despite exhaustive palliative care, patients are restricted to a wheelchair by the age of 12 years and usually succumb to cardiac or respiratory complications in their late 20s. This review provides an update on the current genetically based therapies and clinical trials that target or compensate for the primary defect of this disease. These include dystrophin gene-replacement strategies, genetic modification techniques to restore dystrophin expression, and modulation of the dystrophin homologue, utrophin, as a surrogate to re-establish muscle function. © 2015 The Authors. Experimental Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.

Silencing of the Rice Gene LRR1 Compromises Rice Xa21 Transcript Accumulation and XA21-Mediated Immunity.

PubMed

Caddell, Daniel F; Park, Chang-Jin; Thomas, Nicholas C; Canlas, Patrick E; Ronald, Pamela C

2017-12-01

The rice immune receptor XA21 confers resistance to Xanthomonas oryzae pv. oryzae (Xoo), the causal agent of bacterial leaf blight. We previously demonstrated that an auxilin-like protein, XA21 BINDING PROTEIN 21 (XB21), positively regulates resistance to Xoo. To further investigate the function of XB21, we performed a yeast two-hybrid screen. We identified 22 unique XB21 interacting proteins, including LEUCINE-RICH REPEAT PROTEIN 1 (LRR1), which we selected for further analysis. Silencing of LRR1 in the XA21 genetic background (XA21-LRR1Ri) compromises resistance to Xoo compared with control XA21 plants. XA21-LRR1Ri plants have reduced Xa21 transcript levels and reduced expression of genes that serve as markers of XA21-mediated activation. Overexpression of LRR1 is insufficient to alter resistance to Xoo in rice lines lacking XA21. Taken together, our results indicate that LRR1 is required for wild-type Xa21 transcript expression and XA21-mediated immunity.

Triage of oxidation-prone proteins by Sqstm1/p62 within the mitochondria

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

Lee, Minjung; Shin, Jaekyoon, E-mail: jkshin@med.skku.ac.kr

2011-09-16

Highlights: {yields} The mitochondrion contains its own protein quality control system. {yields} p62 localizes within the mitochondria and forms mega-dalton sized complexes. {yields} p62 interacts with oxidation-prone proteins and the proteins of quality control. {yields} In vitro delivery of p62 improves mitochondrial functions. {yields} p62 is implicated as a participant in mitochondrial protein quality control. -- Abstract: As the mitochondrion is vulnerable to oxidative stress, cells have evolved several strategies to maintain mitochondrial integrity, including mitochondrial protein quality control mechanisms and autophagic removal of damaged mitochondria. Involvement of an autophagy adaptor, Sqstm1/p62, in the latter process has been recently described.more » In the present study, we provide evidence that a portion of p62 directly localizes within the mitochondria and supports stable electron transport by forming heterogeneous protein complexes. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF) of mitochondrial proteins co-purified with p62 revealed that p62 interacts with several oxidation-prone proteins, including a few components of the electron transport chain complexes, as well as multiple chaperone molecules and redox regulatory enzymes. Accordingly, p62-deficient mitochondria exhibited compromised electron transport, and the compromised function was partially restored by in vitro delivery of p62. These results suggest that p62 plays an additional role in maintaining mitochondrial integrity at the vicinity of target machineries through its function in relation to protein quality control.« less

Methionine Sulfoxide Reductase A Knockout Mice Show Progressive Hearing Loss and Sensitivity to Acoustic Trauma.

PubMed

Alqudah, Safa; Chertoff, Mark; Durham, Dianne; Moskovitz, Jackob; Staecker, Hinrich; Peppi, Marcello

2018-06-21

Methionine sulfoxide reductases (MsrA and MsrB) protect the biological activity of proteins from oxidative modifications to methionine residues and are important for protecting against the pathological effects of neurodegenerative diseases. In the current study, we characterized the auditory phenotype of the MsrA knockout mouse. Young MsrA knockout mice showed small high-frequency threshold elevations for auditory brainstem response and distortion product otoacoustic emission compared to those of wild-type mice, which progressively worsened in older MsrA knockout mice. MsrA knockout mice showed an increased sensitivity to noise at young and older ages, suggesting that MsrA is part of a mechanism that protects the cochlea from acoustic damage. MsrA mRNA in the cochlea was increased following acoustic stimulation. Finally, expression of mRNA MsrB1 was compromised at 6 months old, but not in younger MsrA knockout mice (compared to controls). The identification of MsrA in the cochlea as a protective mediator from both early onset hearing loss and acoustic trauma expands our understanding of the pathways that may induce protection from acoustic trauma and foster further studies on how to prevent the damaging effect of noise exposure through Msr-based therapy. © 2018 S. Karger AG, Basel.

Limitations in Bonding to Dentin and Experimental Strategies to Prevent Bond Degradation

PubMed Central

Liu, Y.; Tjäderhane, L.; Breschi, L.; Mazzoni, A.; Li, N.; Mao, J.; Pashley, D.H.; Tay, F.R.

2011-01-01

The limited durability of resin-dentin bonds severely compromises the lifetime of tooth-colored restorations. Bond degradation occurs via hydrolysis of suboptimally polymerized hydrophilic resin components and degradation of water-rich, resin-sparse collagen matrices by matrix metalloproteinases (MMPs) and cysteine cathepsins. This review examined data generated over the past three years on five experimental strategies developed by different research groups for extending the longevity of resin-dentin bonds. They include: (1) increasing the degree of conversion and esterase resistance of hydrophilic adhesives; (2) the use of broad-spectrum inhibitors of collagenolytic enzymes, including novel inhibitor functional groups grafted to methacrylate resins monomers to produce anti-MMP adhesives; (3) the use of cross-linking agents for silencing the activities of MMP and cathepsins that irreversibly alter the 3-D structures of their catalytic/allosteric domains; (4) ethanol wet-bonding with hydrophobic resins to completely replace water from the extrafibrillar and intrafibrillar collagen compartments and immobilize the collagenolytic enzymes; and (5) biomimetic remineralization of the water-filled collagen matrix using analogs of matrix proteins to progressively replace water with intrafibrillar and extrafibrillar apatites to exclude exogenous collagenolytic enzymes and fossilize endogenous collagenolytic enzymes. A combination of several of these strategies should result in overcoming the critical barriers to progress currently encountered in dentin bonding. PMID:21220360

Succination is Increased on Select Proteins in the Brainstem of the NADH dehydrogenase (ubiquinone) Fe-S protein 4 (Ndufs4) Knockout Mouse, a Model of Leigh Syndrome*

PubMed Central

Piroli, Gerardo G.; Manuel, Allison M.; Clapper, Anna C.; Walla, Michael D.; Baatz, John E.; Palmiter, Richard D.; Quintana, Albert; Frizzell, Norma

2016-01-01

Elevated fumarate concentrations as a result of Krebs cycle inhibition lead to increases in protein succination, an irreversible post-translational modification that occurs when fumarate reacts with cysteine residues to generate S-(2-succino)cysteine (2SC). Metabolic events that reduce NADH re-oxidation can block Krebs cycle activity; therefore we hypothesized that oxidative phosphorylation deficiencies, such as those observed in some mitochondrial diseases, would also lead to increased protein succination. Using the Ndufs4 knockout (Ndufs4 KO) mouse, a model of Leigh syndrome, we demonstrate for the first time that protein succination is increased in the brainstem (BS), particularly in the vestibular nucleus. Importantly, the brainstem is the most affected region exhibiting neurodegeneration and astrocyte and microglial proliferation, and these mice typically die of respiratory failure attributed to vestibular nucleus pathology. In contrast, no increases in protein succination were observed in the skeletal muscle, corresponding with the lack of muscle pathology observed in this model. 2D SDS-PAGE followed by immunoblotting for succinated proteins and MS/MS analysis of BS proteins allowed us to identify the voltage-dependent anion channels 1 and 2 as specific targets of succination in the Ndufs4 knockout. Using targeted mass spectrometry, Cys77 and Cys48 were identified as endogenous sites of succination in voltage-dependent anion channels 2. Given the important role of voltage-dependent anion channels isoforms in the exchange of ADP/ATP between the cytosol and the mitochondria, and the already decreased capacity for ATP synthesis in the Ndufs4 KO mice, we propose that the increased protein succination observed in the BS of these animals would further decrease the already compromised mitochondrial function. These data suggest that fumarate is a novel biochemical link that may contribute to the progression of the neuropathology in this mitochondrial disease model. PMID:26450614

Succination is Increased on Select Proteins in the Brainstem of the NADH dehydrogenase (ubiquinone) Fe-S protein 4 (Ndufs4) Knockout Mouse, a Model of Leigh Syndrome.

PubMed

Piroli, Gerardo G; Manuel, Allison M; Clapper, Anna C; Walla, Michael D; Baatz, John E; Palmiter, Richard D; Quintana, Albert; Frizzell, Norma

2016-02-01

Elevated fumarate concentrations as a result of Krebs cycle inhibition lead to increases in protein succination, an irreversible post-translational modification that occurs when fumarate reacts with cysteine residues to generate S-(2-succino)cysteine (2SC). Metabolic events that reduce NADH re-oxidation can block Krebs cycle activity; therefore we hypothesized that oxidative phosphorylation deficiencies, such as those observed in some mitochondrial diseases, would also lead to increased protein succination. Using the Ndufs4 knockout (Ndufs4 KO) mouse, a model of Leigh syndrome, we demonstrate for the first time that protein succination is increased in the brainstem (BS), particularly in the vestibular nucleus. Importantly, the brainstem is the most affected region exhibiting neurodegeneration and astrocyte and microglial proliferation, and these mice typically die of respiratory failure attributed to vestibular nucleus pathology. In contrast, no increases in protein succination were observed in the skeletal muscle, corresponding with the lack of muscle pathology observed in this model. 2D SDS-PAGE followed by immunoblotting for succinated proteins and MS/MS analysis of BS proteins allowed us to identify the voltage-dependent anion channels 1 and 2 as specific targets of succination in the Ndufs4 knockout. Using targeted mass spectrometry, Cys(77) and Cys(48) were identified as endogenous sites of succination in voltage-dependent anion channels 2. Given the important role of voltage-dependent anion channels isoforms in the exchange of ADP/ATP between the cytosol and the mitochondria, and the already decreased capacity for ATP synthesis in the Ndufs4 KO mice, we propose that the increased protein succination observed in the BS of these animals would further decrease the already compromised mitochondrial function. These data suggest that fumarate is a novel biochemical link that may contribute to the progression of the neuropathology in this mitochondrial disease model. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Academic Standards and Regulatory Frameworks: Necessary Compromises?

ERIC Educational Resources Information Center

Stowell, Marie; Falahee, Marie; Woolf, Harvey

2016-01-01

Assessment regulations in higher education, which are important for assuring threshold academic standards, reflect institutional cultures and histories, and are shaped by pragmatic concerns about quality indicators such as retention and progression rates, as well as principles of equity. This paper articulates some of the tensions that confront…

MStern Blotting-High Throughput Polyvinylidene Fluoride (PVDF) Membrane-Based Proteomic Sample Preparation for 96-Well Plates.

PubMed

Berger, Sebastian T; Ahmed, Saima; Muntel, Jan; Cuevas Polo, Nerea; Bachur, Richard; Kentsis, Alex; Steen, Judith; Steen, Hanno

2015-10-01

We describe a 96-well plate compatible membrane-based proteomic sample processing method, which enables the complete processing of 96 samples (or multiples thereof) within a single workday. This method uses a large-pore hydrophobic PVDF membrane that efficiently adsorbs proteins, resulting in fast liquid transfer through the membrane and significantly reduced sample processing times. Low liquid transfer speeds have prevented the useful 96-well plate implementation of FASP as a widely used membrane-based proteomic sample processing method. We validated our approach on whole-cell lysate and urine and cerebrospinal fluid as clinically relevant body fluids. Without compromising peptide and protein identification, our method uses a vacuum manifold and circumvents the need for digest desalting, making our processing method compatible with standard liquid handling robots. In summary, our new method maintains the strengths of FASP and simultaneously overcomes one of the major limitations of FASP without compromising protein identification and quantification. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

A biomimetic approach for enhancing the in vivo half-life of peptides

PubMed Central

Penchala, Sravan C; Miller, Mark R; Pal, Arindom; Dong, Jin; Madadi, Nikhil R.; Xie, Jinghang; Joo, Hyun; Tsai, Jerry; Batoon, Patrick; Samoshin, Vyacheslav; Franz, Andreas; Cox, Trever; Miles, Jesse; Chan, William K; Park, Miki S; Alhamadsheh, Mamoun M

2015-01-01

The tremendous therapeutic potential of peptides has not yet been realized, mainly due to their short in vivo half-life. While conjugation to macromolecules has been a mainstay approach for enhancing the half-life of proteins, the steric hindrance of macromolecules often harms the binding of peptides to target receptors, compromising the in vivo efficacy. Here we report a new strategy for enhancing the in vivo half-life of peptides without compromising their potency. Our approach involves endowing peptides with a small-molecule that binds reversibly to the serum protein, transthyretin. Although there are few reversible albumin-binding molecules, we are unaware of designed small molecules that bind reversibly to other serum proteins and are used for half-life extension in vivo. We show here that our strategy was indeed effective in enhancing the half-life of an agonist for GnRH receptor while maintaining its binding affinity, which was translated into superior in vivo efficacy. PMID:26344696

MStern Blotting–High Throughput Polyvinylidene Fluoride (PVDF) Membrane-Based Proteomic Sample Preparation for 96-Well Plates*

PubMed Central

Berger, Sebastian T.; Ahmed, Saima; Muntel, Jan; Cuevas Polo, Nerea; Bachur, Richard; Kentsis, Alex; Steen, Judith; Steen, Hanno

2015-01-01

We describe a 96-well plate compatible membrane-based proteomic sample processing method, which enables the complete processing of 96 samples (or multiples thereof) within a single workday. This method uses a large-pore hydrophobic PVDF membrane that efficiently adsorbs proteins, resulting in fast liquid transfer through the membrane and significantly reduced sample processing times. Low liquid transfer speeds have prevented the useful 96-well plate implementation of FASP as a widely used membrane-based proteomic sample processing method. We validated our approach on whole-cell lysate and urine and cerebrospinal fluid as clinically relevant body fluids. Without compromising peptide and protein identification, our method uses a vacuum manifold and circumvents the need for digest desalting, making our processing method compatible with standard liquid handling robots. In summary, our new method maintains the strengths of FASP and simultaneously overcomes one of the major limitations of FASP without compromising protein identification and quantification. PMID:26223766

An isoenergetic high-protein, moderate-fat diet does not compromise strength and fatigue during resistance exercise in women.

PubMed

Dipla, Konstantina; Makri, Maria; Zafeiridis, Andreas; Soulas, Dimitrios; Tsalouhidou, Sofia; Mougios, Vassilis; Kellis, Spyros

2008-08-01

Resistance exercise is recommended to individuals following high-protein diets in order to augment changes in body composition. However, alterations in macronutrient composition may compromise physical performance. The present study investigated the effects of an isoenergetic high-protein diet on upper and lower limb strength and fatigue during high-intensity resistance exercise. Ten recreationally active women, aged 25-40 years, followed a control diet (55, 15 and 30 % of energy from carbohydrate, protein and fat, respectively) and a high-protein diet (respective values, 30, 40 and 30) for 7 d each in a random counterbalanced design. Each participant underwent strength testing of upper limb (isometric handgrip strength and endurance) and lower limb (four sets of sixteen maximal knee flexions and extensions on an isokinetic dynamometer) before and after applying each diet. Body weight, body fat and RER were significantly reduced following the high-protein diet (P < 0.05). No differences were found between diets in any of the strength performance parameters (handgrip strength, handgrip endurance, peak torque, total work and fatigue) or the responses of heart rate, systolic and diastolic arterial pressure, blood lactate and blood glucose to exercise. Women on a short-term isoenergetic high-protein, moderate-fat diet maintained muscular strength and endurance of upper and lower limbs during high-intensity resistance exercise without experiencing fatigue earlier compared with a control diet.

Vegetarian diets and bone status.

PubMed

Tucker, Katherine L

2014-07-01

Osteoporosis is a common chronic condition associated with progressive loss of bone mineral density (BMD) and compromised bone strength, with increasing risk of fracture over time. Vegetarian diets have been shown to contain lower amounts of calcium, vitamin D, vitamin B-12, protein, and n-3 (ω-3) fatty acids, all of which have important roles in maintaining bone health. Although zinc intakes are not necessarily lower quantitatively, they are considerably less bioavailable in vegetarian diets, which suggests the need for even higher intakes to maintain adequate status. At the same time, healthy vegetarian diets tend to contain more of several protective nutrients, including magnesium, potassium, vitamin K, and antioxidant and anti-inflammatory phytonutrients. On balance, there is evidence that vegetarians, and particularly vegans, may be at greater risk of lower BMD and fracture. Attention to potential shortfall nutrients through the careful selection of foods or fortified foods or the use of supplements can help ensure healthy bone status to reduce fracture risk in individuals who adhere to vegetarian diets. © 2014 American Society for Nutrition.

Loss of Miro1-directed mitochondrial movement results in a novel murine model for neuron disease

PubMed Central

Nguyen, Tammy T.; Oh, Sang S.; Weaver, David; Lewandowska, Agnieszka; Maxfield, Dane; Schuler, Max-Hinderk; Smith, Nathan K.; Macfarlane, Jane; Saunders, Gerald; Palmer, Cheryl A.; Debattisti, Valentina; Koshiba, Takumi; Pulst, Stefan; Feldman, Eva L.; Hajnóczky, György; Shaw, Janet M.

2014-01-01

Defective mitochondrial distribution in neurons is proposed to cause ATP depletion and calcium-buffering deficiencies that compromise cell function. However, it is unclear whether aberrant mitochondrial motility and distribution alone are sufficient to cause neurological disease. Calcium-binding mitochondrial Rho (Miro) GTPases attach mitochondria to motor proteins for anterograde and retrograde transport in neurons. Using two new KO mouse models, we demonstrate that Miro1 is essential for development of cranial motor nuclei required for respiratory control and maintenance of upper motor neurons required for ambulation. Neuron-specific loss of Miro1 causes depletion of mitochondria from corticospinal tract axons and progressive neurological deficits mirroring human upper motor neuron disease. Although Miro1-deficient neurons exhibit defects in retrograde axonal mitochondrial transport, mitochondrial respiratory function continues. Moreover, Miro1 is not essential for calcium-mediated inhibition of mitochondrial movement or mitochondrial calcium buffering. Our findings indicate that defects in mitochondrial motility and distribution are sufficient to cause neurological disease. PMID:25136135

Fall and Fracture Risk in Sarcopenia and Dynapenia With and Without Obesity: the Role of Lifestyle Interventions.

PubMed

Scott, David; Daly, Robin M; Sanders, Kerrie M; Ebeling, Peter R

2015-08-01

Due to their differing etiologies and consequences, it has been proposed that the term "sarcopenia" should revert to its original definition of age-related muscle mass declines, with a separate term, "dynapenia", describing muscle strength and function declines. There is increasing interest in the interactions of sarcopenia and dynapenia with obesity. Despite an apparent protective effect of obesity on fracture, increased adiposity may compromise bone health, and the presence of sarcopenia and/or dynapenia ("sarcopenic obesity" and "dynapenic obesity") may exacerbate the risk of falls and fracture in obese older adults. Weight loss interventions are likely to be beneficial for older adults with sarcopenic and dynapenic obesity but may result in further reductions in muscle and bone health. The addition of exercise including progressive resistance training and nutritional strategies, including protein and vitamin D supplementation, may optimise body composition and muscle function outcomes thereby reducing falls and fracture risk in this population.

Jellyfish Bioactive Compounds: Methods for Wet-Lab Work

PubMed Central

Frazão, Bárbara; Antunes, Agostinho

2016-01-01

The study of bioactive compounds from marine animals has provided, over time, an endless source of interesting molecules. Jellyfish are commonly targets of study due to their toxic proteins. However, there is a gap in reviewing successful wet-lab methods employed in these animals, which compromises the fast progress in the detection of related biomolecules. Here, we provide a compilation of the most effective wet-lab methodologies for jellyfish venom extraction prior to proteomic analysis—separation, identification and toxicity assays. This includes SDS-PAGE, 2DE, gel chromatography, HPLC, DEAE, LC-MS, MALDI, Western blot, hemolytic assay, antimicrobial assay and protease activity assay. For a more comprehensive approach, jellyfish toxicity studies should further consider transcriptome sequencing. We reviewed such methodologies and other genomic techniques used prior to the deep sequencing of transcripts, including RNA extraction, construction of cDNA libraries and RACE. Overall, we provide an overview of the most promising methods and their successful implementation for optimizing time and effort when studying jellyfish. PMID:27077869

Jellyfish Bioactive Compounds: Methods for Wet-Lab Work.

PubMed

Frazão, Bárbara; Antunes, Agostinho

2016-04-12

The study of bioactive compounds from marine animals has provided, over time, an endless source of interesting molecules. Jellyfish are commonly targets of study due to their toxic proteins. However, there is a gap in reviewing successful wet-lab methods employed in these animals, which compromises the fast progress in the detection of related biomolecules. Here, we provide a compilation of the most effective wet-lab methodologies for jellyfish venom extraction prior to proteomic analysis-separation, identification and toxicity assays. This includes SDS-PAGE, 2DE, gel chromatography, HPLC, DEAE, LC-MS, MALDI, Western blot, hemolytic assay, antimicrobial assay and protease activity assay. For a more comprehensive approach, jellyfish toxicity studies should further consider transcriptome sequencing. We reviewed such methodologies and other genomic techniques used prior to the deep sequencing of transcripts, including RNA extraction, construction of cDNA libraries and RACE. Overall, we provide an overview of the most promising methods and their successful implementation for optimizing time and effort when studying jellyfish.

Enteral nutrition in the chronic obstructive pulmonary disease (COPD) patient.

PubMed

DeBellis, Heather F; Fetterman, James W

2012-12-01

Chronic obstructive pulmonary disease (COPD) is a progressive, chronic disease, in which malnutrition can have an undesirable effect. Therefore, the patient's nutritional status is critical for optimizing outcomes in COPD. The initial nutrition assessment is focused on identifying calorically compromised COPD patients in order to provide them with appropriate nutrition. Nutritional intervention consists of oral supplementation and enteral nutrition to prevent weight loss and muscle mass depletion. Evaluation of nutritional status should include past medical history (medications, lung function, and exercise tolerance) and dietary history (patient's dietary habits, food choices, meal patterns, food allergy information, and malabsorption issues), in addition to physiological stress, visceral proteins, weight, fat-free mass, and body mass index. The current medical literature conflicts regarding the appropriate type of formulation to select for nutritional intervention, especially regarding the amount of calories from fat to provide COPD patients. This review article focuses on the enteral product formulations currently available, and how they are most appropriately utilized in patients with COPD.

FY 2012 Lightweight Materials Annual Report

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

Warren, David C.

2013-04-15

The FY 2012 Annual Progress Report for Lightweight Materials provides a detailed description of the activities and technical accomplishments which focuses on the development and validation of advanced materials and manufacturing technologies to significantly reduce light and heavy duty vehicle weight without compromising other attributes such as safety, performance, recyclability, and cost.

Annual Research Review: Resilient Functioning in Maltreated Children--Past, Present, and Future Perspectives

ERIC Educational Resources Information Center

Cicchetti, Dante

2013-01-01

Background: Through a process of probabilistic epigenesis, child maltreatment progressively contributes to compromised adaptation on a variety of developmental domains central to successful adjustment. These developmental failures pose significant risk for the emergence of psychopathology across the life course. In addition to the psychological…

48 CFR 32.503-14 - Protection of Government title.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 48 Federal Acquisition Regulations System 1 2010-10-01 2010-10-01 false Protection of Government... of Government title. (a) Since the Progress Payments clause gives the Government title to all of the... ensure that the Government title to these inventories is not compromised by other encumbrances...

Speech Intelligibility and Marital Communication in Amyotrophic Lateral Sclerosis: An Exploratory Study

ERIC Educational Resources Information Center

Joubert, Karin; Bornman, Juan; Alant, Erna

2011-01-01

Amyotrophic lateral sclerosis (ALS), a rapidly progressive neuromuscular disease, has a devastating impact not only on individuals diagnosed with ALS but also their spouses. Speech intelligibility, often compromised as a result of dysarthria, affects the couple's ability to maintain effective, intimate communication. The purpose of this…

48 CFR 32.503-14 - Protection of Government title.

Code of Federal Regulations, 2014 CFR

2014-10-01

... 48 Federal Acquisition Regulations System 1 2014-10-01 2014-10-01 false Protection of Government... of Government title. (a) Since the Progress Payments clause gives the Government title to all of the... ensure that the Government title to these inventories is not compromised by other encumbrances...

Introduction to the issue regarding research regarding age related macular degeneration

USDA-ARS?s Scientific Manuscript database

Blindness is the second greatest fear among the elderly. Age-related macular degeneration (AMD) is the leading cause of vision loss among the elderly in most industrialized nations. AMD first compromises central high acuity vision. Subsequently, all vision may be lost. AMD is a progressive retinal d...

Congenital Bone Fractures in Spinal Muscular Atrophy: Functional Role for SMN Protein in Bone Remodeling

PubMed Central

Shanmugarajan, Srinivasan; Swoboda, Kathryn J.; Iannaccone, Susan T.; Ries, William L.; Maria, Bernard L.; Reddy, Sakamuri V.

2009-01-01

Spinal muscular atrophy is the second most common fatal childhood disorder. Core clinical features include muscle weakness caused by degenerating lower motor neurons and a high incidence of bone fractures and hypercalcemia. Fractures further compromise quality of life by progression of joint contractures or additional loss of motor function. Recent observations suggest that bone disease in spinal muscular atrophy may not be attributed entirely to lower motor neuron degeneration. The presence of the spinal muscular atrophy disease-determining survival motor neuron gene (SMN), SMN expression, and differential splicing in bone-resorbing osteoclasts was recently discovered. Its ubiquitous expression and the differential expression of splice variants suggest that SMN has specific roles in bone cell function. SMN protein also interacts with osteoclast stimulatory factor. Mouse models of human spinal muscular atrophy disease suggest a potential role of SMN protein in skeletal development. Dual energy x-ray absorptiometry analysis demonstrated a substantial decrease in total bone area and poorly developed caudal vertebra in the mouse model. These mice also had pelvic bone fractures. Studies delineating SMN signaling mechanisms and gene transcription in a cell-specific manner will provide important molecular insights into the pathogenesis of bone disease in children with spinal muscular atrophy. Moreover, understanding bone remodeling in spinal muscular atrophy may lead to novel therapeutic approaches to enhance skeletal health and quality of life. This article reviews the skeletal complications associated with spinal muscular atrophy and describes a functional role for SMN protein in osteoclast development and bone resorption activity. PMID:17761651

Zinc finger proteins in cancer progression.

PubMed

Jen, Jayu; Wang, Yi-Ching

2016-07-13

Zinc finger proteins are the largest transcription factor family in human genome. The diverse combinations and functions of zinc finger motifs make zinc finger proteins versatile in biological processes, including development, differentiation, metabolism and autophagy. Over the last few decades, increasing evidence reveals the potential roles of zinc finger proteins in cancer progression. However, the underlying mechanisms of zinc finger proteins in cancer progression vary in different cancer types and even in the same cancer type under different types of stress. Here, we discuss general mechanisms of zinc finger proteins in transcription regulation and summarize recent studies on zinc finger proteins in cancer progression. In this review, we also emphasize the importance of further investigations in elucidating the underlying mechanisms of zinc finger proteins in cancer progression.

BTG/Tob family members Tob1 and Tob2 inhibit proliferation of mouse embryonic stem cells via Id3 mRNA degradation

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

Chen, Yuanfan; Wang, Chenchen; Peking University Stem Cell Research Center, China National Center for International Research, Peking University Health Science Center, Beijing 100191

2015-07-03

The mammalian BTG/Tob family is a group of proteins with anti-proliferative ability, and there are six members including BTG1, BTG2/PC3/Tis21, BTG3/ANA, BTG4/PC3B, Tob1/Tob and Tob2. Among them, Tob subfamily members, specifically Tob1/Tob and Tob2, have the most extensive C-terminal regions. As previously reported, overexpression of BTG/Tob proteins is associated with the inhibition of G1 to S-phase cell cycle progression and decreased cell proliferation in a variety of cell types. Tob subfamily proteins have similar anti-proliferative effects on cell cycle progression in cultured tumor cells. An important unresolved question is whether or not they have function in rapidly proliferating cells, suchmore » as embryonic stem cells (ESCs). Tob1 and Tob2 were expressed ubiquitously in mouse ESCs (mESCs), suggesting a possible role in early embryonic development and mESCs. To address the above question and explore the possible functions of the Tob subfamily in ESCs, we established ESCs from different genotypic knockout inner cell mass (ICM). We found that Tob1{sup −/−}, Tob2{sup −/−}, and Tob1/2 double knockout (DKO, Tob1{sup −/−} & Tob2{sup −/−}) ESCs grew faster than wild type (WT) ESCs without losing pluripotency, and we provide a possible mechanistic explanation for these observations: Tob1 and Tob2 inhibit the cell cycle via degradation of Id3 mRNA, which is a set of directly targeted genes of BMP4 signaling in mESCs that play critical roles in the maintenance of ESC properties. Together, our data suggest that BTG/Tob family protein Tob1 and Tob2 regulation cell proliferation does not compromise the basic properties of mESCs. - Highlights: • We established mouse Tob1/2 double knockout embryonic stem cells. • Tob1 and Tob2 inhibit the proliferation of ESCs without effect on pluripotency. • Tob1 and Tob2 involved in the degradation of Id3 in mESCs.« less

N-terminally truncated GADD34 proteins are convenient translation enhancers in a human cell-derived in vitro protein synthesis system.

PubMed

Mikami, Satoshi; Kobayashi, Tominari; Machida, Kodai; Masutani, Mamiko; Yokoyama, Shigeyuki; Imataka, Hiroaki

2010-07-01

Human cell-derived in vitro protein synthesis systems are useful for the production of recombinant proteins. Productivity can be increased by supplementation with GADD34, a protein that is difficult to express in and purify from E. coli. Deletion of the N-terminal 120 or 240 amino acids of GADD34 improves recovery of this protein from E. coli without compromising its ability to boost protein synthesis in an in vitro protein synthesis system. The use of N-terminally truncated GADD34 proteins in place of full-length GADD34 should improve the utility of human cell-based cell-free protein synthesis systems.

Predicting intrapartum fetal compromise using the fetal cerebro-umbilical ratio.

PubMed

Sabdia, S; Greer, R M; Prior, T; Kumar, S

2015-05-01

The aim of this study was to explore the association between the cerebro-umbilical ratio measured at 35-37 weeks and intrapartum fetal compromise. This retrospective cross sectional study was conducted at the Mater Mothers' Hospital in Brisbane, Australia. Maternal demographics and fetal Doppler indices at 35-37 weeks gestation for 1381 women were correlated with intrapartum and neonatal outcomes. Babies born by caesarean section or instrumental delivery for fetal compromise had the lowest median cerebro-umbilical ratio 1.60 (IQR 1.22-2.08) compared to all other delivery groups (vaginal delivery, emergency delivery for failure to progress, emergency caesarean section for other reasons or elective caesarean section). The percentage of infants with a cerebro-umbilical ratio <10th centile that required emergency delivery (caesarean section or instrumental delivery) for fetal compromise was 22%, whereas only 7.3% of infants with a cerebro-umbilical ratio between the 10th-90th centile and 9.6% of infants with a cerebro-umbilical ratio > 90th centile required delivery for the same indication (p < 0.001). A lower cerebro-umbilical ratio was associated with an increased risk of emergency delivery for fetal compromise, OR 2.03 (95% CI 1.41-2.92), p < 0.0001. This study suggests that a low fetal cerebro-umbilical ratio measured at 35-37 weeks is associated with a greater risk of intrapartum compromise. This is a relatively simple technique which could be used to risk stratify women in diverse healthcare settings. Copyright © 2015 Elsevier Ltd. All rights reserved.

Exacerbation of pathology by oxidative stress in respiratory and locomotor muscles with Duchenne muscular dystrophy.

PubMed

Lawler, John M

2011-05-01

Duchenne muscular dystrophy (DMD) is the most devastating type of muscular dystrophy, leading to progressive weakness of respiratory (e.g. diaphragm) and locomotor muscles (e.g. gastrocnemius). DMD is caused by X-linked defects in the gene that encodes for dystrophin, a key scaffolding protein of the dystroglycan complex (DCG) within the sarcolemmal cytoskeleton. As a result of a compromised dystroglycan complex, mechanical integrity is impaired and important signalling proteins (e.g. nNOS, caveolin-3) and pathways are disrupted. Disruption of the dystroglycan complex leads to high susceptibility to injury with repeated, eccentric contractions as well as inflammation, resulting in significant damage and necrosis. Chronic damage and repair cycling leads to fibrosis and weakness. While the link between inflammation with damage and weakness in the DMD diaphragm is unresolved, elevated oxidative stress may contribute to damage, weakness and possibly fibrosis. While utilization of non-specific antioxidant interventions has yielded inconsistent results, recent data suggest that NAD(P)H oxidase could play a pivotal role in elevating oxidative stress via integrated changes in caveolin-3 and stretch-activated channels (SACs). Oxidative stress may act as an amplifier, exacerbating disruption of the dystroglycan complex, upregulation of the inflammatory transcription factor NF-B, and thus functional impairment of force-generating capacity.

Impact of Childhood Malnutrition on Host Defense and Infection.

PubMed

Ibrahim, Marwa K; Zambruni, Mara; Melby, Christopher L; Melby, Peter C

2017-10-01

The global impact of childhood malnutrition is staggering. The synergism between malnutrition and infection contributes substantially to childhood morbidity and mortality. Anthropometric indicators of malnutrition are associated with the increased risk and severity of infections caused by many pathogens, including viruses, bacteria, protozoa, and helminths. Since childhood malnutrition commonly involves the inadequate intake of protein and calories, with superimposed micronutrient deficiencies, the causal factors involved in impaired host defense are usually not defined. This review focuses on literature related to impaired host defense and the risk of infection in primary childhood malnutrition. Particular attention is given to longitudinal and prospective cohort human studies and studies of experimental animal models that address causal, mechanistic relationships between malnutrition and host defense. Protein and micronutrient deficiencies impact the hematopoietic and lymphoid organs and compromise both innate and adaptive immune functions. Malnutrition-related changes in intestinal microbiota contribute to growth faltering and dysregulated inflammation and immune function. Although substantial progress has been made in understanding the malnutrition-infection synergism, critical gaps in our understanding remain. We highlight the need for mechanistic studies that can lead to targeted interventions to improve host defense and reduce the morbidity and mortality of infectious diseases in this vulnerable population. Copyright © 2017 American Society for Microbiology.

Arabidopsis adaptor protein 1G is critical for pollen development.

PubMed

Feng, Chong; Wang, Jia-Gang; Liu, Hai-Hong; Li, Sha; Zhang, Yan

2017-09-01

Pollen development is a pre-requisite for sexual reproduction of angiosperms, during which various cellular activities are involved. Pollen development accompanies dynamic remodeling of vacuoles through fission and fusion, disruption of which often compromises pollen viability. We previously reported that the Y subunit of adaptor protein 1 (AP1G) mediates synergid degeneration during pollen tube reception. Here, we demonstrate that AP1G is essential for pollen development. AP1G loss-of-function resulted in male gametophytic lethality due to defective pollen development. By ultrastructural analysis and fluorescence labeling, we demonstrate that AP1G loss-of-function compromised dynamic vacuolar remodeling during pollen development and impaired vacuolar acidification of pollen. Results presented here support a key role of vacuoles in gametophytic pollen development. © 2017 Institute of Botany, Chinese Academy of Sciences.

Endosome-Associated CRT1 Functions Early in Resistance Gene–Mediated Defense Signaling in Arabidopsis and Tobacco[W

PubMed Central

Kang, Hong-Gu; Oh, Chang-Sik; Sato, Masanao; Katagiri, Fumiaki; Glazebrook, Jane; Takahashi, Hideki; Kachroo, Pradeep; Martin, Gregory B.; Klessig, Daniel F.

2010-01-01

Resistance gene–mediated immunity confers protection against pathogen infection in a wide range of plants. A genetic screen for Arabidopsis thaliana mutants compromised for recognition of turnip crinkle virus previously identified CRT1, a member of the GHKL ATPase/kinase superfamily. Here, we demonstrate that CRT1 interacts with various resistance proteins from different structural classes, and this interaction is disrupted when these resistance proteins are activated. The Arabidopsis mutant crt1-2 crh1-1, which lacks CRT1 and its closest homolog, displayed compromised resistance to avirulent Pseudomonas syringae and Hyaloperonospora arabidopsidis. Additionally, resistance-associated hypersensitive cell death was suppressed in Nicotiana benthamiana silenced for expression of CRT1 homolog(s). Thus, CRT1 appears to be a general factor for resistance gene–mediated immunity. Since elevation of cytosolic calcium triggered by avirulent P. syringae was compromised in crt1-2 crh1-1 plants, but cell death triggered by Nt MEK2DD was unaffected in CRT1-silenced N. benthamiana, CRT1 likely functions at an early step in this pathway. Genome-wide transcriptome analysis led to identification of CRT1-Associated genes, many of which are associated with transport processes, responses to (a)biotic stress, and the endomembrane system. Confocal microscopy and subcellular fractionation revealed that CRT1 localizes to endosome-like vesicles, suggesting a key process in resistance protein activation/signaling occurs in this subcellular compartment. PMID:20332379

Towards an Understanding of Energy Impairment in Huntington’s Disease Brain

PubMed Central

Dubinsky, Janet M.

2017-01-01

This review systematically examines the evidence for shifts in flux through energy generating biochemical pathways in Huntington’s disease (HD) brains from humans and model systems. Compromise of the electron transport chain (ETC) appears not to be the primary or earliest metabolic change in HD pathogenesis. Rather, compromise of glucose uptake facilitates glucose flux through glycolysis and may possibly decrease flux through the pentose phosphate pathway (PPP), limiting subsequent NADPH and GSH production needed for antioxidant protection. As a result, oxidative damage to key glycolytic and tricarboxylic acid (TCA) cycle enzymes further restricts energy production so that while basal needs may be met through oxidative phosphorylation, those of excessive stimulation cannot. Energy production may also be compromised by deficits in mitochondrial biogenesis, dynamics or trafficking. Restrictions on energy production may be compensated for by glutamate oxidation and/or stimulation of fatty acid oxidation. Transcriptional dysregulation generated by mutant huntingtin also contributes to energetic disruption at specific enzymatic steps. Many of the alterations in metabolic substrates and enzymes may derive from normal regulatory feedback mechanisms and appear oscillatory. Fine temporal sequencing of the shifts in metabolic flux and transcriptional and expression changes associated with mutant huntingtin expression remain largely unexplored and may be model dependent. Differences in disease progression among HD model systems at the time of experimentation and their varying states of metabolic compensation may explain conflicting reports in the literature. Progressive shifts in metabolic flux represent homeostatic compensatory mechanisms that maintain the model organism through presymptomatic and symptomatic stages. PMID:29125492

Widespread Volumetric Reductions in Schizophrenia and Schizoaffective Patients Displaying Compromised Cognitive Abilities.

PubMed

Van Rheenen, Tamsyn E; Cropley, Vanessa; Zalesky, Andrew; Bousman, Chad; Wells, Ruth; Bruggemann, Jason; Sundram, Suresh; Weinberg, Danielle; Lenroot, Roshel K; Pereira, Avril; Shannon Weickert, Cynthia; Weickert, Thomas W; Pantelis, Christos

2018-04-06

Progress toward understanding brain mechanisms in psychosis is hampered by failures to account for within-group heterogeneity that exists across neuropsychological domains. We recently identified distinct cognitive subgroups that might assist in identifying more biologically meaningful subtypes of psychosis. In the present study, we examined whether underlying structural brain abnormalities differentiate these cognitively derived subgroups. 1.5T T1 weighted structural scans were acquired for 168 healthy controls and 220 patients with schizophrenia/schizoaffective disorder. Based on previous work, 47 patients were categorized as being cognitively compromised (impaired premorbid and current IQ), 100 as cognitively deteriorated (normal premorbid IQ, impaired current IQ), and 73 as putatively cognitively preserved (premorbid and current IQ within 1 SD of controls). Global, subcortical and cortical volume, thickness, and surface area measures were compared among groups. Whole cortex, subcortical, and regional volume and thickness reductions were evident in all subgroups compared to controls, with the largest effect sizes in the compromised group. This subgroup also showed abnormalities in regions not seen in the other patient groups, including smaller left superior and middle frontal areas, left anterior and inferior temporal areas and right lateral medial and inferior frontal, occipital lobe and superior temporal areas. This pattern of more prominent brain structural abnormalities in the group with the most marked cognitive impairments-both currently and putatively prior to illness onset, is consistent with the concept of schizophrenia as a progressive neurodevelopmental disorder. In this group, neurodevelopmental and neurodegenerative factors may be important for cognitive function.

Focal cartilage defect compromises fluid-pressure dependent load support in the knee joint.

PubMed

Dabiri, Yaghoub; Li, LePing

2015-06-01

A focal cartilage defect involves tissue loss or rupture. Altered mechanics in the affected joint may play an essential role in the onset and progression of osteoarthritis. The objective of the present study was to determine the compromised load support in the human knee joint during defect progression from the cartilage surface to the cartilage-bone interface. Ten normal and defect cases were simulated with a previously tested 3D finite element model of the knee. The focal defects were considered in both condyles within high load-bearing regions. Fluid pressurization, anisotropic fibril-reinforcement, and depth-dependent mechanical properties were considered for the articular cartilages and menisci. The results showed that a small cartilage defect could cause 25% reduction in the load support of the knee joint due to a reduced capacity of fluid pressurization in the defect cartilage. A partial-thickness defect could cause a fluid pressure decrease or increase in the remaining underlying cartilage depending on the defect depth. A cartilage defect also increased the shear strain at the cartilage-bone interface, which was more significant with a full-thickness defect. The effect of cartilage defect on the fluid pressurization also depended on the defect sites and contact conditions. In conclusion, a focal cartilage defect causes a fluid-pressure dependent load reallocation and a compromised load support in the joint, which depend on the defect depth, site, and contact condition. Copyright © 2015 John Wiley & Sons, Ltd.

AMPKα1 controls hepatocyte proliferation independently of energy balance by regulating Cyclin A2 expression.

PubMed

Merlen, Grégory; Gentric, Géraldine; Celton-Morizur, Séverine; Foretz, Marc; Guidotti, Jacques-Emmanuel; Fauveau, Véronique; Leclerc, Jocelyne; Viollet, Benoit; Desdouets, Chantal

2014-01-01

AMP-activated protein kinase (AMPK) is an evolutionarily conserved sensor of cellular energy status that contributes to restoration of energy homeostasis by slowing down ATP-consuming pathways and activating ATP-producing pathways. Unexpectedly, in different systems, AMPK is also required for proper cell division. In the current study, we evaluated the potential effect of the AMPK catalytic subunit, AMPKα1, on hepatocyte proliferation. Hepatocyte proliferation was determined in AMPKα1 knockout and wild-type mice in vivo after two thirds partial hepatectomy, and in vitro in primary hepatocyte cultures. The activities of metabolic and cell cycle-related signaling pathways were measured. After partial hepatectomy, hepatocytes proliferated rapidly, correlating with increased AMPK phosphorylation. Deletion of AMPKα1 delayed liver regeneration by impacting on G1/S transition phase. The proliferative defect of AMPKα1-deficient hepatocytes was cell autonomous, and independent of energy balance. The priming phase, lipid droplet accumulation, protein anabolic responses and growth factor activation after partial hepatectomy occurred normally in the absence of AMPKα1 activity. By contrast, mRNA and protein expression of cyclin A2, a key driver of S phase progression, were compromised in the absence of AMPK activity. Importantly, AMPKα1 controlled cyclin A2 transcription mainly through the ATF/CREB element. Our study highlights a novel role for AMPKα1 as a positive regulator of hepatocyte division occurring independently of energy balance. Copyright © 2013 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Lifting the nebula: novel insights into skeletal muscle contractility.

PubMed

Ottenheijm, Coen A C; Granzier, Henk

2010-10-01

Nebulin is a giant protein and a constituent of the skeletal muscle sarcomere. The name of this protein refers to its unknown (i.e., nebulous) function. However, recent rapid advances reveal that nebulin plays important roles in the regulation of muscle contraction. When these functions of nebulin are compromised, muscle weakness ensues, as is the case in patients with nemaline myopathy.

Red fluorescent protein responsible for pigmentation in trematode-infected Porites compressa tissues.

PubMed

Palmer, Caroline V; Roth, Melissa S; Gates, Ruth D

2009-02-01

Reports of coral disease have increased dramatically over the last decade; however, the biological mechanisms that corals utilize to limit infection and resist disease remain poorly understood. Compromised coral tissues often display non-normal pigmentation that potentially represents an inflammation-like response, although these pigments remain uncharacterized. Using spectral emission analysis and cryo-histological and electrophoretic techniques, we investigated the pink pigmentation associated with trematodiasis, infection with Podocotyloides stenometre larval trematode, in Porites compressa. Spectral emission analysis reveals that macroscopic areas of pink pigmentation fluoresce under blue light excitation (450 nm) and produce a broad emission peak at 590 nm (+/-6) with a 60-nm full width at half maximum. Electrophoretic protein separation of pigmented tissue extract confirms the red fluorescence to be a protein rather than a low-molecular-weight compound. Histological sections demonstrate green fluorescence in healthy coral tissue and red fluorescence in the trematodiasis-compromised tissue. The red fluorescent protein (FP) is limited to the epidermis, is not associated with cells or granules, and appears unstructured. These data collectively suggest that the red FP is produced and localized in tissue infected by larval trematodes and plays a role in the immune response in corals.

Multiplex PCR assay for detection of recombinant genes encoding fatty acid desaturases fused with lichenase reporter protein in GM plants.

PubMed

Berdichevets, Iryna N; Shimshilashvili, Hristina R; Gerasymenko, Iryna M; Sindarovska, Yana R; Sheludko, Yuriy V; Goldenkova-Pavlova, Irina V

2010-07-01

Thermostable lichenase encoded by licB gene of Clostridium thermocellum can be used as a reporter protein in plant, bacterial, yeast, and mammalian cells. It has important advantages of high sensitivity and specificity in qualitative and quantitative assays. Deletion variants of LicB (e.g., LicBM3) retain its enzymatic activity and thermostability and can be expressed in translational fusion with target proteins without compromising with their properties. Fusion with the lichenase reporter is especially convenient for the heterologous expression of proteins whose analysis is difficult or compromised by host enzyme activities, as it is in case of fatty acid desaturases occurring in all groups of organisms. Recombinant desaturase-lichenase genes can be used for creating genetically modified (GM) plants with improved chill tolerance. Development of an analytical method for detection of fused desaturase-lichenase transgenes is necessary both for production of GM plants and for their certification. Here, we report a multiplex polymerase chain reaction method for detection of desA and desC desaturase genes of cyanobacteria Synechocystis sp. PCC6803 and Synechococcus vulcanus, respectively, fused to licBM3 reporter in GM plants.

Intellectual Ability and Executive Function in Pediatric Moyamoya Vasculopathy

ERIC Educational Resources Information Center

Williams, Tricia S.; Westmacott, Robyn; Dlamini, Nomazulu; Granite, Leeor; Dirks, Peter; Askalan, Rand; MacGregor, Daune; Moharir, Mahendranath; Deveber, Gabrielle

2012-01-01

Aim: Moyamoya vasculopathy is characterized by progressive stenosis of the major arteries of the Circle of Willis, resulting in compromised cerebral blood flow and increased risk of stroke. The objectives of the current study were to examine intellectual and executive functioning of children with moyamoya and to evaluate the impact of moyamoya…

FY2013 Lightweight Materials R&D Annual Progress Report

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

none,

2014-02-01

As part of the U.S. Department of Energy’s (DOE’s) Vehicle Technologies Program (VTO), the Lightweight Materials (LM) activity focuses on the development and validation of advanced materials and manufacturing technologies to significantly reduce light and heavy duty vehicle weight without compromising other attributes such as safety, performance, recyclability, and cost.

Cellular Strategies of Protein Quality Control

PubMed Central

Chen, Bryan; Retzlaff, Marco; Roos, Thomas; Frydman, Judith

2011-01-01

Eukaryotic cells must contend with a continuous stream of misfolded proteins that compromise the cellular protein homeostasis balance and jeopardize cell viability. An elaborate network of molecular chaperones and protein degradation factors continually monitor and maintain the integrity of the proteome. Cellular protein quality control relies on three distinct yet interconnected strategies whereby misfolded proteins can either be refolded, degraded, or delivered to distinct quality control compartments that sequester potentially harmful misfolded species. Molecular chaperones play a critical role in determining the fate of misfolded proteins in the cell. Here, we discuss the spatial and temporal organization of cellular quality control strategies and their implications for human diseases linked to protein misfolding and aggregation. PMID:21746797

Efficient sortase-mediated N-terminal labeling of TEV protease cleaved recombinant proteins.

PubMed

Sarpong, Kwabena; Bose, Ron

2017-03-15

A major challenge in attaching fluorophores or other handles to proteins is the availability of a site-specific labeling strategy that provides stoichiometric modification without compromising protein integrity. We developed a simple approach that combines TEV protease cleavage, sortase modification and affinity purification to N-terminally label proteins. To achieve stoichiometrically-labeled protein, we included a short affinity tag in the fluorophore-containing peptide for post-labeling purification of the modified protein. This strategy can be easily applied to any recombinant protein with a TEV site and we demonstrate this on Epidermal Growth Factor Receptor (EGFR) and Membrane Scaffold Protein (MSP) constructs. Copyright © 2017 Elsevier Inc. All rights reserved.

Multidrug-resistant, progressive, invasive diffuse spinal aspergillosis: case report and review of the literature.

PubMed

Andaluz, N; Zuccarello, M

2008-06-01

The authors describe a 65-year-old man who, after 7 years of complete remission from lung cancer, was found on routine oncologic follow imaging to have lesions on several vertebral bodies. Open biopsy of the affected thoracic vertebrae and surrounding soft tissue were negative for neoplasia. Bacteriology cultures revealed colonies of aspergillus fumigatus in all bone samples. Unlike most reported cases in which vertebral compromise rarely extends to more than two adjacent vertebrae, our patient had extensive compromise of the thoracic spine. This infection progressed despite treatment with antifungal regimens known to be effective, even in immunocompromised patients. Invasive aspergillosis of the spine is a rare and typically occurred in terminal patients. However, the spectrum of hosts and clinical presentations of invasive aspergillosis are increasing, due in part to better medical treatments that prolong the survival of patients with cancer, severe infections, and organ failure. In reviewing the literature, the authors discuss the currently available therapies for such infections of the spine, and highlight the growing incidence these and other formerly rare infections.

Diabetes mellitus: novel insights, analysis and interpretation of pathophysiology and complications management with imidazole-containing peptidomimetic antioxidants.

PubMed

Babizhayev, Mark A; Lankin, Vadim Z; Savel'Yeva, Ekaterina L; Deyev, Anatoliy I; Yegorov, Yegor E

2013-12-01

Patients suffering from the severe complications associated with both insulin- (IDDM) and non-insulin-dependent diabetes mellitus (NIDDM): nephropathy, retinopathy, neuropathy, and atherosclerosis are still largely left without a prospect of an efficient treatment. Chronic hyperglycaemia, the primary clinical manifestation of diabetes, is associated with development of certain of the diabetic complications. The accelerated formation of advanced glycation end-products (AGEs) due to elevated glycemia has repeatedly been reported as a central pathogenic factor in the development of diabetic microvascular complications. Glucose and α-dicarbonyl compounds chemically attach to proteins and nucleic acids without the aid of enzymes. Initially, chemically reversible Schiff base and Amadori product adducts form in proportion to glucose concentration. The major biological effects of excessive nonenzymatic glycosylation are leading to increased free radical production and compromised free radical inhibitory and scavenger systems, inactivation of enzymes; inhibition of regulatory molecule binding; crosslinking of glycosylated proteins and trapping of soluble proteins by glycosylated extracellular matrix (both may progress in the absence of glucose); decreased susceptibility to proteolysis; abnormalities of nucleic acid function; altered macromolecular recognition and endocytosis; and increased immunogenicity. The discovery of chemical agents that can inhibit deleterious glycation reactions is potentially of great therapeutic benefit to all diabetes-associated pathologies. This study demonstrates the progress in development of patented carnosine mimetics resistant in formulations to enzymatic hydrolysis with human carnosinases that are acting as a universal form of antioxidant, deglycating and transglycating agents that inhibit sugar-mediated protein cross-linking, chelate or inactivate a number of transition metal ions (including ferrous and copper ions), possess lipid peroxidase type of activity and protection of antioxidant enzymes from inactivation (such as in a case of superoxide dismutase). Carnosine biological mimetics react with methylglyoxal and they are described in this study as a glyoxalase mimetics. The imidazole-containing carnosine biological mimetics can react with a number of deleterious aldehydic products of lipid peroxidation and thereby suppress their toxicity. Carnosine and carcinine can also react with glycated proteins and inhibit advanced glycation end product formation. These studies indicate a therapeutic role for imidazole-containing antioxidants (non-hydrolized carnosine, carcinine, D-carnosine, ophthalmic prodrug N-acetylcarnosine, leucyl-histidylhidrazide and patented formulations thereof) in therapeutic management strategies for Type 2 Diabetes.

Innovative approaches to vitamin A assessment.

PubMed

Craft, N E

2001-05-01

The health and sight of millions of children are compromised each year as a consequence of vitamin A (VA) deficiency. Serum retinol is the most commonly used indicator of VA status. Unfortunately, its use is impractical for national surveys because it involves collection of venous blood, centrifugation and frozen storage before analysis. To make VA assessment more practical, we have developed approaches incorporating dried blood spots (DBS) or portable instrumentation. DBS have been used as a sample matrix to screen neonates for many biochemical compounds. Until recently, it was not thought that VA was stable in DBS. However, we demonstrated that the measure of DBS retinol correlates well with serum retinol in both healthy adults (r(2) = 0.88-0.90) and compromised populations (r(2) = 0.73-0.84). Compared with serum retinol, the sensitivity and specificity of detecting VA deficiency by DBS retinol range from 73 to 93% and from 90 to 100%, respectively. Although few data are available, retinol binding protein (RBP) can also be measured in DBS. RBP has been used as a surrogate marker for serum retinol. Correlations coefficients (r(2)) between serum RBP and serum retinol range from 0.4 to 0.8. In addition, work has been done to develop portable instrumentation to measure VA status in the field. A fluorometer has been optimized for VA fluorescence and is linear into the deficient range for the direct fluorimetric measurement of serum holo-RBP. Progress is being made to use the instrument to directly measure holo-RBP in a drop of whole blood.

FSGS3/CD2AP is a barbed-end capping protein that stabilizes actin and strengthens adherens junctions

PubMed Central

Brieher, William M.

2013-01-01

By combining in vitro reconstitution biochemistry with a cross-linking approach, we have identified focal segmental glomerulosclerosis 3/CD2-associated protein (FSGS3/CD2AP) as a novel actin barbed-end capping protein responsible for actin stability at the adherens junction. FSGS3/CD2AP colocalizes with E-cadherin and α-actinin-4 at the apical junction in polarized Madin-Darby canine kidney (MDCK) cells. Knockdown of FSGS3/CD2AP compromised actin stability and decreased actin accumulation at the adherens junction. Using a novel apparatus to apply mechanical stress to cell–cell junctions, we showed that knockdown of FSGS3/CD2AP compromised adhesive strength, resulting in tearing between cells and disruption of barrier function. Our results reveal a novel function of FSGS3/CD2AP and a previously unrecognized role of barbed-end capping in junctional actin dynamics. Our study underscores the complexity of actin regulation at cell–cell contacts that involves actin activators, inhibitors, and stabilizers to control adhesive strength, epithelial behavior, and permeability barrier integrity. PMID:24322428

Tissue protein nitration and peripheral blood endotoxin activity are indicative of the severity of systemic organ compromise in naturally-occurring clinical cases of bacterial mastitis in Holstein dairy cows

USDA-ARS?s Scientific Manuscript database

The objective of this survey study was to determine a relationship between the intensity of tissue protein tyrosine nitration measured in samples of mammary gland, liver, pancreas and lung compared to estimated blood endotoxin (LPS) activity. Blood was collected from nine multiparous Holstein cows...

Correcting the Record: Understanding the History of Federal Intervention and Failure in Securing U.S. Educational Reform.

ERIC Educational Resources Information Center

Hirschland, Matthew J.; Steinmo, Sven

2003-01-01

Asserts that the debate over the appropriate federal role in education began with the founding of America. Argues that successive waves of educational reform are the products of the compromise between localism and national progressive goals. Consequently, a type of schizophrenia rather than a coherent and stable public policy characterizes…

Sleep and Quality of Life in People With COPD: A Descriptive-Correlational Study.

PubMed

Dignani, Lucia; Toccaceli, Andrea; Lucertini, Carla; Petrucci, Cristina; Lancia, Loreto

2016-08-01

Sleep disorders are very common in patients with chronic obstructive pulmonary disease (COPD). However, it is not clear how sleep disorders and quality of life (QoL) affect each other in the different stages of disease progression. This descriptive-correlational study investigated the relationship between QoL, quality of sleep, and degree of disease progression in 102 outpatients with COPD. The results showed that the QoL in patients with COPD is compromised and worsens with disease progression, and the quality of sleep is significantly associated with QoL and worsened as the disease progressed. The early identification of a risk of alteration of the quality of sleep, especially in nursing care, could facilitate a preventive approach for COPD patients that could positively affect their QoL. © The Author(s) 2015.

Progress toward poliomyelitis eradication - Pakistan, January 2012-September 2013.

PubMed

2013-11-22

Pakistan is one of three countries where transmission of indigenous wild poliovirus (WPV) has never been interrupted. This report describes polio eradication activities and progress in Pakistan during January 2012-September 2013 and updates previous reports. During 2012, 58 WPV cases were reported in selected areas, compared with 198 cases throughout the country in 2011; 52 WPV cases were reported during January-September 2013, compared with 54 cases during the same period in 2012. Of the 110 WPV cases reported since January 2012, 92 cases (84%) occurred in the conflict-affected Federally Administered Tribal Areas (FATA) and in security-compromised Khyber Pakhtunkhwa (KP) Province. WPV type 3 (WPV3) was isolated from only three persons with polio in a single district in 2012; the most recent case occurred in April 2012. During August 2012-September 2013, 52 circulating vaccine-derived poliovirus type 2 (cVDPV2) cases were detected, including 30 cases (58%) identified in FATA during January-September 2013. Approximately 350,000 children in certain districts of FATA have not received polio vaccine during supplementary immunization activities (SIAs) conducted since mid-2012 because local authorities have banned polio vaccination. In some other areas of Pakistan, SIAs have been compromised by attacks targeting polio workers that started in mid-2012. Further efforts to reach children in conflict-affected and security-compromised areas, including vaccinating at transit points and conducting additional short-interval-additional-dose (SIAD) SIAs as areas become accessible, will be necessary to prevent reintroduction of WPV into other areas of Pakistan and other parts of the world.

Alzheimer's as a Systems-Level Disease Involving the Interplay of Multiple Cellular Networks.

PubMed

Castrillo, Juan I; Oliver, Stephen G

2016-01-01

Alzheimer's disease (AD), and many neurodegenerative disorders, are multifactorial in nature. They involve a combination of genomic, epigenomic, interactomic and environmental factors. Progress is being made, and these complex diseases are beginning to be understood as having their origin in altered states of biological networks at the cellular level. In the case of AD, genomic susceptibility and mechanisms leading to (or accompanying) the impairment of the central Amyloid Precursor Protein (APP) processing and tau networks are widely accepted as major contributors to the diseased state. The derangement of these networks may result in both the gain and loss of functions, increased generation of toxic species (e.g., toxic soluble oligomers and aggregates) and imbalances, whose effects can propagate to supra-cellular levels. Although well sustained by empirical data and widely accepted, this global perspective often overlooks the essential roles played by the main counteracting homeostatic networks (e.g., protein quality control/proteostasis, unfolded protein response, protein folding chaperone networks, disaggregases, ER-associated degradation/ubiquitin proteasome system, endolysosomal network, autophagy, and other stress-protective and clearance networks), whose relevance to AD is just beginning to be fully realized. In this chapter, an integrative perspective is presented. Alzheimer's disease is characterized to be a result of: (a) intrinsic genomic/epigenomic susceptibility and, (b) a continued dynamic interplay between the deranged networks and the central homeostatic networks of nerve cells. This interplay of networks will underlie both the onset and rate of progression of the disease in each individual. Integrative Systems Biology approaches are required to effect its elucidation. Comprehensive Systems Biology experiments at different 'omics levels in simple model organisms, engineered to recapitulate the basic features of AD may illuminate the onset and sequence of events underlying AD. Indeed, studies of models of AD in simple organisms, differentiated cells in culture and rodents are beginning to offer hope that the onset and progression of AD, if detected at an early stage, may be stopped, delayed, or even reversed, by activating or modulating networks involved in proteostasis and the clearance of toxic species. In practice, the incorporation of next-generation neuroimaging, high-throughput and computational approaches are opening the way towards early diagnosis well before irreversible cell death. Thus, the presence or co-occurrence of: (a) accumulation of toxic Aβ oligomers and tau species; (b) altered splicing and transcriptome patterns; (c) impaired redox, proteostatic, and metabolic networks together with, (d) compromised homeostatic capacities may constitute relevant 'AD hallmarks at the cellular level' towards reliable and early diagnosis. From here, preventive lifestyle changes and tailored therapies may be investigated, such as combined strategies aimed at both lowering the production of toxic species and potentiating homeostatic responses, in order to prevent or delay the onset, and arrest, alleviate, or even reverse the progression of the disease.

Human Milk for Ill and Medically Compromised Infants: Strategies and Ongoing Innovation.

PubMed

DiLauro, Sara; Unger, Sharon; Stone, Debbie; O'Connor, Deborah L

2016-08-01

The use of human milk (mother's own milk and/or donor milk) in ill or medically compromised infants frequently requires some adaptation to address medical diagnoses and/or altered nutrition requirements. This tutorial describes the nutrition and immunological benefits of breast milk as well as provides evidence for the use of donor milk when mother's own milk is unavailable. Several strategies used to modify human milk to meet the medical and nutrition needs of an ill or medically compromised infant are reviewed. These strategies include (1) the standard fortification of human milk to support adequate growth, (2) the novel concept of target fortification in preterm infants, (3) instructions on how to alter maternal diet to address cow's milk protein intolerance and/or allergy in breast milk-fed infants, and (4) the removal and modification of the fat in breast milk used in infants diagnosed with chylothorax. © 2016 American Society for Parenteral and Enteral Nutrition.

Skeletal muscle contractile properties in a novel murine model for limb girdle muscular dystrophy 2i.

PubMed

Rehwaldt, Jordan D; Rodgers, Buel D; Lin, David C

2017-12-01

Limb-girdle muscular dystrophy (LGMD) 2i results from mutations in fukutin-related protein and aberrant α-dystroglycan glycosylation. Although this significantly compromises muscle function and ambulation, the comprehensive characteristics of contractile dysfunction are unknown. Therefore, we quantified the in situ contractile properties of the medial gastrocnemius in young adult P448L mice, an affected muscle of a novel model of LGMD2i. Normalized maximal twitch force, tetanic force, and power were significantly smaller in P448L mice, compared with sex-matched, wild-type mice. These differences were consistent with the replacement of contractile fibers by passive tissue. The shape of the active force-length relationships were similar in both groups, regardless of sex, consistent with an intact sarcomeric structure in P448L mice. Passive force-length curves normalized to maximal isometric force were steeper in P448L mice, and passive elements contribute disproportionately more to total contractile force in P448L mice. Sex differences were mostly noted in the force-velocity curves, as normalized values for maximal and optimal velocities were significantly slower in P448L males, compared with wild-type, but not in P448L females. This suggests that the dystrophic phenotype, which may include possible changes in cross-bridge kinetics and fiber-type proportions, progresses more quickly in P448L males. These results together indicate that active force and power generation are compromised in both sexes of P448L mice, while passive forces increase. More importantly, the results identified several functional markers of disease pathophysiology that could aid in developing and assessment of novel therapeutics for LGMD2i and possibly other dystroglycanopathies as well. NEW & NOTEWORTHY Comprehensive assessments of muscle contractile function have, until now, never been performed in an animal model for any dystroglycanopathy. This study suggests that skeletal muscle contractile properties are significantly compromised in a recently developed model for limb-girdle muscular dystrophy 2i, the P448L mouse. It further identifies novel pathological markers of muscle function that are suitable for developing therapeutics and for better understanding of disease pathogenesis.

Mechanism of metabolic stroke and spontaneous cerebral hemorrhage in glutaric aciduria type I

PubMed Central

2014-01-01

Background Metabolic stroke is the rapid onset of lasting central neurological deficit associated with decompensation of an underlying metabolic disorder. Glutaric aciduria type I (GA1) is an inherited disorder of lysine and tryptophan metabolism presenting with metabolic stroke in infancy. The clinical presentation includes bilateral striatal necrosis and spontaneous subdural and retinal hemorrhages, which has been frequently misdiagnosed as non-accidental head trauma. The mechanisms underlying metabolic stroke and spontaneous hemorrhage are poorly understood. Results Using a mouse model of GA1, we show that metabolic stroke progresses in the opposite sequence of ischemic stroke, with initial neuronal swelling and vacuole formation leading to cerebral capillary occlusion. Focal regions of cortical followed by striatal capillaries are occluded with shunting to larger non-exchange vessels leading to early filling and dilation of deep cerebral veins. Blood–brain barrier breakdown was associated with displacement of tight-junction protein Occludin. Conclusion Together the current findings illuminate the pathophysiology of metabolic stroke and vascular compromise in GA1, which may translate to other neurometabolic disorders presenting with stroke. PMID:24468193

Mechanism of metabolic stroke and spontaneous cerebral hemorrhage in glutaric aciduria type I.

PubMed

Zinnanti, William J; Lazovic, Jelena; Housman, Cathy; Antonetti, David A; Koeller, David M; Connor, James R; Steinman, Lawrence

2014-01-27

Metabolic stroke is the rapid onset of lasting central neurological deficit associated with decompensation of an underlying metabolic disorder. Glutaric aciduria type I (GA1) is an inherited disorder of lysine and tryptophan metabolism presenting with metabolic stroke in infancy. The clinical presentation includes bilateral striatal necrosis and spontaneous subdural and retinal hemorrhages, which has been frequently misdiagnosed as non-accidental head trauma. The mechanisms underlying metabolic stroke and spontaneous hemorrhage are poorly understood. Using a mouse model of GA1, we show that metabolic stroke progresses in the opposite sequence of ischemic stroke, with initial neuronal swelling and vacuole formation leading to cerebral capillary occlusion. Focal regions of cortical followed by striatal capillaries are occluded with shunting to larger non-exchange vessels leading to early filling and dilation of deep cerebral veins. Blood-brain barrier breakdown was associated with displacement of tight-junction protein Occludin. Together the current findings illuminate the pathophysiology of metabolic stroke and vascular compromise in GA1, which may translate to other neurometabolic disorders presenting with stroke.

Muscle aging is associated with compromised Ca2+ spark signaling and segregated intracellular Ca2+ release

PubMed Central

Weisleder, Noah; Brotto, Marco; Komazaki, Shinji; Pan, Zui; Zhao, Xiaoli; Nosek, Thomas; Parness, Jerome; Takeshima, Hiroshi; Ma, Jianjie

2006-01-01

Reduced homeostatic capacity for intracellular Ca2+ ([Ca2+]i) movement may underlie the progression of sarcopenia and contractile dysfunction during muscle aging. We report two alterations to Ca2+ homeostasis in skeletal muscle that are associated with aging. Ca2+ sparks, which are the elemental units of Ca2+ release from sarcoplasmic reticulum, are silent under resting conditions in young muscle, yet activate in a dynamic manner upon deformation of membrane structures. The dynamic nature of Ca2+ sparks appears to be lost in aged skeletal muscle. Using repetitive voltage stimulation on isolated muscle preparations, we identify a segregated [Ca2+]i reserve that uncouples from the normal excitation–contraction process in aged skeletal muscle. Similar phenotypes are observed in adolescent muscle null for a synaptophysin-family protein named mitsugumin-29 (MG29) that is involved in maintenance of muscle membrane ultrastructure and Ca2+ signaling. This finding, coupled with decreased expression of MG29 in aged skeletal muscle, suggests that MG29 expression is important in maintaining skeletal muscle Ca2+ homeostasis during aging. PMID:16943181

Oxidative Stress and Antioxidant System in Periodontitis

PubMed Central

Wang, Yue; Andrukhov, Oleh; Rausch-Fan, Xiaohui

2017-01-01

Periodontitis is a common inflammatory disease, which is initiated by bacterial infection and subsequently progressed by aberrant host response. It can result in the destruction of teeth supporting tissues and have an influence on systemic health. When periodontitis occurs, reactive oxygen species, which are overproduced mostly by hyperactive neutrophils, could not be balanced by antioxidant defense system and cause tissues damage. This is characterized by increased metabolites of lipid peroxidation, DNA damage and protein damage. Local and systemic activities of antioxidants can also be influenced by periodontitis. Total antioxidant capacity, total oxidant status and oxidative stress index have been used to evaluate the oxidative stress associated with periodontitis. Studies have confirmed that inflammatory response in periodontitis is associated with an increased local and systemic oxidative stress and compromised antioxidant capacity. Our review focuses on increased oxidative stress in periodontal disease, specifically, on the relationship between the local and systemic biomarkers of oxidative stress and periodontitis and their association with the pathogenesis of periodontitis. Also, the relationship between periodontitis and systemic inflammation, and the effects of periodontal therapy on oxidative stress parameters will be discussed. PMID:29180965

Mammalian RAD52 Functions in Break-Induced Replication Repair of Collapsed DNA Replication Forks.

PubMed

Sotiriou, Sotirios K; Kamileri, Irene; Lugli, Natalia; Evangelou, Konstantinos; Da-Ré, Caterina; Huber, Florian; Padayachy, Laura; Tardy, Sebastien; Nicati, Noemie L; Barriot, Samia; Ochs, Fena; Lukas, Claudia; Lukas, Jiri; Gorgoulis, Vassilis G; Scapozza, Leonardo; Halazonetis, Thanos D

2016-12-15

Human cancers are characterized by the presence of oncogene-induced DNA replication stress (DRS), making them dependent on repair pathways such as break-induced replication (BIR) for damaged DNA replication forks. To better understand BIR, we performed a targeted siRNA screen for genes whose depletion inhibited G1 to S phase progression when oncogenic cyclin E was overexpressed. RAD52, a gene dispensable for normal development in mice, was among the top hits. In cells in which fork collapse was induced by oncogenes or chemicals, the Rad52 protein localized to DRS foci. Depletion of Rad52 by siRNA or knockout of the gene by CRISPR/Cas9 compromised restart of collapsed forks and led to DNA damage in cells experiencing DRS. Furthermore, in cancer-prone, heterozygous APC mutant mice, homozygous deletion of the Rad52 gene suppressed tumor growth and prolonged lifespan. We therefore propose that mammalian RAD52 facilitates repair of collapsed DNA replication forks in cancer cells. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

PRP19 Transforms into a Sensor of RPA-ssDNA after DNA Damage and Drives ATR Activation via a Ubiquitin-Mediated Circuitry

PubMed Central

Maréchal, Alexandre; Wu, Ching-Shyi; Yazinski, Stephanie A.; Nguyen, Hai Dang; Liu, Shizhou; Jiménez, Amanda E.; Jin, Jianping; Zou, Lee

2014-01-01

Summary PRP19 is a ubiquitin ligase involved in pre-mRNA splicing and the DNA damage response (DDR). While the role for PRP19 in splicing is well characterized, its role in the DDR remains elusive. Through a proteomic screen for proteins that interact with RPA-coated single-stranded DNA (RPA-ssDNA), we identified PRP19 as a sensor of DNA damage. PRP19 binds RPA directly and localizes to DNA damage sites via RPA, promoting RPA ubiquitylation in a DNA damage-induced manner. PRP19 facilitates the accumulation of ATRIP, the regulatory partner of the ATR kinase, at DNA damage sites. Depletion of PRP19 compromised the phosphorylation of ATR substrates, the recovery of stalled replication forks, and the progression of replication forks on damaged DNA. Importantly, PRP19 mutants that cannot bind RPA or function as an E3 ligase failed to support the ATR response, revealing that PRP19 drives ATR activation by acting as an RPA-ssDNA-sensing ubiquitin ligase during the DDR. PMID:24332808

Triple helix-forming oligonucleotide corresponding to the polypyrimidine sequence in the rat alpha 1(I) collagen promoter specifically inhibits factor binding and transcription.

PubMed

Kovacs, A; Kandala, J C; Weber, K T; Guntaka, R V

1996-01-19

Type I and III fibrillar collagens are the major structural proteins of the extracellular matrix found in various organs including the myocardium. Abnormal and progressive accumulation of fibrillar type I collagen in the interstitial spaces compromises organ function and therefore, the study of transcriptional regulation of this gene and specific targeting of its expression is of major interest. Transient transfection of adult cardiac fibroblasts indicate that the polypurine-polypyrimidine sequence of alpha 1(I) collagen promoter between nucleotides - 200 and -140 represents an overall positive regulatory element. DNase I footprinting and electrophoretic mobility shift assays suggest that multiple factors bind to different elements of this promoter region. We further demonstrate that the unique polypyrimidine sequence between -172 and -138 of the promoter represents a suitable target for a single-stranded polypurine oligonucleotide (TFO) to form a triple helix DNA structure. Modified electrophoretic mobility shift assays show that this TFO specifically inhibits the protein-DNA interaction within the target region. In vitro transcription assays and transient transfection experiments demonstrate that the transcriptional activity of the promoter is inhibited by this oligonucleotide. We propose that TFOs represent a therapeutic potential to specifically influence the expression of alpha 1(I) collagen gene in various disease states where abnormal type I collagen accumulation is known to occur.

Targeting Protein Quality Control Mechanisms by Natural Products to Promote Healthy Ageing.

PubMed

Wedel, Sophia; Manola, Maria; Cavinato, Maria; Trougakos, Ioannis P; Jansen-Dürr, Pidder

2018-05-19

Organismal ageing is associated with increased chance of morbidity or mortality and it is driven by diverse molecular pathways that are affected by both environmental and genetic factors. The progression of ageing correlates with the gradual accumulation of stressors and damaged biomolecules due to the time-dependent decline of stress resistance and functional capacity, which eventually compromise cellular homeodynamics. As protein machines carry out the majority of cellular functions, proteome quality control is critical for cellular functionality and is carried out through the curating activity of the proteostasis network (PN). Key components of the PN are the two main degradation machineries, namely the ubiquitin-proteasome and autophagy-lysosome pathways along with several stress-responsive pathways, such as that of nuclear factor erythroid 2-related factor 2 (Nrf2), which mobilises cytoprotective genomic responses against oxidative and/or xenobiotic damage. Reportedly, genetic or dietary interventions that activate components of the PN delay ageing in evolutionarily diverse organisms. Natural products (extracts or pure compounds) represent an extraordinary inventory of highly diverse structural scaffolds that offer promising activities towards meeting the challenge of increasing healthspan and/or delaying ageing (e.g., spermidine, quercetin or sulforaphane). Herein, we review those natural compounds that have been found to activate proteostatic and/or anti-stress cellular responses and hence have the potential to delay cellular senescence and/or in vivo ageing.

Folate dietary insufficiency and folic acid supplementation similarly impair metabolism and compromise hematopoiesis

PubMed Central

Henry, Curtis J.; Nemkov, Travis; Casás-Selves, Matias; Bilousova, Ganna; Zaberezhnyy, Vadym; Higa, Kelly C.; Serkova, Natalie J.; Hansen, Kirk C.; D’Alessandro, Angelo; DeGregori, James

2017-01-01

While dietary folate deficiency is associated with increased risk for birth defects and other diseases, evidence suggests that supplementation with folic acid can contribute to predisposition to some diseases, including immune dysfunction and cancer. Herein, we show that diets supplemented with folic acid both below and above the recommended levels led to significantly altered metabolism in multiple tissues in mice. Surprisingly, both low and excessive dietary folate induced similar metabolic changes, which were particularly evident for nucleotide biosynthetic pathways in B-progenitor cells. Diet-induced metabolic changes in these cells partially phenocopied those observed in mice treated with anti-folate drugs, suggesting that both deficiency and excessive levels of dietary folic acid compromise folate-dependent biosynthetic pathways. Both folate deficiency and excessive dietary folate levels compromise hematopoiesis, resulting in defective cell cycle progression, persistent DNA damage, and impaired production of lymphocytes. These defects reduce the reconstitution potential in transplantation settings and increase radiation-induced mortality. We conclude that excessive folic acid supplementation can metabolically mimic dietary folate insufficiency, leading to similar functional impairment of hematopoiesis. PMID:28883079

The Regulation of Skeletal Muscle Protein Turnover during the Progression of Cancer Cachexia in the ApcMin/+ Mouse

PubMed Central

White, James P.; Baynes, John W.; Welle, Stephen L.; Kostek, Matthew C.; Matesic, Lydia E.; Sato, Shuichi; Carson, James A.

2011-01-01

Muscle wasting that occurs with cancer cachexia is caused by an imbalance in the rates of muscle protein synthesis and degradation. The ApcMin/+ mouse is a model of colorectal cancer that develops cachexia that is dependent on circulating IL-6. However, the IL-6 regulation of muscle protein turnover during the initiation and progression of cachexia in the ApcMin/+ mouse is not known. Cachexia progression was studied in ApcMin/+ mice that were either weight stable (WS) or had initial (≤5%), intermediate (6–19%), or extreme (≥20%) body weight loss. The initiation of cachexia reduced %MPS 19% and a further ∼50% with additional weight loss. Muscle IGF-1 mRNA expression and mTOR targets were suppressed with the progression of body weight loss, while muscle AMPK phosphorylation (Thr 172), AMPK activity, and raptor phosphorylation (Ser 792) were not increased with the initiation of weight loss, but were induced as cachexia progressed. ATP dependent protein degradation increased during the initiation and progression of cachexia. However, ATP independent protein degradation was not increased until cachexia had progressed beyond the initial phase. IL-6 receptor antibody administration prevented body weight loss and suppressed muscle protein degradation, without any effect on muscle %MPS or IGF-1 associated signaling. In summary, the %MPS reduction during the initiation of cachexia is associated with IGF-1/mTOR signaling repression, while muscle AMPK activation and activation of ATP independent protein degradation occur later in the progression of cachexia. IL-6 receptor antibody treatment blocked cachexia progression through the suppression of muscle protein degradation, while not rescuing the suppression of muscle protein synthesis. Attenuation of IL-6 signaling was effective in blocking the progression of cachexia, but not sufficient to reverse the process. PMID:21949739

E3 Ubiquitin Ligase CHIP and NBR1-Mediated Selective Autophagy Protect Additively against Proteotoxicity in Plant Stress Responses

PubMed Central

Qi, Jingxia; Chi, Yingjin; Fan, Baofang; Yu, Jing-Quan; Chen, Zhixiang

2014-01-01

Plant stress responses require both protective measures that reduce or restore stress-inflicted damage to cellular structures and mechanisms that efficiently remove damaged and toxic macromolecules, such as misfolded and damaged proteins. We have recently reported that NBR1, the first identified plant autophagy adaptor with a ubiquitin-association domain, plays a critical role in plant stress tolerance by targeting stress-induced, ubiquitinated protein aggregates for degradation by autophagy. Here we report a comprehensive genetic analysis of CHIP, a chaperone-associated E3 ubiquitin ligase from Arabidopsis thaliana implicated in mediating degradation of nonnative proteins by 26S proteasomes. We isolated two chip knockout mutants and discovered that they had the same phenotypes as the nbr1 mutants with compromised tolerance to heat, oxidative and salt stresses and increased accumulation of insoluble proteins under heat stress. To determine their functional interactions, we generated chip nbr1 double mutants and found them to be further compromised in stress tolerance and in clearance of stress-induced protein aggregates, indicating additive roles of CHIP and NBR1. Furthermore, stress-induced protein aggregates were still ubiquitinated in the chip mutants. Through proteomic profiling, we systemically identified heat-induced protein aggregates in the chip and nbr1 single and double mutants. These experiments revealed that highly aggregate-prone proteins such as Rubisco activase and catalases preferentially accumulated in the nbr1 mutant while a number of light-harvesting complex proteins accumulated at high levels in the chip mutant after a relatively short period of heat stress. With extended heat stress, aggregates for a large number of intracellular proteins accumulated in both chip and nbr1 mutants and, to a greater extent, in the chip nbr1 double mutant. Based on these results, we propose that CHIP and NBR1 mediate two distinct but complementary anti-proteotoxic pathways and protein's propensity to aggregate under stress conditions is one of the critical factors for pathway selection of protein degradation. PMID:24497840

Human amyloidogenic light chain proteins result in cardiac dysfunction, cell death, and early mortality in zebrafish.

PubMed

Mishra, Shikha; Guan, Jian; Plovie, Eva; Seldin, David C; Connors, Lawreen H; Merlini, Giampaolo; Falk, Rodney H; MacRae, Calum A; Liao, Ronglih

2013-07-01

Systemic amyloid light-chain (AL) amyloidosis is associated with rapidly progressive and fatal cardiomyopathy resulting from the direct cardiotoxic effects of circulating AL light chain (AL-LC) proteins and the indirect effects of AL fibril tissue infiltration. Cardiac amyloidosis is resistant to standard heart failure therapies, and, to date, there are limited treatment options for these patients. The mechanisms underlying the development of cardiac amyloidosis and AL-LC cardiotoxicity are largely unknown, and their study has been limited by the lack of a suitable in vivo model system. Here, we establish an in vivo zebrafish model of human AL-LC-induced cardiotoxicity. AL-LC isolated from AL cardiomyopathy patients or control nonamyloidogenic LC protein isolated from multiple myeloma patients (Con-LC) was directly injected into the circulation of zebrafish at 48 h postfertilization. AL-LC injection resulted in impaired cardiac function, pericardial edema, and increased cell death relative to Con-LC, culminating in compromised survival with 100% mortality within 2 wk, independent of AL fibril deposition. Prior work has implicated noncanonical p38 MAPK activation in the pathogenesis of AL-LC-induced cardiotoxicity, and p38 MAPK inhibition via SB-203580 rescued AL-LC-induced cardiac dysfunction and cell death and attenuated mortality in zebrafish. This in vivo zebrafish model of AL-LC cardiotoxicity demonstrates that antagonism of p38 MAPK within the AL-LC cardiotoxic signaling response may serve to improve cardiac function and mortality in AL cardiomyopathy. Furthermore, this in vivo model system will allow for further study of the molecular underpinnings of AL cardiotoxicity and identification of novel therapeutic strategies.

BRDT is an essential epigenetic regulator for proper chromatin organization, silencing of sex chromosomes and crossover formation in male meiosis

PubMed Central

Oh, Min Young; Garyn, Corey

2018-01-01

The double bromodomain and extra-terminal domain (BET) proteins are critical epigenetic readers that bind to acetylated histones in chromatin and regulate transcriptional activity and modulate changes in chromatin structure and organization. The testis-specific BET member, BRDT, is essential for the normal progression of spermatogenesis as mutations in the Brdt gene result in complete male sterility. Although BRDT is expressed in both spermatocytes and spermatids, loss of the first bromodomain of BRDT leads to severe defects in spermiogenesis without overtly compromising meiosis. In contrast, complete loss of BRDT blocks the progression of spermatocytes into the first meiotic division, resulting in a complete absence of post-meiotic cells. Although BRDT has been implicated in chromatin remodeling and mRNA processing during spermiogenesis, little is known about its role in meiotic processes. Here we report that BRDT is an essential regulator of chromatin organization and reprograming during prophase I of meiosis. Loss of BRDT function disrupts the epigenetic state of the meiotic sex chromosome inactivation in spermatocytes, affecting the synapsis and silencing of the X and Y chromosomes. We also found that BRDT controls the global chromatin organization and histone modifications of the chromatin attached to the synaptonemal complex. Furthermore, the homeostasis of crossover formation and localization during pachynema was altered, underlining a possible epigenetic mechanism by which crossovers are regulated and differentially established in mammalian male genomes. Our observations reveal novel findings about the function of BRDT in meiosis and provide insight into how epigenetic regulators modulate the progression of male mammalian meiosis and the formation of haploid gametes. PMID:29513658

Complementary Kinesiology: Why It Is Not Wise to Choose Sides or Work Alone

ERIC Educational Resources Information Center

Kretchmar, Scott

2014-01-01

In this essay I argue in favor of a holistic vision for our field under the heading of complementary kinesiology. I argue that battles over reified dichotomies and even compromise solutions have impeded our progress as a profession. I describe the theory of complementation as an alternative. I say it is a strange and paradoxical way of…

Response to Industry Canada's Consultation Paper "Seizing Canada's Moment: Moving Forward in Science, Technology and Innovation"

ERIC Educational Resources Information Center

Peters, Wayne D.; Turk, James L.

2014-01-01

According to these authors, Canada is in need of a new science policy and strategy. The current direction of the federal government is threatening to impede scientific progress and compromise the integrity and independence of public science. This is reflected in the government's waning commitment to funding basic research; its attempts to steer…

Teacher Absence as a Factor in Gender Inequalities in Access to Primary Schooling in Rural Pakistan

ERIC Educational Resources Information Center

Ghuman, Sharon; Lloyd, Cynthia

2010-01-01

The presence of a teacher in the classroom is central to the provision of schooling, with accumulating evidence showing that teacher absence compromises student learning. Teacher absence is common in schools in low- and middle-income countries. With much of the developing world making rapid progress in achieving universal primary school enrollment…

NCOA1 promotes angiogenesis in breast tumors by simultaneously enhancing both HIF1α- and AP-1-mediated VEGFa transcription

PubMed Central

Qin, Li; Xu, Yan; Xu, Yixiang; Ma, Gang; Liao, Lan; Wu, Yelin; Li, Yi; Wang, Xian; Wang, Xiaosong; Jiang, Jun; Wang, Jin; Xu, Jianming

2015-01-01

Nuclear receptor coactivator 1 (NCOA1) is overexpressed in a subset of breast cancer and its increased expression positively correlates with disease recurrence and metastasis. Although NCOA1 is known to promote breast cancer metastasis through working with multiple transcription factors to upregulate the expression of Twist1, ITGA5, CSF-1, SDF1 and CXCR4, the role of NCOA1 in breast tumor angiogenesis has not been investigated. In this study, we found that the microvascular density (MVD) was significantly decreased and increased in Ncoa1-knockout and NCOA1-overexpressing mammary tumors, respectively, in several breast cancer mouse models. Knockout or knockdown of NCOA1 in breast cancer cell lines also markedly compromised their capability to induce angiogenesis in Matrigel plugs embedded subcutaneously in mice, while this compromised capability could be rescued by VEGFa treatment. At the molecular level, NCOA1 upregulates VEGFa expression in both mouse mammary tumors and cultured breast cancer cells, and it does so by associating with both c-Fos, which is recruited to the AP-1 site at bp −938 of the VEGFa promoter, and HIF1α, which is recruited to the HIF1α-binding element at bp −979 of the VEGFa promoter, to enhance VEGFa transcription. In 140 human breast tumors, high NCOA1 protein correlates with high MVD and patients with both high NCOA1 and high MVD showed significantly shorter survival time. In summary, this study revealed a novel mechanism that NCOA1 potentiates breast cancer angiogenesis through upregulating HIF1α and AP-1-mediated VEGFa expression, which reinforces the rational of targeting NCOA1 in controlling breast cancer progression and metastasis. PMID:26287601

ESCMID Study Group for Infections in Compromised Hosts (ESGICH) Consensus Document on the safety of targeted and biological therapies: an infectious diseases perspective (Intracellular signaling pathways: tyrosine kinase and mTOR inhibitors).

PubMed

Reinwald, M; Silva, J T; Mueller, N J; Fortún, J; Garzoni, C; de Fijter, J W; Fernández-Ruiz, M; Grossi, P; Aguado, J M

2018-06-01

The present review is part of the European Society of Clinical Microbiology and Infectious Diseases (ESCMID) Study Group for Infections in Compromised Hosts (ESGICH) Consensus Document on the safety of targeted and biologic therapies. To review, from an infectious diseases perspective, the safety profile of therapies targeting different intracellular signaling pathways and to suggest preventive recommendations. Computer-based Medline searches with MeSH terms pertaining to each agent or therapeutic family. Although BCR-ABL tyrosine kinase inhibitors modestly increase the overall risk of infection, dasatinib has been associated with cytomegalovirus and hepatitis B virus reactivation. BRAF/MEK kinase inhibitors do not significantly affect infection susceptibility. The effect of Bruton tyrosine kinase inhibitors (ibrutinib) among patients with B-cell malignancies is difficult to distinguish from that of previous immunosuppression. However, cases of Pneumocystis jirovecii pneumonia (PCP), invasive fungal infection and progressive multifocal leukoencephalopathy have been occasionally reported. Because phosphatidylinositol-3-kinase inhibitors (idelalisib) may predispose to opportunistic infections, anti-Pneumocystis prophylaxis and prevention strategies for cytomegalovirus are recommended. No increased rates of infection have been observed with venetoclax (antiapoptotic protein Bcl-2 inhibitor). Therapy with Janus kinase inhibitors markedly increases the incidence of infection. Pretreatment screening for chronic hepatitis B virus and latent tuberculosis infection must be performed, and anti-Pneumocystis prophylaxis should be considered for patients with additional risk factors. Cancer patients receiving mTOR inhibitors face an increased incidence of overall infection, especially those with additional risk factors (prior therapies or delayed wound healing). Specific preventive approaches are warranted in view of the increased risk of infection associated with some of the reviewed agents. Copyright © 2018. Published by Elsevier Ltd.

Conversion of amino-acid sequence in proteins to classical music: search for auditory patterns

PubMed Central

2007-01-01

We have converted genome-encoded protein sequences into musical notes to reveal auditory patterns without compromising musicality. We derived a reduced range of 13 base notes by pairing similar amino acids and distinguishing them using variations of three-note chords and codon distribution to dictate rhythm. The conversion will help make genomic coding sequences more approachable for the general public, young children, and vision-impaired scientists. PMID:17477882

Staurosporine Induces Necroptotic Cell Death under Caspase-Compromised Conditions in U937 Cells

PubMed Central

Dunai, Zsuzsanna A.; Imre, Gergely; Barna, Gabor; Korcsmaros, Tamas; Petak, Istvan; Bauer, Pal I.; Mihalik, Rudolf

2012-01-01

For a long time necrosis was thought to be an uncontrolled process but evidences recently have revealed that necrosis can also occur in a regulated manner. Necroptosis, a type of programmed necrosis is defined as a death receptor-initiated process under caspase-compromised conditions. The process requires the kinase activity of receptor-interacting protein kinase 1 and 3 (RIPK1 and RIPK3) and mixed lineage kinase domain-like protein (MLKL), as a substrate of RIPK3. The further downstream events remain elusive. We applied known inhibitors to characterize the contributing enzymes in necroptosis and their effect on cell viability and different cellular functions were detected mainly by flow cytometry. Here we report that staurosporine, the classical inducer of intrinsic apoptotic pathway can induce necroptosis under caspase-compromised conditions in U937 cell line. This process could be hampered at least partially by the RIPK1 inhibitor necrotstin-1 and by the heat shock protein 90 kDa inhibitor geldanamycin. Moreover both the staurosporine-triggered and the classical death ligand-induced necroptotic pathway can be effectively arrested by a lysosomal enzyme inhibitor CA-074-OMe and the recently discovered MLKL inhibitor necrosulfonamide. We also confirmed that the enzymatic role of poly(ADP-ribose)polymerase (PARP) is dispensable in necroptosis but it contributes to membrane disruption in secondary necrosis. In conclusion, we identified a novel way of necroptosis induction that can facilitate our understanding of the molecular mechanisms of necroptosis. Our results shed light on alternative application of staurosporine, as a possible anticancer therapeutic agent. Furthermore, we showed that the CA-074-OMe has a target in the signaling pathway leading to necroptosis. Finally, we could differentiate necroptotic and secondary necrotic processes based on participation of PARP enzyme. PMID:22860037

Compromised Mitochondrial Fatty Acid Synthesis in Transgenic Mice Results in Defective Protein Lipoylation and Energy Disequilibrium

PubMed Central

Smith, Stuart; Witkowski, Andrzej; Moghul, Ayesha; Yoshinaga, Yuko; Nefedov, Michael; de Jong, Pieter; Feng, Dejiang; Fong, Loren; Tu, Yiping; Hu, Yan; Young, Stephen G.; Pham, Thomas; Cheung, Carling; Katzman, Shana M.; Brand, Martin D.; Quinlan, Casey L.; Fens, Marcel; Kuypers, Frans; Misquitta, Stephanie; Griffey, Stephen M.; Tran, Son; Gharib, Afshin; Knudsen, Jens; Hannibal-Bach, Hans Kristian; Wang, Grace; Larkin, Sandra; Thweatt, Jennifer; Pasta, Saloni

2012-01-01

A mouse model with compromised mitochondrial fatty acid synthesis has been engineered in order to assess the role of this pathway in mitochondrial function and overall health. Reduction in the expression of mitochondrial malonyl CoA-acyl carrier protein transacylase, a key enzyme in the pathway encoded by the nuclear Mcat gene, was achieved to varying extents in all examined tissues employing tamoxifen-inducible Cre-lox technology. Although affected mice consumed more food than control animals, they failed to gain weight, were less physically active, suffered from loss of white adipose tissue, reduced muscle strength, kyphosis, alopecia, hypothermia and shortened lifespan. The Mcat-deficient phenotype is attributed primarily to reduced synthesis, in several tissues, of the octanoyl precursors required for the posttranslational lipoylation of pyruvate and α-ketoglutarate dehydrogenase complexes, resulting in diminished capacity of the citric acid cycle and disruption of energy metabolism. The presence of an alternative lipoylation pathway that utilizes exogenous free lipoate appears restricted to liver and alone is insufficient for preservation of normal energy metabolism. Thus, de novo synthesis of precursors for the protein lipoylation pathway plays a vital role in maintenance of mitochondrial function and overall vigor. PMID:23077570

The plant-specific transcription factors CBP60g and SARD1 are targeted by a Verticillium secretory protein VdSCP41 to modulate immunity

PubMed Central

Qin, Jun; Wang, Kailun; Sun, Lifan; Xing, Haiying; Wang, Sheng; Li, Lin; Chen, She

2018-01-01

The vascular pathogen Verticillium dahliae infects the roots of plants to cause Verticillium wilt. The molecular mechanisms underlying V. dahliae virulence and host resistance remain elusive. Here, we demonstrate that a secretory protein, VdSCP41, functions as an intracellular effector that promotes V. dahliae virulence. The Arabidopsis master immune regulators CBP60g and SARD1 and cotton GhCBP60b are targeted by VdSCP41. VdSCP41 binds the C-terminal portion of CBP60g to inhibit its transcription factor activity. Further analyses reveal a transcription activation domain within CBP60g that is required for VdSCP41 targeting. Mutations in both CBP60g and SARD1 compromise Arabidopsis resistance against V. dahliae and partially impair VdSCP41-mediated virulence. Moreover, virus-induced silencing of GhCBP60b compromises cotton resistance to V. dahliae. This work uncovers a virulence strategy in which the V. dahliae secretory protein VdSCP41 directly targets plant transcription factors to inhibit immunity, and reveals CBP60g, SARD1 and GhCBP60b as crucial components governing V. dahliae resistance. PMID:29757140

Pakistan and the Millennium Development Goals for Maternal and Child Health: progress and the way forward.

PubMed

Rizvi, Arjumand; Bhatti, Zaid; Das, Jai K; Bhutta, Zulfiqar A

2015-01-01

The world has made substantial progress in reducing maternal and child mortality, but many countries are projected to fall short of achieving their Millennium Development Goals (MDGs) 4 and 5 targets. The major objective of this paper is to examine progress in Pakistan in reducing maternal and child mortality and malnutrition over the last two decades. Data from recent national and international surveys suggest that Pakistan lags behind on all of its MDGs related to maternal and child health and, for some indicators especially related to nutrition, the situation has worsened from the baseline of 1990. Progress in addressing key social determinants such as poverty, female education and empowerment has also been slow and unregulated population growth has further compromised progress. There is a need to integrate the various different sectors and programmes to achieve the desired results effectively and efficiently as many of the determinants and influencing factors are outside the health sector.

Identification of immunodominant proteins from Mannheimia haemolytica and Histophilus somni by an immunoproteomic approach.

PubMed

Alvarez, Angel H; Gutiérrez-Ortega, Abel; Hernández-Gutiérrez, Rodolfo

2015-10-01

Mannheimia haemolytica and Histophilus somni are frequently isolated from diseased cattle with bovine respiratory disease (BRD). They compromise animal lung function and the immune responses generated are not sufficient to limit infection. Identification of specific immunogenic antigens for vaccine development represents a great challenge. Immunogenic proteins were identified by immunoproteomic approach with sera from cattle immunized with a commercial cellular vaccine of M. haemolytica and H. somni. Proteins of M. haemolytica were identified as solute ABC transporter, iron-binding protein, and hypothetical protein of capsular biosynthesis. Histophilus somni proteins correspond to porin, amino acid ABC transporter, hypothetical outer membrane protein, cysteine synthase, and outer membrane protein P6. Although these antigens share strong similarities with other proteins from animal pathogens, the ABC system proteins have been associated with virulence and these proteins could be considered as potential vaccine candidates for BRD.

Identification of immunodominant proteins from Mannheimia haemolytica and Histophilus somni by an immunoproteomic approach

PubMed Central

Alvarez, Angel H.; Gutiérrez-Ortega, Abel; Hernández-Gutiérrez, Rodolfo

2015-01-01

Mannheimia haemolytica and Histophilus somni are frequently isolated from diseased cattle with bovine respiratory disease (BRD). They compromise animal lung function and the immune responses generated are not sufficient to limit infection. Identification of specific immunogenic antigens for vaccine development represents a great challenge. Immunogenic proteins were identified by immunoproteomic approach with sera from cattle immunized with a commercial cellular vaccine of M. haemolytica and H. somni. Proteins of M. haemolytica were identified as solute ABC transporter, iron-binding protein, and hypothetical protein of capsular biosynthesis. Histophilus somni proteins correspond to porin, amino acid ABC transporter, hypothetical outer membrane protein, cysteine synthase, and outer membrane protein P6. Although these antigens share strong similarities with other proteins from animal pathogens, the ABC system proteins have been associated with virulence and these proteins could be considered as potential vaccine candidates for BRD. PMID:26424916

Early detection of health and welfare compromises through automated detection of behavioural changes in pigs.

PubMed

Matthews, Stephen G; Miller, Amy L; Clapp, James; Plötz, Thomas; Kyriazakis, Ilias

2016-11-01

Early detection of health and welfare compromises in commercial piggeries is essential for timely intervention to enhance treatment success, reduce impact on welfare, and promote sustainable pig production. Behavioural changes that precede or accompany subclinical and clinical signs may have diagnostic value. Often referred to as sickness behaviour, this encompasses changes in feeding, drinking, and elimination behaviours, social behaviours, and locomotion and posture. Such subtle changes in behaviour are not easy to quantify and require lengthy observation input by staff, which is impractical on a commercial scale. Automated early-warning systems may provide an alternative by objectively measuring behaviour with sensors to automatically monitor and detect behavioural changes. This paper aims to: (1) review the quantifiable changes in behaviours with potential diagnostic value; (2) subsequently identify available sensors for measuring behaviours; and (3) describe the progress towards automating monitoring and detection, which may allow such behavioural changes to be captured, measured, and interpreted and thus lead to automation in commercial, housed piggeries. Multiple sensor modalities are available for automatic measurement and monitoring of behaviour, which require humans to actively identify behavioural changes. This has been demonstrated for the detection of small deviations in diurnal drinking, deviations in feeding behaviour, monitoring coughs and vocalisation, and monitoring thermal comfort, but not social behaviour. However, current progress is in the early stages of developing fully automated detection systems that do not require humans to identify behavioural changes; e.g., through automated alerts sent to mobile phones. Challenges for achieving automation are multifaceted and trade-offs are considered between health, welfare, and costs, between analysis of individuals and groups, and between generic and compromise-specific behaviours. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Profiling global kinome signatures of the radioresistant MCF-7/C6 breast cancer cells using MRM-based targeted proteomics.

PubMed

Guo, Lei; Xiao, Yongsheng; Fan, Ming; Li, Jian Jian; Wang, Yinsheng

2015-01-02

Ionizing radiation is widely used in cancer therapy; however, cancer cells often develop radioresistance, which compromises the efficacy of cancer radiation therapy. Quantitative assessment of the alteration of the entire kinome in radioresistant cancer cells relative to their radiosensitive counterparts may provide important knowledge to define the mechanism(s) underlying tumor adaptive radioresistance and uncover novel target(s) for effective prevention and treatment of tumor radioresistance. By employing a scheduled multiple-reaction monitoring analysis in conjunction with isotope-coded ATP affinity probes, we assessed the global kinome of radioresistant MCF-7/C6 cells and their parental MCF-7 human breast cancer cells. We rigorously quantified 120 kinases, of which (1)/3 exhibited significant differences in expression levels or ATP binding affinities. Several kinases involved in cell cycle progression and DNA damage response were found to be overexpressed or hyperactivated, including checkpoint kinase 1 (CHK1), cyclin-dependent kinases 1 and 2 (CDK1 and CDK2), and the catalytic subunit of DNA-dependent protein kinase. The elevated expression of CHK1, CDK1, and CDK2 in MCF-7/C6 cells was further validated by Western blot analysis. Thus, the altered kinome profile of radioresistant MCF-7/C6 cells suggests the involvement of kinases on cell cycle progression and DNA repair in tumor adaptive radioresistance. The unique kinome profiling results also afforded potential effective targets for resensitizing radioresistant cancer cells and counteracting deleterious effects of ionizing radiation exposure.

Postoperative Aortic Neck Dilation: Myth or Fact?

PubMed

Ribner, A S; Tassiopoulos, A K

2018-06-01

The abdominal aorta is the most common site of an aortic aneurysm. The visceral and most proximal infrarenal segment (aneurysm neck) are usually spared and considered more resistant to aneurysmal degeneration. However, if an abdominal aortic aneurysm (AAA) is left untreated, the natural history of the aortic neck is progressive dilatation and shortening. This may have significant implications for patients undergoing endovascular repair of AAAs (EVAR) as endograft stability and integrity of the repair are dependent on an intact proximal seal zone. Compromised seal zones, caused by progressive diameter enlargement and foreshortening of the aortic neck, may lead to distal endograft migration, type Ia endoleak, aortic sac repressurization, and, ultimately, aortic rupture.

[Engineering and expertise in sustainable development in hospitals].

PubMed

Barat, Pascal

2012-01-01

Sustainable development is nowadays a concept shared by most people even if it is sometimes hard to grasp. Combining good economic management, social progress and the protection of the environment is not always easy. It is however an essential exercise in order not to compromise the ability of future generations to meet their requirements. Tours University Hospital embarked on a sustainable development strategy in 2008.

The interleukin-20 receptor axis in early rheumatoid arthritis: novel links between disease-associated autoantibodies and radiographic progression.

PubMed

Kragstrup, Tue Wenzel; Greisen, Stinne Ravn; Nielsen, Morten Aagaard; Rhodes, Christopher; Stengaard-Pedersen, Kristian; Hetland, Merete Lund; Hørslev-Petersen, Kim; Junker, Peter; Østergaard, Mikkel; Hvid, Malene; Vorup-Jensen, Thomas; Robinson, William H; Sokolove, Jeremy; Deleuran, Bent

2016-03-11

Rheumatoid arthritis (RA) is often characterized by the presence of rheumatoid factor, anti-citrullinated protein antibodies, and bone erosions. Current therapies can compromise immunity, leading to risk of infection. The interleukin-20 receptor (IL-20R) axis comprising IL-19, IL-20, and IL-24 and their shared receptors activates tissue homeostasis processes but not the immune system. Consequently, modulation of the IL-20R axis may not lead to immunosuppression, making it an interesting drug target. We evaluated the role of the IL-20R axis in RA and associations between plasma cytokine levels and clinical disease. Plasma IL-19, IL-20, and IL-24 levels were measured in early RA patients during a treat-to-target strategy by enzyme-linked immunosorbent assays. The IL-20R1 and IL-22R1 levels in paired peripheral blood mononuclear cells and synovial fluid mononuclear cells from a different cohort of RA patients were evaluated by flow cytometry and confocal microscopy. Monocytes/macrophages were stimulated with heat-aggregated human immunoglobulin immune complexes and immune complexes containing citrullinated fibrinogen, and osteoclasts were incubated with IL-19, IL-20, and IL-24. The plasma concentrations of IL-20 and IL-24 (but not IL-19) were increased in early RA patients compared with healthy controls (both P < 0.002) and decreased after 6 months of treatment (both P < 0.0001). The expression of IL-22R1 (but not IL-20R1) was increased on monocytes from RA synovial fluid compared with monocytes from both RA and healthy control peripheral blood. The plasma concentrations of IL-20 and IL-24 were increased in rheumatoid factor and anti-citrullinated protein antibody positive compared with negative early RA patients (all P < 0.0001). Immune complexes stimulated the production of the IL-20R cytokines by monocytes/macrophages. Increased baseline plasma concentrations of IL-20 and IL-24 were associated with Sharp-van der Heijde score progression after 24 months (Spearman's rho = 0.19 and 0.26, both P < 0.05) in the early RA patients. The IL-22R1 was expressed by osteoclast precursors and in multinucleated osteoclasts. IL-20 and IL-24 increased the secretion of monocyte chemoattractant protein 1 by these cells. This study suggests that IL-20 and IL-24 link RA-associated autoantibodies with radiographic progression via the IL-22R1. Modulation of this axis holds promise as feasible anti-erosive treatment modalities in seropositive RA.

IDENTIFICATION OF PROTEINS INVOLVED IN TESTICULAR TOXICITY INDUCED BY HALOACID BY-PRODUCTS OF DRINKING WATER DISINFECTION

EPA Science Inventory

Dibromoacetic acid (DBA), a prevalent disinfection by-product in drinking water, perturbs spermiogenesis in adult rats suggesting that Sertoli-germ cell communication is compromised. When isolated seminiferous tubules from rats exposed to DBA in vivo were cultured, quantitative a...

Membrane protein separation and analysis by supercritical fluid chromatography-mass spectrometry.

PubMed

Zhang, Xu; Scalf, Mark; Westphall, Michael S; Smith, Lloyd M

2008-04-01

Membrane proteins comprise 25-30% of the human genome and play critical roles in a wide variety of important biological processes. However, their hydrophobic nature has compromised efforts at structural characterization by both X-ray crystallography and mass spectrometry. The detergents that are generally used to solubilize membrane proteins interfere with the crystallization process essential to X-ray studies and cause severe ion suppression effects that hinder mass spectrometric analysis. In this report, the use of supercritical fluid chromatography-mass spectrometry for the separation and analysis of integral membrane proteins and hydrophobic peptides is investigated. It is shown that detergents are rapidly and effectively separated from the proteins and peptides, yielding them in a state suitable for direct mass spectrometric analysis.

Models for evaluation of relative immunogenic potential of protein particles in biopharmaceutical protein formulations.

PubMed

Johnson, Richard; Jiskoot, Wim

2012-10-01

An immune response to a therapeutic protein that compromises the biopharmaceutical activity or cross-reacts with an endogenous protein is a serious clinical event. The role of protein aggregates and particles in biopharmaceutical formulations in mediating this immune response has gained considerable attention over the recent past. Model systems that could consistently and reliably predict the relative immunogenicity of biopharmaceutical protein formulations would be extremely valuable. Several approaches have been developed in an attempt to provide this insight, including in silico algorithms, in vitro tests utilizing human leukocytes and in vivo animal models. This commentary provides an update of these various approaches as well as the author's perspectives on the pros and cons of these different methods. Copyright © 2012 Wiley Periodicals, Inc.

Protein and glucogenic precursor supplementation: A nutritional strategy to increase reproductive and economic output

USDA-ARS?s Scientific Manuscript database

Reproductive performance in young beef cows is often compromised due to a mismatch of physiological demands and suboptimal environmental conditions. Studies conducted at the Corona Range and Livestock Research Center from 2000 to 2007 evaluated 3 postpartum supplement strategies that varied in the ...

[Effect of inhibitors serine/threonine protein kinases and protein phosphatases on mitosis progression of synchronized tobacco by-2 cells].

PubMed

Sheremet, Ia A; Emets, A I; Azmi, A; Vissenberg, K; Verbelen, J-P; Blium, Ia B

2012-01-01

In order to investigate the role of various serine/ threonine protein kinases and protein phosphatases in the regulation of mitosis progression in plant cells the influence of cyclin-dependent (olomoucine) and Ca2+ -calmodulin-dependent (W7) protein kinases inhibitors, as well as protein kinase C inhibitors (H7 and staurosporine) and protein phosphatases inhibitor (okadaic acid) on mitosis progression in synchronized tobacco BY-2 cells has been studied. It was found that BY-2 culture treatment with inhibitors of cyclin dependent protein kinases and protein kinase C causes prophase delay, reduces the mitotic index and displaces of mitotic peak as compare with control cells. Inhibition of Ca2+ -calmodulin dependent protein kinases enhances the cell entry into prophase and delays their exit from mitosis. Meanwhile inhibition of serine/threonine protein phosphatases insignificantly enhances of synchronized BY-2 cells entering into all phases of mitosis.

Sugar reduction without compromising sensory perception. An impossible dream?

PubMed

Hutchings, Scott C; Low, Julia Y Q; Keast, Russell S J

2018-03-21

Sugar reduction is a major technical challenge for the food industry to address in response to public health concerns regarding the amount of added sugars in foods. This paper reviews sweet taste perception, sensory methods to evaluate sugar reduction and the merits of different techniques available to reduce sugar content. The use of sugar substitutes (non-nutritive sweeteners, sugar alcohols, and fibres) can achieve the greatest magnitude of sugar and energy reduction, however bitter side tastes and varying temporal sweet profiles are common issues. The use of multisensory integration principles (particularly aroma) can be an effective approach to reduce sugar content, however the magnitude of sugar reduction is small. Innovation in food structure (modifying the sucrose distribution, serum release and fracture mechanics) offers a new way to reduce sugar without significant changes in food composition, however may be difficult to implement in food produced on a large scale. Gradual sugar reduction presents difficulties for food companies from a sales perspective if acceptability is compromised. Ultimately, a holistic approach where food manufacturers integrate a range of these techniques is likely to provide the best progress. However, substantial reduction of sugar in processed foods without compromising sensory properties may be an impossible dream.

Effect of polyethyleneglycol (PEG) chain length on the bio-nano-interactions between PEGylated lipid nanoparticles and biological fluids: from nanostructure to uptake in cancer cells

NASA Astrophysics Data System (ADS)

Pozzi, Daniela; Colapicchioni, Valentina; Caracciolo, Giulio; Piovesana, Susy; Capriotti, Anna Laura; Palchetti, Sara; de Grossi, Stefania; Riccioli, Anna; Amenitsch, Heinz; Laganà, Aldo

2014-02-01

When nanoparticles (NPs) enter a physiological environment, medium components compete for binding to the NP surface leading to formation of a rich protein shell known as the ``protein corona''. Unfortunately, opsonins are also adsorbed. These proteins are immediately recognized by the phagocyte system with rapid clearance of the NPs from the bloodstream. Polyethyleneglycol (PEG) coating of NPs (PEGylation) is the most efficient anti-opsonization strategy. Linear chains of PEG, grafted onto the NP surface, are able to create steric hindrance, resulting in a significant inhibition of protein adsorption and less recognition by macrophages. However, excessive PEGylation can lead to a strong inhibition of cellular uptake and less efficient binding with protein targets, reducing the potential of the delivery system. To reach a compromise in this regard we employed a multi-component (MC) lipid system with uncommon properties of cell uptake and endosomal escape and increasing length of PEG chains. Nano liquid chromatography coupled with tandem mass spectrometry (nanoLC-MS/MS) analysis allowed us to accurately determine the corona composition showing that apolipoproteins are the most abundant class in the corona and that increasing the PEG length reduced the protein adsorption and the liposomal surface affinity for apolipoproteins. Due to the abundance of apolipoproteins, we exploited the ``protein corona effect'' to deliver cationic liposome-human plasma complexes to human prostate cancer PC3 cells that express a high level of scavenger receptor class B type 1 in order to evaluate the cellular uptake efficiency of the systems used. Combining laser scanning confocal microscopy with flow cytometry analysis in PC3 cells we demonstrated that MC-PEG2k is the best compromise between an anti-opsonization strategy and active targeting and could be a promising candidate to treat prostate cancer in vivo.When nanoparticles (NPs) enter a physiological environment, medium components compete for binding to the NP surface leading to formation of a rich protein shell known as the ``protein corona''. Unfortunately, opsonins are also adsorbed. These proteins are immediately recognized by the phagocyte system with rapid clearance of the NPs from the bloodstream. Polyethyleneglycol (PEG) coating of NPs (PEGylation) is the most efficient anti-opsonization strategy. Linear chains of PEG, grafted onto the NP surface, are able to create steric hindrance, resulting in a significant inhibition of protein adsorption and less recognition by macrophages. However, excessive PEGylation can lead to a strong inhibition of cellular uptake and less efficient binding with protein targets, reducing the potential of the delivery system. To reach a compromise in this regard we employed a multi-component (MC) lipid system with uncommon properties of cell uptake and endosomal escape and increasing length of PEG chains. Nano liquid chromatography coupled with tandem mass spectrometry (nanoLC-MS/MS) analysis allowed us to accurately determine the corona composition showing that apolipoproteins are the most abundant class in the corona and that increasing the PEG length reduced the protein adsorption and the liposomal surface affinity for apolipoproteins. Due to the abundance of apolipoproteins, we exploited the ``protein corona effect'' to deliver cationic liposome-human plasma complexes to human prostate cancer PC3 cells that express a high level of scavenger receptor class B type 1 in order to evaluate the cellular uptake efficiency of the systems used. Combining laser scanning confocal microscopy with flow cytometry analysis in PC3 cells we demonstrated that MC-PEG2k is the best compromise between an anti-opsonization strategy and active targeting and could be a promising candidate to treat prostate cancer in vivo. Electronic supplementary information (ESI) available: Table S1. The slope of the lines fitting the temporal evolution of size and zeta-potential of MC, MC-PEG1k, MC-PEG2k and MC-PEG5k liposomes. Table S2. The full list of the most abundant corona proteins associated with MC, MC-PEG1k, MC-PEG2k and MC-PEG5k liposomes as identified by NanoLC-MS/MS. See DOI: 10.1039/c3nr05559k

Active photosynthetic inhibition mediated by MPK3/MPK6 is critical to effector-triggered immunity

PubMed Central

Su, Jianbin; Yang, Liuyi; Zhu, Qiankun; Wu, Hongjiao; He, Yi; Liu, Yidong; Xu, Juan; Jiang, Dean

2018-01-01

Extensive research revealed tremendous details about how plants sense pathogen effectors during effector-triggered immunity (ETI). However, less is known about downstream signaling events. In this report, we demonstrate that prolonged activation of MPK3 and MPK6, two Arabidopsis pathogen-responsive mitogen-activated protein kinases (MPKs), is essential to ETI mediated by both coiled coil-nucleotide binding site-leucine rich repeats (CNLs) and toll/interleukin-1 receptor nucleotide binding site-leucine rich repeats (TNLs) types of R proteins. MPK3/MPK6 activation rapidly alters the expression of photosynthesis-related genes and inhibits photosynthesis, which promotes the accumulation of superoxide (O2•−) and hydrogen peroxide (H2O2), two major reactive oxygen species (ROS), in chloroplasts under light. In the chemical-genetically rescued mpk3 mpk6 double mutants, ETI-induced photosynthetic inhibition and chloroplastic ROS accumulation are compromised, which correlates with delayed hypersensitive response (HR) cell death and compromised resistance. Furthermore, protection of chloroplasts by expressing a plastid-targeted cyanobacterial flavodoxin (pFLD) delays photosynthetic inhibition and compromises ETI. Collectively, this study highlights a critical role of MPK3/MPK6 in manipulating plant photosynthetic activities to promote ROS accumulation in chloroplasts and HR cell death, which contributes to the robustness of ETI. Furthermore, the dual functionality of MPK3/MPK6 cascade in promoting defense and inhibiting photosynthesis potentially allow it to orchestrate the trade-off between plant growth and defense in plant immunity. PMID:29723186

Differential and Conditional Activation of PKC-Isoforms Dictates Cardiac Adaptation during Physiological to Pathological Hypertrophy

PubMed Central

Naskar, Shaon; Datta, Kaberi; Mitra, Arkadeep; Pathak, Kanchan; Datta, Ritwik; Bansal, Trisha; Sarkar, Sagartirtha

2014-01-01

A cardiac hypertrophy is defined as an increase in heart mass which may either be beneficial (physiological hypertrophy) or detrimental (pathological hypertrophy). This study was undertaken to establish the role of different protein kinase-C (PKC) isoforms in the regulation of cardiac adaptation during two types of cardiac hypertrophy. Phosphorylation of specific PKC-isoforms and expression of their downstream proteins were studied during physiological and pathological hypertrophy in 24 week male Balb/c mice (Mus musculus) models, by reverse transcriptase-PCR, western blot analysis and M-mode echocardiography for cardiac function analysis. PKC-δ was significantly induced during pathological hypertrophy while PKC-α was exclusively activated during physiological hypertrophy in our study. PKC-δ activation during pathological hypertrophy resulted in cardiomyocyte apoptosis leading to compromised cardiac function and on the other hand, activation of PKC-α during physiological hypertrophy promoted cardiomyocyte growth but down regulated cellular apoptotic load resulting in improved cardiac function. Reversal in PKC-isoform with induced activation of PKC-δ and simultaneous inhibition of phospho-PKC-α resulted in an efficient myocardium to deteriorate considerably resulting in compromised cardiac function during physiological hypertrophy via augmentation of apoptotic and fibrotic load. This is the first report where PKC-α and -δ have been shown to play crucial role in cardiac adaptation during physiological and pathological hypertrophy respectively thereby rendering compromised cardiac function to an otherwise efficient heart by conditional reversal of their activation. PMID:25116170

The archetypal R90C CADASIL-NOTCH3 mutation retains NOTCH3 function in vivo.

PubMed

Monet, Marie; Domenga, Valérie; Lemaire, Barbara; Souilhol, Céline; Langa, Francina; Babinet, Charles; Gridley, Thomas; Tournier-Lasserve, Elisabeth; Cohen-Tannoudji, Michel; Joutel, Anne

2007-04-15

Cerebral Autosomal Dominant Arteriopathy with Subcortical infarcts and Leukoencephalopathy (CADASIL) is the most prominent known cause of inherited stroke and vascular dementia in human adult. The disease gene, NOTCH3, encodes a transmembrane receptor primarily expressed in arterial smooth muscle cells (SMC). Pathogenic mutations lead to an odd number of cysteine residues within the NOTCH3 extracellular domain (NOTCH3(ECD)), and are associated with progressive accumulation of NOTCH3(ECD) at the SMC plasma membrane. The murine homolog, Notch3, is dispensable for viability but required post-natally for the elaboration and maintenance of arteries. How CADASIL-associated mutations impact NOTCH3 function remains a fundamental, yet unresolved issue. Particularly, whether NOTCH3(ECD) accumulation may titrate the ligand and inhibit the normal pathway is unknown. Herein, using genetic analyses in the mouse, we assessed the functional significance of an archetypal CADASIL-associated mutation (R90C), in vivo, in brain arteries. We show that transgenic mouse lines expressing either the wild-type human NOTCH3 or the mutant R90C human NOTCH3, at comparable and physiological levels, can rescue the arterial defects of Notch3-/- mice to similar degrees. In vivo assessment of NOTCH3/RBP-Jk activity provides evidence that the mutant NOTCH3 protein exhibits normal level of activity in brain arteries. Remarkably, the mutant NOTCH3 protein remains functional and does not exhibit dominant negative interfering activity, even when NOTCH3(ECD) accumulates. Collectively, these data suggest a model that invokes novel pathogenic roles for the mutant NOTCH3 protein rather than compromised NOTCH3 function as the primary determinant of the CADASIL arteriopathy.

Massive plexiform neurofibromas in childhood: natural history and management issues.

PubMed

Serletis, Demitre; Parkin, Patricia; Bouffet, Eric; Shroff, Manohar; Drake, James M; Rutka, James T

2007-05-01

The authors review their experience with massive plexiform neurofibromas (PNs) in patients with pediatric neurofibromatosis Type 1 (NF1) to better characterize the natural history and management of these complex lesions. The authors performed a retrospective review of data obtained in seven patients with NF1 in whom massive PNs were diagnosed at The Hospital for Sick Children in Toronto, Ontario, Canada. These patients attended routine follow-up examinations conducted by a number of specialists, and serial neuroimaging studies were obtained to monitor disease progression. The most common presenting feature of PN was that of a painful, expanding lesion. Furthermore, two patients harbored multiple, distinct PNs affecting different body sites. With respect to management, two patients were simply observed, undergoing serial neuroimaging studies; two patients underwent biopsy sampling of their plexiform lesions; two patients underwent attempted medical treatment (farnesyl transferase inhibitor, R11577, and cyclophosphamide chemotherapy); and three patients required surgical debulking of their PNs because the massive growth of these tumors caused functional compromise. Ultimately, one patient died of respiratory complications due to progressive growth of the massive PN lesion. In this review of their experience, the authors found certain features that underscore the presentation and natural history of PNs. The management of these complex lesions, however, remains unclear. Slow-growing PNs may be observed conservatively, but the authors' experience suggests that resection should be considered in selected cases involving significant deterioration or functional compromise. Nevertheless, patients with massive PNs will benefit from close surveillance by a team of specialists to monitor for ongoing disease progression.

Attenuation of Recombinant Yellow Fever 17D Viruses Expressing Foreign Protein Epitopes at the Surface

PubMed Central

Bonaldo, Myrna C.; Garratt, Richard C.; Marchevsky, Renato S.; Coutinho, Evandro S. F.; Jabor, Alfredo V.; Almeida, Luís F. C.; Yamamura, Anna M. Y.; Duarte, Adriana S.; Oliveira, Prisciliana J.; Lizeu, Jackeline O. P.; Camacho, Luiz A. B.; Freire, Marcos S.; Galler, Ricardo

2005-01-01

The yellow fever (YF) 17D vaccine is a live attenuated virus. Three-dimensional (3D) homology modeling of the E protein structure from YF 17D virus and its comparison with that from tick-borne encephalitis virus revealed that it is possible to accommodate inserts of different sizes and amino acid compositions in the flavivirus E protein fg loop. This is consistent with the 3D structures of both the dimeric and trimeric forms in which the fg loop lies exposed to solvents. We demonstrate here that YF 17D viruses bearing foreign humoral (17D/8) and T-cell (17D/13) epitopes, which vary in sequence and length, displayed growth restriction. It is hypothesized that interference with the dimer-trimer transition and with the formation of a ring of such trimers in order to allow fusion compromises the capability of the E protein to induce fusion of viral and endosomal membranes, and a slower rate of fusion may delay the extent of virus production. This would account for the lower levels of replication in cultured cells and of viremia in monkeys, as well as for the more attenuated phenotype of the recombinant viruses in monkeys. Testing of both recombinant viruses (17D/8 and 17D/13) for monkey neurovirulence also suggests that insertion at the 17D E protein fg loop does not compromise the attenuated phenotype of YF 17D virus, further confirming the potential use of this site for the development of new live attenuated 17D virus-based vaccines. PMID:15956601

In serum veritas—in serum sanitas? Cell non-autonomous aging compromises differentiation and survival of mesenchymal stromal cells via the oxidative stress pathway

PubMed Central

Geißler, S; Textor, M; Schmidt-Bleek, K; Klein, O; Thiele, M; Ellinghaus, A; Jacobi, D; Ode, A; Perka, C; Dienelt, A; Klose, J; Kasper, G; Duda, G N; Strube, P

2013-01-01

Even tissues capable of complete regeneration, such as bone, show an age-related reduction in their healing capacity. Here, we hypothesized that this decline is primarily due to cell non-autonomous (extrinsic) aging mediated by the systemic environment. We demonstrate that culture of mesenchymal stromal cells (MSCs) in serum from aged Sprague–Dawley rats negatively affects their survival and differentiation ability. Proteome analysis and further cellular investigations strongly suggest that serum from aged animals not only changes expression of proteins related to mitochondria, unfolded protein binding or involved in stress responses, it also significantly enhances intracellular reactive oxygen species production and leads to the accumulation of oxidatively damaged proteins. Conversely, reduction of oxidative stress levels in vitro markedly improved MSC function. These results were validated in an in vivo model of compromised bone healing, which demonstrated significant increase regeneration in aged animals following oral antioxidant administration. These observations indicate the high impact of extrinsic aging on cellular functions and the process of endogenous (bone) regeneration. Thus, addressing the cell environment by, for example, systemic antioxidant treatment is a promising approach to enhance tissue regeneration and to regain cellular function especially in elderly patients. PMID:24357801

The Fusarium oxysporum effector Six6 contributes to virulence and suppresses I-2-mediated cell death.

PubMed

Gawehns, F; Houterman, P M; Ichou, F Ait; Michielse, C B; Hijdra, M; Cornelissen, B J C; Rep, M; Takken, F L W

2014-04-01

Plant pathogens secrete effectors to manipulate their host and facilitate colonization. Fusarium oxysporum f. sp. lycopersici is the causal agent of Fusarium wilt disease in tomato. Upon infection, F. oxysporum f. sp. lycopersici secretes numerous small proteins into the xylem sap (Six proteins). Most Six proteins are unique to F. oxysporum, but Six6 is an exception; a homolog is also present in two Colletotrichum spp. SIX6 expression was found to require living host cells and a knockout of SIX6 in F. oxysporum f. sp. lycopersici compromised virulence, classifying it as a genuine effector. Heterologous expression of SIX6 did not affect growth of Agrobacterium tumefaciens in Nicotiana benthamiana leaves or susceptibility of Arabidopsis thaliana toward Verticillium dahliae, Pseudomonas syringae, or F. oxysporum, suggesting a specific function for F. oxysporum f. sp. lycopersici Six6 in the F. oxysporum f. sp. lycopersici- tomato pathosystem. Remarkably, Six6 was found to specifically suppress I-2-mediated cell death (I2CD) upon transient expression in N. benthamiana, whereas it did not compromise the activity of other cell-death-inducing genes. Still, this I2CD suppressing activity of Six6 does not allow the fungus to overcome I-2 resistance in tomato, suggesting that I-2-mediated resistance is independent from cell death.

The Epithelial Cell Adhesion Molecule EpCAM Is Required for Epithelial Morphogenesis and Integrity during Zebrafish Epiboly and Skin Development

PubMed Central

Slanchev, Krasimir; Carney, Thomas J.; Stemmler, Marc P.; Koschorz, Birgit; Amsterdam, Adam; Schwarz, Heinz; Hammerschmidt, Matthias

2009-01-01

The aberrant expression of the transmembrane protein EpCAM is associated with tumor progression, affecting different cellular processes such as cell–cell adhesion, migration, proliferation, differentiation, signaling, and invasion. However, the in vivo function of EpCAM still remains elusive due to the lack of genetic loss-of-function studies. Here, we describe epcam (tacstd) null mutants in zebrafish. Maternal-zygotic mutants display compromised basal protrusive activity and epithelial morphogenesis in cells of the enveloping layer (EVL) during epiboly. In partial redundancy with E-cadherin (Ecad), EpCAM made by EVL cells is further required for cell–cell adhesion within the EVL and, possibly, for proper attachment of underlying deep cells to the inner surface of the EVL, thereby also affecting deep cell epiboly movements. During later development, EpCAM per se becomes indispensable for epithelial integrity within the periderm of the skin, secondarily leading to disrupted morphology of the underlying basal epidermis and moderate hyper-proliferation of skin cells. On the molecular level, EVL cells of epcam mutant embryos display reduced levels of membranous Ecad, accompanied by an enrichment of tight junction proteins and a basal extension of apical junction complexes (AJCs). Our data suggest that EpCAM acts as a partner of E-cadherin to control adhesiveness and integrity as well as plasticity and morphogenesis within simple epithelia. In addition, EpCAM is required for the interaction of the epithelia with underlying cell layers. PMID:19609345

Up-Regulation of Soluble Axl and Mer Receptor Tyrosine Kinases Negatively Correlates with Gas6 in Established Multiple Sclerosis Lesions

PubMed Central

Weinger, Jason G.; Omari, Kakuri M.; Marsden, Kurt; Raine, Cedric S.; Shafit-Zagardo, Bridget

2009-01-01

Multiple sclerosis is a disease that is characterized by inflammation, demyelination, and axonal damage; it ultimately forms gliotic scars and lesions that severely compromise the function of the central nervous system. Evidence has shown previously that altered growth factor receptor signaling contributes to lesion formation, impedes recovery, and plays a role in disease progression. Growth arrest-specific protein 6 (Gas6), the ligand for the TAM receptor tyrosine kinase family, consisting of Tyro3, Axl, and Mer, is important for cell growth, survival, and clearance of debris. In this study, we show that levels of membrane-bound Mer (205 kd), soluble Mer (∼150 kd), and soluble Axl (80 kd) were all significantly elevated in homogenates from established multiple sclerosis lesions comprised of both chronic active and chronic silent lesions. Whereas in normal tissue Gas6 positively correlated with soluble Axl and Mer, there was a negative correlation between Gas6 and soluble Axl and Mer in established multiple sclerosis lesions. In addition, increased levels of soluble Axl and Mer were associated with increased levels of mature ADAM17, mature ADAM10, and Furin, proteins that are associated with Axl and Mer solubilization. Soluble Axl and Mer are both known to act as decoy receptors and block Gas6 binding to membrane-bound receptors. These data suggest that in multiple sclerosis lesions, dysregulation of protective Gas6 receptor signaling may prolong lesion activity. PMID:19541935

Parkinson's disease.

PubMed

Thomas, Bobby; Beal, M Flint

2007-10-15

Parkinson's disease (PD) is a chronic progressive neurodegenerative movement disorder characterized by a profound and selective loss of nigrostriatal dopaminergic neurons. Clinical manifestations of this complex disease include motor impairments involving resting tremor, bradykinesia, postural instability, gait difficulty and rigidity. Current medications only provide symptomatic relief and fail to halt the death of dopaminergic neurons. A major hurdle in development of neuroprotective therapies are due to limited understanding of disease processes leading to death of dopaminergic neurons. While the etiology of dopaminergic neuronal demise is elusive, a combination of genetic susceptibilities and environmental factors seems to play a critical role. The majority of PD cases are sporadic however, the discovery of genes linked to rare familial forms of disease (encoding alpha-synuclein, parkin, DJ-1, PINK-1 and LRRK2) and studies from experimental animal models has provided crucial insights into molecular mechanisms in disease pathogenesis and identified probable targets for therapeutic intervention. Recent findings implicate mitochondrial dysfunction, oxidative damage, abnormal protein accumulation and protein phosphorylation as key molecular mechanisms compromising dopamine neuronal function and survival as the underlying cause of pathogenesis in both sporadic and familial PD. In this review we provide an overview of the most relevant findings made by the PD research community in the last year and discuss how these significant findings improved our understanding of events leading to nigrostriatal dopaminergic degeneration, and identification of potential cell survival pathways that could serve as targets for neuroprotective therapies in preventing this disabling neurological illness.

Emerging (and converging) pathways in Parkinson's disease: keeping mitochondrial wellness

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

Cieri, Domenico; Brini, Marisa; Calì, Tito

The selective cell loss in the ventral component of the substantia nigra pars compacta and the presence of alpha-synuclein (α-syn)-rich intraneuronal inclusions called Lewy bodies are the pathological hallmarks of Parkinson's disease (PD), the most common motor system disorder whose aetiology remains largely elusive. Although most cases of PD are idiopathic, there are rare familial forms of the disease that can be traced to single gene mutations that follow Mendelian inheritance pattern. The study of several nuclear encoded proteins whose mutations are linked to the development of autosomal recessive and dominant forms of familial PD enhanced our understanding of biochemicalmore » and cellular mechanisms contributing to the disease and suggested that many signs of neurodegeneration result from compromised mitochondrial function. Here we present an overview of the current understanding of PD-related mitochondrial dysfunction including defects in bioenergetics and Ca{sup 2+} homeostasis, mitochondrial DNA mutations, altered mitochondrial dynamics and autophagy. We emphasize, in particular, the convergence of many “apparently” different pathways towards a common route involving mitochondria. Understanding whether mitochondrial dysfunction in PD represents the cause or the consequence of the disease is challenging and will help to define the pathogenic processes at the basis of the PD onset and progression. - Highlights: • Mitochondrial dysfunctions are a common feature of neurodegenerative diseases. • Many familial PD related proteins ensure mitochondrial function. • Mutations in PD genes differently affect mitochondria related activities.« less

Increased O-GlcNAcylation of Endothelial Nitric Oxide Synthase Compromises the Anti-contractile Properties of Perivascular Adipose Tissue in Metabolic Syndrome.

PubMed

da Costa, Rafael M; da Silva, Josiane F; Alves, Juliano V; Dias, Thiago B; Rassi, Diane M; Garcia, Luis V; Lobato, Núbia de Souza; Tostes, Rita C

2018-01-01

Under physiological conditions, the perivascular adipose tissue (PVAT) negatively modulates vascular contractility. This property is lost in experimental and human obesity and in the metabolic syndrome, indicating that changes in PVAT function may contribute to vascular dysfunction associated with increased body weight and hyperglycemia. The O -linked β-N-acetylglucosamine ( O -GlcNAc) modification of proteins ( O -GlcNAcylation) is a unique posttranslational process that integrates glucose metabolism with intracellular protein activity. Increased flux of glucose through the hexosamine biosynthetic pathway and the consequent increase in tissue-specific O -GlcNAc modification of proteins have been linked to multiple facets of vascular dysfunction in diabetes and other pathological conditions. We hypothesized that chronic consumption of glucose, a condition that progresses to metabolic syndrome, leads to increased O -GlcNAc modification of proteins in the PVAT, decreasing its anti-contractile effects. Therefore, the current study was devised to determine whether a high-sugar diet increases O -GlcNAcylation in the PVAT and how increased O -GlcNAc interferes with PVAT vasorelaxant function. To assess molecular mechanisms by which O -GlcNAc contributes to PVAT dysfunction, thoracic aortas surrounded by PVAT were isolated from Wistar rats fed either a control or high sugar diet, for 10 and 12 weeks. Rats chronically fed a high sugar diet exhibited metabolic syndrome features, increased O -GlcNAcylated-proteins in the PVAT and loss of PVAT anti-contractile effect. PVAT from high sugar diet-fed rats for 12 weeks exhibited decreased NO formation, reduced expression of endothelial nitric oxide synthase (eNOS) and increased O -GlcNAcylation of eNOS. High sugar diet also decreased OGA activity and increased superoxide anion generation in the PVAT. Visceral adipose tissue samples from hyperglycemic patients showed increased levels of O -GlcNAc-modified proteins, increased ROS generation and decreased OGA activity. These data indicate that O -GlcNAcylation contributes to metabolic syndrome-induced PVAT dysfunction and that O -GlcNAcylation of eNOS may be targeted in the development of novel therapies for vascular dysfunction in conditions associated with hyperglycemia.

Direct digestion of proteins in living cells into peptides for proteomic analysis.

PubMed

Chen, Qi; Yan, Guoquan; Gao, Mingxia; Zhang, Xiangmin

2015-01-01

To analyze the proteome of an extremely low number of cells or even a single cell, we established a new method of digesting whole cells into mass-spectrometry-identifiable peptides in a single step within 2 h. Our sampling method greatly simplified the processes of cell lysis, protein extraction, protein purification, and overnight digestion, without compromising efficiency. We used our method to digest hundred-scale cells. As far as we know, there is no report of proteome analysis starting directly with as few as 100 cells. We identified an average of 109 proteins from 100 cells, and with three replicates, the number of proteins rose to 204. Good reproducibility was achieved, showing stability and reliability of the method. Gene Ontology analysis revealed that proteins in different cellular compartments were well represented.

Poly(zwitterionic)protein conjugates offer increased stability without sacrificing binding affinity or bioactivity

NASA Astrophysics Data System (ADS)

Keefe, Andrew J.; Jiang, Shaoyi

2012-01-01

Treatment with therapeutic proteins is an attractive approach to targeting a number of challenging diseases. Unfortunately, the native proteins themselves are often unstable in physiological conditions, reducing bioavailability and therefore increasing the dose that is required. Conjugation with poly(ethylene glycol) (PEG) is often used to increase stability, but this has a detrimental effect on bioactivity. Here, we introduce conjugation with zwitterionic polymers such as poly(carboxybetaine). We show that poly(carboxybetaine) conjugation improves stability in a manner similar to PEGylation, but that the new conjugates retain or even improve the binding affinity as a result of enhanced protein-substrate hydrophobic interactions. This chemistry opens a new avenue for the development of protein therapeutics by avoiding the need to compromise between stability and affinity.

Influence of maternal hyperthyroidism in the rat on the expression of neuronal and astrocytic cytoskeletal proteins in fetal brain.

PubMed

Evans, I M; Pickard, M R; Sinha, A K; Leonard, A J; Sampson, D C; Ekins, R P

2002-12-01

Maternal hypothyroidism during pregnancy impairs brain function in human and rat offspring, but little is known regarding the influence of maternal hyperthyroidism on neurodevelopment. We have previously shown that the expression of neuronal and glial differentiation markers in fetal brain is compromised in hypothyroid rat dam pregnancies and have now therefore extended this investigation to hyperthyroid rat dams. Study groups comprised partially thyroidectomised dams, implanted with osmotic pumps infusing either vehicle (TX dams) or a supraphysiological dose of thyroxine (T4) (HYPER dams), and euthyroid dams infused with vehicle (N dams). Cytoskeletal protein abundance was determined in fetal brain at 21 days of gestation by immunoblot analysis. Relative to N dams, circulating total T4 levels were reduced to around one-third in TX dams but were doubled in HYPER dams. Fetal brain weight was increased in HYPER dams, whereas litter size and fetal body weight were reduced in TX dams. Glial fibrillary acidic protein expression was similar in HYPER and TX dams, being reduced in both cases relative to N dams. alpha-Internexin (INX) abundance was reduced in HYPER dams and increased in TX dams, whereas neurofilament 68 (NF68) exhibited increased abundance in HYPER dams. Furthermore, INX was inversely related to - and NF68 directly related to - maternal serum total T4 levels, independently of fetal brain weight. In conclusion, maternal hyperthyroidism compromises the expression of neuronal cytoskeletal proteins in late fetal brain, suggestive of a pattern of accelerated neuronal differentiation.

A targeted IL-15 fusion protein with potent anti-tumor activity

PubMed Central

Chen, Siqi; Huang, Qiang; Liu, Jiayu; Xing, Jieyu; Zhang, Ning; Liu, Yawei; Wang, Zhong; Li, Qing

2015-01-01

IL-15 has been actively investigated for its potential in tumor immunotherapy. To enhance the anti-tumor activity of IL-15, the novel PFC-1 construct was designed, which comprises the following 3 parts: (1) IL-15Rα fused with IL-15 to enhance IL-15 activity, (2) an Fc fragment to increase protein half-life, and (3) an integrin-targeting RGD peptide to enhance tumor targeting. PFC-1 showed tumor cell targeting without compromising IL-15 activity. PFC-1 also had potent anti-tumor activities in xenograft models, suggesting the potential application of this multi-functional fusion protein in tumor therapy. PMID:26176990

Metalloproteinases and atherothrombosis: MMP-10 mediates vascular remodeling promoted by inflammatory stimuli.

PubMed

Rodriguez, Jose A; Orbe, Josune; Martinez de Lizarrondo, Sara; Calvayrac, Olivier; Rodriguez, Cristina; Martinez-Gonzalez, Jose; Paramo, Jose A

2008-01-01

Atherosclerosis is the common pathophysiological substrate of ischemic vascular diseases and their thrombotic complications. The unbalance between matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) has been hypothesized to be involved in the growth, destabilization, and eventual rupture of atherosclerotic lesions. Different MMPs have been assigned relevant roles in the pathology of vascular diseases and MMP-10 (stromelysin-2) has been involved in vascular development and atherogenesis. This article examines the pathophysiological role of MMPs, particularly MMP-10, in the onset and progression of vascular diseases and their regulation by pro-inflammatory stimuli. MMP-10 over-expression has been shown to compromise vascular integrity and it has been associated with aortic aneurysms. MMP-10 is induced by C-reactive protein in endothelial cells, and it is over-expressed in atherosclerotic lesions. Additionally, higher MMP-10 serum levels are associated with inflammatory markers, increased carotid intima-media thickness and the presence of atherosclerotic plaques. We have cloned the promoter region of the MMP-10 gene and studied the effect of inflammatory stimuli on MMP-10 transcriptional regulation, providing evidences further supporting the involvement of MMP-10 in the pathophysiology of atherothrombosis.

Smoking, p53 Mutation, and Lung Cancer

PubMed Central

Gibbons, Don L.; Byers, Lauren A.; Kurie, Jonathan M.

2014-01-01

This issue marks the 50th Anniversary of the release of the U.S. Surgeon General’s Report on Smoking and Health. Perhaps no other singular event has done more to highlight the effects of smoking on the development of cancer. Tobacco exposure is the leading cause of cancers involving the oral cavity, conductive airways and the lung. Owing to the many carcinogens in tobacco smoke, smoking-related malignancies have a high genome-wide burden of mutations, including in the gene encoding for p53. The p53 protein is the most frequently mutated tumor suppressor in cancer, responsible for a range of critical cellular functions that are compromised by the presence of a mutation. Herein we review the epidemiologic connection between tobacco exposure and cancer, the molecular basis of p53 mutation in lung cancer, and the normal molecular and cellular roles of p53 that are abrogated during lung tumor development and progression as defined by in vitro and in vivo studies. We also consider the therapeutic potential of targeting mutant p53 in a clinical setting based upon the cellular role of mutant p53 and data from genetic murine models. PMID:24442106

miR-300 mediates Bmi1 function and regulates differentiation in primitive cardiac progenitors

PubMed Central

Cruz, F M; Tomé, M; Bernal, J A; Bernad, A

2015-01-01

B lymphoma Mo-MLV insertion region 1 (Bmi1) is a polycomb-family transcriptional factor critical for self-renewal in many adult stem cells and human neoplasia. We sought to identify microRNAs regulated by Bmi1 that could play a role in multipotent cardiac progenitor cell (CPC) decisions. We found that miR-300, a poorly characterized microRNA mapping in the Dlk1-Dio3 microRNA cluster, was positively regulated by Bmi1 in CPCs. Forced expression of miR-300 in CPCs promoted an improved stemness signature with a significant increase in Oct4 levels, a reduction in senescence progression and an enhanced proliferative status via p19 activation and inhibition of p16 accumulation. Endothelial and cardiogenic differentiation were clearly compromised by sustained miR-300 expression. Additionally, RNA and protein analysis revealed a significant reduction in key cardiac transcription factors, including Nkx2.5 and Tbx5. Collectively, these results suggest that some functions attributed to Bmi1 are due to induction of miR-300, which decreases the cardiogenic differentiation potential of multipotent CPCs in vitro and promotes self-renewal. PMID:26512961

PRP19 transforms into a sensor of RPA-ssDNA after DNA damage and drives ATR activation via a ubiquitin-mediated circuitry.

PubMed

Maréchal, Alexandre; Li, Ju-Mei; Ji, Xiao Ye; Wu, Ching-Shyi; Yazinski, Stephanie A; Nguyen, Hai Dang; Liu, Shizhou; Jiménez, Amanda E; Jin, Jianping; Zou, Lee

2014-01-23

PRP19 is a ubiquitin ligase involved in pre-mRNA splicing and the DNA damage response (DDR). Although the role for PRP19 in splicing is well characterized, its role in the DDR remains elusive. Through a proteomic screen for proteins that interact with RPA-coated single-stranded DNA (RPA-ssDNA), we identified PRP19 as a sensor of DNA damage. PRP19 directly binds RPA and localizes to DNA damage sites via RPA, promoting RPA ubiquitylation in a DNA-damage-induced manner. PRP19 facilitates the accumulation of ATRIP, the regulatory partner of the ataxia telangiectasia mutated and Rad3-related (ATR) kinase, at DNA damage sites. Depletion of PRP19 compromised the phosphorylation of ATR substrates, recovery of stalled replication forks, and progression of replication forks on damaged DNA. Importantly, PRP19 mutants that cannot bind RPA or function as an E3 ligase failed to support the ATR response, revealing that PRP19 drives ATR activation by acting as an RPA-ssDNA-sensing ubiquitin ligase during the DDR. Copyright © 2014 Elsevier Inc. All rights reserved.

Parenteral Nutrition-Associated Liver Disease: The Role of the Gut Microbiota.

PubMed

Cahova, Monika; Bratova, Miriam; Wohl, Petr

2017-09-07

Parenteral nutrition (PN) provides life-saving nutritional support in situations where caloric supply via the enteral route cannot cover the necessary needs of the organism. However, it does have serious adverse effects, including parenteral nutrition-associated liver disease (PNALD). The development of liver injury associated with PN is multifactorial, including non-specific intestine inflammation, compromised intestinal permeability, and barrier function associated with increased bacterial translocation, primary and secondary cholangitis, cholelithiasis, short bowel syndrome, disturbance of hepatobiliary circulation, lack of enteral nutrition, shortage of some nutrients (proteins, essential fatty acids, choline, glycine, taurine, carnitine, etc.), and toxicity of components within the nutrition mixture itself (glucose, phytosterols, manganese, aluminium, etc.). Recently, an increasing number of studies have provided evidence that some of these factors are directly or indirectly associated with microbial dysbiosis in the intestine. In this review, we focus on PN-induced changes in the taxonomic and functional composition of the microbiome. We also discuss immune cell and microbial crosstalk during parenteral nutrition, and the implications for the onset and progression of PNALD. Finally, we provide an overview of recent advances in the therapeutic utilisation of pro- and prebiotics for the mitigation of PN-associated liver complications.

Neutrophil-derived JAML Inhibits Repair of Intestinal Epithelial Injury During Acute Inflammation

PubMed Central

Weber, Dominique A.; Sumagin, Ronen; McCall, Ingrid C.; Leoni, Giovanna; Neumann, Philipp A.; Andargachew, Rakieb; Brazil, Jennifer C.; Medina-Contreras, Oscar; Denning, Timothy L.; Nusrat, Asma; Parkos, Charles A.

2014-01-01

Neutrophil transepithelial migration (TEM) during acute inflammation is associated with mucosal injury. Using models of acute mucosal injury in-vitro and in-vivo, we describe a new mechanism by which neutrophils infiltrating the intestinal mucosa disrupt epithelial homeostasis. We report that junctional adhesion molecule-like protein (JAML) is cleaved from neutrophil surface by zinc-metalloproteases during TEM. Neutrophil-derived soluble JAML bound to the epithelial tight junction protein coxsackie-adenovirus receptor (CAR) resulting in compromised barrier and inhibition of wound repair, through decreased epithelial proliferation. The deleterious effects of JAML on barrier and wound repair were reversed with an anti-JAML mAb that inhibits JAML-CAR binding. Thus, JAML released from transmigrating neutrophils across inflamed epithelia can promote recruitment of leukocytes and aid in clearance of invading microorganisms. However, sustained release of JAML under pathologic conditions associated with persistence of large numbers of infiltrated neutrophil would compromise intestinal barrier and inhibit mucosal healing. Targeting JAML-CAR interactions may thus improve mucosal healing responses under conditions of dysregulated neutrophil recruitment. PMID:24621992

Cervical spondylosis anatomy: pathophysiology and biomechanics.

PubMed

Shedid, Daniel; Benzel, Edward C

2007-01-01

Cervical spondylosis is the most common progressive disorder in the aging cervical spine. It results from the process of degeneration of the intervertebral discs and facet joints of the cervical spine. Biomechanically, the disc and the facets are the connecting structures between the vertebrae for the transmission of external forces. They also facilitate cervical spine mobility. Symptoms related to myelopathy and radiculopathy are caused by the formation of osteophytes, which compromise the diameter of the spinal canal. This compromise may also be partially developmental. The developmental process, together with the degenerative process, may cause mechanical pressure on the spinal cord at one or multiple levels. This pressure may produce direct neurological damage or ischemic changes and, thus, lead to spinal cord disturbances. A thorough understanding of the biomechanics, the pathology, the clinical presentation, the radiological evaluation, as well as the surgical indications of cervical spondylosis, is essential for the management of patients with cervical spondylosis.

Current pathophysiological concepts and management of pulmonary hypertension.

PubMed

Lourenço, André P; Fontoura, Dulce; Henriques-Coelho, Tiago; Leite-Moreira, Adelino F

2012-03-22

Pulmonary hypertension (PH), increasingly recognized as a major health burden, remains underdiagnosed due mainly to the unspecific symptoms. Pulmonary arterial hypertension (PAH) has been extensively investigated. Pathophysiological knowledge derives mostly from experimental models. Paradoxically, common non-PAH PH forms remain largely unexplored. Drugs targeting lung vascular tonus became available during the last two decades, notwithstanding the disease progresses in many patients. The aim of this review is to summarize recent advances in epidemiology, pathophysiology and management with particular focus on associated myocardial and systemic compromise and experimental therapeutic possibilities. PAH, currently viewed as a panvasculopathy, is due to a crosstalk between endothelial and smooth muscle cells, inflammatory activation and altered subcellular pathways. Cardiac cachexia and right ventricular compromise are fundamental determinants of PH prognosis. Combined vasodilator therapy is already mainstay for refractory cases, but drugs directed at these new pathophysiological pathways may constitute a significant advance. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

Development of a transgenic Plasmodium berghei line (Pb pfpkg) expressing the P. falciparum cGMP-dependent protein kinase, a novel antimalarial drug target.

PubMed

Tewari, Rita; Patzewitz, Eva-Maria; Poulin, Benoit; Stewart, Lindsay; Baker, David A

2014-01-01

With the inevitable selection of resistance to antimalarial drugs in treated populations, there is a need for new medicines to enter the clinic and new targets to progress through the drug discovery pipeline. In this study we set out to develop a transgenic rodent model for testing inhibitors of the Plasmodium falciparum cyclic GMP-dependent kinase in vivo. A model was needed that would allow us to investigate whether differences in amino acid sequence of this enzyme between species influences in vivo efficacy. Here we report the successful development of a transgenic P. berghei line in which the cyclic GMP-dependent protein kinase (PKG) was replaced by the P. falciparum orthologue. We demonstrate that the P. falciparum orthologue was able to functionally complement the endogenous P. berghei pkg gene throughout blood stage development and early sexual development. However, subsequent development in the mosquito was severely compromised. We show that this is due to a defect in the female lineage of the transgenic by using genetic crosses with both male and female deficient P. berghei lines. This defect could be due to expression of a female-specific target in the mosquito stages of P. berghei that cannot be phosphorylated by the P. falciparum kinase. Using a previously reported anti-coccidial inhibitor of the cyclic GMP-dependent protein kinase, we show no difference in in vivo efficacy between the transgenic and control P. berghei lines. This in vivo model will be useful for screening future generations of cyclic GMP-dependent protein kinase inhibitors and allowing us to overcome any species-specific differences in the enzyme primary sequence that would influence in vivo efficacy in the rodent model. The approach will also be applicable to in vivo testing of other antimalarial compounds where the target is known.

NUP-1 Is a Large Coiled-Coil Nucleoskeletal Protein in Trypanosomes with Lamin-Like Functions

PubMed Central

DuBois, Kelly N.; Alsford, Sam; Holden, Jennifer M.; Buisson, Johanna; Swiderski, Michal; Bart, Jean-Mathieu; Ratushny, Alexander V.; Wan, Yakun; Bastin, Philippe; Barry, J. David; Navarro, Miguel; Horn, David; Aitchison, John D.; Rout, Michael P.; Field, Mark C.

2012-01-01

A unifying feature of eukaryotic nuclear organization is genome segregation into transcriptionally active euchromatin and transcriptionally repressed heterochromatin. In metazoa, lamin proteins preserve nuclear integrity and higher order heterochromatin organization at the nuclear periphery, but no non-metazoan lamin orthologues have been identified, despite the likely presence of nucleoskeletal elements in many lineages. This suggests a metazoan-specific origin for lamins, and therefore that distinct protein elements must compose the nucleoskeleton in other lineages. The trypanosomatids are highly divergent organisms and possess well-documented but remarkably distinct mechanisms for control of gene expression, including polycistronic transcription and trans-splicing. NUP-1 is a large protein localizing to the nuclear periphery of Trypanosoma brucei and a candidate nucleoskeletal component. We sought to determine if NUP-1 mediates heterochromatin organization and gene regulation at the nuclear periphery by examining the influence of NUP-1 knockdown on morphology, chromatin positioning, and transcription. We demonstrate that NUP-1 is essential and part of a stable network at the inner face of the trypanosome nuclear envelope, since knockdown cells have abnormally shaped nuclei with compromised structural integrity. NUP-1 knockdown also disrupts organization of nuclear pore complexes and chromosomes. Most significantly, we find that NUP-1 is required to maintain the silenced state of developmentally regulated genes at the nuclear periphery; NUP-1 knockdown results in highly specific mis-regulation of telomere-proximal silenced variant surface glycoprotein (VSG) expression sites and procyclin loci, indicating a disruption to normal chromatin organization essential to life-cycle progression. Further, NUP-1 depletion leads to increased VSG switching and therefore appears to have a role in control of antigenic variation. Thus, analogous to vertebrate lamins, NUP-1 is a major component of the nucleoskeleton with key roles in organization of the nuclear periphery, heterochromatin, and epigenetic control of developmentally regulated loci. PMID:22479148

A Novel Algorithm for Detecting Protein Complexes with the Breadth First Search

PubMed Central

Tang, Xiwei; Wang, Jianxin; Li, Min; He, Yiming; Pan, Yi

2014-01-01

Most biological processes are carried out by protein complexes. A substantial number of false positives of the protein-protein interaction (PPI) data can compromise the utility of the datasets for complexes reconstruction. In order to reduce the impact of such discrepancies, a number of data integration and affinity scoring schemes have been devised. The methods encode the reliabilities (confidence) of physical interactions between pairs of proteins. The challenge now is to identify novel and meaningful protein complexes from the weighted PPI network. To address this problem, a novel protein complex mining algorithm ClusterBFS (Cluster with Breadth-First Search) is proposed. Based on the weighted density, ClusterBFS detects protein complexes of the weighted network by the breadth first search algorithm, which originates from a given seed protein used as starting-point. The experimental results show that ClusterBFS performs significantly better than the other computational approaches in terms of the identification of protein complexes. PMID:24818139

Cardiac transplant in young female patient diagnosed with diffuse systemic sclerosis.

PubMed

Bennasar, Guillermo; Carlevaris, Leandro; Secco, Anastasia; Romanini, Felix; Mamani, Marta

2016-01-01

Systemic sclerosis (SS) in a multifactorial and systemic, chronic, autoimmune disease that affects the connective tissue. We present this clinical case given the low prevalence of diffuse SS with early and progressive cardiac compromise in a young patient, and treatment with cardiac transplantation. Copyright © 2015 Elsevier España, S.L.U. and Sociedad Española de Reumatología y Colegio Mexicano de Reumatología. All rights reserved.

The Mustard Consortium’s Elucidation of the Pathophysiology of Sulfur Mustard and Antidote Development

DTIC Science & Technology

2006-09-01

these tissues. There was essentially no change in the tocopherol levels. These results imply that CEES produces systemic oxidative stress at a...of CEES-induced acute lung injury, even though little is currently known about how CEES produces acute and progressive lung injury. Body...Medical School). 3. Pulmonary clearance of Pseudomonas aeruginosa in CEES treated rats. It is not known if exposure to CEES compromises the ability

Implication of a Central Cysteine Residue and the HHCC Domain of Moloney Murine Leukemia Virus Integrase Protein in Functional Multimerization

PubMed Central

Donzella, George A.; Leon, Oscar; Roth, Monica J.

1998-01-01

Moloney murine leukemia virus (M-MuLV) IN-IN protein interactions important for catalysis of strand transfer and unimolecular and bimolecular disintegration reactions were investigated by using a panel of chemically modified M-MuLV IN proteins. Functional complementation of an HHCC-deleted protein (NΔ105) by an independent HHCC domain (CΔ232) was severely compromised by NEM modification of either subunit. Productive NΔ105 IN-DNA interactions with a disintegration substrate lacking a long terminal repeat 5′-single-stranded tail also required complementation by a functional HHCC domain. Virus encoding the C209A M-MuLV IN mutation exhibited delayed virion production and replication kinetics. PMID:9445080

Distinctive Roles for Periplasmic Proteases in the Maintenance of Essential Outer Membrane Protein Assembly.

PubMed

Soltes, Garner R; Martin, Nicholas R; Park, Eunhae; Sutterlin, Holly A; Silhavy, Thomas J

2017-10-15

Outer membrane protein (OMP) biogenesis in Escherichia coli is a robust process essential to the life of the organism. It is catalyzed by the β-barrel assembly machine (Bam) complex, and a number of quality control factors, including periplasmic chaperones and proteases, maintain the integrity of this trafficking pathway. Little is known, however, about how periplasmic proteases recognize and degrade OMP substrates when assembly is compromised or whether different proteases recognize the same substrate at distinct points in the assembly pathway. In this work, we use well-defined assembly-defective mutants of LptD, the essential lipopolysaccharide assembly translocon, to show that the periplasmic protease DegP degrades substrates with assembly defects that prevent or impair initial contact with Bam, causing the mutant protein to accumulate in the periplasm. In contrast, another periplasmic protease, BepA, degrades a LptD mutant substrate that has engaged the Bam complex and formed a nearly complete barrel. Furthermore, we describe the role of the outer membrane lipoprotein YcaL, a protease of heretofore unknown function, in the degradation of a LptD substrate that has engaged the Bam complex but is stalled at an earlier step in the assembly process that is not accessible to BepA. Our results demonstrate that multiple periplasmic proteases monitor OMPs at distinct points in the assembly process. IMPORTANCE OMP assembly is catalyzed by the essential Bam complex and occurs in a cellular environment devoid of energy sources. Assembly intermediates that misfold can compromise this essential molecular machine. Here we demonstrate distinctive roles for three different periplasmic proteases that can clear OMP substrates with folding defects that compromise assembly at three different stages. These quality control factors help ensure the integrity of the permeability barrier that contributes to the intrinsic resistance of Gram-negative organisms to many antibiotics. Copyright © 2017 American Society for Microbiology.

Proteomic analysis reveals the important roles of alpha-5-collagen and ATP5β during skin ulceration syndrome progression of sea cucumber Apostichopus japonicus.

PubMed

Zhao, Zelong; Jiang, Jingwei; Pan, Yongjia; Sun, Hongjuan; Guan, Xiaoyan; Gao, Shan; Chen, Zhong; Dong, Ying; Zhou, Zunchun

2018-03-20

Apostichopus japonicus is one of the most important aquaculture species in China. Skin ulceration syndrome (SUS) of sea cucumber is a common and serious disease affected the development of A. japonicus culture industry. To better understand the response mechanisms of A. japonicus during SUS progression, the protein variations in the body wall of A. japonicus at different stages of SUS were investigated by a comparative proteomic approach based on isobaric tags for relative and absolute quantification. A total of 1449 proteins were identified from the samples at different SUS stages. Among these proteins, 145 proteins were differentially expressed in the SUS-related samples compared to those of healthy A. japonicus. These differentially expressed proteins involved a wide range of functions. Among these differentially expressed proteins, only two proteins, alpha-5-collagen and an unknown function protein, were differentially expressed during the whole progression of SUS compared with healthy A. japonicus. In addition, ATP synthase subunit beta (ATP5β) interacted with a variety of proteins with different functions during the SUS progression. These results implied that alpha-5-collagen and ATP5β could play important roles during the SUS progression of A. japonicus. Our study provided a new sight to understand the molecular responses of sea cucumber during the SUS progression and accumulated data for the prevention of SUS in sea cucumber aquaculture. The current study aimed to reveal how the body wall of Apostichopus japonicus response to skin ulceration syndrome (SUS). To the best of our knowledge, this is the first proteomic study analyzing the differences in protein profile of sea cucumber during the whole SUS progression. By analyzing the expression differences of the proteome via isobaric labeling-based quantitative proteomic, we identified some proteins which may play important roles during the SUS progression. According to the enrichment analyses of these proteins based on Gene Ontology and Kyoto Encyclopedia of Genes and Genomes, a draft view of how the sea cucumber affected by SUS has been drawn. The common and unique differentially expressed proteins by Venn analysis showed that alpha-5-collagen was down-regulated at all stages of SUS, which had the potential as a target component for the host-directed SUS therapy. In addition, ATP5β, a subunit of mitochondrial ATP synthase, interacting with a variety of proteins with different functions during the SUS progression. This result illustrated that energy production and metabolism could play an important role in the formation of skin ulceration and resistance to pathogens in sea cucumber. The results of this study will be helpful for researchers to gain insights into the complex molecular mechanism of SUS in sea cucumber. Copyright © 2018 Elsevier B.V. All rights reserved.

Protein Mediated Oxidative Stress in Patients with Diabetes and its Associated Neuropathy: Correlation with Protein Carbonylation and Disease Activity Markers

PubMed Central

Almogbel, Ebtehal

2017-01-01

Introduction Free radicals have been implicated as Diabetes Mellitus (DM) contributors in type 2 DM and its associated Diabetes Mellitus Neuropathy (DMN). However, the potential for protein mediated oxidative stress to contribute disease pathogenesis remains largely unexplored. Aim To investigate the status and contribution of protein mediated oxidative stress in patients with DM or DMN and to explore whether oxidative protein modification has a role in DM progression to DM associated neuropathy. Materials and Methods Sera from 42 DM and 37 DMN patients with varying levels of disease activities biomarkers (HbA1C, patients’ age or disease duration) and 21 age- and sex-matched healthy controls were evaluated for serum levels of protein mediated oxidative stress. Results Serum analysis showed significantly higher levels of protein carbonyl contents in both DM and DMN patients compared with healthy controls. Importantly, not only was there an increased number of subjects positive for protein carbonylation, but also the levels of protein carbonyl contents were significantly higher among DM and DMN patients, whose HbA1C were ≥8.8 as compared with patients with lower HbA1C (HbA1C<8.8). Similar pattern of protein carbonyls formation was also observed with patients’ ages or with patient’s disease durations, suggesting a possible relationship between protein oxidation and disease progression. Furthermore, sera from DMN patients had higher levels of protein carbonylation compared with non-neuropathic DM patients’ sera, suggesting an involvement of protein oxidation in the progression of diabetes to diabetes neuropathy. Conclusion These findings support an association between protein oxidation and DM or DMN progression. The stronger response observed in patients with higher HbA1C or patients’ ages or disease durations suggests, that protein mediated oxidative stress may be useful in evaluating the progression of DM and its associated DMN and in elucidating the mechanisms of these disorders pathogenesis. PMID:28384853

Extracellular deoxyribonuclease made by group A Streptococcus assists pathogenesis by enhancing evasion of the innate immune response.

PubMed

Sumby, Paul; Barbian, Kent D; Gardner, Donald J; Whitney, Adeline R; Welty, Diane M; Long, R Daniel; Bailey, John R; Parnell, Michael J; Hoe, Nancy P; Adams, Gerald G; Deleo, Frank R; Musser, James M

2005-02-01

Many pathogenic bacteria produce extracellular DNase, but the benefit of this enzymatic activity is not understood. For example, all strains of the human bacterial pathogen group A Streptococcus (GAS) produce at least one extracellular DNase, and most strains make several distinct enzymes. Despite six decades of study, it is not known whether production of DNase by GAS enhances virulence. To test the hypothesis that extracellular DNase is required for normal progression of GAS infection, we generated seven isogenic mutant strains in which the three chromosomal- and prophage-encoded DNases made by a contemporary serotype M1 GAS strain were inactivated. Compared to the wild-type parental strain, the isogenic triple-mutant strain was significantly less virulent in two mouse models of invasive infection. The triple-mutant strain was cleared from the skin injection site significantly faster than the wild-type strain. Preferential clearance of the mutant strain was related to the differential extracellular killing of the mutant and wild-type strains, possibly through degradation of neutrophil extracellular traps, innate immune structures composed of chromatin and granule proteins. The triple-mutant strain was also significantly compromised in its ability to cause experimental pharyngeal disease in cynomolgus macaques. Comparative analysis of the seven DNase mutant strains strongly suggested that the prophage-encoded SdaD2 enzyme is the major DNase that contributes to virulence in this clone. We conclude that extracellular DNase activity made by GAS contributes to disease progression, thereby resolving a long-standing question in bacterial pathogenesis research.

Overview of the cellular and molecular basis of kidney fibrosis

PubMed Central

Eddy, Allison A

2014-01-01

The common pathogenetic pathway of progressive injury in patients with chronic kidney disease (CKD) is epitomized as normal kidney parenchymal destruction due to scarring (fibrosis). Understanding the fundamental pathways that lead to renal fibrosis is essential in order to develop better therapeutic options for human CKD. Although complex, four cellular responses are pivotal. (1) An interstitial inflammatory response that has multiple consequences—some harmful and others healing. (2) The appearance of a unique interstitial cell population of myofibroblasts, primarily derived from kidney stromal cells (fibroblasts and pericytes), that are the primary source of the various extracellular matrix proteins that form interstitial scars. (3) Tubular epithelial cells that have variable and time-dependent roles as early responders to injury and later as victims of fibrosis due to the loss of their regenerative abilities. (4) Loss of interstitial capillary integrity that compromises oxygen delivery and leads to a vicious cascade of hypoxia–oxidant stress that accentuates injury and fibrosis. In the absence of adequate angiogenic responses, a healthy interstitial capillary network is not maintained. The fibrotic ‘scar' that typifies CKD is an interesting consortium of multifunctional macromolecules that not only change in composition and structure over time, but can be degraded via extracellular and intracellular proteases. Although transforming growth factor beta appears to be the primary driver of kidney fibrosis, a vast array of additional molecules may have modulating roles. The importance of genetic and epigenetic factors is increasingly appreciated. An intriguing but incompletely understood cardiorenal syndrome underlies the high morbidity and mortality rates that develop in association with progressive kidney fibrosis. PMID:25401038

Pharmacologic inhibition of Pim kinases alters prostate cancer cell growth and resensitizes chemoresistant cells to taxanes.

PubMed

Mumenthaler, Shannon M; Ng, Patricia Y B; Hodge, Amanda; Bearss, David; Berk, Gregory; Kanekal, Sarath; Redkar, Sanjeev; Taverna, Pietro; Agus, David B; Jain, Anjali

2009-10-01

The serine/threonine family of Pim kinases function as oncogenes and have been implicated in prostate cancer progression, particularly in hormone-refractory prostate disease, as a result of their antiapoptotic function. In this study, we used a pharmacologic inhibitor targeting the Pim family members, SGI-1776, to determine whether modulation of Pim kinase activity could alter prostate cancer cell survival and modulate chemotherapy resistance. Extensive biochemical characterization of SGI-1776 confirmed its specificity for the three isoforms of the Pim family. Treatment of prostate cancer cells with SGI-1776 resulted in a dose-dependent reduction in phosphorylation of known Pim kinase substrates that are involved in cell cycle progression and apoptosis (p21(Cip1/WAF1) and Bad). Consequently, SGI-1776 compromised overall cell viability by inducing G(1) cell cycle arrest and triggering apoptosis. Overexpression of recombinant Pim-1 markedly increased sensitivity of SGI-1776-mediated prostate cancer cell apoptosis and p21(Cip1/WAF1) phosphorylation inhibition, reinforcing the specificity of SGI-1776. An additional cytotoxic effect was observed when SGI-1776 was combined with taxane-based chemotherapy agents. SGI-1776 was able to reduce cell viability in a multidrug resistance 1 protein-based taxane-refractory prostate cancer cell line. In addition, SGI-1776 treatment was able to resensitize chemoresistant cells to taxane-based therapies by inhibiting multidrug resistance 1 activity and inducing apoptosis. These findings support the idea that inhibiting Pim kinases, in combination with a chemotherapeutic agent, could play an important role in prostate cancer treatment by targeting the clinical problem of chemoresistance.

A peptide-retrieval strategy enables significant improvement of quantitative performance without compromising confidence of identification.

PubMed

Tu, Chengjian; Shen, Shichen; Sheng, Quanhu; Shyr, Yu; Qu, Jun

2017-01-30

Reliable quantification of low-abundance proteins in complex proteomes is challenging largely owing to the limited number of spectra/peptides identified. In this study we developed a straightforward method to improve the quantitative accuracy and precision of proteins by strategically retrieving the less confident peptides that were previously filtered out using the standard target-decoy search strategy. The filtered-out MS/MS spectra matched to confidently-identified proteins were recovered, and the peptide-spectrum-match FDR were re-calculated and controlled at a confident level of FDR≤1%, while protein FDR maintained at ~1%. We evaluated the performance of this strategy in both spectral count- and ion current-based methods. >60% increase of total quantified spectra/peptides was respectively achieved for analyzing a spike-in sample set and a public dataset from CPTAC. Incorporating the peptide retrieval strategy significantly improved the quantitative accuracy and precision, especially for low-abundance proteins (e.g. one-hit proteins). Moreover, the capacity of confidently discovering significantly-altered proteins was also enhanced substantially, as demonstrated with two spike-in datasets. In summary, improved quantitative performance was achieved by this peptide recovery strategy without compromising confidence of protein identification, which can be readily implemented in a broad range of quantitative proteomics techniques including label-free or labeling approaches. We hypothesize that more quantifiable spectra and peptides in a protein, even including less confident peptides, could help reduce variations and improve protein quantification. Hence the peptide retrieval strategy was developed and evaluated in two spike-in sample sets with different LC-MS/MS variations using both MS1- and MS2-based quantitative approach. The list of confidently identified proteins using the standard target-decoy search strategy was fixed and more spectra/peptides with less confidence matched to confident proteins were retrieved. However, the total peptide-spectrum-match false discovery rate (PSM FDR) after retrieval analysis was still controlled at a confident level of FDR≤1%. As expected, the penalty for occasionally incorporating incorrect peptide identifications is negligible by comparison with the improvements in quantitative performance. More quantifiable peptides, lower missing value rate, better quantitative accuracy and precision were significantly achieved for the same protein identifications by this simple strategy. This strategy is theoretically applicable for any quantitative approaches in proteomics and thereby provides more quantitative information, especially on low-abundance proteins. Published by Elsevier B.V.

Detection and quantification of proteins and cells by use of elemental mass spectrometry: progress and challenges.

PubMed

Yan, Xiaowen; Yang, Limin; Wang, Qiuquan

2013-07-01

Much progress has been made in identification of the proteins in proteomes, and quantification of these proteins has attracted much interest. In addition to popular tandem mass spectrometric methods based on soft ionization, inductively coupled plasma mass spectrometry (ICPMS), a typical example of mass spectrometry based on hard ionization, usually used for analysis of elements, has unique advantages in absolute quantification of proteins by determination of an element with a definite stoichiometry in a protein or attached to the protein. In this Trends article, we briefly describe state-of-the-art ICPMS-based methods for quantification of proteins, emphasizing protein-labeling and element-tagging strategies developed on the basis of chemically selective reactions and/or biospecific interactions. Recent progress from protein to cell quantification by use of ICPMS is also discussed, and the possibilities and challenges of ICPMS-based protein quantification for universal, selective, or targeted quantification of proteins and cells in a biological sample are also discussed critically. We believe ICPMS-based protein quantification will become ever more important in targeted quantitative proteomics and bioanalysis in the near future.

Autophagy Deficiency Compromises Alternative Pathways of Respiration following Energy Deprivation in Arabidopsis thaliana.

PubMed

Barros, Jessica A S; Cavalcanti, João Henrique F; Medeiros, David B; Nunes-Nesi, Adriano; Avin-Wittenberg, Tamar; Fernie, Alisdair R; Araújo, Wagner L

2017-09-01

Under heterotrophic conditions, carbohydrate oxidation inside the mitochondrion is the primary energy source for cellular metabolism. However, during energy-limited conditions, alternative substrates are required to support respiration. Amino acid oxidation in plant cells plays a key role in this by generating electrons that can be transferred to the mitochondrial electron transport chain via the electron transfer flavoprotein/ubiquinone oxidoreductase system. Autophagy, a catabolic mechanism for macromolecule and protein recycling, allows the maintenance of amino acid pools and nutrient remobilization. Although the association between autophagy and alternative respiratory substrates has been suggested, the extent to which autophagy and primary metabolism interact to support plant respiration remains unclear. To investigate the metabolic importance of autophagy during development and under extended darkness, Arabidopsis ( Arabidopsis thaliana ) mutants with disruption of autophagy ( atg mutants) were used. Under normal growth conditions, atg mutants showed lower growth and seed production with no impact on photosynthesis. Following extended darkness, atg mutants were characterized by signatures of early senescence, including decreased chlorophyll content and maximum photochemical efficiency of photosystem II coupled with increases in dark respiration. Transcript levels of genes involved in alternative pathways of respiration and amino acid catabolism were up-regulated in atg mutants. The metabolite profiles of dark-treated leaves revealed an extensive metabolic reprogramming in which increases in amino acid levels were partially compromised in atg mutants. Although an enhanced respiration in atg mutants was observed during extended darkness, autophagy deficiency compromises protein degradation and the generation of amino acids used as alternative substrates to the respiration. © 2017 American Society of Plant Biologists. All Rights Reserved.

Zebrafish Staufen1 and Staufen2 are required for the survival and migration of primordial germ cells.

PubMed

Ramasamy, Srinivas; Wang, Hui; Quach, Helen Ngoc Bao; Sampath, Karuna

2006-04-15

In sexually reproducing organisms, primordial germ cells (PGCs) give rise to the cells of the germ line, the gametes. In many animals, PGCs are set apart from somatic cells early during embryogenesis. Work in Drosophila, C. elegans, Xenopus, and zebrafish has shown that maternally provided localized cytoplasmic determinants specify the germ line in these organisms (Raz, E., 2003. Primordial germ-cell development: the zebrafish perspective. Nat. Rev., Genet. 4, 690--700; Santos, A.C., Lehmann, R., 2004. Germ cell specification and migration in Drosophila and beyond. Curr. Biol. 14, R578-R589). The Drosophila RNA-binding protein, Staufen is required for germ cell formation, and mutations in stau result in a maternal effect grandchild-less phenotype (Schupbach,T., Weischaus, E., 1989. Female sterile mutations on the second chromosome of Drosophila melanogaster:1. Maternal effect mutations. Genetics 121, 101-17). Here we describe the functions of two zebrafish Staufen-related proteins, Stau1 and Stau2. When Stau1 or Stau2 functions are compromised in embryos by injecting antisense morpholino modified oligonucleotides or dominant-negative Stau peptides, germ layer patterning is not affected. However, expression of the PGC marker vasa is not maintained. Furthermore, expression of a green fluorescent protein (GFP):nanos 3'UTR fusion protein in germ cells shows that PGC migration is aberrant, and the mis-migrating PGCs do not survive in Stau-compromised embryos. Stau2 is also required for survival of neurons in the central nervous system (CNS). These phenotypes are rescued by co-injection of Drosophila stau mRNA. Thus, staufen has an evolutionarily conserved function in germ cells. In addition, we have identified a function for Stau proteins in PGC migration.

The fluorescence intensities ratio is not a reliable parameter for evaluation of protein unfolding transitions.

PubMed

Žoldák, Gabriel; Jancura, Daniel; Sedlák, Erik

2017-06-01

Monitoring the fluorescence of proteins, particularly the fluorescence of intrinsic tryptophan residues, is a popular method often used in the analysis of unfolding transitions (induced by temperature, chemical denaturant, and pH) in proteins. The tryptophan fluorescence provides several suitable parameters, such as steady-state fluorescence intensity, apparent quantum yield, mean fluorescence lifetime, position of emission maximum that are often utilized for the observation of the conformational/unfolding transitions of proteins. In addition, the fluorescence intensities ratio at different wavelengths (usually at 330 nm and 350 nm) is becoming an increasingly popular parameter for the evaluation of thermal transitions. We show that, under certain conditions, the use of this parameter for the analysis of unfolding transitions leads to the incorrect determination of thermodynamic parameters characterizing unfolding transitions in proteins (e.g., melting temperature) and, hence, can compromise the hit identification during high-throughput drug screening campaigns. © 2017 The Protein Society.

MitoCPR-A surveillance pathway that protects mitochondria in response to protein import stress.

PubMed

Weidberg, Hilla; Amon, Angelika

2018-04-13

Mitochondrial functions are essential for cell viability and rely on protein import into the organelle. Various disease and stress conditions can lead to mitochondrial import defects. We found that inhibition of mitochondrial import in budding yeast activated a surveillance mechanism, mitoCPR, that improved mitochondrial import and protected mitochondria during import stress. mitoCPR induced expression of Cis1, which associated with the mitochondrial translocase to reduce the accumulation of mitochondrial precursor proteins at the mitochondrial translocase. Clearance of precursor proteins depended on the Cis1-interacting AAA + adenosine triphosphatase Msp1 and the proteasome, suggesting that Cis1 facilitates degradation of unimported proteins. mitoCPR was required for maintaining mitochondrial functions when protein import was compromised, demonstrating the importance of mitoCPR in protecting the mitochondrial compartment. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

Histologic Changes Associated With Placental Separation in Gilts Infected with Porcine Reproductive and Respiratory Syndrome Virus.

PubMed

Novakovic, Predrag; Detmer, Susan E; Suleman, Muhammad; Malgarin, Carol M; MacPhee, Daniel J; Harding, John C S

2018-07-01

The placenta is a vital organ providing the developing fetus with nutrient and gas exchange, thermoregulation, and waste elimination necessary for fetal development, as well as producing hormones to maintain pregnancy. It is hypothesized that fetal pig death in porcine reproductive and respiratory syndrome may be attributed to pathology of the maternal-fetal interface leading to premature placental separation. This study was designed to evaluate the chronologic progression of porcine reproductive and respiratory syndrome virus (PRRSV)-induced lesions at the maternal-fetal interface, with particular focus on placental separation in experimentally challenged third-trimester gilts. Fifteen gilts were inoculated with a virulent strain of PRRSV-2 on gestation day 86 ± 0.4. On multiple days postinoculation, 3 gilts along with 1 sham-inoculated control per time point were euthanized, and uterine and fetal placental tissues corresponding to each fetus were collected for histopathologic evaluation. The presence of any fetal lesion was 23 times more likely in compromised (meconium-stained and decomposed) compared with viable fetuses ( P < .001). In PRRSV-infected gilts, endometritis was more severe than placentitis, and the severity of endometrial inflammation and vasculitis increased progressively from 2 to 14 days postinoculation. Neither placental vasculitis nor a chronologic progression in the severity of placental detachment was observed. Severe placental detachment was more frequently present in PRRSV-infected compared with noninfected samples and was most significantly associated with placental inflammation, compared with other uterine lesions, viral load, or termination day. The results of this study suggest that placental separation by itself is not sufficient to significantly compromise fetal viability in reproductive porcine reproductive and respiratory syndrome.

DIBROMOACETIC ACID, A PREVALENT BY-PRODUCT OF DRINKING WATER DISINFECTION, COMPROMISES THE SYNTHESIS OF SPECIFIC SEMINFEROUS TUBULE PROTEINS FOLLOWING BOTH IN VIVO AND IN VITRO EXPOSURES

EPA Science Inventory

ABSTRACT
Dibromoacetic acid(DBA) is a byproduct of drinking water disinfection that alters spermatogenesis in adult male rats. To identify a mechanism by which DBA alters spermatogenesis, seminiferous tubules representing specific groups of spermatogenic stages were expos...

Autophagy and oxidative stress in non-communicable diseases: A matter of the inflammatory state?

PubMed

Peña-Oyarzun, Daniel; Bravo-Sagua, Roberto; Diaz-Vega, Alexis; Aleman, Larissa; Chiong, Mario; Garcia, Lorena; Bambs, Claudia; Troncoso, Rodrigo; Cifuentes, Mariana; Morselli, Eugenia; Ferreccio, Catterina; Quest, Andrew F G; Criollo, Alfredo; Lavandero, Sergio

2018-05-30

Non-communicable diseases (NCDs), also known as chronic diseases, are long-lasting conditions that affect millions of people around the world. Different factors contribute to their genesis and progression; however they share common features, which are critical for the development of novel therapeutic strategies. A persistently altered inflammatory response is typically observed in many NCDs together with redox imbalance. Additionally, dysregulated proteostasis, mainly derived as a consequence of compromised autophagy, is a common feature of several chronic diseases. In this review, we discuss the crosstalk among inflammation, autophagy and oxidative stress, and how they participate in the progression of chronic diseases such as cancer, cardiovascular diseases, obesity and type II diabetes mellitus. Copyright © 2018 Elsevier Inc. All rights reserved.

The Global Sustainability Index: An Instrument For Assessing The Progress Towards The Sustainable Organization

NASA Astrophysics Data System (ADS)

Grecu, Valentin

2015-09-01

There is rarely an optimal solution in sustainable development but most frequently a need to build compromises between conflicting aspects such as economic, social and environmental ones and different expectations of stakeholders. Moreover, information is rarely available and precise. This paper will focus on how to use indicators to monitor sustainable development, integrating the information provided by many of them into a complex general sustainability index. Having this general indicator is essential for decision makers as it is very complicated to evaluate the performance of the organization based on multiple indicators. The objective of this paper is to find mathematical algorithms for simplifying the decision-making process by offering an instrument for the evaluation of the sustainability progress.

High Fat Diets Induce Colonic Epithelial Cell Stress and Inflammation that is Reversed by IL-22

PubMed Central

Gulhane, Max; Murray, Lydia; Lourie, Rohan; Tong, Hui; Sheng, Yong H.; Wang, Ran; Kang, Alicia; Schreiber, Veronika; Wong, Kuan Yau; Magor, Graham; Denman, Stuart; Begun, Jakob; Florin, Timothy H.; Perkins, Andrew; Cuív, Páraic Ó.; McGuckin, Michael A.; Hasnain, Sumaira Z.

2016-01-01

Prolonged high fat diets (HFD) induce low-grade chronic intestinal inflammation in mice, and diets high in saturated fat are a risk factor for the development of human inflammatory bowel diseases. We hypothesized that HFD-induced endoplasmic reticulum (ER)/oxidative stress occur in intestinal secretory goblet cells, triggering inflammatory signaling and reducing synthesis/secretion of proteins that form the protective mucus barrier. In cultured intestinal cells non-esterified long-chain saturated fatty acids directly increased oxidative/ER stress leading to protein misfolding. A prolonged HFD elevated the intestinal inflammatory cytokine signature, alongside compromised mucosal barrier integrity with a decrease in goblet cell differentiation and Muc2, a loss in the tight junction protein, claudin-1 and increased serum endotoxin levels. In Winnie mice, that develop spontaneous colitis, HFD-feeding increased ER stress, further compromised the mucosal barrier and increased the severity of colitis. In obese mice IL-22 reduced ER/oxidative stress and improved the integrity of the mucosal barrier, and reversed microbial changes associated with obesity with an increase in Akkermansia muciniphila. Consistent with epidemiological studies, our experiments suggest that HFDs are likely to impair intestinal barrier function, particularly in early life, which partially involves direct effects of free-fatty acids on intestinal cells, and this can be reversed by IL-22 therapy. PMID:27350069

Ion Exchangers NHX1 and NHX2 Mediate Active Potassium Uptake into Vacuoles to Regulate Cell Turgor and Stomatal Function in Arabidopsis[W][OA

PubMed Central

Barragán, Verónica; Leidi, Eduardo O.; Andrés, Zaida; Rubio, Lourdes; De Luca, Anna; Fernández, José A.; Cubero, Beatriz; Pardo, José M.

2012-01-01

Intracellular NHX proteins are Na+,K+/H+ antiporters involved in K+ homeostasis, endosomal pH regulation, and salt tolerance. Proteins NHX1 and NHX2 are the two major tonoplast-localized NHX isoforms. Here, we show that NHX1 and NHX2 have similar expression patterns and identical biochemical activity, and together they account for a significant amount of the Na+,K+/H+ antiport activity in tonoplast vesicles. Reverse genetics showed functional redundancy of NHX1 and NHX2 genes. Growth of the double mutant nhx1 nhx2 was severely impaired, and plants were extremely sensitive to external K+. By contrast, nhx1 nhx2 mutants showed similar sensitivity to salinity stress and even greater rates of Na+ sequestration than the wild type. Double mutants had reduced ability to create the vacuolar K+ pool, which in turn provoked greater K+ retention in the cytosol, impaired osmoregulation, and compromised turgor generation for cell expansion. Genes NHX1 and NHX2 were highly expressed in guard cells, and stomatal function was defective in mutant plants, further compromising their ability to regulate water relations. Together, these results show that tonoplast-localized NHX proteins are essential for active K+ uptake at the tonoplast, for turgor regulation, and for stomatal function. PMID:22438021

Etiopathology of chronic tubular, glomerular and renovascular nephropathies: Clinical implications

PubMed Central

2011-01-01

Chronic kidney disease (CKD) comprises a group of pathologies in which the renal excretory function is chronically compromised. Most, but not all, forms of CKD are progressive and irreversible, pathological syndromes that start silently (i.e. no functional alterations are evident), continue through renal dysfunction and ends up in renal failure. At this point, kidney transplant or dialysis (renal replacement therapy, RRT) becomes necessary to prevent death derived from the inability of the kidneys to cleanse the blood and achieve hydroelectrolytic balance. Worldwide, nearly 1.5 million people need RRT, and the incidence of CKD has increased significantly over the last decades. Diabetes and hypertension are among the leading causes of end stage renal disease, although autoimmunity, renal atherosclerosis, certain infections, drugs and toxins, obstruction of the urinary tract, genetic alterations, and other insults may initiate the disease by damaging the glomerular, tubular, vascular or interstitial compartments of the kidneys. In all cases, CKD eventually compromises all these structures and gives rise to a similar phenotype regardless of etiology. This review describes with an integrative approach the pathophysiological process of tubulointerstitial, glomerular and renovascular diseases, and makes emphasis on the key cellular and molecular events involved. It further analyses the key mechanisms leading to a merging phenotype and pathophysiological scenario as etiologically distinct diseases progress. Finally clinical implications and future experimental and therapeutic perspectives are discussed. PMID:21251296

Stabilization of a protein conferred by an increase in folded state entropy.

PubMed

Dagan, Shlomi; Hagai, Tzachi; Gavrilov, Yulian; Kapon, Ruti; Levy, Yaakov; Reich, Ziv

2013-06-25

Entropic stabilization of native protein structures typically relies on strategies that serve to decrease the entropy of the unfolded state. Here we report, using a combination of experimental and computational approaches, on enhanced thermodynamic stability conferred by an increase in the configurational entropy of the folded state. The enhanced stability is observed upon modifications of a loop region in the enzyme acylphosphatase and is achieved despite significant enthalpy losses. The modifications that lead to increased stability, as well as those that result in destabilization, however, strongly compromise enzymatic activity, rationalizing the preservation of the native loop structure even though it does not provide the protein with maximal stability or kinetic foldability.

Age-dependent increase of blood-brain barrier permeability and neuron-binding autoantibodies in S100B knockout mice.

PubMed

Wu, Hao; Brown, Eric V; Acharya, Nimish K; Appelt, Denah M; Marks, Alexander; Nagele, Robert G; Venkataraman, Venkat

2016-04-15

S100B is a calcium-sensor protein that impacts multiple signal transduction pathways. It is widely considered to be an important biomarker for several neuronal diseases as well as blood-brain barrier (BBB) breakdown. In this report, we demonstrate a BBB deficiency in mice that lack S100B through detection of leaked Immunoglobulin G (IgG) in the brain parenchyma. IgG leaks and IgG-binding to selected neurons were observed in S100B knockout (S100BKO) mice at 6 months of age but not at 3 months. By 9 months, IgG leaks persisted and the density of IgG-bound neurons increased significantly. These results reveal a chronic increase in BBB permeability upon aging in S100BKO mice for the first time. Moreover, coincident with the increase in IgG-bound neurons, autoantibodies targeting brain proteins were detected in the serum via western blots. These events were concurrent with compromise of neurons, increase of activated microglia and lack of astrocytic activation as evidenced by decreased expression of microtubule-associated protein type 2 (MAP2), elevated number of CD68 positive cells and unaltered expression of glial fibrillary acidic protein (GFAP) respectively. Results suggest a key role for S100B in maintaining BBB functional integrity and, further, propose the S100BKO mouse as a valuable model system to explore the link between chronic functional compromise of the BBB, generation of brain-reactive autoantibodies and neuronal dysfunctions. Copyright © 2016. Published by Elsevier B.V.

Role of the hydrophilic channels of simian virus 40 T-antigen helicase in DNA replication.

PubMed

Wang, Weiping; Manna, David; Simmons, Daniel T

2007-05-01

The simian virus 40 (SV40) hexameric helicase consists of a central channel and six hydrophilic channels located between adjacent large tier domains within each hexamer. To study the function of the hydrophilic channels in SV40 DNA replication, a series of single-point substitutions were introduced at sites not directly involved in protein-protein contacts. The mutants were characterized biochemically in various ways. All mutants oligomerized normally in the absence of DNA. Interestingly, 8 of the 10 mutants failed to unwind an origin-containing DNA fragment and nine of them were totally unable to support SV40 DNA replication in vitro. The mutants fell into four classes based on their biochemical properties. Class A mutants bound DNA normally and had normal ATPase and helicase activities but failed to unwind origin DNA and support SV40 DNA replication. Class B mutants were compromised in single-stranded DNA and origin DNA binding at low protein concentrations. They were defective in helicase activity and unwinding of the origin and in supporting DNA replication. Class C and D mutants possessed higher-than-normal single-stranded DNA binding activity at low protein concentrations. The class C mutants failed to separate origin DNA and support DNA replication. The class D mutants unwound origin DNA normally but were compromised in their ability to support DNA replication. Taken together, these results suggest that the hydrophilic channels have an active role in the unwinding of SV40 DNA from the origin and the placement of the resulting single strands within the helicase.

Progress towards the development of SH2 domain inhibitors.

PubMed

Kraskouskaya, Dziyana; Duodu, Eugenia; Arpin, Carolynn C; Gunning, Patrick T

2013-04-21

Src homology 2 (SH2) domains are 100 amino acid modular units, which recognize and bind to tyrosyl-phosphorylated peptide sequences on their target proteins, and thereby mediate intracellular protein-protein interactions. This review summarizes the progress towards the development of synthetic agents that disrupt the function of the SH2 domains in different proteins as well as the clinical relevance of targeting a specific SH2 domain. Since 1986, SH2 domains have been identified in over 110 human proteins, including kinases, transcription factors, and adaptor proteins. A number of these proteins are over-activated in many diseases, including cancer, and their function is highly dependent on their SH2 domain. Thus, inhibition of a protein's function through disrupting that of its SH2 domain has emerged as a promising approach towards the development of novel therapeutic modalities. Although targeting the SH2 domain is a challenging task in molecular recognition, the progress reported here demonstrates the feasibility of such an approach.

Overcoming bottlenecks in the membrane protein structural biology pipeline.

PubMed

Hardy, David; Bill, Roslyn M; Jawhari, Anass; Rothnie, Alice J

2016-06-15

Membrane proteins account for a third of the eukaryotic proteome, but are greatly under-represented in the Protein Data Bank. Unfortunately, recent technological advances in X-ray crystallography and EM cannot account for the poor solubility and stability of membrane protein samples. A limitation of conventional detergent-based methods is that detergent molecules destabilize membrane proteins, leading to their aggregation. The use of orthologues, mutants and fusion tags has helped improve protein stability, but at the expense of not working with the sequence of interest. Novel detergents such as glucose neopentyl glycol (GNG), maltose neopentyl glycol (MNG) and calixarene-based detergents can improve protein stability without compromising their solubilizing properties. Styrene maleic acid lipid particles (SMALPs) focus on retaining the native lipid bilayer of a membrane protein during purification and biophysical analysis. Overcoming bottlenecks in the membrane protein structural biology pipeline, primarily by maintaining protein stability, will facilitate the elucidation of many more membrane protein structures in the near future. © 2016 The Author(s). published by Portland Press Limited on behalf of the Biochemical Society.

Pakistan and the Millennium Development Goals for Maternal and Child Health: progress and the way forward.

PubMed

Rizvi, Arjumand; Bhatti, Zaid; Das, Jai K; Bhutta, Zulfiqar A

2015-04-03

The world has made substantial progress in reducing maternal and child mortality, but many countries are projected to fall short of achieving their Millennium Development Goals (MDGs) 4 and 5 targets. The major objective of this paper is to examine progress in Pakistan in reducing maternal and child mortality and malnutrition over the last two decades. Data from recent national and international surveys suggest that Pakistan lags behind on all of its MDGs related to maternal and child health and, for some indicators especially related to nutrition, the situation has worsened from the baseline of 1990. Progress in addressing key social determinants such as poverty, female education and empowerment has also been slow and unregulated population growth has further compromised progress. There is a need to integrate the various different sectors and programmes to achieve the desired results effectively and efficiently as many of the determinants and influencing factors are outside the health sector. Pakistan has to accelerate improvement of access to maternal health services, particularly contraception, emergency obstetric care and skilled birth attendance; the need to improve maternal and child nutrition cannot be over-emphasised.

Comprehensive proteomic profiles of mouse AApoAII amyloid fibrils provide insights into the involvement of lipoproteins in the pathology of amyloidosis.

PubMed

Miyahara, Hiroki; Sawashita, Jinko; Ishikawa, Eri; Yang, Mu; Ding, Xin; Liu, Yingye; Hachiya, Naomi; Kametani, Fuyuki; Yazaki, Masahide; Mori, Masayuki; Higuchi, Keiichi

2018-02-10

Amyloidosis is a disorder characterized by extracellular fibrillar deposits of misfolded proteins. The amyloid deposits commonly contain several non-fibrillar proteins as amyloid-associated proteins, but their roles in amyloidosis pathology are still unknown. In mouse senile amyloidosis, apolipoprotein A-II (ApoA-II) forms extracellular amyloid fibril (AApoAII) deposits with other proteins (AApoAII-associated proteins) in many organs. We previously reported that R1.P1-Apoa2 c mice provide a reproducible model of AApoAII amyloidosis. In order to investigate the sequential alterations of AApoAII-associated protein, we performed a proteomic analysis of amyloid fibrils extracted from mouse liver tissues that contained different levels of AApoAII deposition. We identified 6 AApoAII-associated proteins that constituted 20 of the top-ranked proteins in mice with severe AApoAII deposition. Although the amount of AApoAII-associated proteins increased with the progression of amyloidosis, the relative abundance of AApoAII-associated proteins changed little throughout the progression of amyloidosis. On the other hand, plasma levels of these proteins showed dramatic changes during the progression of amyloidosis. In addition, we confirmed that AApoAII-associated proteins were significantly associated with lipid metabolism based on functional enrichment analysis, and lipids were co-deposited with AApoAII fibrils from early stages of development of amyloidosis. Thus, these results demonstrate that lipoproteins are involved in AApoAII amyloidosis pathology. This study presented proteomic profiles of AApoAII amyloidosis during disease progression and it revealed co-deposition of lipids with AApoAII deposits based on functional analyses. The relative abundance of AApoAII-associated proteins in the amyloid fibril fractions did not change over the course of development of AApoAII amyloidosis pathology. However, their concentrations in plasma changed dramatically with progression of the disease. Interestingly, several AApoAII-associated proteins have been found as constituents of lipid-rich lesions of other degenerative diseases, such as atherosclerosis and age-related macular degeneration. The common protein components among these diseases with lipid-rich deposits could be accounted for by a lipoprotein retention model. Copyright © 2017 Elsevier B.V. All rights reserved.

A Sensitive Sensor Cell Line for the Detection of Oxidative Stress Responses in Cultured Human Keratinocytes

PubMed Central

Hofmann, Ute; Priem, Melanie; Bartzsch, Christine; Winckler, Thomas; Feller, Karl-Heinz

2014-01-01

In the progress of allergic and irritant contact dermatitis, chemicals that cause the generation of reactive oxygen species trigger a heat shock response in keratinocytes. In this study, an optical sensor cell line based on cultured human keratinocytes (HaCaT cells) expressing green fluorescent protein (GFP) under the control of the stress-inducible HSP70B' promoter were constructed. Exposure of HaCaT sensor cells to 25 μM cadmium, a model substance for oxidative stress induction, provoked a 1.7-fold increase in total glutathione and a ∼300-fold induction of transcript level of the gene coding for heat shock protein HSP70B'. An extract of Arnica montana flowers resulted in a strong induction of the HSP70B' gene and a pronounced decrease of total glutathione in keratinocytes. The HSP70B' promoter-based sensor cells conveniently detected cadmium-induced stress using GFP fluorescence as read-out with a limit of detection of 6 μM cadmium. In addition the sensor cells responded to exposure of cells to A. montana extract with induction of GFP fluorescence. Thus, the HaCaT sensor cells provide a means for the automated detection of the compromised redox status of keratinocytes as an early indicator of the development of human skin disorders and could be applied for the prediction of skin irritation in more complex in vitro 3D human skin models and in the development of micro-total analysis systems (μTAS) that may be utilized in dermatology, toxicology, pharmacology and drug screenings. PMID:24967604

Cathepsin S Contributes to the Pathogenesis of Muscular Dystrophy in Mice.

PubMed

Tjondrokoesoemo, Andoria; Schips, Tobias G; Sargent, Michelle A; Vanhoutte, Davy; Kanisicak, Onur; Prasad, Vikram; Lin, Suh-Chin J; Maillet, Marjorie; Molkentin, Jeffery D

2016-05-06

Duchenne muscular dystrophy (DMD) is an X-linked recessive disease caused by mutations in the gene encoding dystrophin. Loss of dystrophin protein compromises the stability of the sarcolemma membrane surrounding each muscle cell fiber, leading to membrane ruptures and leakiness that induces myofiber necrosis, a subsequent inflammatory response, and progressive tissue fibrosis with loss of functional capacity. Cathepsin S (Ctss) is a cysteine protease that is actively secreted in areas of tissue injury and ongoing inflammation, where it participates in extracellular matrix remodeling and healing. Here we show significant induction of Ctss expression and proteolytic activity following acute muscle injury or in muscle from mdx mice, a model of DMD. To examine the functional ramifications associated with greater Ctss expression, the Ctss gene was deleted in the mdx genetic background, resulting in protection from muscular dystrophy pathogenesis that included reduced myofiber turnover and histopathology, reduced fibrosis, and improved running capacity. Mechanistically, deletion of the Ctss gene in the mdx background significantly increased myofiber sarcolemmal membrane stability with greater expression and membrane localization of utrophin, integrins, and β-dystroglycan, which anchor the membrane to the basal lamina and underlying cytoskeletal proteins. Consistent with these results, skeletal muscle-specific transgenic mice overexpressing Ctss showed increased myofiber necrosis, muscle histopathology, and a functional deficit reminiscent of muscular dystrophy. Hence, Ctss induction during muscular dystrophy is a pathologic event that partially underlies disease pathogenesis, and its inhibition might serve as a new therapeutic strategy in DMD. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Intergenic Transcriptional Interference Is Blocked by RNA Polymerase III Transcription Factor TFIIIB in Saccharomyces cerevisiae

PubMed Central

Korde, Asawari; Rosselot, Jessica M.; Donze, David

2014-01-01

The major function of eukaryotic RNA polymerase III is to transcribe transfer RNA, 5S ribosomal RNA, and other small non-protein-coding RNA molecules. Assembly of the RNA polymerase III complex on chromosomal DNA requires the sequential binding of transcription factor complexes TFIIIC and TFIIIB. Recent evidence has suggested that in addition to producing RNA transcripts, chromatin-assembled RNA polymerase III complexes may mediate additional nuclear functions that include chromatin boundary, nucleosome phasing, and general genome organization activities. This study provides evidence of another such “extratranscriptional” activity of assembled RNA polymerase III complexes, which is the ability to block progression of intergenic RNA polymerase II transcription. We demonstrate that the RNA polymerase III complex bound to the tRNA gene upstream of the Saccharomyces cerevisiae ATG31 gene protects the ATG31 promoter against readthrough transcriptional interference from the upstream noncoding intergenic SUT467 transcription unit. This protection is predominately mediated by binding of the TFIIIB complex. When TFIIIB binding to this tRNA gene is weakened, an extended SUT467–ATG31 readthrough transcript is produced, resulting in compromised ATG31 translation. Since the ATG31 gene product is required for autophagy, strains expressing the readthrough transcript exhibit defective autophagy induction and reduced fitness under autophagy-inducing nitrogen starvation conditions. Given the recent discovery of widespread pervasive transcription in all forms of life, protection of neighboring genes from intergenic transcriptional interference may be a key extratranscriptional function of assembled RNA polymerase III complexes and possibly other DNA binding proteins. PMID:24336746

Chondrocytes, synoviocytes and dermal fibroblasts all express PH-20, a hyaluronidase active at neutral pH

PubMed Central

El Hajjaji, Hafida; Cole, Ada Asbury; Manicourt, Daniel-Henri

2005-01-01

Hyaluronan (HA), an important component of connective tissues, is highly metabolically active, but the mechanisms involved in its catabolism are still largely unknown. We hypothesized that a protein similar to sperm PH-20, the only mammalian hyaluronidase known to be active at neutral pH, could be expressed in connective tissue cells. An mRNA transcript similar to that of PH-20 was found in chondrocytes, synoviocytes, and dermal fibroblasts, and its levels were enhanced upon stimulation with IL-1. In cell layers extracted with Triton X-100 – but not with octylglucoside – and in culture media, a polyclonal antipeptide anti-PH-20 antibody identified protein bands with a molecular weight similar to that of sperm PH-20 (60 to 65 kDa) and exhibiting a hyaluronidase activity at neutral pH. Further, upon stimulation with IL-1, the amounts of the neutral-active hyaluronidase increased in both cell layers and culture media. These findings contribute potential important new insights into the biology of connective tissues. It is likely that PH-20 facilitates cell-receptor-mediated uptake of HA, while overexpression or uncontrolled expression of the enzyme can cause great havoc to connective tissues: not only does HA fragmentation compromise the structural integrity of tissues, but also the HA fragments generated are highly angiogenic and are potent inducers of proinflammatory cytokines. On the other hand, the enzyme activity may account for the progressive depletion of HA seen in osteoarthritis cartilage, a depletion that is believed to play an important role in the apparent irreversibility of this disease process. PMID:15987477

Cranial irradiation compromises neuronal architecture in the hippocampus.

PubMed

Parihar, Vipan Kumar; Limoli, Charles L

2013-07-30

Cranial irradiation is used routinely for the treatment of nearly all brain tumors, but may lead to progressive and debilitating impairments of cognitive function. Changes in synaptic plasticity underlie many neurodegenerative conditions that correlate to specific structural alterations in neurons that are believed to be morphologic determinants of learning and memory. To determine whether changes in dendritic architecture might underlie the neurocognitive sequelae found after irradiation, we investigated the impact of cranial irradiation (1 and 10 Gy) on a range of micromorphometric parameters in mice 10 and 30 d following exposure. Our data revealed significant reductions in dendritic complexity, where dendritic branching, length, and area were routinely reduced (>50%) in a dose-dependent manner. At these same doses and times we found significant reductions in the number (20-35%) and density (40-70%) of dendritic spines on hippocampal neurons of the dentate gyrus. Interestingly, immature filopodia showed the greatest sensitivity to irradiation compared with more mature spine morphologies, with reductions of 43% and 73% found 30 d after 1 and 10 Gy, respectively. Analysis of granule-cell neurons spanning the subfields of the dentate gyrus revealed significant reductions in synaptophysin expression at presynaptic sites in the dentate hilus, and significant increases in postsynaptic density protein (PSD-95) were found along dendrites in the granule cell and molecular layers. These findings are unique in demonstrating dose-responsive changes in dendritic complexity, synaptic protein levels, spine density and morphology, alterations induced in hippocampal neurons by irradiation that persist for at least 1 mo, and that resemble similar types of changes found in many neurodegenerative conditions.

Use of principal components analysis and protein microarray to explore the association of HIV-1-specific IgG responses with disease progression.

PubMed

Gerns Storey, Helen L; Richardson, Barbra A; Singa, Benson; Naulikha, Jackie; Prindle, Vivian C; Diaz-Ochoa, Vladimir E; Felgner, Phil L; Camerini, David; Horton, Helen; John-Stewart, Grace; Walson, Judd L

2014-01-01

The role of HIV-1-specific antibody responses in HIV disease progression is complex and would benefit from analysis techniques that examine clusterings of responses. Protein microarray platforms facilitate the simultaneous evaluation of numerous protein-specific antibody responses, though excessive data are cumbersome in analyses. Principal components analysis (PCA) reduces data dimensionality by generating fewer composite variables that maximally account for variance in a dataset. To identify clusters of antibody responses involved in disease control, we investigated the association of HIV-1-specific antibody responses by protein microarray, and assessed their association with disease progression using PCA in a nested cohort design. Associations observed among collections of antibody responses paralleled protein-specific responses. At baseline, greater antibody responses to the transmembrane glycoprotein (TM) and reverse transcriptase (RT) were associated with higher viral loads, while responses to the surface glycoprotein (SU), capsid (CA), matrix (MA), and integrase (IN) proteins were associated with lower viral loads. Over 12 months greater antibody responses were associated with smaller decreases in CD4 count (CA, MA, IN), and reduced likelihood of disease progression (CA, IN). PCA and protein microarray analyses highlighted a collection of HIV-specific antibody responses that together were associated with reduced disease progression, and may not have been identified by examining individual antibody responses. This technique may be useful to explore multifaceted host-disease interactions, such as HIV coinfections.

Deep dysphasic performance in non-fluent progressive aphasia: a case study.

PubMed

Tree, J J; Perfect, T J; Hirsh, K W; Copstick, S

2001-01-01

We present a patient (PW) with non-fluent progressive aphasia, characterized by severe word finding difficulties and frequent phonemic paraphasias in spontaneous speech. It has been suggested that such patients have insufficient access to phonological information for output and cannot construct the appropriate sequence of selected phonemes for articulation. Consistent with such a proposal, we found that PW was impaired on a variety of verbal tasks that demand access to phonological representations (reading, repetition, confrontational naming and rhyme judgement); she also demonstrated poor performance on syntactic and grammatical processing tasks. However, examination of PW's repetition performance also revealed that she made semantic paraphasias and that her performance was influenced by imageability and lexical status. Her auditory-verbal short-term memory was also severely compromised. These features are consistent with 'deep dysphasia', a disorder reported in patients suffering from stroke or cerebrovascular accident, and rarely reported in the context of non-fluent progressive aphasia. PW's pattern of performance is evaluated in terms of current models of both non-fluent progressive aphasia and deep dysphasia.

Liver Inflammation and Metabolic Signaling in ApcMin/+ Mice: The Role of Cachexia Progression

PubMed Central

Narsale, Aditi A.; Enos, Reilly T.; Puppa, Melissa J.; Chatterjee, Saurabh; Murphy, E. Angela; Fayad, Raja; Pena, Majorette O’; Durstine, J. Larry; Carson, James A.

2015-01-01

The ApcMin/+ mouse exhibits an intestinal tumor associated loss of muscle and fat that is accompanied by chronic inflammation, insulin resistance and hyperlipidemia. Since the liver governs systemic energy demands through regulation of glucose and lipid metabolism, it is likely that the liver is a pathological target of cachexia progression in the ApcMin/+ mouse. The purpose of this study was to determine if cancer and the progression of cachexia affected liver endoplasmic reticulum (ER)-stress, inflammation, metabolism, and protein synthesis signaling. The effect of cancer (without cachexia) was examined in wild-type and weight-stable ApcMin/+ mice. Cachexia progression was examined in weight-stable, pre-cachectic, and severely-cachectic ApcMin/+ mice. Livers were analyzed for morphology, glycogen content, ER-stress, inflammation, and metabolic changes. Cancer induced hepatic expression of ER-stress markers BiP (binding immunoglobulin protein), IRE-1α (endoplasmic reticulum to nucleus signaling 1), and inflammatory intermediate STAT-3 (signal transducer and activator of transcription 3). While gluconeogenic enzyme phosphoenolpyruvate carboxykinase (PEPCK) mRNA expression was suppressed by cancer, glycogen content or protein synthesis signaling remained unaffected. Cachexia progression depleted liver glycogen content and increased mRNA expression of glycolytic enzyme PFK (phosphofrucktokinase) and gluconeogenic enzyme PEPCK. Cachexia progression further increased pSTAT-3 but suppressed p-65 and JNK (c-Jun NH2-terminal kinase) activation. Interestingly, progression of cachexia suppressed upstream ER-stress markers BiP and IRE-1α, while inducing its downstream target CHOP (DNA-damage inducible transcript 3). Cachectic mice exhibited a dysregulation of protein synthesis signaling, with an induction of p-mTOR (mechanistic target of rapamycin), despite a suppression of Akt (thymoma viral proto-oncogene 1) and S6 (ribosomal protein S6) phosphorylation. Thus, cancer induced ER-stress markers in the liver, however cachexia progression further deteriorated liver ER-stress, disrupted protein synthesis regulation and caused a differential inflammatory response related to STAT-3 and NF-κB (Nuclear factor—κB) signaling. PMID:25789991

Mass spectrometry-based carboxyl footprinting of proteins: Method evaluation

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

Zhang, Hao; Wen, Jianzhong; Huang, Richard Y-C.

2012-02-01

Protein structure determines function in biology, and a variety of approaches have been employed to obtain structural information about proteins. Mass spectrometry-based protein footprinting is one fast-growing approach. One labeling-based footprinting approach is the use of a water-soluble carbodiimide, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and glycine ethyl ester (GEE) to modify solvent-accessible carboxyl groups on glutamate (E) and aspartate (D). This paper describes method development of carboxyl-group modification in protein footprinting. The modification protocol was evaluated by using the protein calmodulin as a model. Because carboxyl-group modification is a slow reaction relative to protein folding and unfolding, there is an issue that modificationsmore » at certain sites may induce protein unfolding and lead to additional modification at sites that are not solvent-accessible in the wild-type protein. We investigated this possibility by using hydrogen deuterium amide exchange (H/DX). The study demonstrated that application of carboxyl group modification in probing conformational changes in calmodulin induced by Ca{sup 2+} binding provides useful information that is not compromised by modification-induced protein unfolding.« less

Validation of a single biopsy approach and bolus protein feeding to determine myofibrillar protein synthesis in stable isotope tracer studies in humans

PubMed Central

2011-01-01

Background Minimizing the number of muscle biopsies has important methodological implications and minimizes subject discomfort during a stable isotope amino acid infusion. We aimed to determine the reliability of obtaining a single muscle biopsy for the calculation of muscle protein fractional synthetic rate (FSR) as well as the amount of incorporation time necessary to obtain that biopsy after initiating a stable isotope infusion (Study 1). The calculation of muscle protein FSR requires tracer steady-state during the stable isotope infusion. Therefore, a second aim was to examine if steady-state conditions are compromised in the precursor pools (plasma free or muscle intracellular [IC]) after ingestion of a tracer enriched protein drink and after resistance exercise (Study 2). Methods Sixteen men (23 ± 3 years; BMI = 23.8 ± 2.2 kg/m2, means ± SD) were randomized to perform Study 1 or Study 2 (n = 8, per study). Subjects received a primed, constant infusion of L-[ring-13C6]phenylalanine coupled with muscle biopsies of the vastus lateralis to measure rates of myofibrillar protein synthesis (MPS). Subjects in Study 2 were fed 25 g of whey protein immediately after an acute bout of unilateral resistance exercise. Results There was no difference (P = 0.3) in rates of MPS determined using the steady-state precursor-product equation and determination of tracer incorporation between sequential biopsies 150 min apart or using plasma protein as the baseline enrichment, provided the infusion length was sufficient (230 ± 0.3 min). We also found that adding a modest amount of tracer (4% enriched), calculated based on the measured phenylalanine content of the protein (3.5%) in the drink, did not compromise steady-state conditions (slope of the enrichment curve not different from zero) in the plasma free or, more importantly, the IC pool (both P > 0.05). Conclusions These data demonstrate that the single biopsy approach yields comparable rates of muscle protein synthesis, provided a longer incorporation time is utilized, to that seen with a traditional two biopsy approach. In addition, we demonstrate that enriching protein-containing drinks with tracer does not disturb isotopic steady-state and thus both are reliable techniques to determine rates of MPS in humans. PMID:21388545

Metazoan Hsp70 machines use Hsp110 to power protein disaggregation.

PubMed

Rampelt, Heike; Kirstein-Miles, Janine; Nillegoda, Nadinath B; Chi, Kang; Scholz, Sebastian R; Morimoto, Richard I; Bukau, Bernd

2012-11-05

Accumulation of aggregation-prone misfolded proteins disrupts normal cellular function and promotes ageing and disease. Bacteria, fungi and plants counteract this by solubilizing and refolding aggregated proteins via a powerful cytosolic ATP-dependent bichaperone system, comprising the AAA+ disaggregase Hsp100 and the Hsp70-Hsp40 system. Metazoa, however, lack Hsp100 disaggregases. We show that instead the Hsp110 member of the Hsp70 superfamily remodels the human Hsp70-Hsp40 system to efficiently disaggregate and refold aggregates of heat and chemically denatured proteins in vitro and in cell extracts. This Hsp110 effect relies on nucleotide exchange, not on ATPase activity, implying ATP-driven chaperoning is not required. Knock-down of nematode Caenorhabditis elegans Hsp110, but not an unrelated nucleotide exchange factor, compromises dissolution of heat-induced protein aggregates and severely shortens lifespan after heat shock. We conclude that in metazoa, Hsp70-Hsp40 powered by Hsp110 nucleotide exchange represents the crucial disaggregation machinery that reestablishes protein homeostasis to counteract protein unfolding stress.

Annexin A2 Mediates the Localization of Measles Virus Matrix Protein at the Plasma Membrane.

PubMed

Koga, Ritsuko; Kubota, Marie; Hashiguchi, Takao; Yanagi, Yusuke; Ohno, Shinji

2018-02-28

Annexins are a family of structurally related proteins that bind negatively charged membrane phospholipids in a Ca 2+ -dependent manner. Annexin A2 (AnxA2), a member of the family, has been implicated in a variety of cellular functions including the organization of membrane domains, vesicular trafficking and cell-cell adhesion. AnxA2 generally forms the heterotetrameric complex with a small Ca 2+ -binding protein S100A10. Measles virus (MV), a member of the family Paramyxoviridae , is an enveloped virus with a nonsegmented negative strand RNA genome. Knockdown of AnxA2 greatly reduced MV growth in cells, without affecting its entry and viral RNA production. In MV-infected, AnxA2-knockdown cells, the expression level of the matrix (M) protein, but not other viral proteins, was reduced compared with that in control cells, and the distribution of the M protein at the plasma membrane was decreased. The M protein lines the inner surface of the envelope and plays an important role in virus assembly by connecting the nucleocapsid to the envelope proteins. The M protein bound to AnxA2 independently of AnxA2's phosphorylation or its association with S100A10, and was co-localized with AnxA2 within cells. Truncation of the N-terminal 10 amino acid residues, but not the N-terminal 5 residues, compromised the ability of the M protein to interact with AnxA2 and localize at the plasma membrane. These results indicate that AnxA2 mediates the localization of the MV M protein at the plasma membrane by interacting with its N-terminal region (especially residues at positions 6-10), thereby aiding in MV assembly. IMPORTANCE Measles virus (MV) is an important human pathogen, still claiming ∼ 100,000 lives per year despite the presence of effective vaccines, and causes occasional outbreaks even in developed countries. Replication of viruses largely relies on the functions of host cells. Our study revealed that the reduction of the host protein annexin A2 compromises the replication of MV within the cell. Further studies demonstrated that annexin A2 interacts with the MV matrix (M) protein and mediates the localization of the M protein at the plasma membrane where MV particles are formed. The M protein lines the inner surface of the MV envelope membrane and plays a role in MV particle formation. Our results provide useful information for the understanding of the MV replication process and potential development of anti-viral agents. Copyright © 2018 American Society for Microbiology.

Linearization of the bradford protein assay.

PubMed

Ernst, Orna; Zor, Tsaffrir

2010-04-12

Determination of microgram quantities of protein in the Bradford Coomassie brilliant blue assay is accomplished by measurement of absorbance at 590 nm. This most common assay enables rapid and simple protein quantification in cell lysates, cellular fractions, or recombinant protein samples, for the purpose of normalization of biochemical measurements. However, an intrinsic nonlinearity compromises the sensitivity and accuracy of this method. It is shown that under standard assay conditions, the ratio of the absorbance measurements at 590 nm and 450 nm is strictly linear with protein concentration. This simple procedure increases the accuracy and improves the sensitivity of the assay about 10-fold, permitting quantification down to 50 ng of bovine serum albumin. Furthermore, the interference commonly introduced by detergents that are used to create the cell lysates is greatly reduced by the new protocol. A linear equation developed on the basis of mass action and Beer's law perfectly fits the experimental data.

Spontaneous tension haemopneumothorax.

PubMed

Patterson, Benjamin Oliver; Itam, Sarah; Probst, Fey

2008-10-31

We present a patient with sudden onset progressive shortness of breath and no history of trauma, who rapidly became haemodynamically compromised with a pneumothorax and pleural effusion seen on chest radiograph. He was treated for spontaneous tension pneumothorax but this was soon revealed to be a tension haemopneumothorax. He underwent urgent thoracotomy after persistent bleeding to explore an apical vascular abnormality seen on CT scanning. To our knowledge this is the first such case reported.Aetiology and current approach to spontaneous haemothorax are discussed briefly.

The Arabidopsis KINβγ Subunit of the SnRK1 Complex Regulates Pollen Hydration on the Stigma by Mediating the Level of Reactive Oxygen Species in Pollen

PubMed Central

Zhao, Ting Ting; Li, Fei; Jia, Xiao Na; Zhao, Xin-Ying; Zhang, Xian Sheng

2016-01-01

Pollen–stigma interactions are essential for pollen germination. The highly regulated process of pollen germination includes pollen adhesion, hydration, and germination on the stigma. However, the internal signaling of pollen that regulates pollen–stigma interactions is poorly understood. KINβγ is a plant-specific subunit of the SNF1-related protein kinase 1 complex which plays important roles in the regulation of plant development. Here, we showed that KINβγ was a cytoplasm- and nucleus-localized protein in the vegetative cells of pollen grains in Arabidopsis. The pollen of the Arabidopsis kinβγ mutant could not germinate on stigma, although it germinated normally in vitro. Further analysis revealed the hydration of kinβγ mutant pollen on the stigma was compromised. However, adding water to the stigma promoted the germination of the mutant pollen in vivo, suggesting that the compromised hydration of the mutant pollen led to its defective germination. In kinβγ mutant pollen, the structure of the mitochondria and peroxisomes was destroyed, and their numbers were significantly reduced compared with those in the wild type. Furthermore, we found that the kinβγ mutant exhibited reduced levels of reactive oxygen species (ROS) in pollen. The addition of H2O2 in vitro partially compensated for the reduced water absorption of the mutant pollen, and reducing ROS levels in pollen by overexpressing Arabidopsis CATALASE 3 resulted in compromised hydration of pollen on the stigma. These results indicate that Arabidopsis KINβγ is critical for the regulation of ROS levels by mediating the biogenesis of mitochondria and peroxisomes in pollen, which is required for pollen–stigma interactions during pollination. PMID:27472382

Antagonism of CD11b with neutrophil inhibitory factor (NIF) inhibits vascular lesions in diabetic retinopathy.

PubMed

Veenstra, Alexander A; Tang, Jie; Kern, Timothy S

2013-01-01

Leukocytes and proteins that govern leukocyte adhesion to endothelial cells play a causal role in retinal abnormalities characteristic of the early stages of diabetic retinopathy, including diabetes-induced degeneration of retinal capillaries. Leukocyte integrin αmβ2 (CD11b/CD18, MAC1), a protein mediating adhesion, has been shown to mediate damage to endothelial cells by activated leukocytes in vitro. We hypothesized that Neutrophil Inhibitory Factor (NIF), a selective antagonist of integrin αmβ2, would inhibit the diabetes-induced degeneration of retinal capillaries by inhibiting the excessive interaction between leukocytes and retinal endothelial cells in diabetes. Wild type animals and transgenic animals expressing NIF were made diabetic with streptozotocin and assessed for diabetes-induced retinal vascular abnormalities and leukocyte activation. To assess if the leukocyte blocking therapy compromised the immune system, animals were challenged with bacteria. Retinal superoxide production, leukostasis and leukocyte superoxide production were increased in wild type mice diabetic for 10 weeks, as was the ability of leukocytes isolated from diabetic animals to kill retinal endothelial cells in vitro. Retinal capillary degeneration was significantly increased in wild type mice diabetic 40 weeks. In contrast, mice expressing NIF did not develop any of these abnormalities, with the exception that non-diabetic and diabetic mice expressing NIF generated greater amounts of superoxide than did similar mice not expressing NIF. Importantly, NIF did not significantly impair the ability of mice to clear an opportunistic bacterial challenge, suggesting that NIF did not compromise immune surveillance. We conclude that antagonism of CD11b (integrin αmβ2) by NIF is sufficient to inhibit early stages of diabetic retinopathy, while not compromising the basic immune response.

Antagonism of CD11b with Neutrophil Inhibitory Factor (NIF) Inhibits Vascular Lesions in Diabetic Retinopathy

PubMed Central

Veenstra, Alexander A.; Tang, Jie; Kern, Timothy S.

2013-01-01

Leukocytes and proteins that govern leukocyte adhesion to endothelial cells play a causal role in retinal abnormalities characteristic of the early stages of diabetic retinopathy, including diabetes-induced degeneration of retinal capillaries. Leukocyte integrin αmβ2 (CD11b/CD18, MAC1), a protein mediating adhesion, has been shown to mediate damage to endothelial cells by activated leukocytes in vitro. We hypothesized that Neutrophil Inhibitory Factor (NIF), a selective antagonist of integrin αmβ2, would inhibit the diabetes-induced degeneration of retinal capillaries by inhibiting the excessive interaction between leukocytes and retinal endothelial cells in diabetes. Wild type animals and transgenic animals expressing NIF were made diabetic with streptozotocin and assessed for diabetes-induced retinal vascular abnormalities and leukocyte activation. To assess if the leukocyte blocking therapy compromised the immune system, animals were challenged with bacteria. Retinal superoxide production, leukostasis and leukocyte superoxide production were increased in wild type mice diabetic for 10 weeks, as was the ability of leukocytes isolated from diabetic animals to kill retinal endothelial cells in vitro. Retinal capillary degeneration was significantly increased in wild type mice diabetic 40 weeks. In contrast, mice expressing NIF did not develop any of these abnormalities, with the exception that non-diabetic and diabetic mice expressing NIF generated greater amounts of superoxide than did similar mice not expressing NIF. Importantly, NIF did not significantly impair the ability of mice to clear an opportunistic bacterial challenge, suggesting that NIF did not compromise immune surveillance. We conclude that antagonism of CD11b (integrin αmβ2) by NIF is sufficient to inhibit early stages of diabetic retinopathy, while not compromising the basic immune response. PMID:24205223

Toward high-resolution computational design of helical membrane protein structure and function

PubMed Central

Barth, Patrick; Senes, Alessandro

2016-01-01

The computational design of α-helical membrane proteins is still in its infancy but has made important progress. De novo design has produced stable, specific and active minimalistic oligomeric systems. Computational re-engineering can improve stability and modulate the function of natural membrane proteins. Currently, the major hurdle for the field is not computational, but the experimental characterization of the designs. The emergence of new structural methods for membrane proteins will accelerate progress PMID:27273630

Toward high-resolution computational design of the structure and function of helical membrane proteins.

PubMed

Barth, Patrick; Senes, Alessandro

2016-06-07

The computational design of α-helical membrane proteins is still in its infancy but has already made great progress. De novo design allows stable, specific and active minimal oligomeric systems to be obtained. Computational reengineering can improve the stability and function of naturally occurring membrane proteins. Currently, the major hurdle for the field is the experimental characterization of the designs. The emergence of new structural methods for membrane proteins will accelerate progress.

The sterol-binding activity of PATHOGENESIS-RELATED PROTEIN 1 reveals the mode of action of an antimicrobial protein.

PubMed

Gamir, Jordi; Darwiche, Rabih; Van't Hof, Pieter; Choudhary, Vineet; Stumpe, Michael; Schneiter, Roger; Mauch, Felix

2017-02-01

Pathogenesis-related proteins played a pioneering role 50 years ago in the discovery of plant innate immunity as a set of proteins that accumulated upon pathogen challenge. The most abundant of these proteins, PATHOGENESIS-RELATED 1 (PR-1) encodes a small antimicrobial protein that has become, as a marker of plant immune signaling, one of the most referred to plant proteins. The biochemical activity and mode of action of PR-1 proteins has remained elusive, however. Here, we provide genetic and biochemical evidence for the capacity of PR-1 proteins to bind sterols, and demonstrate that the inhibitory effect on pathogen growth is caused by the sequestration of sterol from pathogens. In support of our findings, sterol-auxotroph pathogens such as the oomycete Phytophthora are particularly sensitive to PR-1, whereas sterol-prototroph fungal pathogens become highly sensitive only when sterol biosynthesis is compromised. Our results are in line with previous findings showing that plants with enhanced PR-1 expression are particularly well protected against oomycete pathogens. © 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.

Dynamic Fluctuations of Protein-Carbohydrate Interactions Promote Protein Aggregation

PubMed Central

Voynov, Vladimir; Chennamsetty, Naresh; Kayser, Veysel; Helk, Bernhard; Forrer, Kurt; Zhang, Heidi; Fritsch, Cornelius; Heine, Holger; Trout, Bernhardt L.

2009-01-01

Protein-carbohydrate interactions are important for glycoprotein structure and function. Antibodies of the IgG class, with increasing significance as therapeutics, are glycosylated at a conserved site in the constant Fc region. We hypothesized that disruption of protein-carbohydrate interactions in the glycosylated domain of antibodies leads to the exposure of aggregation-prone motifs. Aggregation is one of the main problems in protein-based therapeutics because of immunogenicity concerns and decreased efficacy. To explore the significance of intramolecular interactions between aromatic amino acids and carbohydrates in the IgG glycosylated domain, we utilized computer simulations, fluorescence analysis, and site-directed mutagenesis. We find that the surface exposure of one aromatic amino acid increases due to dynamic fluctuations. Moreover, protein-carbohydrate interactions decrease upon stress, while protein-protein and carbohydrate-carbohydrate interactions increase. Substitution of the carbohydrate-interacting aromatic amino acids with non-aromatic residues leads to a significantly lower stability than wild type, and to compromised binding to Fc receptors. Our results support a mechanism for antibody aggregation via decreased protein-carbohydrate interactions, leading to the exposure of aggregation-prone regions, and to aggregation. PMID:20037630

An alternative easy method for antibody purification and analysis of protein-protein interaction using GST fusion proteins immobilized onto glutathione-agarose.

PubMed

Zalazar, L; Alonso, C A I; De Castro, R E; Cesari, A

2014-01-01

Immobilization of small proteins designed to perform protein-protein assays can be a difficult task. Often, the modification of reactive residues necessary for the interaction between the immobilized protein and the matrix compromises the interaction between the protein and its target. In these cases, glutathione-S-transferase (GST) is a valuable tag providing a long arm that makes the bait protein accessible to the mobile flow phase of the chromatography. In the present report, we used a GST fusion version of the 8-kDa protein serine protease inhibitor Kazal-type 3 (SPINK3) as the bait to purify anti-SPINK3 antibodies from a rabbit crude serum. The protocol for immobilization of GST-SPINK3 to glutathione-agarose beads was modified from previously reported protocols by using an alternative bifunctional cross-linker (dithiobis(succinimidyl propionate)) in a very simple procedure and by using simple buffers under physiological conditions. We concluded that the immobilized protein remained bound to the column after elution with low pH, allowing the reuse of the column for alternative uses, such as screening for other protein-protein interactions using SPINK3 as the bait.

Abnormal splicing switch of DMD's penultimate exon compromises muscle fibre maintenance in myotonic dystrophy.

PubMed

Rau, Frédérique; Lainé, Jeanne; Ramanoudjame, Laetitita; Ferry, Arnaud; Arandel, Ludovic; Delalande, Olivier; Jollet, Arnaud; Dingli, Florent; Lee, Kuang-Yung; Peccate, Cécile; Lorain, Stéphanie; Kabashi, Edor; Athanasopoulos, Takis; Koo, Taeyoung; Loew, Damarys; Swanson, Maurice S; Le Rumeur, Elisabeth; Dickson, George; Allamand, Valérie; Marie, Joëlle; Furling, Denis

2015-05-28

Myotonic Dystrophy type 1 (DM1) is a dominant neuromuscular disease caused by nuclear-retained RNAs containing expanded CUG repeats. These toxic RNAs alter the activities of RNA splicing factors resulting in alternative splicing misregulation and muscular dysfunction. Here we show that the abnormal splicing of DMD exon 78 found in dystrophic muscles of DM1 patients is due to the functional loss of MBNL1 and leads to the re-expression of an embryonic dystrophin in place of the adult isoform. Forced expression of embryonic dystrophin in zebrafish using an exon-skipping approach severely impairs the mobility and muscle architecture. Moreover, reproducing Dmd exon 78 missplicing switch in mice induces muscle fibre remodelling and ultrastructural abnormalities including ringed fibres, sarcoplasmic masses or Z-band disorganization, which are characteristic features of dystrophic DM1 skeletal muscles. Thus, we propose that splicing misregulation of DMD exon 78 compromises muscle fibre maintenance and contributes to the progressive dystrophic process in DM1.

Abnormal splicing switch of DMD's penultimate exon compromises muscle fibre maintenance in myotonic dystrophy

PubMed Central

Rau, Frédérique; Lainé, Jeanne; Ramanoudjame, Laetitita; Ferry, Arnaud; Arandel, Ludovic; Delalande, Olivier; Jollet, Arnaud; Dingli, Florent; Lee, Kuang-Yung; Peccate, Cécile; Lorain, Stéphanie; Kabashi, Edor; Athanasopoulos, Takis; Koo, Taeyoung; Loew, Damarys; Swanson, Maurice S.; Le Rumeur, Elisabeth; Dickson, George; Allamand, Valérie; Marie, Joëlle; Furling, Denis

2015-01-01

Myotonic Dystrophy type 1 (DM1) is a dominant neuromuscular disease caused by nuclear-retained RNAs containing expanded CUG repeats. These toxic RNAs alter the activities of RNA splicing factors resulting in alternative splicing misregulation and muscular dysfunction. Here we show that the abnormal splicing of DMD exon 78 found in dystrophic muscles of DM1 patients is due to the functional loss of MBNL1 and leads to the re-expression of an embryonic dystrophin in place of the adult isoform. Forced expression of embryonic dystrophin in zebrafish using an exon-skipping approach severely impairs the mobility and muscle architecture. Moreover, reproducing Dmd exon 78 missplicing switch in mice induces muscle fibre remodelling and ultrastructural abnormalities including ringed fibres, sarcoplasmic masses or Z-band disorganization, which are characteristic features of dystrophic DM1 skeletal muscles. Thus, we propose that splicing misregulation of DMD exon 78 compromises muscle fibre maintenance and contributes to the progressive dystrophic process in DM1. PMID:26018658

[Sarcopenia intervention with progressive resistance training and protein nutritional supplements].

PubMed

Palop Montoro, M Victoria; Párraga Montilla, Juan Antonio; Lozano Aguilera, Emilio; Arteaga Checa, Milagros

2015-04-01

Aging is accompanied by changes in body composition among which is a progressive reduction in muscle mass, which may contribute to the development of functional limitations in older people, and where the lifestyle plays a particularly important role. To test the effectiveness of progressive resistance training, protein nutritional supplements and both interventions combined in the treatment of sarcopenia. Review of literature in Medline, ScienceDirect, CINAHL, ISI WOK and PEDro data by combining the descriptors of Medical Subject Headings (MeSH) concerning sarcopenia, progressive resistance training, protein supplements and seniors. A total of 147 studies were found which resistance exercise performed by sessions 45-60 minutes, 2-3 times a week, and 3-4 sets of 8 repetitions, to an increasing intensity. This exercise resulted in increased muscle mass and strength, and increased skeletal muscle protein synthesis and muscle fiber size. Nutritional supplements such as beta-hydroxy-beta-methylbutyrate, leucine and essential amino acids produced gains in muscle mass. All supplements increased strength, especially when combined with resistance exercise. The combination of progressive resistance training and protein included in the diet, either in the form of nutritional supplements, strengthens the impact that each of these interventions can have on the treatment of sarcopenia in the elderly. Copyright AULA MEDICA EDICIONES 2014. Published by AULA MEDICA. All rights reserved.

Declines in arrestin and rhodopsin in the macula with progression of age-related macular degeneration.

PubMed

Ethen, Cheryl M; Feng, Xiao; Olsen, Timothy W; Ferrington, Deborah A

2005-03-01

Biochemical analysis of age-related macular degeneration (AMD) at distinct stages of the disease will help further understanding of the molecular events associated with disease progression. This study was conducted to determine the ability of a new grading system for eye bank eyes, the Minnesota Grading System (MGS), to discern distinct stages of AMD so that retinal region-specific changes in rod photoreceptor protein expression from donors could be determined. Donor eyes were assigned to a specific level of AMD by using the MGS. Expression of the rod photoreceptor proteins rhodopsin and arrestin was evaluated by Western immunoblot analysis in the macular and peripheral regions of the neurosensory retina from donors at different stages of AMD. A significant linear decline in both arrestin and rhodopsin content correlated with progressive MGS levels in the macula. In contrast, the peripheral region showed no significant correlation between MGS level and the content of either protein. The statistically significant relationship between decreasing macular rod photoreceptor proteins and progressive MGS levels of AMD demonstrates the utility of the clinically based MGS to correspond with specific protein changes found at known, progressive stages of degeneration. Future biochemical analysis of clinically characterized donor eyes will further understanding of the pathobiochemistry of AMD.

Reduced volatile organic compound (VOC) ammunition coatings. Progress report, October 1994-September 1995

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

Duncan, J.L.

1996-05-01

Production requirements and film thickness limitations typically require that ammunition coating systems consist of a single film. This single film must provide the corrosion resistance of a primer plus such properties as color, gloss, and solvent resistance that are required of a topcoat, a compromise at best. Federal and local regulations resulting from the Clean Air Act and its amendments restrict the amount of VOC emitted during the application of protective coatings, and regulations on worker safety restrict exposure to hazardous materials such as chromates. These materials also generate hazardous wastes and the associated high disposal costs. This report summarizesmore » progress in developing ammunition coatings that perform as well as or better than current systems, but at reduced VOC levels with chromate-free pigmentation.« less

Traffic safety for the cell: influence of cyclin-dependent kinase activity on genomic stability.

PubMed

Enders, Greg H; Maude, Shannon L

2006-04-12

Genomic instability has long been considered a key factor in tumorigenesis. Recent evidence suggests that DNA damage may be widespread in early pre-neoplastic states, with deregulation of cyclin-dependent kinase (Cdk) activity a driving force. Increased Cdk activity may critically reduce licensing of origins of DNA replication, drive re-replication, or mediate overexpression of checkpoint proteins, inducing deleterious cell cycle delay. Conversely, inhibition of Cdk activity may compromise replication efficiency, expression of checkpoint proteins, or activation of DNA repair proteins. These vital functions point to the impact of Cdk activity on the stability of the genome. Insight into these pathways may improve our understanding of tumorigenesis and lead to more rational cancer therapies.

Uterine deletion of Trp53 compromises antioxidant responses in mouse decidua

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

Burnum, Kristin E.; Hirota, Yasushi; Baker, Erin Shammel

2012-09-01

Preterm birth is a global health issue impacting both mothers and children. However, the etiology of preterm birth is not clearly understood. From our recent finding that premature decidual senescence with terminal differentiation is a cause of preterm birth in mice with uterine Trp53 deletion, encoding p53 protein, led us to explore other potential factors that are related to preterm birth. Utilizing proteomics approaches, here we show that 183 candidate proteins cause significant changes in decidua with Trp53 deletion as compared to normal decidua. Functional categorization of these proteins unveiled new pathways that are influenced by p53. In particular, downregulationmore » of a cluster of antioxidant proteins in p53 deficient decidua suggests that increased oxidative stress could be one cause of preterm birth in mice with uterine deletion of Trp53.« less

Ultrastructural identification of peripheral myelin proteins by a pre-embedding immunogold labeling method.

PubMed

Canron, Marie-Hélène; Bouillot, Sandrine; Favereaux, Alexandre; Petry, Klaus G; Vital, Anne

2003-03-01

Ultrastructural immunolabeling of peripheral nervous system components is an important tool to study the relation between structure and function. Owing to the scarcity of certain antigens and the dense structure of the peripheral nerve, a pre-embedding technique is likely appropriate. After several investigations on procedures for pre-embedding immunolabeling, we propose a method that offers a good compromise between detection of antigenic sites and preservation of morphology at the ultrastructural level, and that is easy to use and suitable for investigations on peripheral nerve biopsies from humans. Pre-fixation by immersion in paraformaldehyde/glutaraldehyde is necessary to stabilize the ultrastructure. Then, ultrasmall gold particles with silver enhancement are advised. Antibodies against myelin protein zero and myelin basic protein were chosen for demonstration. The same technique was applied to localize a 35 kDa myelin protein.

The Retinoblastoma Tumor Suppressor Regulates a Xenobiotic Detoxification Pathway

PubMed Central

Sáenz Robles, Maria Teresa; Case, Ashley; Chong, Jean-Leon; Leone, Gustavo; Pipas, James M.

2011-01-01

The retinoblastoma tumor suppressor (pRb) regulates cell cycle entry, progression and exit by controlling the activity of the E2F-family of transcription factors. During cell cycle exit pRb acts as a transcriptional repressor by associating with E2F proteins and thereby inhibiting their ability to stimulate the expression of genes required for S phase. Indeed, many tumors harbor mutations in the RB gene and the pRb-E2F pathway is compromised in nearly all types of cancers. In this report we show that both pRb and its interacting partners, the transcriptional factors E2F1-2-3, act as positive modulators of detoxification pathways important for metabolizing and clearing xenobiotics—such as toxins and drugs—from the body. Using a combination of conventional molecular biology techniques and microarray analysis of specific cell populations, we have analyzed the detoxification pathway in murine samples in the presence or absence of pRb and/or E2F1-2-3. In this report, we show that both pRb and E2F1-2-3 act as positive modulators of detoxification pathways in mice, challenging the conventional view of E2F1-2-3 as transcriptional repressors negatively regulated by pRb. These results suggest that mutations altering the pRb-E2F axis may have consequences beyond loss of cell cycle control by altering the ability of tissues to remove toxins and to properly metabolize anticancer drugs, and might help to understand the formation and progression rates of different types of cancer, as well as to better design appropriate therapies based on the particular genetic composition of the tumors. PMID:22022495

A system identification approach for developing model predictive controllers of antibody quality attributes in cell culture processes.

PubMed

Downey, Brandon; Schmitt, John; Beller, Justin; Russell, Brian; Quach, Anthony; Hermann, Elizabeth; Lyon, David; Breit, Jeffrey

2017-11-01

As the biopharmaceutical industry evolves to include more diverse protein formats and processes, more robust control of Critical Quality Attributes (CQAs) is needed to maintain processing flexibility without compromising quality. Active control of CQAs has been demonstrated using model predictive control techniques, which allow development of processes which are robust against disturbances associated with raw material variability and other potentially flexible operating conditions. Wide adoption of model predictive control in biopharmaceutical cell culture processes has been hampered, however, in part due to the large amount of data and expertise required to make a predictive model of controlled CQAs, a requirement for model predictive control. Here we developed a highly automated, perfusion apparatus to systematically and efficiently generate predictive models using application of system identification approaches. We successfully created a predictive model of %galactosylation using data obtained by manipulating galactose concentration in the perfusion apparatus in serialized step change experiments. We then demonstrated the use of the model in a model predictive controller in a simulated control scenario to successfully achieve a %galactosylation set point in a simulated fed-batch culture. The automated model identification approach demonstrated here can potentially be generalized to many CQAs, and could be a more efficient, faster, and highly automated alternative to batch experiments for developing predictive models in cell culture processes, and allow the wider adoption of model predictive control in biopharmaceutical processes. © 2017 The Authors Biotechnology Progress published by Wiley Periodicals, Inc. on behalf of American Institute of Chemical Engineers Biotechnol. Prog., 33:1647-1661, 2017. © 2017 The Authors Biotechnology Progress published by Wiley Periodicals, Inc. on behalf of American Institute of Chemical Engineers.

Dry age-related macular degeneration: mechanisms, therapeutic targets, and imaging.

PubMed

Bowes Rickman, Catherine; Farsiu, Sina; Toth, Cynthia A; Klingeborn, Mikael

2013-12-13

Age-related macular degeneration is the leading cause of irreversible visual dysfunction in individuals over 65 in Western Society. Patients with AMD are classified as having early stage disease (early AMD), in which visual function is affected, or late AMD (generally characterized as either "wet" neovascular AMD, "dry" atrophic AMD or both), in which central vision is severely compromised or lost. Until recently, there have been no therapies available to treat the disorder(s). Now, the most common wet form of late-stage AMD, choroidal neovascularization, generally responds to treatment with anti-vascular endothelial growth factor therapies. Nevertheless, there are no current therapies to restore lost vision in eyes with advanced atrophic AMD. Oral supplementation with the Age-Related Eye Disease Study (AREDS) or AREDS2 formulation (antioxidant vitamins C and E, lutein, zeaxanthin, and zinc) has been shown to reduce the risk of progression to advanced AMD, although the impact was in neovascular rather than atrophic AMD. Recent findings, however, have demonstrated several features of early AMD that are likely to be druggable targets for treatment. Studies have established that much of the genetic risk for AMD is associated with complement genes. Consequently, several complement-based therapeutic treatment approaches are being pursued. Potential treatment strategies against AMD deposit formation and protein and/or lipid deposition will be discussed, including anti-amyloid therapies. In addition, the role of autophagy in AMD and prevention of oxidative stress through modulation of the antioxidant system will be explored. Finally, the success of these new therapies in clinical trials and beyond relies on early detection, disease typing, and predicting disease progression, areas that are currently being rapidly transformed by improving imaging modalities and functional assays.

SI113, a SGK1 inhibitor, potentiates the effects of radiotherapy, modulates the response to oxidative stress and induces cytotoxic autophagy in human glioblastoma multiforme cells

PubMed Central

Talarico, Cristina; Dattilo, Vincenzo; D'Antona, Lucia; Barone, Agnese; Amodio, Nicola; Belviso, Stefania; Musumeci, Francesca; Abbruzzese, Claudia; Bianco, Cataldo; Trapasso, Francesco; Schenone, Silvia; Alcaro, Stefano; Ortuso, Francesco; Florio, Tullio; Paggi, Marco G.; Perrotti, Nicola; Amato, Rosario

2016-01-01

Glioblastoma multiforme (GBM) is the most aggressive CNS tumor and is characterized by a very high frequency of clinical relapse after therapy and thus by a dismal prognosis, which strongly compromises patients survival. We have recently identified the small molecule SI113, as a potent and selective inhibitor of SGK1, a serine/threonine protein kinase, that modulates several oncogenic signaling cascades. The SI113-dependent SGK1 inhibition induces cell death, blocks proliferation and perturbs cell cycle progression by modulating SGK1-related substrates. SI113 is also able to strongly and consistently block, in vitro and in vivo, growth and survival of human hepatocellular-carcinomas, either used as a single agent or in combination with ionizing radiations. In the present paper we aim to study the effect of SI113 on human GBM cell lines with variable p53 expression. Cell viability, cell death, caspase activation and cell cycle progression were then analyzed by FACS and WB-based assays, after exposure to SI113, with or without oxidative stress and ionizing radiations. Moreover, autophagy and related reticulum stress response were evaluated. We show here, that i) SGK1 is over-expressed in highly malignant gliomas and that the treatment with SI113 leads to ii) significant increase in caspase-mediated apoptotic cell death in GBM cell lines but not in normal fibroblasts; iii)enhancement of the effects of ionizing radiations; iv) modulation of the response to oxidative reticulum stress; v) induction of cytotoxic autophagy. Evidence reported here underlines the therapeutic potential of SI113 in GBM, suggesting a new therapeutic strategy either alone or in combination with radiotherapy. PMID:26908461

SI113, a SGK1 inhibitor, potentiates the effects of radiotherapy, modulates the response to oxidative stress and induces cytotoxic autophagy in human glioblastoma multiforme cells.

PubMed

Talarico, Cristina; Dattilo, Vincenzo; D'Antona, Lucia; Barone, Agnese; Amodio, Nicola; Belviso, Stefania; Musumeci, Francesca; Abbruzzese, Claudia; Bianco, Cataldo; Trapasso, Francesco; Schenone, Silvia; Alcaro, Stefano; Ortuso, Francesco; Florio, Tullio; Paggi, Marco G; Perrotti, Nicola; Amato, Rosario

2016-03-29

Glioblastoma multiforme (GBM) is the most aggressive CNS tumor and is characterized by a very high frequency of clinical relapse after therapy and thus by a dismal prognosis, which strongly compromises patients survival. We have recently identified the small molecule SI113, as a potent and selective inhibitor of SGK1, a serine/threonine protein kinase, that modulates several oncogenic signaling cascades. The SI113-dependent SGK1 inhibition induces cell death, blocks proliferation and perturbs cell cycle progression by modulating SGK1-related substrates. SI113 is also able to strongly and consistently block, in vitro and in vivo, growth and survival of human hepatocellular-carcinomas, either used as a single agent or in combination with ionizing radiations. In the present paper we aim to study the effect of SI113 on human GBM cell lines with variable p53 expression. Cell viability, cell death, caspase activation and cell cycle progression were then analyzed by FACS and WB-based assays, after exposure to SI113, with or without oxidative stress and ionizing radiations. Moreover, autophagy and related reticulum stress response were evaluated. We show here, that i) SGK1 is over-expressed in highly malignant gliomas and that the treatment with SI113 leads to ii) significant increase in caspase-mediated apoptotic cell death in GBM cell lines but not in normal fibroblasts; iii)enhancement of the effects of ionizing radiations; iv) modulation of the response to oxidative reticulum stress; v) induction of cytotoxic autophagy. Evidence reported here underlines the therapeutic potential of SI113 in GBM, suggesting a new therapeutic strategy either alone or in combination with radiotherapy.

Critical functions of RhoB in support of glioblastoma tumorigenesis

PubMed Central

Ma, Yufang; Gong, Yuanying; Cheng, Zhixiang; Loganathan, Sudan; Kao, Crystal; Sarkaria, Jann N.; Abel, Ty W.; Wang, Jialiang

2015-01-01

Background RhoB is a member of the Rho small GTPase family that regulates cytoskeletal dynamics and vesicle trafficking. The RhoB homologs, RhoA and RhoC, have been shown to promote cancer progression and metastasis. In contrast, the functions of RhoB in human cancers are context dependent. Although expression of RhoB inversely correlates with disease progression in several epithelial cancers, recent data suggest that RhoB may support malignant phenotypes in certain cancer types. Methods We assessed RhoB protein levels in glioma surgical specimens and patient-derived xenografts. The roles of RhoB in glioblastoma were determined by loss-of-function and gain-of-function assays in vitro and in vivo. The impact on p53 and STAT3 signaling was investigated. Results RhoB expression was similar in tumor specimens compared with normal neural tissues obtained from epilepsy surgery. RhoB was expressed in the vast majority of xenograft tumors and spheroid cultures. Knockdown of RhoB induced cell-cycle arrest and apoptosis and compromised in vivo tumorigenic potential. However, overexpression of wild-type RhoB or a constitutively active mutant (RhoB-V14) did not significantly affect cell growth, which suggests that RhoB is not a rate-limiting oncogenic factor and is consistent with the scarcity of RhoB mutations in human cancer. Knockdown of RhoB reduced basal STAT3 activity and impaired cytokine-induced STAT3 activation. In glioblastoma tumors retaining wild-type p53, depletion of RhoB also activated p53 and induced expression of p21CIP1/WAF1. Conclusions Our data suggest that RhoB belongs to an emerging class of “nononcogene addiction” factors that are essential for maintenance of malignant phenotypes in human cancers. PMID:25216671

Nonalcoholic steatohepatitic (NASH) mice are protected from higher hepatotoxicity of acetaminophen upon induction of PPAR{alpha} with clofibrate

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

Donthamsetty, Shashikiran; Bhave, Vishakha S.; Mitra, Mayurranjan S.

2008-08-01

The objective was to investigate if the hepatotoxic sensitivity in nonalcoholic steatohepatitic mice to acetaminophen (APAP) is due to downregulation of nuclear receptor PPAR{alpha} via lower cell division and tissue repair. Male Swiss Webster mice fed methionine and choline deficient diet for 31 days exhibited NASH. On the 32nd day, a marginally toxic dose of APAP (360 mg/kg, ip) yielded 70% mortality in steatohepatitic mice, while all non steatohepatitic mice receiving the same dose survived. {sup 14}C-APAP covalent binding, CYP2E1 protein, and enzyme activity did not differ from the controls, obviating increased APAP bioactivation as the cause of amplified APAPmore » hepatotoxicity. Liver injury progressed only in steatohepatitic livers between 6 and 24 h. Cell division and tissue repair assessed by {sup 3}H-thymidine incorporation and PCNA were inhibited only in the steatohepatitic mice given APAP suggesting that higher sensitivity of NASH liver to APAP-induced hepatotoxicity was due to lower tissue repair. The hypothesis that impeded liver tissue repair in steatohepatitic mice was due to downregulation of PPAR{alpha} was tested. PPAR{alpha} was downregulated in NASH. To investigate whether downregulation of PPAR{alpha} in NASH is the critical mechanism of compromised liver tissue repair, PPAR{alpha} was induced in steatohepatitic mice with clofibrate (250 mg/kg for 3 days, ip) before injecting APAP. All clofibrate pretreated steatohepatitic mice receiving APAP exhibited lower liver injury, which did not progress and the mice survived. The protection was not due to lower bioactivation of APAP but due to higher liver tissue repair. These findings suggest that inadequate PPAR{alpha} expression in steatohepatitic mice sensitizes them to APAP hepatotoxicity.« less

Immune checkpoint inhibitor-related myocarditis.

PubMed

Tajiri, Kazuko; Aonuma, Kazutaka; Sekine, Ikuo

2018-01-01

Immune checkpoint inhibitors have demonstrated significant clinical benefit in many cancers. The clinical benefit afforded by these treatments can be accompanied by a unique and distinct spectrum of adverse events. Recently, several fatal cases of immune checkpoint inhibitor-related myocarditis were reported. Although its frequency is comparatively lower than that of other immune-related adverse events, myocarditis can lead to circulatory collapse and lethal ventricular arrhythmia. Immune checkpoints, cytotoxic T-lymphocyte antigen 4 (CTLA-4) and programmed cell death protein 1 (PD-1), play important roles in establishing peripheral tolerance to the heart. Evidence from studies using genetically engineered mouse models suggests that CTLA-4 signaling terminates proliferation and promotes anergy during the primary response to cardiac self-peptide recognition. PD-1 signaling restrains autoreactive T cells that enter the peripheral tissues and recognize cardiac-peptide, maintaining them in an anergic state. Patients affected by immune checkpoint inhibitor-related myocarditis often experience rapid onset of profound hemodynamic compromise progressing to cardiogenic shock. Early diagnosis is mandatory to address specific therapy and correct the timing of circulatory support. However, the diagnosis of myocarditis is challenging due to the heterogeneity of clinical presentations. Owing to its early onset, nonspecific symptomatology and fulminant progression, especially when these drugs are used in combination, oncologists should be vigilant for immune checkpoint inhibitor-related myocarditis. With many questions yet to be answered, from basic immune biology to clinical management, future research should aim to optimize the use of these drugs by identifying predictive biomarkers of either a response to therapy or the risks of myocarditis development. © The Author 2017. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Dry Age-Related Macular Degeneration: Mechanisms, Therapeutic Targets, and Imaging

PubMed Central

Bowes Rickman, Catherine; Farsiu, Sina; Toth, Cynthia A.; Klingeborn, Mikael

2013-01-01

Age-related macular degeneration is the leading cause of irreversible visual dysfunction in individuals over 65 in Western Society. Patients with AMD are classified as having early stage disease (early AMD), in which visual function is affected, or late AMD (generally characterized as either “wet” neovascular AMD, “dry” atrophic AMD or both), in which central vision is severely compromised or lost. Until recently, there have been no therapies available to treat the disorder(s). Now, the most common wet form of late-stage AMD, choroidal neovascularization, generally responds to treatment with anti–vascular endothelial growth factor therapies. Nevertheless, there are no current therapies to restore lost vision in eyes with advanced atrophic AMD. Oral supplementation with the Age-Related Eye Disease Study (AREDS) or AREDS2 formulation (antioxidant vitamins C and E, lutein, zeaxanthin, and zinc) has been shown to reduce the risk of progression to advanced AMD, although the impact was in neovascular rather than atrophic AMD. Recent findings, however, have demonstrated several features of early AMD that are likely to be druggable targets for treatment. Studies have established that much of the genetic risk for AMD is associated with complement genes. Consequently, several complement-based therapeutic treatment approaches are being pursued. Potential treatment strategies against AMD deposit formation and protein and/or lipid deposition will be discussed, including anti-amyloid therapies. In addition, the role of autophagy in AMD and prevention of oxidative stress through modulation of the antioxidant system will be explored. Finally, the success of these new therapies in clinical trials and beyond relies on early detection, disease typing, and predicting disease progression, areas that are currently being rapidly transformed by improving imaging modalities and functional assays. PMID:24335072

Thiols of flagellar proteins are essential for progressive motility in human spermatozoa.

PubMed

Cabrillana, María Eugenia; Monclus, María de Los Ángeles; Lancellotti, Tania Estefania Sáez; Boarelli, Paola Vanina; Vincenti, Amanda Edith; Fornés, Miguel Matias; Sanabria, Eduardo Alfredo; Fornés, Miguel Walter

2017-07-01

Male infertility is a disorder of the reproductive system defined by the failure to achieve a clinical pregnancy after 12 months or more of regular unprotected sexual intercourse. The presence of low-motile or immotile spermatozoa is one of many causes of infertility; however, this observation provides little or no information regarding the pathogenesis of the malfunction. Good sperm motility depends on correct assembly of the sperm tail in the testis and efficient maturation during epididymal transit. Thiols of flagellar proteins, such as outer dense fibre protein 1 (ODF1), are oxidised to form disulfides during epididymal transit and the spermatozoa become motile. This study was designed to determine how oxidative changes in protein thiol status affect progressive motility in human spermatozoa. Monobromobimane (mBBr) was used as a specific thiol marker and disruptor of sperm progressive motility. When mBBr was blocked by dithiothreitol it did not promote motility changes. The analysis of mBBr-treated spermatozoa revealed a reduction of progressive motility and an increased number of spermatozoa with non-progressive motility without affecting ATP production. Laser confocal microscopy and western blot analysis showed that one of the mBBr-positive proteins reacted with an antibody to ODF1. Monobromobimane fluorescence intensity of the sperm tail was lower in normozoospermic than asthenozoospermic men, suggesting that thiol oxidation in spermatozoa of asthenozoospermic men is incomplete. Our findings indicate that mBBr affects the thiol status of ODF1 in human spermatozoa and interferes with progressive motility.

Albumin-bound fatty acids but not albumin itself alter redox balance in tubular epithelial cells and induce a peroxide-mediated redox-sensitive apoptosis

PubMed Central

Ruggiero, Christine; Elks, Carrie M.; Kruger, Claudia; Cleland, Ellen; Addison, Kaity; Noland, Robert C.

2014-01-01

Albuminuria is associated with metabolic syndrome and diabetes. It correlates with the progression of chronic kidney disease, particularly with tubular atrophy. The fatty acid load on albumin significantly increases in obesity, presenting a proinflammatory environment to the proximal tubules. However, little is known about changes in the redox milieu during fatty acid overload and how redox-sensitive mechanisms mediate cell death. Here, we show that albumin with fatty acid impurities or conjugated with palmitate but not albumin itself compromised mitochondrial and cell viability, membrane potential and respiration. Fatty acid overload led to a redox imbalance which deactivated the antioxidant protein peroxiredoxin 2 and caused a peroxide-mediated apoptosis through the redox-sensitive pJNK/caspase-3 pathway. Transfection of tubular cells with peroxiredoxin 2 was protective and mitigated apoptosis. Mitochondrial fatty acid entry and ceramide synthesis modulators suggested that mitochondrial β oxidation but not ceramide synthesis may modulate lipotoxic effects on tubular cell survival. These results suggest that albumin overloaded with fatty acids but not albumin itself changes the redox environment in the tubules, inducing a peroxide-mediated redox-sensitive apoptosis. Thus, mitigating circulating fatty acid levels may be an important factor in both preserving redox balance and preventing tubular cell damage in proteinuric diseases. PMID:24500687

Conditional abatement of tissue fibrosis using nucleoside analogs to selectively corrupt DNA replication in transgenic fibroblasts.

PubMed

Iwano, M; Fischer, A; Okada, H; Plieth, D; Xue, C; Danoff, T M; Neilson, E G

2001-02-01

Progressive tissue fibrosis can compromise epithelial function resulting in organ failure. Appreciating evidence suggests that fibroblasts provide fibrogenic collagens during such injury. We further tested this notion by attempting to reduce the physiologic consequences of organ fibrosis through the selective killing of fibroblasts at sites of injury. Here, we report the conditional reduction of tissue fibroblasts using the coding sequence for herpesvirus thymidine kinase (DeltaTK) put under the control of a cell-specific promoter from the gene encoding fibroblast-specific protein 1 (FSP1). Transgenic fibroblasts from mice carrying FSP1.DeltaTK minigenes expressed thymidine kinase concordantly with native FSP1 and, compared to transgenic epithelium, were selectively susceptible to the lethal effects of nucleoside analogs either in culture or during experimental renal fibrosis. The numbers of fibroblasts in fibrogenic kidney tissue were reduced on exposure to nucleoside analogs as was the degree of type I collagen deposition and the extent of fibrosis. Fibroblast reduction following the stress of DNA chain termination highlights the important contribution of cell division during fibrogenesis. Our findings convey a proof of principle regarding the importance of FSP1(+) fibroblasts in fibrosis as well as providing a new approach to treating the relentless scarification of tissue.

A Feline HFpEF Model with Pulmonary Hypertension and Compromised Pulmonary Function.

PubMed

Wallner, Markus; Eaton, Deborah M; Berretta, Remus M; Borghetti, Giulia; Wu, Jichuan; Baker, Sandy T; Feldsott, Eric A; Sharp, Thomas E; Mohsin, Sadia; Oyama, Mark A; von Lewinski, Dirk; Post, Heiner; Wolfson, Marla R; Houser, Steven R

2017-11-29

Heart Failure with preserved Ejection Fraction (HFpEF) represents a major public health problem. The causative mechanisms are multifactorial and there are no effective treatments for HFpEF, partially attributable to the lack of well-established HFpEF animal models. We established a feline HFpEF model induced by slow-progressive pressure overload. Male domestic short hair cats (n = 20), underwent either sham procedures (n = 8) or aortic constriction (n = 12) with a customized pre-shaped band. Pulmonary function, gas exchange, and invasive hemodynamics were measured at 4-months post-banding. In banded cats, echocardiography at 4-months revealed concentric left ventricular (LV) hypertrophy, left atrial (LA) enlargement and dysfunction, and LV diastolic dysfunction with preserved systolic function, which subsequently led to elevated LV end-diastolic pressures and pulmonary hypertension. Furthermore, LV diastolic dysfunction was associated with increased LV fibrosis, cardiomyocyte hypertrophy, elevated NT-proBNP plasma levels, fluid and protein loss in pulmonary interstitium, impaired lung expansion, and alveolar-capillary membrane thickening. We report for the first time in HFpEF perivascular fluid cuff formation around extra-alveolar vessels with decreased respiratory compliance. Ultimately, these cardiopulmonary abnormalities resulted in impaired oxygenation. Our findings support the idea that this model can be used for testing novel therapeutic strategies to treat the ever growing HFpEF population.

Male contraception: another Holy Grail.

PubMed

Murdoch, Fern E; Goldberg, Erwin

2014-01-15

The idea that men should participate in family planning by playing an active role in contraception has become more acceptable in recent years. Up to the present the condom and vasectomy have been the main methods of male contraception. There have been and continue to be efforts to develop an acceptable hormonal contraceptive involving testosterone (T) suppression. However the off target affects, delivery of the analogs and the need for T replacement have proven difficult obstacles to this technology. Research into the development of non-hormonal contraception for men is progressing in several laboratories and this will be the subject of the present review. A number of promising targets for the male pill are being investigated. These involve disruption of spermatogenesis by compromising the integrity of the germinal epithelium, interfering with sperm production at the level of meiosis, attacking specific sperm proteins to disrupt fertilizing ability, or interfering with the assembly of seminal fluid components required by ejaculated sperm for acquisition of motility. Blocking contractility of the vas deferens smooth muscle vasculature to prevent ejaculation is a unique approach that prevents sperm from reaching the egg. We shall note the lack of interest by big pharma with most of the support for male contraception provided by the NIH. Copyright © 2013 The Authors. Published by Elsevier Ltd.. All rights reserved.

Patient-Specific Induced Pluripotent Stem Cell as a Model for Familial Dilated Cardiomyopathy

PubMed Central

Sun, Ning; Yazawa, Masayuki; Liu, Jianwei; Han, Leng; Sanchez-Freire, Veronica; Abilez, Oscar J.; Navarrete, Enrique G.; Hu, Shijun; Wang, Li; Lee, Andrew; Pavlovic, Aleksandra; Lin, Shin; Chen, Rui; Hajjar, Roger J.; Snyder, Michael P.; Dolmetsch, Ricardo E.; Butte, Manish J.; Ashley, Euan A.; Longaker, Michael T.; Robbins, Robert C.; Wu, Joseph C.

2013-01-01

Dilated cardiomyopathy (DCM) is the most common cardiomyopathy, characterized by ventricular dilatation, systolic dysfunction, and progressive heart failure. DCM is the most common diagnosis leading to heart transplantation and places a significant burden on healthcare worldwide. The advent of induced pluripotent stem cells (iPSCs) offers an exceptional opportunity for creating disease-specific models, investigating underlying mechanisms, and optimizing therapy. Here we generated cardiomyocytes (CMs) from iPSCs derived from patients of a DCM family carrying a point mutation (R173W) in the gene encoding sarcomeric protein cardiac troponin T. Compared to the control healthy individuals in the same family cohort, DCM iPSC-CMs exhibited altered Ca2+ handling, decreased contractility, and abnormal sarcomeric α-actinin distribution. When stimulated with β-adrenergic agonist, DCM iPSC-CMs showed characteristics of failure such as reduced beating rates, compromised contraction, and significantly more cells with abnormal sarcomeric α-actinin distribution. β-adrenergic blocker treatment and over-expression of sarcoplasmic reticulum Ca2+ ATPase (Serca2a) improved DCM iPSC-CMs function. Our study demonstrated that human DCM iPSC-CMs recapitulated to some extent the disease phenotypes morphologically and functionally, and thus can serve as a useful platform for exploring molecular and cellular mechanisms and optimizing treatment of this particular disease. PMID:22517884

Does advancing male age influence the expression levels and localisation patterns of phospholipase C zeta (PLCζ) in human sperm?

PubMed Central

Yeste, Marc; Jones, Celine; Amdani, Siti Nornadhirah; Yelumalai, Suseela; Mounce, Ginny; da Silva, Sarah J. Martins; Child, Tim; Coward, Kevin

2016-01-01

Socio-economic factors have led to an increasing trend for couples to delay parenthood. However, advancing age exerts detrimental effects upon gametes which can have serious consequences upon embryo viability. While such effects are well documented for the oocyte, relatively little is known with regard to the sperm. One fundamental role of sperm is to activate the oocyte at fertilisation, a process initiated by phospholipase C zeta (PLCζ), a sperm-specific protein. While PLCζ deficiency can lead to oocyte activation deficiency and infertility, it is currently unknown whether the expression or function of PLCζ is compromised by advancing male age. Here, we evaluate sperm motility and the proportion of sperm expressing PLCζ in 71 males (22–54 years; 44 fertile controls and 27 infertile patients), along with total levels and localisation patterns of PLCζ within the sperm head. Three different statistical approaches were deployed with male age considered both as a categorical and a continuous factor. While progressive motility was negatively correlated with male age, all three statistical models concurred that no PLCζ–related parameter was associated with male age, suggesting that advancing male age is unlikely to cause problems in terms of the sperm’s fundamental ability to activate an oocyte. PMID:27270687

Exosomal DNMT1 mediates cisplatin resistance in ovarian cancer.

PubMed

Cao, Ya-Lei; Zhuang, Ting; Xing, Bao-Heng; Li, Na; Li, Qin

2017-08-01

Ovarian cancer is the most common malignancy in women. Owing to late syndromic presentation and lack of efficient early detection, most cases are diagnosed at advanced stages. Surgery and platinum-based chemotherapy are still the standard care currently. However, resistance invoked often compromises the clinical value of the latter. Expression of DNA methyltransferase 1 (DNMT1) was analysed by gene array. Protein was determined by immunoblotting. Exosome was isolated with commercial kit. Cell proliferation was measured by CCK8 method. Annexin V-PI double staining was performed for apoptosis evaluation. Xenograft model was established and administrated with exosome. Tumour growth and overall survival were monitored. We demonstrated the upregulation of DNMT1 in both tumour and derived cell line. DNMT1 transcripts were highly enriched in exosomes from conditioned medium of ovarian cells. Co-incubation with exosomes stimulated endogenous expression and rendered host cell the resistance to cytotoxicity of cisplatin. In vivo administration of DNMT1-containing exosomes exacerbated xenograft progression and reduced overall survival significantly. Moreover, treatment with exosome inhibitor GW4869 almost completely restored sensitivity in resistant cells. Our data elucidated an unappreciated mechanism of exosomal DNMT1 in cisplatin resistance in ovarian cancer, also indicating the potential of the combination of exosome inhibitor with cisplatin in resistant patients. Copyright © 2017 John Wiley & Sons, Ltd.

Serine/Threonine Protein Phosphatase PstP of Mycobacterium tuberculosis Is Necessary for Accurate Cell Division and Survival of Pathogen*

PubMed Central

Sharma, Aditya K.; Arora, Divya; Singh, Lalit K.; Gangwal, Aakriti; Sajid, Andaleeb; Molle, Virginie; Singh, Yogendra; Nandicoori, Vinay Kumar

2016-01-01

Protein phosphatases play vital roles in phosphorylation-mediated cellular signaling. Although there are 11 serine/threonine protein kinases in Mycobacterium tuberculosis, only one serine/threonine phosphatase, PstP, has been identified. Although PstP has been biochemically characterized and multiple in vitro substrates have been identified, its physiological role has not yet been elucidated. In this study, we have investigated the impact of PstP on cell growth and survival of the pathogen in the host. Overexpression of PstP led to elongated cells and partially compromised survival. We find that depletion of PstP is detrimental to cell survival, eventually leading to cell death. PstP depletion results in elongated multiseptate cells, suggesting a role for PstP in regulating cell division events. Complementation experiments performed with PstP deletion mutants revealed marginally compromised survival, suggesting that all of the domains, including the extracellular domain, are necessary for complete rescue. On the other hand, the catalytic activity of PstP is absolutely essential for the in vitro growth. Mice infection experiments establish a definitive role for PstP in pathogen survival within the host. Depletion of PstP from established infections causes pathogen clearance, indicating that the continued presence of PstP is necessary for pathogen survival. Taken together, our data suggest an important role for PstP in establishing and maintaining infection, possibly via the modulation of cell division events. PMID:27758870

Cyclic AMP-receptor proteins in heart muscle of rats flown on Cosmos 1887

NASA Technical Reports Server (NTRS)

Mednieks, Maija I.; Popova, Irina A.; Grindeland, Richard E.

1991-01-01

The cellular compartmentalization of the cyclic AMP-receptor proteins in heart ventricular tissue obtained from rats flown on the Cosmos 1887 is determined. Photoaffinity labeling of soluble and particular cell fractions with a (32P)-8-azido analog of cyclic AMP is followed by electrophoretic separation of the proteins and by autoradiographic identification of the labeled isoforms of cAPK R subunits. It is shown that RII in the particulate subcellular fraction was significantly decreased in heart cells from rats in the flight group when compared to controls. Protein banding patterns in both the cytoplasmic fraction and in a fraction enriched in chromatin-bound proteins exhibited some variability in tissues of individual animals, but showed no changes that could be directly attributed to flight conditions. No significant change was apparent in the distribution of RI or RII cyclic AMP binding in the soluble fractions. It is inferred that the cardiac cell integrity or its protein content is not compromised under flight conditions.

A high speed multifocal multiphoton fluorescence lifetime imaging microscope for live-cell FRET imaging

PubMed Central

Poland, Simon P.; Krstajić, Nikola; Monypenny, James; Coelho, Simao; Tyndall, David; Walker, Richard J.; Devauges, Viviane; Richardson, Justin; Dutton, Neale; Barber, Paul; Li, David Day-Uei; Suhling, Klaus; Ng, Tony; Henderson, Robert K.; Ameer-Beg, Simon M.

2015-01-01

We demonstrate diffraction limited multiphoton imaging in a massively parallel, fully addressable time-resolved multi-beam multiphoton microscope capable of producing fluorescence lifetime images with sub-50ps temporal resolution. This imaging platform offers a significant improvement in acquisition speed over single-beam laser scanning FLIM by a factor of 64 without compromising in either the temporal or spatial resolutions of the system. We demonstrate FLIM acquisition at 500 ms with live cells expressing green fluorescent protein. The applicability of the technique to imaging protein-protein interactions in live cells is exemplified by observation of time-dependent FRET between the epidermal growth factor receptor (EGFR) and the adapter protein Grb2 following stimulation with the receptor ligand. Furthermore, ligand-dependent association of HER2-HER3 receptor tyrosine kinases was observed on a similar timescale and involved the internalisation and accumulation or receptor heterodimers within endosomes. These data demonstrate the broad applicability of this novel FLIM technique to the spatio-temporal dynamics of protein-protein interaction. PMID:25780724

Hacking the Cell: Network Intrusion and Exploitation by Adenovirus E1A.

PubMed

King, Cason R; Zhang, Ali; Tessier, Tanner M; Gameiro, Steven F; Mymryk, Joe S

2018-05-01

As obligate intracellular parasites, viruses are dependent on their infected hosts for survival. Consequently, viruses are under enormous selective pressure to utilize available cellular components and processes to their own advantage. As most, if not all, cellular activities are regulated at some level via protein interactions, host protein interaction networks are particularly vulnerable to viral exploitation. Indeed, viral proteins frequently target highly connected "hub" proteins to "hack" the cellular network, defining the molecular basis for viral control over the host. This widespread and successful strategy of network intrusion and exploitation has evolved convergently among numerous genetically distinct viruses as a result of the endless evolutionary arms race between pathogens and hosts. Here we examine the means by which a particularly well-connected viral hub protein, human adenovirus E1A, compromises and exploits the vulnerabilities of eukaryotic protein interaction networks. Importantly, these interactions identify critical regulatory hubs in the human proteome and help define the molecular basis of their function. Copyright © 2018 King et al.

Hacking the Cell: Network Intrusion and Exploitation by Adenovirus E1A

PubMed Central

King, Cason R.; Zhang, Ali; Tessier, Tanner M.; Gameiro, Steven F.

2018-01-01

ABSTRACT As obligate intracellular parasites, viruses are dependent on their infected hosts for survival. Consequently, viruses are under enormous selective pressure to utilize available cellular components and processes to their own advantage. As most, if not all, cellular activities are regulated at some level via protein interactions, host protein interaction networks are particularly vulnerable to viral exploitation. Indeed, viral proteins frequently target highly connected “hub” proteins to “hack” the cellular network, defining the molecular basis for viral control over the host. This widespread and successful strategy of network intrusion and exploitation has evolved convergently among numerous genetically distinct viruses as a result of the endless evolutionary arms race between pathogens and hosts. Here we examine the means by which a particularly well-connected viral hub protein, human adenovirus E1A, compromises and exploits the vulnerabilities of eukaryotic protein interaction networks. Importantly, these interactions identify critical regulatory hubs in the human proteome and help define the molecular basis of their function. PMID:29717008

Balance of power in host-virus arms races.

PubMed

Kok, Kin-Hang; Jin, Dong-Yan

2013-07-17

The sensing of viral RNA by the host innate immune system is mediated by RIG-I and its partner PACT. In this issue of Cell Host & Microbe, Luthra et al. (2013) show that the Ebola virus VP35 protein counteracts the action of PACT at the cost of compromising its own function in viral replication. Copyright © 2013 Elsevier Inc. All rights reserved.

Determining the relative contribution and hierarchy of qseBC and hha in the regulation of flagellar motility of Escherichia coli O157:H7

USDA-ARS?s Scientific Manuscript database

In a recent study we demonstrated that in comparison to the wild-type enterohemorrhagic Escherichia coli (EHEC) O157:H7, a motility-compromised hha deletion mutant with an up-regulated type III secretion system and increased secretion of adherence proteins showed reduced fecal shedding in cattle. In...

DNA-PKcs deficiency leads to persistence of oxidatively-induced clustered DNA lesions in human tumor cells

PubMed Central

Peddi, Prakash; Loftin, Charles W.; Dickey, Jennifer S.; Hair, Jessica M.; Burns, Kara J.; Aziz, Khaled; Francisco, Dave C.; Panayiotidis, Mihalis I.; Sedelnikova, Olga A.; Bonner, William M.; Winters, Thomas A.; Georgakilas, Alexandros G.

2010-01-01

DNA-dependent protein kinase (DNA-PK) is a key non-homologous end joining (NHEJ) nuclear serine/threonine protein kinase involved in various DNA metabolic and damage signaling pathways contributing to the maintenance of genomic stability and prevention of cancer. In order to examine the role of DNA-PK in processing of non-DSB clustered DNA damage, we have used three different models of DNA-PK deficiency i.e. chemical inactivation of its kinase activity by novel inhibitors IC86621 and NU7026, knock-down and complete absence of the protein in human breast cancer (MCF-7) and glioblastoma cell lines (MO59-J/K). Compromised DNA-PK repair pathway has lead to accumulation of clustered DNA lesions induced by γ-rays. Tumor cells lacking protein expression or with inhibited kinase activity showed a marked decrease in their ability to process oxidatively-induced non-DSB clustered DNA lesions measured using a modified version of pulsed field gel electrophoresis or single cell gel electrophoresis (Comet assay). In all cases, DNA-PK inactivation lead to a higher level of lesion persistence even after 24–72 hrs of repair. We suggest a model in which DNA-PK deficiency affects the processing of these clusters by first compromising base excision repair and second by the presence of catalytically inactive DNA-PK inhibiting the efficient processing of these lesions due to the failure of DNA-PK to disassociate from the DNA ends. The information rendered will be important not only for understating cancer etiology in the presence of a NHEJ deficiency but also lead to a better understanding of cancer treatments based on the induction of oxidative stress and inhibition of cluster repair. PMID:20193758

Progesterone receptor expression during prostate cancer progression suggests a role of this receptor in stromal cell differentiation.

PubMed

Yu, Yue; Yang, Ou; Fazli, Ladan; Rennie, Paul S; Gleave, Martin E; Dong, Xuesen

2015-07-01

The progesterone receptor, like the androgen receptor, belongs to the steroid receptor superfamily. Our previous studies have reported that the PR is expressed specifically in prostate stroma. PR inhibits proliferation of, and regulates cytokine secretion by stromal cells. However, PR protein expression in cancer-associated stroma during prostate cancer progression has not been profiled. Since the phenotypes of prostate stromal cells change dynamically as tumors progress, whether the PR plays a role in regulating stromal cell differentiation needs to be investigated. Immunohistochemistry assays measured PR protein levels on human prostate tissue microarrays containing 367 tissue cores from benign prostate, prostate tumors with different Gleason scores, tumors under various durations of castration therapy, and tumors at the castration-resistant stage. Immunoblotting assays determined whether PR regulated the expression of alpha smooth muscle actin (α-SMA), vimentin, and fibroblast specific protein (FSP) in human prostate stromal cells. PR protein levels decreased in cancer-associated stroma when compared with that in benign prostate stroma. This reduction in PR expression was not correlated with Gleason scores. PR protein levels were elevated by castration therapy, but reduced to pre-castration levels when tumors progressed to the castration-resistant stage. Enhanced PR expression in human prostate stromal cells increased α-SMA, but decreased vimentin and FSP protein levels ligand-independently. These results suggest that PR plays an active role in regulating stromal cell phenotypes during prostate cancer progression. © 2015 Wiley Periodicals, Inc.

Improved Soybean Oil for Biodiesel Fuel

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

Tom Clemente; Jon Van Gerpen

2007-11-30

The goal of this program was to generate information on the utility of soybean germplasm that produces oil, high in oleic acid and low in saturated fatty acids, for its use as a biodiesel. Moreover, data was ascertained on the quality of the derived soybean meal (protein component), and the agronomic performance of this novel soybean germplasm. Gathering data on these later two areas is critical, with respect to the first, soybean meal (protein) component is a major driver for commodity soybean, which is utilized as feed supplements in cattle, swine, poultry and more recently aquaculture production. Hence, it ismore » imperative that the resultant modulation in the fatty acid profile of the oil does not compromise the quality of the derived meal, for if it does, the net value of the novel soybean will be drastically reduced. Similarly, if the improved oil trait negative impacts the agronomics (i.e. yield) of the soybean, this in turn will reduce the value of the trait. Over the course of this program oil was extruded from approximately 350 bushels of soybean designated 335-13, which produces oil high in oleic acid (>85%) and low in saturated fatty acid (<6%). As predicted improvement in cold flow parameters were observed as compared to standard commodity soybean oil. Moreover, engine tests revealed that biodiesel derived from this novel oil mitigated NOx emissions. Seed quality of this soybean was not compromised with respect to total oil and protein, nor was the amino acid profile of the derived meal as compared to the respective control soybean cultivar with a conventional fatty acid profile. Importantly, the high oleic acid/low saturated fatty acids oil trait was not impacted by environment and yield was not compromised. Improving the genetic potential of soybean by exploiting the tools of biotechnology to improve upon the lipid quality of the seed for use in industrial applications such as biodiesel will aid in expanding the market for the crop. This in turn, may lead to job creation in rural areas of the country and help stimulate the agricultural economy. Moreover, production of soybean with enhanced oil quality for biodiesel may increase the attractiveness of this renewable, environmentally friendly fuel.« less

An Upgrade on the Rabbit Model of Anthracycline-Induced Cardiomyopathy: Shorter Protocol, Reduced Mortality, and Higher Incidence of Overt Dilated Cardiomyopathy

PubMed Central

Talavera, Jesús; Fernández-Del-Palacio, María Josefa; García-Nicolás, Obdulio; Seva, Juan; Brooks, Gavin; Moraleda, Jose M.

2015-01-01

Current protocols of anthracycline-induced cardiomyopathy in rabbits present with high premature mortality and nephrotoxicity, thus rendering them unsuitable for studies requiring long-term functional evaluation of myocardial function (e.g., stem cell therapy). We compared two previously described protocols to an in-house developed protocol in three groups: Group DOX2 received doxorubicin 2 mg/kg/week (8 weeks); Group DAU3 received daunorubicin 3 mg/kg/week (10 weeks); and Group DAU4 received daunorubicin 4 mg/kg/week (6 weeks). A cohort of rabbits received saline (control). Results of blood tests, cardiac troponin I, echocardiography, and histopathology were analysed. Whilst DOX2 and DAU3 rabbits showed high premature mortality (50% and 33%, resp.), DAU4 rabbits showed 7.6% premature mortality. None of DOX2 rabbits developed overt dilated cardiomyopathy; 66% of DAU3 rabbits developed overt dilated cardiomyopathy and quickly progressed to severe congestive heart failure. Interestingly, 92% of DAU4 rabbits showed overt dilated cardiomyopathy and 67% developed congestive heart failure exhibiting stable disease. DOX2 and DAU3 rabbits showed alterations of renal function, with DAU3 also exhibiting hepatic function compromise. Thus, a shortened protocol of anthracycline-induced cardiomyopathy as in DAU4 group results in high incidence of overt dilated cardiomyopathy, which insidiously progressed to congestive heart failure, associated to reduced systemic compromise and very low premature mortality. PMID:26788502

Knowledge-based computational intelligence development for predicting protein secondary structures from sequences.

PubMed

Shen, Hong-Bin; Yi, Dong-Liang; Yao, Li-Xiu; Yang, Jie; Chou, Kuo-Chen

2008-10-01

In the postgenomic age, with the avalanche of protein sequences generated and relatively slow progress in determining their structures by experiments, it is important to develop automated methods to predict the structure of a protein from its sequence. The membrane proteins are a special group in the protein family that accounts for approximately 30% of all proteins; however, solved membrane protein structures only represent less than 1% of known protein structures to date. Although a great success has been achieved for developing computational intelligence techniques to predict secondary structures in both globular and membrane proteins, there is still much challenging work in this regard. In this review article, we firstly summarize the recent progress of automation methodology development in predicting protein secondary structures, especially in membrane proteins; we will then give some future directions in this research field.

Identification of Glutathione S-Transferase Pi as a Protein Involved in Parkinson Disease Progression

PubMed Central

Shi, Min; Bradner, Joshua; Bammler, Theo K.; Eaton, David L.; Zhang, JianPeng; Ye, ZuCheng; Wilson, Angela M.; Montine, Thomas J.; Pan, Catherine; Zhang, Jing

2009-01-01

Parkinson disease (PD) typically affects the cortical regions during the later stages of disease, with neuronal loss, gliosis, and formation of diffuse cortical Lewy bodies in a significant portion of patients with dementia. To identify novel proteins involved in PD progression, we prepared synaptosomal fractions from the frontal cortices of pathologically verified PD patients at different stages along with age-matched controls. Protein expression profiles were compared using a robust quantitative proteomic technique. Approximately 100 proteins displayed significant differences in their relative abundances between PD patients at various stages and controls; three of these proteins were validated using independent techniques. One of the confirmed proteins, glutathione S-transferase Pi, was further investigated in cellular models of PD, demonstrating that its level was intimately associated with several critical cellular processes that are directly related to neurodegeneration in PD. These results have, for the first time, suggested that the levels of glutathione S-transferase Pi may play an important role in modulating the progression of PD. PMID:19498008

Identification and functional analysis of a core gene module associated with hepatitis C virus-induced human hepatocellular carcinoma progression.

PubMed

Bai, Gaobo; Zheng, Wenling; Ma, Wenli

2018-05-01

Hepatitis C virus (HCV)-induced human hepatocellular carcinoma (HCC) progression may be due to a complex multi-step processes. The developmental mechanism of these processes is worth investigating for the prevention, diagnosis and therapy of HCC. The aim of the present study was to investigate the molecular mechanism underlying the progression of HCV-induced hepatocarcinogenesis. First, the dynamic gene module, consisting of key genes associated with progression between the normal stage and HCC, was identified using the Weighted Gene Co-expression Network Analysis tool from R language. By defining those genes in the module as seeds, the change of co-expression in differentially expressed gene sets in two consecutive stages of pathological progression was examined. Finally, interaction pairs of HCV viral proteins and their directly targeted proteins in the identified module were extracted from the literature and a comprehensive interaction dataset from yeast two-hybrid experiments. By combining the interactions between HCV and their targets, and protein-protein interactions in the Search Tool for the Retrieval of Interacting Genes database (STRING), the HCV-key genes interaction network was constructed and visualized using Cytoscape software 3.2. As a result, a module containing 44 key genes was identified to be associated with HCC progression, due to the dynamic features and functions of those genes in the module. Several important differentially co-expressed gene pairs were identified between non-HCC and HCC stages. In the key genes, cyclin dependent kinase 1 (CDK1), NDC80, cyclin A2 (CCNA2) and rac GTPase activating protein 1 (RACGAP1) were shown to be targeted by the HCV nonstructural proteins NS5A, NS3 and NS5B, respectively. The four genes perform an intermediary role between the HCV viral proteins and the dysfunctional module in the HCV key genes interaction network. These findings provided valuable information for understanding the mechanism of HCV-induced HCC progression and for seeking drug targets for the therapy and prevention of HCC.

RETREG1 (FAM134B): A new player in human diseases: 15 years after the discovery in cancer.

PubMed

Islam, Farhadul; Gopalan, Vinod; Lam, Alfred King-Yin

2018-06-01

FAM134B (family with sequence similarity 134, member B)/RETREG1 and its functional roles are relatively new in human diseases. This review aimed to summarize various functions of FAM134B since our first discovery of the gene in 2001. The protein encoded by FAM134B is a reticulophagy receptor that regulates turnover of the endoplasmic reticulum (ER) by selective phagocytosis. Absence or non-functional expression of FAM134B protein impairs ER-turnover and thereby is involved in the pathogenesis of some human diseases. FAM134B inhibition contributes to impair proteostasis in the ER due to the accumulation of misfolded or aggregated proteins, which in turn leads to compromised neuronal survival and progressive neuronal degenerative diseases. Mutations in FAM134B associated with hereditary sensory and autonomic neuropathy type IIB (HSAN IIB). Selective cleavage of FAM134B by Dengue, Zika, and West Nile virus encoded protease NS2B3 leads to the increased production of infection units, whereas upregulation of FAM134B inhibits viral replication. In cancer, FAM134B acts as a tumor suppressor and inhibit cancer growth both in-vitro and in-vivo. Pharmacological upregulation of FAM134B resulted in reduced cancer cell growth and proliferation. In addition, FAM134B mutations are common in patients with colorectal adenocarcinoma, and oesophageal squamous cell carcinoma. These mutations and expression changes of FAM134B were associated with the biological aggressiveness of these cancers. FAM134B also plays a role in allergic rhinitis, vascular dementia, and identification of stem cells. Taken together, information available in the literature suggests that FAM134B plays critical roles in human diseases, by interacting with different biological and chemical mediators, which are primarily regulated by ER turnover. © 2017 Wiley Periodicals, Inc.

Role of Protein Synthesis Initiation Factors in Dietary Soy Isoflavone-Mediated Effects on Breast Cancer Progression

DTIC Science & Technology

2012-03-01

After 1 week of tumor inoculation, vehicle (10% ethanol, 90% corn oil ), 10 mg/kg body weight (BW) of daidzein, or combined soy isoflavones 10 mg/kg BW...Dietary Soy Isoflavone-Mediated Effects on Breast Cancer Progression. PRINCIPAL INVESTIGATOR: Columba de la Parra Simental CONTRACTING...00935 Role of Protein Synthesis Initiation Factors in Dietary Soy Isoflavone-Mediated Effects on Breast Cancer Progression Columba de la Parra Simental

Progression of 3D Protein Structure and Dynamics Measurements

NASA Astrophysics Data System (ADS)

Sato-Tomita, Ayana; Sekiguchi, Hiroshi; Sasaki, Yuji C.

2018-06-01

New measurement methodologies have begun to be proposed with the recent progress in the life sciences. Here, we introduce two new methodologies, X-ray fluorescence holography for protein structural analysis and diffracted X-ray tracking (DXT), to observe the dynamic behaviors of individual single molecules.

Immunological roulette: Luck or something more? Considering the connections between host and environment in TB.

PubMed

Pearl, John E; Das, Mrinal; Cooper, Andrea M

2018-03-01

Accurate prediction of which patient will progress from a sub-clinical Mycobacterium tuberculosis infection to active tuberculosis represents an elusive, yet critical, clinical research objective. From the individual perspective, progression can be considered to be the product of a series of unfortunate events or even a run of bad luck. Here, we identify the subtle physiological relationships that can influence the odds of progression to active TB and how this progression may reflect directed dysbiosis in a number of interrelated systems. Most infected individuals who progress to disease have apparently good immune responses, but these responses are, at times, compromised by either local or systemic environmental factors. Obvious disease promoting processes, such as tissue-damaging granulomata, usually manifest in the lung, but illness is systemic. This apparent dichotomy between local and systemic reflects a clear need to define the factors that promote progression to active disease within the context of the body as a physiological whole. We discuss aspects of the host environment that can impact expression of immunity, including the microbiome, glucocorticoid-mediated regulation, catecholamines and interaction between the gut, liver and lung. We suggest the importance of integrating precision medicine into our analyses of experimental outcomes such that apparently conflicting results are not contentious, but rather reflect the impact of these subtle relationships with our environment and microbiota.

Probiotics promote endocytic allergen degradation in gut epithelial cells

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

Song, Chun-Hua; Liu, Zhi-Qiang; Department of Pathology and Molecular Medicine, McMaster University, Hamilton, ON

Highlights: Black-Right-Pointing-Pointer Knockdown of A20 compromised the epithelial barrier function. Black-Right-Pointing-Pointer The fusion of endosome/lysosome was disturbed in the A20-deficient HT-29 cells. Black-Right-Pointing-Pointer Antigens transported across A20-deficient HT-29 monolayers conserved antigenicity. Black-Right-Pointing-Pointer Probiotic proteins increased the expression of A20 in HT-29 cells. -- Abstract: Background and aims: Epithelial barrier dysfunction plays a critical role in the pathogenesis of allergic diseases; the mechanism is to be further understood. The ubiquitin E3 ligase A20 (A20) plays a role in the endocytic protein degradation in the cells. This study aims to elucidate the role of A20 in the maintenance of gut epithelial barriermore » function. Methods: Gut epithelial cell line, HT-29 cell, was cultured into monolayers to evaluate the barrier function in transwells. RNA interference was employed to knock down the A20 gene in HT-29 cells to test the role of A20 in the maintenance of epithelial barrier function. Probiotic derived proteins were extracted from the culture supernatants using to enhance the expression of A20 in HT-29 cells. Results: The results showed that the knockdown of A20 compromised the epithelial barrier function in HT-29 monolayers, mainly increased the intracellular permeability. The fusion of endosome/lysosome was disturbed in the A20-deficient HT-29 cells. Allergens collected from the transwell basal chambers of A20-deficient HT-29 monolayers still conserved functional antigenicity. Treating with probiotic derived proteins increased the expression of A20 in HT-29 cells and promote the barrier function. Conclusion: A20 plays an important role in the maintenance of epithelial barrier function as shown by HT-29 monolayer. Probiotic derived protein increases the expression of A20 and promote the HT-29 monolayer barrier function.« less

Multiplex single-molecule interaction profiling of DNA-barcoded proteins.

PubMed

Gu, Liangcai; Li, Chao; Aach, John; Hill, David E; Vidal, Marc; Church, George M

2014-11-27

In contrast with advances in massively parallel DNA sequencing, high-throughput protein analyses are often limited by ensemble measurements, individual analyte purification and hence compromised quality and cost-effectiveness. Single-molecule protein detection using optical methods is limited by the number of spectrally non-overlapping chromophores. Here we introduce a single-molecular-interaction sequencing (SMI-seq) technology for parallel protein interaction profiling leveraging single-molecule advantages. DNA barcodes are attached to proteins collectively via ribosome display or individually via enzymatic conjugation. Barcoded proteins are assayed en masse in aqueous solution and subsequently immobilized in a polyacrylamide thin film to construct a random single-molecule array, where barcoding DNAs are amplified into in situ polymerase colonies (polonies) and analysed by DNA sequencing. This method allows precise quantification of various proteins with a theoretical maximum array density of over one million polonies per square millimetre. Furthermore, protein interactions can be measured on the basis of the statistics of colocalized polonies arising from barcoding DNAs of interacting proteins. Two demanding applications, G-protein coupled receptor and antibody-binding profiling, are demonstrated. SMI-seq enables 'library versus library' screening in a one-pot assay, simultaneously interrogating molecular binding affinity and specificity.

Multiplex single-molecule interaction profiling of DNA barcoded proteins

PubMed Central

Gu, Liangcai; Li, Chao; Aach, John; Hill, David E.; Vidal, Marc; Church, George M.

2014-01-01

In contrast with advances in massively parallel DNA sequencing1, high-throughput protein analyses2-4 are often limited by ensemble measurements, individual analyte purification and hence compromised quality and cost-effectiveness. Single-molecule (SM) protein detection achieved using optical methods5 is limited by the number of spectrally nonoverlapping chromophores. Here, we introduce a single molecular interaction-sequencing (SMI-Seq) technology for parallel protein interaction profiling leveraging SM advantages. DNA barcodes are attached to proteins collectively via ribosome display6 or individually via enzymatic conjugation. Barcoded proteins are assayed en masse in aqueous solution and subsequently immobilized in a polyacrylamide (PAA) thin film to construct a random SM array, where barcoding DNAs are amplified into in situ polymerase colonies (polonies)7 and analyzed by DNA sequencing. This method allows precise quantification of various proteins with a theoretical maximum array density of over one million polonies per square millimeter. Furthermore, protein interactions can be measured based on the statistics of colocalized polonies arising from barcoding DNAs of interacting proteins. Two demanding applications, G-protein coupled receptor (GPCR) and antibody binding profiling, were demonstrated. SMI-Seq enables “library vs. library” screening in a one-pot assay, simultaneously interrogating molecular binding affinity and specificity. PMID:25252978

Deregulation of F-box proteins and its consequence on cancer development, progression and metastasis

PubMed Central

Heo, Jinho; Eki, Rebeka; Abbas, Tarek

2015-01-01

F-box proteins are substrate receptors of the SCF (SKP1-Cullin 1-F-box protein) E3 ubiquitin ligase that play important roles in a number of physiological processes and activities. Through their ability to assemble distinct E3 ubiquitin ligases and target key regulators of cellular activities for ubiquitylation and degradation, this versatile group of proteins is able to regulate the abundance of cellular proteins whose deregulated expression or activity contributes to disease. In this review, we describe the important roles of select F-box proteins in regulating cellular activities, the perturbation of which contributes to the initiation and progression of a number of human malignancies. PMID:26432751

Alternative Splicing May Not Be the Key to Proteome Complexity.

PubMed

Tress, Michael L; Abascal, Federico; Valencia, Alfonso

2017-02-01

Alternative splicing is commonly believed to be a major source of cellular protein diversity. However, although many thousands of alternatively spliced transcripts are routinely detected in RNA-seq studies, reliable large-scale mass spectrometry-based proteomics analyses identify only a small fraction of annotated alternative isoforms. The clearest finding from proteomics experiments is that most human genes have a single main protein isoform, while those alternative isoforms that are identified tend to be the most biologically plausible: those with the most cross-species conservation and those that do not compromise functional domains. Indeed, most alternative exons do not seem to be under selective pressure, suggesting that a large majority of predicted alternative transcripts may not even be translated into proteins. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Tight junctions in inflammatory bowel diseases and inflammatory bowel disease associated colorectal cancer

PubMed Central

Landy, Jonathan; Ronde, Emma; English, Nick; Clark, Sue K; Hart, Ailsa L; Knight, Stella C; Ciclitira, Paul J; Al-Hassi, Hafid Omar

2016-01-01

Inflammatory bowel diseases are characterised by inflammation that compromises the integrity of the epithelial barrier. The intestinal epithelium is not only a static barrier but has evolved complex mechanisms to control and regulate bacterial interactions with the mucosal surface. Apical tight junction proteins are critical in the maintenance of epithelial barrier function and control of paracellular permeability. The characterisation of alterations in tight junction proteins as key players in epithelial barrier function in inflammatory bowel diseases is rapidly enhancing our understanding of critical mechanisms in disease pathogenesis as well as novel therapeutic opportunities. Here we give an overview of recent literature focusing on the role of tight junction proteins, in particular claudins, in inflammatory bowel diseases and inflammatory bowel disease associated colorectal cancer. PMID:27003989

Variation in Protein Origin and Utilization: Research and Clinical Application.

PubMed

Paddon-Jones, Douglas; Coss-Bu, Jorge A; Morris, Claudia R; Phillips, Stuart M; Wernerman, Jan

2017-04-01

Muscle health can be rapidly compromised in clinical environments. Modifiable strategies to preserve metabolic homeostasis in adult patient populations include physical activity and pharmacologic support; however, optimizing dietary practices, or more specifically protein intake, is a necessary prerequisite for any other treatment strategy to be fully effective. Simply increasing protein intake is a well-intentioned but often unfocused strategy to protect muscle health in an intensive care setting. Protein quality is a frequently overlooked factor with the potential to differentially influence health outcomes. Quality can be assessed by a variety of techniques, with digestible indispensable amino acid score being the current and most comprehensive technique endorsed by the Food and Agriculture Organization. In practical terms, animal-based proteins are consistently scored higher in quality compared with incomplete proteins, regardless of the assessment method. Consequently, choosing parenteral and/or enteral feeding options that contain high-quality proteins, rich in the branched-chain amino acid leucine, may help establish a dietary framework with the potential to support clinical practice and improve health outcomes in critically ill patients.

The molecular and cellular response of normal and progressed human bronchial epithelial cells to HZE particles

NASA Astrophysics Data System (ADS)

Story, Michael; Ding, Liang-Hao; Minna, John; Park, Seong-mi; Larsen, Jill

We have used a model of non-oncogenically immortalized normal human bronchial epithelial cells to determine the response of such cells to particles found outside the protection of the earth’s electromagnetic field. We have identified an enhanced frequency of cellular transformation, as measured by growth in soft agar, for both 56Fe and 28Si (1 GeV/n) that is maximal (4-6 fold) at 0.25 Gy and 0.40 Gy, respectively. At 4 months post-irradiation 38 individual soft agar clones were isolated. These clones were characterized extensively for cellular and molecular changes. Gene expression analysis suggested that these clones had down-regulated several genes associated with anti-oxidant pathways including GLS2, GPX1 and 4, SOD2, PIG3, and NQO1 amongst others. As a result, many of these transformed clones were exposed to high levels of intracellular radical oxygen species (ROS), although there appeared not to be any enhanced mitochondrial ROS. DNA repair pathways associated with ATM/ATR signaling were also upregulated. However, these transformants do not develop into tumors when injected into immune-compromised mice, suggesting that they have not progressed sufficiently to become oncogenic. Therefore we chose 6 soft agar clones for continuous culture for an additional 14 months. Amongst the 6 clones, only one clone showed any significant change in phenotype. Clone 3kt-ff.2a, propagated for 18 months, were 2-fold more radioresistant, had a shortened doubling time and the background rate of transformation more than doubled. Furthermore, the morphology of transformed clones changed. Clones from this culture are being compared to the original clone as well as the parental HBEC3KT and will be injected into immune-compromised mice for oncogenic potential. Oncogenically progressed HBECs, HBEC3KT cells that overexpress a mutant RAS gene and where p53 has been knocked down, designated HBEC3KTR53, responded quite differently to HZE particle exposure. First, these cells are more radioresistant to all radiations used when compared to the parental cell line HBEC3KT. Furthermore, within days of their exposure to low and high LET radiations they exhibit enhanced cellular transformation over the parental cells. Moreover, HZE radiations are many fold more effective at initiating cellular transformation. Gene expression analysis identified several pathways that support oncogenic growth as overrepresented in the progressed cells. With continual culture some clones undergo epithelial to mesenchymal transition, change morphology and express markers associated with EMT. And, at least one clone is oncogenic forming highly aggressive tumors in an immune compromised mouse strain. It is important to note that HBEC3KTR53 cells will not form tumors in mice, however, this irradiated clone has moved through the multi-step process of carcinogenesis. We are now examining the molecular alterations that led to oncogenesis in this clone.

Prevention of Age-Related Macular Degeneration.

PubMed

Singh, Niharika; Srinivasan, Sangeetha; Muralidharan, Vinata; Roy, Rupak; V, Jayprakash; Raman, Rajiv

2017-01-01

Age-related macular degeneration (AMD) compromises quality of life. However, the available therapeutic options are limited. This has led to the identification of modifiable risk factors to prevent the development or alter the natural course and prognosis of AMD. The identification and modification of risk factors has the potential for greater public health impact on reducing morbidity from AMD. Likewise, identifying the imaging clues and genetic clues could serve as a guide to recognizing the propensity for progression to severe and end stages of the disease. Several attempts, both successful and unsuccessful, have been made for interventions that could delay the progression of AMD. Of these, pharmacological interventions have shown promising results. The Age-Related Eye Disease Study 1 and 2 have shown the beneficial role of antioxidants in a selected group of patients. Copyright 2017 Asia-Pacific Academy of Ophthalmology.

Armc5 deletion causes developmental defects and compromises T-cell immune responses

PubMed Central

Hu, Yan; Lao, Linjiang; Mao, Jianning; Jin, Wei; Luo, Hongyu; Charpentier, Tania; Qi, Shijie; Peng, Junzheng; Hu, Bing; Marcinkiewicz, Mieczyslaw Martin; Lamarre, Alain; Wu, Jiangping

2017-01-01

Armadillo repeat containing 5 (ARMC5) is a cytosolic protein with no enzymatic activities. Little is known about its function and mechanisms of action, except that gene mutations are associated with risks of primary macronodular adrenal gland hyperplasia. Here we map Armc5 expression by in situ hybridization, and generate Armc5 knockout mice, which are small in body size. Armc5 knockout mice have compromised T-cell proliferation and differentiation into Th1 and Th17 cells, increased T-cell apoptosis, reduced severity of experimental autoimmune encephalitis, and defective immune responses to lymphocytic choriomeningitis virus infection. These mice also develop adrenal gland hyperplasia in old age. Yeast 2-hybrid assays identify 16 ARMC5-binding partners. Together these data indicate that ARMC5 is crucial in fetal development, T-cell function and adrenal gland growth homeostasis, and that the functions of ARMC5 probably depend on interaction with multiple signalling pathways. PMID:28169274

DMP-1-mediated Ghr gene recombination compromises skeletal development and impairs skeletal response to intermittent PTH.

PubMed

Liu, Zhongbo; Kennedy, Oran D; Cardoso, Luis; Basta-Pljakic, Jelena; Partridge, Nicola C; Schaffler, Mitchell B; Rosen, Clifford J; Yakar, Shoshana

2016-02-01

Bone minerals are acquired during growth and are key determinants of adult skeletal health. During puberty, the serum levels of growth hormone (GH) and its downstream effector IGF-1 increase and play critical roles in bone acquisition. The goal of the current study was to determine how bone cells integrate signals from the GH/IGF-1 to enhance skeletal mineralization and strength during pubertal growth. Osteocytes, the most abundant bone cells, were shown to orchestrate bone modeling during growth. We used dentin matrix protein (Dmp)-1-mediated Ghr knockout (DMP-GHRKO) mice to address the role of the GH/IGF axis in osteocytes. We found that DMP-GHRKO did not affect linear growth but compromised overall bone accrual. DMP-GHRKO mice exhibited reduced serum inorganic phosphate and parathyroid hormone (PTH) levels and decreased bone formation indices and were associated with an impaired response to intermittent PTH treatment. Using an osteocyte-like cell line along with in vivo studies, we found that PTH sensitized the response of bone to GH by increasing Janus kinase-2 and IGF-1R protein levels. We concluded that endogenously secreted PTH and GHR signaling in bone are necessary to establish radial bone growth and optimize mineral acquisition during growth. © FASEB.

Poly(GR) in C9ORF72-Related ALS/FTD Compromises Mitochondrial Function and Increases Oxidative Stress and DNA Damage in iPSC-Derived Motor Neurons.

PubMed

Lopez-Gonzalez, Rodrigo; Lu, Yubing; Gendron, Tania F; Karydas, Anna; Tran, Helene; Yang, Dejun; Petrucelli, Leonard; Miller, Bruce L; Almeida, Sandra; Gao, Fen-Biao

2016-10-19

GGGGCC repeat expansions in C9ORF72 are the most common genetic cause of both ALS and FTD. To uncover underlying pathogenic mechanisms, we found that DNA damage was greater, in an age-dependent manner, in motor neurons differentiated from iPSCs of multiple C9ORF72 patients than control neurons. Ectopic expression of the dipeptide repeat (DPR) protein (GR) 80 in iPSC-derived control neurons increased DNA damage, suggesting poly(GR) contributes to DNA damage in aged C9ORF72 neurons. Oxidative stress was also increased in C9ORF72 neurons in an age-dependent manner. Pharmacological or genetic reduction of oxidative stress partially rescued DNA damage in C9ORF72 neurons and control neurons expressing (GR) 80 or (GR) 80 -induced cellular toxicity in flies. Moreover, interactome analysis revealed that (GR) 80 preferentially bound to mitochondrial ribosomal proteins and caused mitochondrial dysfunction. Thus, poly(GR) in C9ORF72 neurons compromises mitochondrial function and causes DNA damage in part by increasing oxidative stress, revealing another pathogenic mechanism in C9ORF72-related ALS and FTD. Copyright © 2016 Elsevier Inc. All rights reserved.

Dehydration decreases saliva antimicrobial proteins important for mucosal immunity.

PubMed

Fortes, Matthew B; Diment, Bethany C; Di Felice, Umberto; Walsh, Neil P

2012-10-01

The aim of the study was to investigate the effect of exercise-induced dehydration and subsequent overnight fluid restriction on saliva antimicrobial proteins important for host defence (secretory IgA (SIgA), α-amylase, and lysozyme). On two randomized occasions, 13 participants exercised in the heat, either without fluid intake to evoke progressive body mass losses (BML) of 1%, 2%, and 3% with subsequent overnight fluid restriction until 0800 h in the following morning (DEH) or with fluids to offset losses (CON). Participants in the DEH trial rehydrated from 0800 h until 1100 h on day 2. BML, plasma osmolality (Posm), and urine specific gravity (USG) were assessed as hydration indices. Unstimulated saliva samples were assessed for flow rate (SFR), SIgA, α-amylase, and lysozyme concentrations. Posm and USG increased during dehydration and remained elevated after overnight fluid restriction (BML = 3.5% ± 0.3%, Posm = 297 ± 6 mosmol·kg⁻¹, and USG = 1.026 ± 0.002; P < 0.001). Dehydration decreased SFR (67% at 3% BML, 70% at 0800 h; P < 0.01) and increased SIgA concentration, with no effect on SIgA secretion rate. SFR and SIgA responses remained unchanged in the CON trial. Dehydration did not affect α-amylase or lysozyme concentration but decreased secretion rates of α-amylase (44% at 3% BML, 78% at 0800 h; P < 0.01) and lysozyme (46% at 3% BML, 61% at 0800 h; P < 0.01), which were lower than in CON at these time points (P < 0.05). Rehydration returned all saliva variables to baseline. In conclusion, modest dehydration (~3% BML) decreased SFR, α-amylase, and lysozyme secretion rates. Whether the observed magnitude of decrease in saliva AMPs during dehydration compromises host defence remains to be shown.

The germacranolide sesquiterpene lactone neurolenin B of the medicinal plant Neurolaena lobata (L.) R.Br. ex Cass inhibits NPM/ALK-driven cell expansion and NF-κB-driven tumour intravasation.

PubMed

Unger, Christine; Kiss, Izabella; Vasas, Andrea; Lajter, Ildikó; Kramer, Nina; Atanasov, Atanas Georgiev; Nguyen, Chi Huu; Chatuphonprasert, Waranya; Brenner, Stefan; Krieger, Sigurd; McKinnon, Ruxandra; Peschel, Andrea; Kain, Renate; Saiko, Philipp; Szekeres, Thomas; Kenner, Lukas; Hassler, Melanie R; Diaz, Rene; Frisch, Richard; Dirsch, Verena M; Jäger, Walter; de Martin, Rainer; Bochkov, Valery N; Passreiter, Claus M; Peter-Vörösmarty, Barbara; Mader, Robert M; Grusch, Michael; Dolznig, Helmut; Kopp, Brigitte; Zupko, Istvan; Hohmann, Judit; Krupitza, Georg

2015-08-15

The t(2;5)(p23;q35) chromosomal translocation results in the expression of the fusion protein NPM/ALK that when expressed in T-lymphocytes gives rise to anaplastic large cell lymphomas (ALCL). In search of new therapy options the dichloromethane extract of the ethnomedicinal plant Neurolaena lobata (L.) R.Br. ex Cass was shown to inhibit NPM/ALK expression. Therefore, we analysed whether the active principles that were recently isolated and found to inhibit inflammatory responses specifically inhibit growth of NPM/ALK+ ALCL, leukaemia and breast cancer cells, but not of normal cells, and the intravasation through the lymphendothelial barrier. ALCL, leukaemia and breast cancer cells, and normal peripheral blood mononuclear cells (PBMCs) were treated with isolated sesquiterpene lactones and analysed for cell cycle progression, proliferation, mitochondrial activity, apoptosis, protein and mRNA expression, NF-κB and cytochrome P450 activity, 12(S)-HETE production and lymphendothelial intravasation. In vitro treatment of ALCL by neurolenin B suppressed NPM/ALK, JunB and PDGF-Rβ expression, inhibited the growth of ALCL cells late in M phase, and induced apoptosis via caspase 3 without compromising mitochondrial activity (as a measure of general exogenic toxicity). Moreover, neurolenin B attenuated tumour spheroid intravasation probably through inhibition of NF-κB and CYP1A1. Neurolenin B specifically decreased pro-carcinogenic NPM/ALK expression in ALK+ ALCL cells and, via the inhibition of NF-kB signalling, attenuated tumour intra/extravasation into the lymphatics. Hence, neurolenin B may open new options to treat ALCL and to manage early metastatic processes to which no other therapies exist. Copyright © 2015 Elsevier GmbH. All rights reserved.

Lethal cutaneous disease in transgenic mice conditionally expressing type I human T cell leukemia virus Tax.

PubMed

Kwon, Hakju; Ogle, Louise; Benitez, Bobby; Bohuslav, Jan; Montano, Mauricio; Felsher, Dean W; Greene, Warner C

2005-10-21

Type I human T cell leukemia virus (HTLV-I) is etiologically linked with adult T cell leukemia, an aggressive and usually fatal expansion of activated CD4+ T lymphocytes that frequently traffic to skin. T cell transformation induced by HTLV-I involves the action of the 40-kDa viral Tax transactivator protein. Tax both stimulates the HTLV-I long terminal repeat and deregulates the expression of select cellular genes by altering the activity of specific host transcription factors, including cyclic AMP-responsive element-binding protein (CREB)/activating transcription factor, NF-kappaB/Rel, and serum response factor. To study initiating events involved in HTLV-I Tax-induced T cell transformation, we generated "Tet-off" transgenic mice conditionally expressing in a lymphocyte-restricted manner (EmuSR alpha promoter-enhancer) either wild-type Tax or mutant forms of Tax that selectively compromise the NF-kappaB (M22) or CREB/activating transcription factor (M47) activation pathways. Wild-type Tax and M47 Tax-expressing mice, but not M22-Tax expressing mice, developed progressive alopecia, hyperkeratosis, and skin lesions containing profuse activated CD4 T cell infiltrates with evidence of deregulated inflammatory cytokine production. In addition, these animals displayed systemic lymphadenopathy and splenomegaly. These findings suggest that Tax-mediated activation of NF-kappaB plays a key role in the development of this aggressive skin disease that shares several features in common with the skin disease occurring during the preleukemic stage in HTLV-I-infected patients. Of note, this skin disease completely resolved when Tax transgene expression was suppressed by administration of doxycycline, emphasizing the key role played by this viral oncoprotein in the observed pathology.

Inhibition of activated NR2B gene- and caspase-3 protein-expression by glutathione following traumatic brain injury in a rat model

PubMed Central

Arifin, Muhammad Zafrullah; Faried, Ahmad; Shahib, Muhammad Nurhalim; Wiriadisastra, Kahdar; Bisri, Tatang

2011-01-01

Background. Traumatic brain injury (TBI) remains a major cause of death and disability. Oxidative stress is an important element of the injury cascade following TBI. Progressive compromise of antioxidant defenses and free radical-mediated lipid peroxidation are one of the major mechanisms of secondary TBI. NR2B is a glutamate receptor and its activation is caused by TBI increasing a brain cell death, along with caspase-3 as a hall mark of apoptosis. Glutathione is a potent free radical scavenger that might prevent secondary TBI damage and inhibited apoptosis. Materials and Methods. In the present study, it aims to demonstrate the effect of glutathione on inhibition of brain oxidative damage in a TBI rat model. Results. In this study, the expressions of mRNA NR2B in placebo group and groups with glutathione administration at 0, 3, and 6 hours after TBI were 328.14, 229.90, 178.50, and 136.14, respectively (P<0.001). The highest caspase-3 expression was shown in placebo group with 66.7% showing strong positive results (>80%); as expected, glutathione administered in 0, 3, and 6 hours groups had lower strong positive results of 50%, 16.7%, and 16.7%, respectively, (P=0.025). Conclusion. In conclusion, this study showed that glutathione administration in a TBI rat model decreased NR2B gene- and caspase-3 protein-expression that lead to the inhibition of brain cell death. Our results suggest that glutathione, as a potent free radical scavenger, has a brain cell protective effect against oxidative damage and cell death induced by TBI in rat model. PMID:22347327

Application of the International Classification of Functioning, Disability and Health system to symptoms of the Duchenne and Becker muscular dystrophies.

PubMed

Conway, Kristin M; Ciafaloni, Emma; Matthews, Dennis; Westfield, Chris; James, Kathy; Paramsothy, Pangaja; Romitti, Paul A

2018-07-01

Duchenne and Becker muscular dystrophies, collectively referred to as dystrophinopathies, are X-linked recessive diseases that affect dystrophin production resulting in compromised muscle function across multiple systems. The International Classification of Functioning, Disability and Health provides a systematic classification scheme from which body functions affected by a dystrophinopathy can be identified and used to examine functional health. The infrastructure of the Muscular Dystrophy Surveillance, Tracking, and Research Network was used to identify commonly affected body functions and link selected functions to clinical surveillance data collected through medical record abstraction. Seventy-one (24 second-, 41 third- and 7 fourth-level) body function categories were selected via clinician review and consensus. Of these, 15 of 24 retained second-level categories were linked to data elements from the Muscular Dystrophy Surveillance, Tracking, and Research Network surveillance database. Our findings support continued development of a core set of body functions from the International Classification of Functioning, Disability and Health system that are representative of disease progression in dystrophinopathies and the incorporation of these functions in standardized evaluations of functional health and implementation of individualized rehabilitation care plans. Implications for Rehabilitation Duchenne and Becker muscular dystrophies, collectively referred to as dystrophinopathies, are X-linked recessive disorders that affect the production of dystrophin resulting in compromised muscle function across multiple systems. The severity and progressive nature of dystrophinopathies can have considerable impact on a patient's participation in activities across multiple life domains. Our findings support continued development of an International Classification of Functioning, Disability and Health core set for childhood-onset dystrophinopathies. A standardized dystrophinopathy International Classification of Functioning, Disability and Health documentation form can be used as a screening tool by rehabilitation professionals and for patient goal setting when developing rehabilitation plans. Patient reports of perceived functional health should be incorporated into the rehabilitation plan and therapeutic progress monitored by a standardized form.

Inhibition of E2F1 activity and cell cycle progression by arsenic via retinoblastoma protein.

PubMed

Sheldon, Lynn A

2017-01-01

The regulation of cell cycle progression by steroid hormones and growth factors is important for maintaining normal cellular processes including development and cell proliferation. Deregulated progression through the G1/S and G2/M cell cycle transitions can lead to uncontrolled cell proliferation and cancer. The transcription factor E2F1, a key cell cycle regulator, targets genes encoding proteins that regulate cell cycle progression through the G1/S transition as well as proteins important in DNA repair and apoptosis. E2F1 expression and activity is inhibited by inorganic arsenic (iAs) that has a dual role as a cancer therapeutic and as a toxin that leads to diseases including cancer. An understanding of what underlies this dichotomy will contribute to understanding how to use iAs as a more effective therapeutic and also how to treat cancers that iAs promotes. Here, we show that quiescent breast adenocarcinoma MCF-7 cells treated with 17-β estradiol (E2) progress through the cell cycle, but few cells treated with E2 + iAs progress from G1 into S-phase due to a block in cell cycle progression. Our data support a model in which iAs inhibits the dissociation of E2F1 from the tumor suppressor, retinoblastoma protein (pRB) due to changes in pRB phosphorylation which leads to decreased E2F1 transcriptional activity. These findings present an explanation for how iAs can disrupt cell cycle progression through E2F1-pRB and has implications for how iAs acts as a cancer therapeutic as well as how it may promote tumorigenesis through decreased DNA repair.

Characterization of a nuclear localization signal in the C-terminus of the adeno-associated virus Rep68/78 proteins

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

Cassell, Geoffrey D.; Weitzman, Matthew D.

2004-10-01

Adeno-associated virus (AAV) replicates in the nucleus of infected cells, and therefore multiple nuclear import events are required for productive infection. We analyzed nuclear import of the viral Rep proteins and characterized a nuclear localization signal (NLS) in the C-terminus. We demonstrate that basic residues in this region constitute an NLS that is transferable and mediates interaction with the nuclear import receptor importin {alpha} in vitro. Mutant Rep proteins are predominantly cytoplasmic and are severely compromised for interactions with importin {alpha}, but retain their enzymatic functions in vitro. Interestingly, mutations of the NLS had significantly less effect on importin {alpha}more » interaction and replication in the context of Rep78 than when incorporated into the Rep68 protein. Together, our results demonstrate that a bipartite NLS exists in the shared part of Rep68 and Rep78, and suggest that an alternate entry mechanism may also contribute to nuclear localization of the Rep78 protein.« less

Glycation of H1 Histone by 3-Deoxyglucosone: Effects on Protein Structure and Generation of Different Advanced Glycation End Products

PubMed Central

Ashraf, Jalaluddin Mohammad; Rabbani, Gulam; Ahmad, Saheem; Hasan, Qambar; Khan, Rizwan Hasan; Alam, Khursheed; Choi, Inho

2015-01-01

Advanced glycation end products (AGEs) culminate from the non-enzymatic reaction between a free carbonyl group of a reducing sugar and free amino group of proteins. 3-deoxyglucosone (3-DG) is one of the dicarbonyl species that rapidly forms several protein-AGE complexes that are believed to be involved in the pathogenesis of several diseases, particularly diabetic complications. In this study, the generation of AGEs (Nε-carboxymethyl lysine and pentosidine) by 3-DG in H1 histone protein was characterized by evaluating extent of side chain modification (lysine and arginine) and formation of Amadori products as well as carbonyl contents using several physicochemical techniques. Results strongly suggested that 3-DG is a potent glycating agent that forms various intermediates and AGEs during glycation reactions and affects the secondary structure of the H1 protein. Structural changes and AGE formation may influence the function of H1 histone and compromise chromatin structures in cases of secondary diabetic complications. PMID:26121680

Genetic Engineering of the Phosphocarrier Protein NPr of the Escherichia coli Phosphotransferase System Selectively Improves Sugar Uptake Activity*

PubMed Central

Lopez-de los Santos, Yossef; Chan, Henry; Cantu, Vito A.; Rettner, Rachael; Sanchez, Filiberto; Zhang, Zhongge; Saier, Milton H.; Soberon, Xavier

2012-01-01

The Escherichia coli phosphoenolpyruvate:sugar phosphotransferase system (PTS) in prokaryotes mediates the uptake and phosphorylation of its numerous substrates through a phosphoryl transfer chain where a phosphoryl transfer protein, HPr, transfers its phosphoryl group to any of several sugar-specific Enzyme IIA proteins in preparation for sugar transport. A phosphoryl transfer protein of the PTS, NPr, homologous to HPr, functions to regulate nitrogen metabolism and shows virtually no enzymatic cross-reactivity with HPr. Here we describe the genetic engineering of a “chimeric” HPr/NPr protein, termed CPr14 because 14 amino acid residues of the interface were replaced. CPr14 shows decreased activity with most PTS permeases relative to HPr, but increases activity with the broad specificity mannose permease. The results lead to the proposal that HPr is not optimal for most PTS permeases but instead represents a compromise with suboptimal activity for most PTS permeases. The evolutionary implications are discussed. PMID:22767600

Balancing energy and entropy: A minimalist model for the characterization of protein folding landscapes

PubMed Central

Das, Payel; Matysiak, Silvina; Clementi, Cecilia

2005-01-01

Coarse-grained models have been extremely valuable in promoting our understanding of protein folding. However, the quantitative accuracy of existing simplified models is strongly hindered either from the complete removal of frustration (as in the widely used Gō-like models) or from the compromise with the minimal frustration principle and/or realistic protein geometry (as in the simple on-lattice models). We present a coarse-grained model that “naturally” incorporates sequence details and energetic frustration into an overall minimally frustrated folding landscape. The model is coupled with an optimization procedure to design the parameters of the protein Hamiltonian to fold into a desired native structure. The application to the study of src-Src homology 3 domain shows that this coarse-grained model contains the main physical-chemical ingredients that are responsible for shaping the folding landscape of this protein. The results illustrate the importance of nonnative interactions and energetic heterogeneity for a quantitative characterization of folding mechanisms. PMID:16006532

Inflammation disrupts the LDL receptor pathway and accelerates the progression of vascular calcification in ESRD patients.

PubMed

Liu, Jing; Ma, Kun Ling; Gao, Min; Wang, Chang Xian; Ni, Jie; Zhang, Yang; Zhang, Xiao Liang; Liu, Hong; Wang, Yan Li; Liu, Bi Cheng

2012-01-01

Chronic inflammation plays a crucial role in the progression of vascular calcification (VC). This study was designed to investigate whether the low-density lipoprotein receptor (LDLr) pathway is involved in the progression of VC in patients with end-stage renal disease (ESRD) during inflammation. Twenty-eight ESRD patients were divided into control and inflamed groups according to plasma C-reactive protein (CRP) level. Surgically removed tissues from the radial arteries of patients receiving arteriovenostomy were used in the experiments. The expression of tumour necrosis factor-α (TNF-α) and monocyte chemotactic protein-1 (MCP-1) of the radial artery were increased in the inflamed group. Hematoxylin-eosin and alizarin red S staining revealed parallel increases in foam cell formation and calcium deposit formation in continuous cross-sections of radial arteries in the inflamed group compared to the control, which were closely correlated with increased LDLr, sterol regulatory element binding protein-2 (SREBP-2), bone morphogenetic proteins-2 (BMP-2), and collagen I protein expression, as shown by immunohistochemical and immunofluorescent staining. Confocal microscopy confirmed that inflammation enhanced the translocation of the SREBP cleavage-activating protein (SCAP)/SREBP-2 complex from the endoplasmic reticulum to the Golgi, thereby activating LDLr gene transcription. Inflammation increased alkaline phosphatase protein expression and reduced α-smooth muscle actin protein expression, contributing to the conversion of the vascular smooth muscle cells in calcified vessels from the fibroblastic to the osteogenic phenotype; osteogenic cells are the main cellular components involved in VC. Further analysis showed that the inflammation-induced disruption of the LDLr pathway was significantly associated with enhanced BMP-2 and collagen I expression. Inflammation accelerated the progression of VC in ESRD patients by disrupting the LDLr pathway, which may represent a novel mechanism involved in the progression of both VC and atherosclerosis.

Quantitative Proteomic Analysis of the Influenza A Virus Nonstructural Proteins NS1 and NS2 during Natural Cell Infection Identifies PACT as an NS1 Target Protein and Antiviral Host Factor

PubMed Central

Tawaratsumida, Kazuki; Phan, Van; Hrincius, Eike R.; High, Anthony A.; Webby, Richard; Redecke, Vanessa

2014-01-01

ABSTRACT Influenza A virus (IAV) replication depends on the interaction of virus proteins with host factors. The viral nonstructural protein 1 (NS1) is essential in this process by targeting diverse cellular functions, including mRNA splicing and translation, cell survival, and immune defense, in particular the type I interferon (IFN-I) response. In order to identify host proteins targeted by NS1, we established a replication-competent recombinant IAV that expresses epitope-tagged forms of NS1 and NS2, which are encoded by the same gene segment, allowing purification of NS proteins during natural cell infection and analysis of interacting proteins by quantitative mass spectrometry. We identified known NS1- and NS2-interacting proteins but also uncharacterized proteins, including PACT, an important cofactor for the IFN-I response triggered by the viral RNA-sensor RIG-I. We show here that NS1 binds PACT during virus replication and blocks PACT/RIG-I-mediated activation of IFN-I, which represents a critical event for the host defense. Protein interaction and interference with IFN-I activation depended on the functional integrity of the highly conserved RNA binding domain of NS1. A mutant virus with deletion of NS1 induced high levels of IFN-I in control cells, as expected; in contrast, shRNA-mediated knockdown of PACT compromised IFN-I activation by the mutant virus, but not wild-type virus, a finding consistent with the interpretation that PACT (i) is essential for IAV recognition and (ii) is functionally compromised by NS1. Together, our data describe a novel approach to identify virus-host protein interactions and demonstrate that NS1 interferes with PACT, whose function is critical for robust IFN-I production. IMPORTANCE Influenza A virus (IAV) is an important human pathogen that is responsible for annual epidemics and occasional devastating pandemics. Viral replication and pathogenicity depends on the interference of viral factors with components of the host defense system, particularly the type I interferon (IFN-I) response. The viral NS1 protein is known to counteract virus recognition and IFN-I production, but the molecular mechanism is only partially defined. We used a novel proteomic approach to identify host proteins that are bound by NS1 during virus replication and identified the protein PACT, which had previously been shown to be involved in virus-mediated IFN-I activation. We find that NS1 prevents PACT from interacting with an essential component of the virus recognition pathway, RIG-I, thereby disabling efficient IFN-I production. These observations provide an important piece of information on how IAV efficiently counteracts the host immune defense. PMID:24899174

Single-column purification of the tag-free, recombinant form of the neuronal calcium sensor protein, hippocalcin expressed in Escherichia coli.

PubMed

Krishnan, Anuradha; Viviano, Jeffrey; Morozov, Yaroslav; Venkataraman, Venkat

2016-07-01

Hippocalcin is a 193 aa protein that is a member of the neuronal calcium sensor protein family, whose functions are regulated by calcium. Mice that lack the function of this protein are compromised in the long term potentiation aspect of memory generation. Recently, mutations in the gene have been linked with dystonia in human. The protein has no intrinsic enzyme activity but is known to bind to variety of target proteins. Very little information is available on how the protein executes its critical role in signaling pathways, except that it is regulated by binding of calcium. Further delineation of its function requires large amounts of pure protein. In this report, we present a single-step purification procedure that yields high quantities of the bacterially expressed, recombinant protein. The procedure may be adapted to purify the protein from inclusion bodies or cytosol in its myristoylated or non-myristoylated forms. MALDI-MS (in source decay) analyses demonstrates that the myristoylation occurs at the glycine residue. The protein is also biologically active as measured through tryptophan fluorescence, mobility shift and guanylate cyclase activity assays. Thus, further analyses of hippocalcin, both structural and functional, need no longer be limited by protein availability. Copyright © 2016 Elsevier Inc. All rights reserved.

Molecular chaperones and protein folding as therapeutic targets in Parkinson's disease and other synucleinopathies.

PubMed

Ebrahimi-Fakhari, Darius; Saidi, Laiq-Jan; Wahlster, Lara

2013-12-05

Changes in protein metabolism are key to disease onset and progression in many neurodegenerative diseases. As a prime example, in Parkinson's disease, folding, post-translational modification and recycling of the synaptic protein α-synuclein are clearly altered, leading to a progressive accumulation of pathogenic protein species and the formation of intracellular inclusion bodies. Altered protein folding is one of the first steps of an increasingly understood cascade in which α-synuclein forms complex oligomers and finally distinct protein aggregates, termed Lewy bodies and Lewy neurites. In neurons, an elaborated network of chaperone and co-chaperone proteins is instrumental in mediating protein folding and re-folding. In addition to their direct influence on client proteins, chaperones interact with protein degradation pathways such as the ubiquitin-proteasome-system or autophagy in order to ensure the effective removal of irreversibly misfolded and potentially pathogenic proteins. Because of the vital role of proper protein folding for protein homeostasis, a growing number of studies have evaluated the contribution of chaperone proteins to neurodegeneration. We herein review our current understanding of the involvement of chaperones, co-chaperones and chaperone-mediated autophagy in synucleinopathies with a focus on the Hsp90 and Hsp70 chaperone system. We discuss genetic and pathological studies in Parkinson's disease as well as experimental studies in models of synucleinopathies that explore molecular chaperones and protein degradation pathways as a novel therapeutic target. To this end, we examine the capacity of chaperones to prevent or modulate neurodegeneration and summarize the current progress in models of Parkinson's disease and related neurodegenerative disorders.

Phosphorus and Nutrition in Chronic Kidney Disease

PubMed Central

González-Parra, Emilio; Gracia-Iguacel, Carolina; Egido, Jesús; Ortiz, Alberto

2012-01-01

Patients with renal impairment progressively lose the ability to excrete phosphorus. Decreased glomerular filtration of phosphorus is initially compensated by decreased tubular reabsorption, regulated by PTH and FGF23, maintaining normal serum phosphorus concentrations. There is a close relationship between protein and phosphorus intake. In chronic renal disease, a low dietary protein content slows the progression of kidney disease, especially in patients with proteinuria and decreases the supply of phosphorus, which has been directly related with progression of kidney disease and with patient survival. However, not all animal proteins and vegetables have the same proportion of phosphorus in their composition. Adequate labeling of food requires showing the phosphorus-to-protein ratio. The diet in patients with advanced-stage CKD has been controversial, because a diet with too low protein content can favor malnutrition and increase morbidity and mortality. Phosphorus binders lower serum phosphorus and also FGF23 levels, without decreasing diet protein content. But the interaction between intestinal dysbacteriosis in dialysis patients, phosphate binder efficacy, and patient tolerance to the binder could reduce their efficiency. PMID:22701173

Novel CXCR3/CXCR7-Directed Biological Antagonist for Inhibition of Breast Cancer Progression

DTIC Science & Technology

2012-09-01

neutropenia ), hearing loss, and death. Hence, therapies that simultaneously prevent BrCa progression and improve docetaxel efficacy are greatly needed...kinase (RAFTK/PYK2), Crk and Paxillin. Crk, which belongs to the adaptor family of proteins composed of SH2 (Src Homology 2) and SH3 domains, has a...Phosphorylated Akt in turn phosphorylates Bad, a pro-apoptotic protein belonging to the Bcl- 2 family . When phosphorylated, the cytoplasmic protein

New candidate markers of head and neck squamous cell carcinoma progression

NASA Astrophysics Data System (ADS)

Kakurina, G. V.; Kolegova, E. S.; Cheremisina, O. V.; Kulbakin, D. E.; Choinzonov, E. L.

2017-09-01

The tumor progression in head and neck squamous cell carcinoma (HNSCC) is one of the main causes of high mortality of the patients with HNSCC. The tumor progression, particularly the metastasis, is characterized by the changes in the composition, functions and structure of different proteins. We have previously shown that serum of HNSCC patients contains the proteins which regulate various cellular processes—adenylyl cyclase associated protein 1 (CAP1), protein phosphatase 1 B (PPM1B), etc. The levels of CAP1 and PPM1B were determined using the enzyme immunoassay. The results of this study show that CAP1 and PPM1B take a part in the progression of HNSCC. The levels of CAP1 and PPM1B in the tumor and in morphologically normal tissue depended on the prevalence of the tumor process. The CAP1 and PPM1B levels were significantly higher in tumor tissue of the patients with regional metastasis. Our data allow assuming the potential possibility for predicting the outcome of the HNSCC measuring the level of tissue CAP1.

Surfing the Big WAVE: Insights into the Role of WAVE3 as a Driving Force in Cancer Progression and Metastasis

PubMed Central

Sossey-Alaoui, Khalid

2014-01-01

WAVE3 belongs to the WASP/WAVE family of actin cytoskeleton remodeling proteins. These proteins are known to be involved in several biological functions ranging from controlling cell shape and movement, to being closely associated with pathological conditions such as cancer progression and metastasis. Last decade has seen an explosion in the literature reporting significant scientific advances on the molecular mechanisms whereby the WASP/WAVE proteins are regulated both in normal physiological as well as pathological conditions. The purpose of this review is to present the major findings pertaining to how WAVE3 has become a critical player in the regulation of signaling pathways involved in cancer progression and metastasis. The review will conclude with suggesting options for the potential use of WAVE3 as a therapeutic target to prevent the progression of cancer to the lethal stage that is the metastatic disease. PMID:23116924

Mammalian O-Mannosylation Pathway: Glycan Structures, Enzymes, and Protein Substrates

PubMed Central

2015-01-01

The mammalian O-mannosylation pathway for protein post-translational modification is intricately involved in modulating cell–matrix interactions in the musculature and nervous system. Defects in enzymes of this biosynthetic pathway are causative for multiple forms of congenital muscular dystophy. The application of advanced genetic and biochemical technologies has resulted in remarkable progress in this field over the past few years, culminating with the publication of three landmark papers in 2013 alone. In this review, we will highlight recent progress focusing on the dramatic expansion of the set of genes known to be involved in O-mannosylation and disease processes, the concurrent acceleration of the rate of O-mannosylation pathway protein functional assignments, the tremendous increase in the number of proteins now known to be modified by O-mannosylation, and the recent progress in protein O-mannose glycan quantification and site assignment. Also, we attempt to highlight key outstanding questions raised by this abundance of new information. PMID:24786756

The Rho GTPase effector ROCK regulates cyclin A, cyclin D1, and p27Kip1 levels by distinct mechanisms.

PubMed

Croft, Daniel R; Olson, Michael F

2006-06-01

The members of the Rho GTPase family are well known for their regulation of actin cytoskeletal structures. In addition, they influence progression through the cell cycle. The RhoA and RhoC proteins regulate numerous effector proteins, with a central and vital signaling role mediated by the ROCK I and ROCK II serine/threonine kinases. The requirement for ROCK function in the proliferation of numerous cell types has been revealed by studies utilizing ROCK-selective inhibitors such as Y-27632. However, the mechanisms by which ROCK signaling promotes cell cycle progression have not been thoroughly characterized. Using a conditionally activated ROCK-estrogen receptor fusion protein, we found that ROCK activation is sufficient to stimulate G1/S cell cycle progression in NIH 3T3 mouse fibroblasts. Further analysis revealed that ROCK acts via independent pathways to alter the levels of cell cycle regulatory proteins: cyclin D1 and p21(Cip1) elevation via Ras and the mitogen-activated protein kinase pathway, increased cyclin A via LIM kinase 2, and reduction of p27(Kip1) protein levels. Therefore, the influence of ROCK on cell cycle regulatory proteins occurs by multiple independent mechanisms.

A Novel Interaction of Ecdysoneless (ECD) Protein with R2TP Complex Component RUVBL1 Is Required for the Functional Role of ECD in Cell Cycle Progression.

PubMed

Mir, Riyaz A; Bele, Aditya; Mirza, Sameer; Srivastava, Shashank; Olou, Appolinaire A; Ammons, Shalis A; Kim, Jun Hyun; Gurumurthy, Channabasavaiah B; Qiu, Fang; Band, Hamid; Band, Vimla

2015-12-28

Ecdysoneless (ECD) is an evolutionarily conserved protein whose germ line deletion is embryonic lethal. Deletion of Ecd in cells causes cell cycle arrest, which is rescued by exogenous ECD, demonstrating a requirement of ECD for normal mammalian cell cycle progression. However, the exact mechanism by which ECD regulates cell cycle is unknown. Here, we demonstrate that ECD protein levels and subcellular localization are invariant during cell cycle progression, suggesting a potential role of posttranslational modifications or protein-protein interactions. Since phosphorylated ECD was recently shown to interact with the PIH1D1 adaptor component of the R2TP cochaperone complex, we examined the requirement of ECD phosphorylation in cell cycle progression. Notably, phosphorylation-deficient ECD mutants that failed to bind to PIH1D1 in vitro fully retained the ability to interact with the R2TP complex and yet exhibited a reduced ability to rescue Ecd-deficient cells from cell cycle arrest. Biochemical analyses demonstrated an additional phosphorylation-independent interaction of ECD with the RUVBL1 component of the R2TP complex, and this interaction is essential for ECD's cell cycle progression function. These studies demonstrate that interaction of ECD with RUVBL1, and its CK2-mediated phosphorylation, independent of its interaction with PIH1D1, are important for its cell cycle regulatory function. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

A Novel Interaction of Ecdysoneless (ECD) Protein with R2TP Complex Component RUVBL1 Is Required for the Functional Role of ECD in Cell Cycle Progression

PubMed Central

Mir, Riyaz A.; Bele, Aditya; Mirza, Sameer; Srivastava, Shashank; Olou, Appolinaire A.; Ammons, Shalis A.; Kim, Jun Hyun; Gurumurthy, Channabasavaiah B.; Qiu, Fang; Band, Hamid

2015-01-01

Ecdysoneless (ECD) is an evolutionarily conserved protein whose germ line deletion is embryonic lethal. Deletion of Ecd in cells causes cell cycle arrest, which is rescued by exogenous ECD, demonstrating a requirement of ECD for normal mammalian cell cycle progression. However, the exact mechanism by which ECD regulates cell cycle is unknown. Here, we demonstrate that ECD protein levels and subcellular localization are invariant during cell cycle progression, suggesting a potential role of posttranslational modifications or protein-protein interactions. Since phosphorylated ECD was recently shown to interact with the PIH1D1 adaptor component of the R2TP cochaperone complex, we examined the requirement of ECD phosphorylation in cell cycle progression. Notably, phosphorylation-deficient ECD mutants that failed to bind to PIH1D1 in vitro fully retained the ability to interact with the R2TP complex and yet exhibited a reduced ability to rescue Ecd-deficient cells from cell cycle arrest. Biochemical analyses demonstrated an additional phosphorylation-independent interaction of ECD with the RUVBL1 component of the R2TP complex, and this interaction is essential for ECD's cell cycle progression function. These studies demonstrate that interaction of ECD with RUVBL1, and its CK2-mediated phosphorylation, independent of its interaction with PIH1D1, are important for its cell cycle regulatory function. PMID:26711270

The Ustilago maydis a2 Mating-Type Locus Genes lga2 and rga2 Compromise Pathogenicity in the Absence of the Mitochondrial p32 Family Protein Mrb1

PubMed Central

Bortfeld, Miriam; Auffarth, Kathrin; Kahmann, Regine; Basse, Christoph W.

2004-01-01

The Ustilago maydis mrb1 gene specifies a mitochondrial matrix protein with significant similarity to mitochondrial p32 family proteins known from human and many other eukaryotic species. Compatible mrb1 mutant strains were able to mate and form dikaryotic hyphae; however, proliferation within infected tissue and the ability to induce tumor development of infected maize (Zea mays) plants were drastically impaired. Surprisingly, manifestation of the mrb1 mutant phenotype selectively depended on the a2 mating type locus. The a2 locus contains, in addition to pheromone signaling components, the genes lga2 and rga2 of unknown function. Deletion of lga2 in an a2Δmrb1 strain fully restored pathogenicity, whereas pathogenicity was partially regained in an a2Δmrb1Δrga2 strain, implicating a concerted action between Lga2 and Rga2 in compromising pathogenicity in Δmrb1 strains. Lga2 and Rga2 localized to mitochondria and Mrb1 interacted with Rga2 in the yeast two-hybrid system. Conditional expression of lga2 in haploid cells reduced vegetative growth, conferred mitochondrial fragmentation and mitochondrial DNA degradation, and interfered with respiratory activity. The consequences of lga2 overexpression depended on the expression strength and were greatly exacerbated in Δmrb1 mutants. We propose that Lga2 interferes with mitochondrial fusion and that Mrb1 controls this activity, emphasizing a critical link between mitochondrial morphology and pathogenicity. PMID:15273296

Maintenance of mitochondrial DNA copy number and expression are essential for preservation of mitochondrial function and cell growth.

PubMed

Jeng, Jaan-Yeh; Yeh, Tien-Shun; Lee, Jing-Wen; Lin, Shyh-Hsiang; Fong, Tsorng-Han; Hsieh, Rong-Hong

2008-02-01

To examine whether a reduction in the mtDNA level will compromise mitochondrial biogenesis and mitochondrial function, we created a cell model with depleted mtDNA. Stable transfection of small interfering (si)RNA of mitochondrial transcription factor A (Tfam) was used to interfere with Tfam gene expression. Selected stable clones showed 60-95% reduction in Tfam gene expression and 50-90% reduction in cytochrome b (Cyt b) gene expression. Tfam gene knockdown clones also showed decreased mtDNA-encoded cytochrome c oxidase subunit I (COX I) protein expression. However, no significant differences in protein expression were observed in nuclear DNA (nDNA)-encoded mitochondrial respiratory enzyme subunits. The cell morphology changed from a rhombus-like to a spindle-like form as determined in clones with decreased expressions of Tfam, mtRNA, and mitochondrial proteins. The mitochondrial respiratory enzyme activities and ATP production in such clones were significantly lower. The proportions of mtDNA mutations including 8-hydroxy-2'-deoxyguanosine (8-OHdG), a 4,977-bp deletion, and a 3,243-point mutation were also examined in these clones. No obvious increase in mtDNA mutations was observed in mitochondrial dysfunctional cell clones. The mitochondrial respiratory activity and ATP production ability recovered in cells with increased mtDNA levels after removal of the specific siRNA treatment. These experimental results provide direct evidence to substantiate that downregulation of mtDNA copy number and expression may compromise mitochondrial function and subsequent cell growth and morphology. (c) 2007 Wiley-Liss, Inc.

The Batten disease gene CLN3 is required for the response to oxidative stress

PubMed Central

Tuxworth, Richard I.; Chen, Haiyang; Vivancos, Valerie; Carvajal, Nancy; Huang, Xun; Tear, Guy

2011-01-01

Mutations in the CLN3 gene cause juvenile neuronal ceroid lipofuscinosis (JNCL or Batten disease), an early onset neurodegenerative disorder. JNCL is the most common of the NCLs, a group of disorders with infant or childhood onset that are caused by single gene mutations. The NCLs, although relatively rare, share many pathological and clinical similarities with the more common late-onset neurodegenerative disorders, while their simple genetic basis makes them an excellent paradigm. The early onset and rapid disease progression in the NCLs suggests that one or more key cellular processes are severely compromised. To identify the functional pathways compromised in JNCL, we have performed a gain-of-function modifier screen in Drosophila. We find that CLN3 interacts genetically with the core stress signalling pathways and components of stress granules, suggesting a function in stress responses. In support of this, we find that Drosophila lacking CLN3 function are hypersensitive to oxidative stress yet they respond normally to other physiological stresses. Overexpression of CLN3 is sufficient to confer increased resistance to oxidative stress. We find that CLN3 mutant flies perceive conditions of increased oxidative stress correctly but are unable to detoxify reactive oxygen species, suggesting that their ability to respond is compromised. Together, our data suggest that the lack of CLN3 function leads to a failure to manage the response to oxidative stress and this may be the key deficit in JNCL that leads to neuronal degeneration. PMID:21372148

A Novel Approach to the Identification of Compromised Pulmonary Systems in Smokers by Exploiting Tidal Breathing Patterns.

PubMed

Rakshit, Raj; Khasnobish, Anwesha; Chowdhury, Arijit; Sinharay, Arijit; Pal, Arpan; Chakravarty, Tapas

2018-04-25

Smoking causes unalterable physiological abnormalities in the pulmonary system. This is emerging as a serious threat worldwide. Unlike spirometry, tidal breathing does not require subjects to undergo forceful breathing maneuvers and is progressing as a new direction towards pulmonary health assessment. The aim of the paper is to evaluate whether tidal breathing signatures can indicate deteriorating adult lung condition in an otherwise healthy person. If successful, such a system can be used as a pre-screening tool for all people before some of them need to undergo a thorough clinical checkup. This work presents a novel systematic approach to identify compromised pulmonary systems in smokers from acquired tidal breathing patterns. Tidal breathing patterns are acquired during restful breathing of adult participants. Thereafter, physiological attributes are extracted from the acquired tidal breathing signals. Finally, a unique classification approach of locally weighted learning with ridge regression (LWL-ridge) is implemented, which handles the subjective variations in tidal breathing data without performing feature normalization. The LWL-ridge classifier recognized compromised pulmonary systems in smokers with an average classification accuracy of 86.17% along with a sensitivity of 80% and a specificity of 92%. The implemented approach outperformed other variants of LWL as well as other standard classifiers and generated comparable results when applied on an external cohort. This end-to-end automated system is suitable for pre-screening people routinely for early detection of lung ailments as a preventive measure in an infrastructure-agnostic way.

The therapeutic potential of cell cycle targeting in multiple myeloma.

PubMed

Maes, Anke; Menu, Eline; Veirman, Kim De; Maes, Ken; Vand Erkerken, Karin; De Bruyne, Elke

2017-10-27

Proper cell cycle progression through the interphase and mitosis is regulated by coordinated activation of important cell cycle proteins (including cyclin-dependent kinases and mitotic kinases) and several checkpoint pathways. Aberrant activity of these cell cycle proteins and checkpoint pathways results in deregulation of cell cycle progression, which is one of the key hallmarks of cancer. Consequently, intensive research on targeting these cell cycle regulatory proteins identified several candidate small molecule inhibitors that are able to induce cell cycle arrest and even apoptosis in cancer cells. Importantly, several of these cell cycle regulatory proteins have also been proposed as therapeutic targets in the plasma cell malignancy multiple myeloma (MM). Despite the enormous progress in the treatment of MM the past 5 years, MM still remains most often incurable due to the development of drug resistance. Deregulated expression of the cyclins D is observed in virtually all myeloma patients, emphasizing the potential therapeutic interest of cyclin-dependent kinase inhibitors in MM. Furthermore, other targets have also been identified in MM, such as microtubules, kinesin motor proteins, aurora kinases, polo-like kinases and the anaphase promoting complex/cyclosome. This review will provide an overview of the cell cycle proteins and checkpoint pathways deregulated in MM and discuss the therapeutic potential of targeting proteins or protein complexes involved in cell cycle control in MM.

Investigating the usefulness of a cluster-based trend analysis to detect visual field progression in patients with open-angle glaucoma.

PubMed

Aoki, Shuichiro; Murata, Hiroshi; Fujino, Yuri; Matsuura, Masato; Miki, Atsuya; Tanito, Masaki; Mizoue, Shiro; Mori, Kazuhiko; Suzuki, Katsuyoshi; Yamashita, Takehiro; Kashiwagi, Kenji; Hirasawa, Kazunori; Shoji, Nobuyuki; Asaoka, Ryo

2017-12-01

To investigate the usefulness of the Octopus (Haag-Streit) EyeSuite's cluster trend analysis in glaucoma. Ten visual fields (VFs) with the Humphrey Field Analyzer (Carl Zeiss Meditec), spanning 7.7 years on average were obtained from 728 eyes of 475 primary open angle glaucoma patients. Mean total deviation (mTD) trend analysis and EyeSuite's cluster trend analysis were performed on various series of VFs (from 1st to 10th: VF1-10 to 6th to 10th: VF6-10). The results of the cluster-based trend analysis, based on different lengths of VF series, were compared against mTD trend analysis. Cluster-based trend analysis and mTD trend analysis results were significantly associated in all clusters and with all lengths of VF series. Between 21.2% and 45.9% (depending on VF series length and location) of clusters were deemed to progress when the mTD trend analysis suggested no progression. On the other hand, 4.8% of eyes were observed to progress using the mTD trend analysis when cluster trend analysis suggested no progression in any two (or more) clusters. Whole field trend analysis can miss local VF progression. Cluster trend analysis appears as robust as mTD trend analysis and useful to assess both sectorial and whole field progression. Cluster-based trend analyses, in particular the definition of two or more progressing cluster, may help clinicians to detect glaucomatous progression in a timelier manner than using a whole field trend analysis, without significantly compromising specificity. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

GSTM3 and GSTP1: novel players driving tumor progression in cervical cancer.

PubMed

Checa-Rojas, Alberto; Delgadillo-Silva, Luis Fernando; Velasco-Herrera, Martín Del Castillo; Andrade-Domínguez, Andrés; Gil, Jeovanis; Santillán, Orlando; Lozano, Luis; Toledo-Leyva, Alfredo; Ramírez-Torres, Alberto; Talamas-Rohana, Patricia; Encarnación-Guevara, Sergio

2018-04-24

The molecular processes and proteomic markers leading to tumor progression (TP) in cervical cancer (CC) are either unknown or only partially understood. TP affects metabolic and regulatory mechanisms that can be identified as proteomic changes. To identify which proteins are differentially expressed and to understand the mechanisms of cancer progression, we analyzed the dynamics of the tumor proteome in CC cell lines. This analysis revealed two proteins that are up-regulated during TP, GSTM3 and GSTP1. These proteins are involved in cell maintenance, cell survival and the cellular stress response via the NF-κB and MAP kinase pathways during TP. Furthermore, GSTM3 and GSTP1 knockdown showed that evasion of apoptosis was affected, and tumor proliferation was significantly reduced. Our data indicate the critical role of GST proteins in the regulation and progression of cervical cancer cells. Hence, we suggest GSTM3 and GSTP1 as novel biomarkers and potential therapeutic targets for treating cervical cancer. CC is particularly hazardous in the advanced stages, and there are few therapeutic strategies specifically targeting these stages. We performed analyses on CC tumor proteome dynamics and identified GSTM3 and GSTP1 as novel potential therapeutic targets. Knockdown of these proteins showed that they are involved in cell survival, cell proliferation and cellular evasion of apoptosis.

Proteomic Analysis of Cerebrospinal Fluid in a Fulminant Case of Multiple Sclerosis

PubMed Central

Füvesi, Judit; Hanrieder, Jörg; Bencsik, Krisztina; Rajda, Cecilia; Kovács, S. Krisztián; Kaizer, László; Beniczky, Sándor; Vécsei, László; Bergquist, Jonas

2012-01-01

Multiple Sclerosis (MS) is a chronic disease, but in rare fulminant cases rapid progression may lead to death shortly after diagnosis. Currently there is no diagnostic test to predict disease course. The aim of this study was to identify potential biomarkers/proteins related to rapid progression. We present the case history of a 15-year-old male MS patient. Cerebrospinal fluid (CSF) was taken at diagnosis and at the time of rapid progression leading to the patient’s death. Using isobaric tag labeling and nanoflow liquid chromatography in conjunction with matrix assisted laser desorption/ionization time of flight tandem mass spectrometry we quantitatively analyzed the protein content of two CSF samples from the patient with fulminant MS as well as one relapsing-remitting (RR) MS patient and one control headache patient, whose CSF analysis was normal. Seventy-eight proteins were identified and seven proteins were found to be more abundant in both fulminant MS samples but not in the RR MS sample compared to the control. These proteins are involved in the immune response, blood coagulation, cell proliferation and cell adhesion. In conclusion, in this pilot study we were able to show differences in the CSF proteome of a rapidly progressing MS patient compared to a more typical clinical form of MS and a control subject. PMID:22837721

Evaluation of task shifting in community-based DOTS program as an effective control strategy for tuberculosis.

PubMed

Gabriel, André P; Mercado, Charles P

2011-01-01

Tuberculosis (TB) remains to be the most prevalent and debilitating pulmonary (PTB) infection in the world today, affecting about one-third of the world's population. TB is an endemic disease in many developing countries, and efforts at eliminating the disease remain futile. While the course of the disease is indolent with years of latency, the reactivation of the disease can pose serious pulmonary and systemic infections that compromise multiple organ functions which lead to respiratory failure or end-organ damage. Despite attempts to control and eradicate the mycobacterium, the prevalence of the disease remains high due to increasing population rate, persistence of poverty and poor health care, treatment failure, increasing multidrug resistance as a consequence of treatment failure and poor compliance, and existence of comorbid conditions that compromise immune response. Limited government resources to screen and monitor disease progression of TB in third world countries hamper the eradication of the disease. In response, we have evaluated the efficiency and effectivity of a Community-Based Directly Observed Treatment, Short-Course (CB-DOTS), which is an equally effective alternative strategy to health center DOTS.

The Safety Dance: Biophysics of Membrane Protein Folding and Misfolding in a Cellular Context

PubMed Central

Schlebach, Jonathan P.; Sanders, Charles R.

2015-01-01

Most biological processes require the production and degradation of proteins, a task that weighs heavily on the cell. Mutations that compromise the conformational stability of proteins place both specific and general burdens on cellular protein homeostasis (proteostasis) in ways that contribute to numerous diseases. Efforts to elucidate the chain of molecular events responsible for diseases of protein folding address one of the foremost challenges in biomedical science. However, relatively little is known about the processes by which mutations prompt the misfolding of α-helical membrane proteins, which rely on an intricate network of cellular machinery to acquire and maintain their functional structures within cellular membranes. In this review, we summarize the current understanding of the physical principles that guide membrane protein biogenesis and folding in the context of mammalian cells. Additionally, we explore how pathogenic mutations that influence biogenesis may differ from those that disrupt folding and assembly, as well as how this may relate to disease mechanisms and therapeutic intervention. These perspectives indicate an imperative for the use of information from structural, cellular, and biochemical studies of membrane proteins in the design of novel therapeutics and in personalized medicine. PMID:25420508

Foxi3 deficiency compromises hair follicle stem cell specification and activation

PubMed Central

Shirokova, Vera; Biggs, Leah C.; Jussila, Maria; Ohyama, Takahiro; Groves, Andrew K.; Mikkola, Marja L.

2017-01-01

The hair follicle is an ideal system to study stem cell specification and homeostasis due to its well characterized morphogenesis and stereotypic cycles of stem cell activation upon each hair cycle to produce a new hair shaft. The adult hair follicle stem cell niche consists of two distinct populations, the bulge and the more activation-prone secondary hair germ. Hair follicle stem cells are set aside during early stages of morphogenesis. This process is known to depend on the Sox9 transcription factor, but otherwise the establishment of the hair follicle stem cell niche is poorly understood. Here we show that that mutation of Foxi3, a Forkhead family transcription factor mutated in several hairless dog breeds, compromises stem cell specification. Further, loss of Foxi3 impedes hair follicle downgrowth and progression of the hair cycle. Genome-wide profiling revealed a number of downstream effectors of Foxi3 including transcription factors with a recognized function in hair follicle stem cells such as Lhx2, Runx1, and Nfatc1, suggesting that the Foxi3 mutant phenotype results from simultaneous downregulation of several stem cell signature genes. We show that Foxi3 displays a highly dynamic expression pattern during hair morphogenesis and cycling, and identify Foxi3 as a novel secondary hair germ marker. Absence of Foxi3 results in poor hair regeneration upon hair plucking, and a sparse fur phenotype in unperturbed mice that exacerbates with age, caused by impaired secondary hair germ activation leading to progressive depletion of stem cells. Thus, Foxi3 regulates multiple aspects of hair follicle development and homeostasis. PMID:26992132

MORC Proteins: Novel Players in Plant and Animal Health

PubMed Central

Koch, Aline; Kang, Hong-Gu; Steinbrenner, Jens; Dempsey, D'Maris A.; Klessig, Daniel F.; Kogel, Karl-Heinz

2017-01-01

Microrchidia (MORC) proteins comprise a family of proteins that have been identified in prokaryotes and eukaryotes. They are defined by two hallmark domains: a GHKL-type ATPase and an S5 fold. MORC proteins in plants were first discovered via a genetic screen for Arabidopsis mutants compromised for resistance to a viral pathogen. Subsequent studies expanded their role in plant immunity and revealed their involvement in gene silencing and transposable element repression. Emerging data suggest that MORC proteins also participate in pathogen-induced chromatin remodeling and epigenetic gene regulation. In addition, biochemical analyses recently demonstrated that plant MORCs have topoisomerase II (topo II)-like DNA modifying activities that may be important for their function. Interestingly, animal MORC proteins exhibit many parallels with their plant counterparts, as they have been implicated in disease development and gene silencing. In addition, human MORCs, like plant MORCs, bind salicylic acid and this inhibits some of their topo II-like activities. In this review, we will focus primarily on plant MORCs, although relevant comparisons with animal MORCs will be provided. PMID:29093720

MORC Proteins: Novel Players in Plant and Animal Health.

PubMed

Koch, Aline; Kang, Hong-Gu; Steinbrenner, Jens; Dempsey, D'Maris A; Klessig, Daniel F; Kogel, Karl-Heinz

2017-01-01

Microrchidia (MORC) proteins comprise a family of proteins that have been identified in prokaryotes and eukaryotes. They are defined by two hallmark domains: a GHKL-type ATPase and an S5 fold. MORC proteins in plants were first discovered via a genetic screen for Arabidopsis mutants compromised for resistance to a viral pathogen. Subsequent studies expanded their role in plant immunity and revealed their involvement in gene silencing and transposable element repression. Emerging data suggest that MORC proteins also participate in pathogen-induced chromatin remodeling and epigenetic gene regulation. In addition, biochemical analyses recently demonstrated that plant MORCs have topoisomerase II (topo II)-like DNA modifying activities that may be important for their function. Interestingly, animal MORC proteins exhibit many parallels with their plant counterparts, as they have been implicated in disease development and gene silencing. In addition, human MORCs, like plant MORCs, bind salicylic acid and this inhibits some of their topo II-like activities. In this review, we will focus primarily on plant MORCs, although relevant comparisons with animal MORCs will be provided.

Collapse of proteostasis represents an early molecular event in Caenorhabditis elegans aging.

PubMed

Ben-Zvi, Anat; Miller, Elizabeth A; Morimoto, Richard I

2009-09-01

Protein damage contributes prominently to cellular aging. To address whether this occurs at a specific period during aging or accumulates gradually, we monitored the biochemical, cellular, and physiological properties of folding sensors expressed in different tissues of C. elegans. We observed the age-dependent misfolding and loss of function of diverse proteins harboring temperature-sensitive missense mutations in all somatic tissues at the permissive condition. This widespread failure in proteostasis occurs rapidly at an early stage of adulthood, and coincides with a severely reduced activation of the cytoprotective heat shock response and the unfolded protein response. Enhancing stress responsive factors HSF-1 or DAF-16 suppresses misfolding of these metastable folding sensors and restores the ability of the cell to maintain a functional proteome. This suggests that a compromise in the regulation of proteostatic stress responses occurs early in adulthood and tips the balance between the load of damaged proteins and the proteostasis machinery. We propose that the collapse of proteostasis represents an early molecular event of aging that amplifies protein damage in age-associated diseases of protein conformation.

Tobacco BY-2 Media Component Optimization for a Cost-Efficient Recombinant Protein Production.

PubMed

Häkkinen, Suvi T; Reuter, Lauri; Nuorti, Ninni; Joensuu, Jussi J; Rischer, Heiko; Ritala, Anneli

2018-01-01

Plant cells constitute an attractive platform for production of recombinant proteins as more and more animal-free products and processes are desired. One of the challenges in using plant cells as production hosts has been the costs deriving from expensive culture medium components. In this work, the aim was to optimize the levels of most expensive components in the nutrient medium without compromising the accumulation of biomass and recombinant protein yields. Wild-type BY-2 culture and transgenic tobacco BY-2 expressing green fluorescent protein-Hydrophobin I (GFP-HFBI) fusion protein were used to determine the most inexpensive medium composition. One particularly high-accumulating BY-2 clone, named 'Hulk,' produced 1.1 ± 0.2 g/l GFP-HFBI in suspension and kept its high performance during prolonged subculturing. In addition, both cultures were successfully cryopreserved enabling truly industrial application of this plant cell host. With the optimized culture medium, 43-55% cost reduction with regard to biomass and up to 69% reduction with regard to recombinant protein production was achieved.

Protein misfolding specifies recruitment to cytoplasmic inclusion bodies.

PubMed

Bersuker, Kirill; Brandeis, Michael; Kopito, Ron R

2016-04-25

Inclusion bodies (IBs) containing aggregated disease-associated proteins and polyubiquitin (poly-Ub) conjugates are universal histopathological features of neurodegenerative diseases. Ub has been proposed to target proteins to IBs for degradation via autophagy, but the mechanisms that govern recruitment of ubiquitylated proteins to IBs are not well understood. In this paper, we use conditionally destabilized reporters that undergo misfolding and ubiquitylation upon removal of a stabilizing ligand to examine the role of Ub conjugation in targeting proteins to IBs that are composed of an N-terminal fragment of mutant huntingtin, the causative protein of Huntington's disease. We show that reporters are excluded from IBs in the presence of the stabilizing ligand but are recruited to IBs after ligand washout. However, we find that Ub conjugation is not necessary to target reporters to IBs. We also report that forced Ub conjugation by the Ub fusion degradation pathway is not sufficient for recruitment to IBs. Finally, we find that reporters and Ub conjugates are stable at IBs. These data indicate that compromised folding states, rather than conjugation to Ub, can specify recruitment to IBs. © 2016 Bersuker et al.

Sequence variation and structural conservation allows development of novel function and immune evasion in parasite surface protein families

PubMed Central

Higgins, Matthew K; Carrington, Mark

2014-01-01

Trypanosoma and Plasmodium species are unicellular, eukaryotic pathogens that have evolved the capacity to survive and proliferate within a human host, causing sleeping sickness and malaria, respectively. They have very different survival strategies. African trypanosomes divide in blood and extracellular spaces, whereas Plasmodium species invade and proliferate within host cells. Interaction with host macromolecules is central to establishment and maintenance of an infection by both parasites. Proteins that mediate these interactions are under selection pressure to bind host ligands without compromising immune avoidance strategies. In both parasites, the expansion of genes encoding a small number of protein folds has established large protein families. This has permitted both diversification to form novel ligand binding sites and variation in sequence that contributes to avoidance of immune recognition. In this review we consider two such parasite surface protein families, one from each species. In each case, known structures demonstrate how extensive sequence variation around a conserved molecular architecture provides an adaptable protein scaffold that the parasites can mobilise to mediate interactions with their hosts. PMID:24442723

General overview on structure prediction of twilight-zone proteins.

PubMed

Khor, Bee Yin; Tye, Gee Jun; Lim, Theam Soon; Choong, Yee Siew

2015-09-04

Protein structure prediction from amino acid sequence has been one of the most challenging aspects in computational structural biology despite significant progress in recent years showed by critical assessment of protein structure prediction (CASP) experiments. When experimentally determined structures are unavailable, the predictive structures may serve as starting points to study a protein. If the target protein consists of homologous region, high-resolution (typically <1.5 Å) model can be built via comparative modelling. However, when confronted with low sequence similarity of the target protein (also known as twilight-zone protein, sequence identity with available templates is less than 30%), the protein structure prediction has to be initiated from scratch. Traditionally, twilight-zone proteins can be predicted via threading or ab initio method. Based on the current trend, combination of different methods brings an improved success in the prediction of twilight-zone proteins. In this mini review, the methods, progresses and challenges for the prediction of twilight-zone proteins were discussed.

Connective tissue growth factor (CTGF) and cancer progression.

PubMed

Chu, Chia-Yu; Chang, Cheng-Chi; Prakash, Ekambaranellore; Kuo, Min-Liang

2008-11-01

Connective tissue growth factor (CTGF) is a member of the CCN family of secreted, matrix-associated proteins encoded by immediate early genes that play various roles in angiogenesis and tumor growth. CCN family proteins share uniform modular structure which mediates various cellular functions such as regulation of cell division, chemotaxis, apoptosis, adhesion, motility, angiogenesis, neoplastic transformation, and ion transport. Recently, CTGF expression has been shown to be associated with tumor development and progression. There is growing body of evidence that CTGF may regulate cancer cell migration, invasion, angiogenesis, and anoikis. In this review, we will highlight the influence of CTGF expression on the biological behavior and progression of various cancer cells, as well as its regulation on various types of protein signals and their mechanisms.

Association of protein kinase FA/GSK-3alpha (a proline-directed kinase and a regulator of protooncogenes) with human cervical carcinoma dedifferentiation/progression.

PubMed

Yang, S D; Yu, J S; Lee, T T; Ni, M H; Yang, C C; Ho, Y S; Tsen, T Z

1995-10-01

Computer analysis of protein phosphorylation-sites sequence revealed that most transcriptional factors and viral oncoproteins are prime targets for regulation of proline-directed protein phosphorylation, suggesting an association of proline-directed protein kinase (PDPK) family with neoplastic transformation and tumorigenesis. In this report, an immunoprecipitate activity assay of protein kinase FA/glycogen synthase kinase-3alpha (kinase FA/GSK-3alpha) (a particular member of PDPK family) has been optimized for human cervical tissue and used to demonstrate for the first time significantly increased (P < 0.001) activity in poorly differentiated cervical carcinoma (82.8 +/- 6.6 U/mg of protein), moderately differentiated carcinoma (36.2 +/- 3.4 U/mg of protein), and well-differentiated carcinoma (18.3 +/- 2.4 U/mg of protein) from 36 human cervical carcinoma samples when compared to 12 normal controls (4.9 +/- 0.6 U/mg of protein). Immunoblotting analysis further revealed that increased activity of kinase FA/GSK-3alpha in cervical carcinoma is due to overexpression of protein synthesis of the kinase. Taken together, the results provide initial evidence that overexpression of protein synthesis and cellular activity of kinase FA/GSK-3alpha may be involved in human cervical carcinoma dedifferentiation/progression, supporting an association of proline-directed protein kinase with neoplastic transformation and tumorigenesis. Since protein kinase FA/GSK-3alpha may function as a possible regulator of transcription factors/proto-oncogenes, the results further suggest that kinase FA/GSK-3alpha may play a potential role in human cervical carcinogenesis, especially in its dedifferentiation and progression.

Are the Gut Bacteria Telling Us to Eat or Not to Eat? Reviewing the Role of Gut Microbiota in the Etiology, Disease Progression and Treatment of Eating Disorders

PubMed Central

Lam, Yan Y.; Maguire, Sarah; Palacios, Talia; Caterson, Ian D.

2017-01-01

Traditionally recognized as mental illnesses, eating disorders are increasingly appreciated to be biologically-driven. There is a growing body of literature that implicates a role of the gut microbiota in the etiology and progression of these conditions. Gut bacteria may act on the gut–brain axis to alter appetite control and brain function as part of the genesis of eating disorders. As the illnesses progress, extreme feeding patterns and psychological stress potentially feed back to the gut ecosystem that can further compromise physiological, cognitive, and social functioning. Given the established causality between dysbiosis and metabolic diseases, an altered gut microbial profile is likely to play a role in the co-morbidities of eating disorders with altered immune function, short-chain fatty acid production, and the gut barrier being the key mechanistic links. Understanding the role of the gut ecosystem in the pathophysiology of eating disorders will provide critical insights into improving current treatments and developing novel microbiome-based interventions that will benefit patients with eating disorders. PMID:28613252

Rod Migration Into the Spinal Canal After Posterior Instrumented Fusion Causing Late-Onset Neurological Symptoms.

PubMed

Canavese, Federico; Dmitriev, Petru; Deslandes, Jacques; Samba, Antoine; Dimeglio, Alain; Mansour, Mounira; Rousset, Marie; Dubousset, Jean

2017-01-01

Rod migration into the spinal canal after posterior instrumented fusion is a rare complication causing late-onset neurological symptoms. The purpose of the present study is to report a case of a 13-year-old boy with spastic cerebral palsy and related neuromuscular kyphoscoliosis who developed late-onset neurological deterioration secondary to progressive implant migration into the spinal canal over a 5-year period. A decision was made to remove both rods to achieve decompression. Intraoperative findings were consistent with information gained from preoperative imaging. The rods were found to have an intracanal trajectory at T9-T10 for the right rod and T12-L2 for the left rod. The cause of implant migration, with progressive laminar erosion slow enough to generate a solid mass behind, was progressive kyphosis in a skeletally immature patient with neuromuscular compromise. Fixation type, early surgery, and spasticity management contributed significantly to the presenting condition. Mechanical factors and timing of surgery played a decisive role in this particular presentation. Level IV--Case report and review of the literature.

Are the Gut Bacteria Telling Us to Eat or Not to Eat? Reviewing the Role of Gut Microbiota in the Etiology, Disease Progression and Treatment of Eating Disorders.

PubMed

Lam, Yan Y; Maguire, Sarah; Palacios, Talia; Caterson, Ian D

2017-06-14

Traditionally recognized as mental illnesses, eating disorders are increasingly appreciated to be biologically-driven. There is a growing body of literature that implicates a role of the gut microbiota in the etiology and progression of these conditions. Gut bacteria may act on the gut-brain axis to alter appetite control and brain function as part of the genesis of eating disorders. As the illnesses progress, extreme feeding patterns and psychological stress potentially feed back to the gut ecosystem that can further compromise physiological, cognitive, and social functioning. Given the established causality between dysbiosis and metabolic diseases, an altered gut microbial profile is likely to play a role in the co-morbidities of eating disorders with altered immune function, short-chain fatty acid production, and the gut barrier being the key mechanistic links. Understanding the role of the gut ecosystem in the pathophysiology of eating disorders will provide critical insights into improving current treatments and developing novel microbiome-based interventions that will benefit patients with eating disorders.

Uterine Deletion of Trp53 Compromises Antioxidant Responses in the Mouse Decidua

PubMed Central

Burnum, Kristin E.; Hirota, Yasushi; Baker, Erin S.; Yoshie, Mikihiro; Ibrahim, Yehia M.; Monroe, Matthew E.; Anderson, Gordon A.; Smith, Richard D.; Daikoku, Takiko

2012-01-01

Preterm birth is a global health issue impacting millions of mothers and babies. However, the etiology of preterm birth is not clearly understood. Our recent finding that premature decidual senescence with terminal differentiation is a cause of preterm birth in mice with uterine Trp53 deletion, encoding p53 protein, led us to explore other potential factors that are related to preterm birth. Using proteomics approaches, here, we show that 183 candidate proteins show significant changes in deciduae with Trp53 deletion as compared with normal deciduae. Functional categorization of these proteins unveiled new pathways that are influenced by p53. In particular, down-regulation of a cluster of antioxidant enzymes in p53-deficient deciduae suggests that increased oxidative stress could be one cause of preterm birth in mice harboring uterine deletion of Trp53. PMID:22759378

Designing proteins for therapeutic applications.

PubMed

Lazar, Greg A; Marshall, Shannon A; Plecs, Joseph J; Mayo, Stephen L; Desjarlais, John R

2003-08-01

Protein design is becoming an increasingly useful tool for optimizing protein drugs and creating novel biotherapeutics. Recent progress includes the engineering of monoclonal antibodies, cytokines, enzymes and viral fusion inhibitors.

A Nonsense Mutation in TMEM95 Encoding a Nondescript Transmembrane Protein Causes Idiopathic Male Subfertility in Cattle

PubMed Central

Pausch, Hubert; Kölle, Sabine; Wurmser, Christine; Schwarzenbacher, Hermann; Emmerling, Reiner; Jansen, Sandra; Trottmann, Matthias; Fuerst, Christian; Götz, Kay-Uwe; Fries, Ruedi

2014-01-01

Genetic variants underlying reduced male reproductive performance have been identified in humans and model organisms, most of them compromising semen quality. Occasionally, male fertility is severely compromised although semen analysis remains without any apparent pathological findings (i.e., idiopathic subfertility). Artificial insemination (AI) in most cattle populations requires close examination of all ejaculates before insemination. Although anomalous ejaculates are rejected, insemination success varies considerably among AI bulls. In an attempt to identify genetic causes of such variation, we undertook a genome-wide association study (GWAS). Imputed genotypes of 652,856 SNPs were available for 7962 AI bulls of the Fleckvieh (FV) population. Male reproductive ability (MRA) was assessed based on 15.3 million artificial inseminations. The GWAS uncovered a strong association signal on bovine chromosome 19 (P = 4.08×10−59). Subsequent autozygosity mapping revealed a common 1386 kb segment of extended homozygosity in 40 bulls with exceptionally poor reproductive performance. Only 1.7% of 35,671 inseminations with semen samples of those bulls were successful. None of the bulls with normal reproductive performance was homozygous, indicating recessive inheritance. Exploiting whole-genome re-sequencing data of 43 animals revealed a candidate causal nonsense mutation (rs378652941, c.483C>A, p.Cys161X) in the transmembrane protein 95 encoding gene TMEM95 which was subsequently validated in 1990 AI bulls. Immunohistochemical investigations evidenced that TMEM95 is located at the surface of spermatozoa of fertile animals whereas it is absent in spermatozoa of subfertile animals. These findings imply that integrity of TMEM95 is required for an undisturbed fertilisation. Our results demonstrate that deficiency of TMEM95 severely compromises male reproductive performance in cattle and reveal for the first time a phenotypic effect associated with genomic variation in TMEM95. PMID:24391514

A Novel indole compound that inhibits Pseudomonas aeruginosa growth by targeting MreB is a substrate for MexAB-OprM.

PubMed

Robertson, Gregory T; Doyle, Timothy B; Du, Qun; Duncan, Leonard; Mdluli, Khisimuzi E; Lynch, A Simon

2007-10-01

Drug efflux systems contribute to the intrinsic resistance of Pseudomonas aeruginosa to many antibiotics and biocides and hamper research focused on the discovery and development of new antimicrobial agents targeted against this important opportunistic pathogen. Using a P. aeruginosa PAO1 derivative bearing deletions of opmH, encoding an outer membrane channel for efflux substrates, and four efflux pumps belonging to the resistance nodulation/cell division class including mexAB-oprM, we identified a small-molecule indole-class compound (CBR-4830) that is inhibitory to growth of this efflux-compromised strain. Genetic studies established MexAB-OprM as the principal pump for CBR-4830 and revealed MreB, a prokaryotic actin homolog, as the proximal cellular target of CBR-4830. Additional studies establish MreB as an essential protein in P. aeruginosa, and efflux-compromised strains treated with CBR-4830 transition to coccoid shape, consistent with MreB inhibition or depletion. Resistance genetics further suggest that CBR-4830 interacts with the putative ATP-binding pocket in MreB and demonstrate significant cross-resistance with A22, a structurally unrelated compound that has been shown to promote rapid dispersion of MreB filaments in vivo. Interestingly, however, ATP-dependent polymerization of purified recombinant P. aeruginosa MreB is blocked in vitro in a dose-dependent manner by CBR-4830 but not by A22. Neither compound exhibits significant inhibitory activity against mutant forms of MreB protein that bear mutations identified in CBR-4830-resistant strains. Finally, employing the strains and reagents prepared and characterized during the course of these studies, we have begun to investigate the ability of analogues of CBR-4830 to inhibit the growth of both efflux-proficient and efflux-compromised P. aeruginosa through specific inhibition of MreB function.

In female rat heart mitochondria, oophorectomy results in loss of oxidative phosphorylation.

PubMed

Pavón, Natalia; Cabrera-Orefice, Alfredo; Gallardo-Pérez, Juan Carlos; Uribe-Alvarez, Cristina; Rivero-Segura, Nadia A; Vazquez-Martínez, Edgar Ricardo; Cerbón, Marco; Martínez-Abundis, Eduardo; Torres-Narvaez, Juan Carlos; Martínez-Memije, Raúl; Roldán-Gómez, Francisco-Javier; Uribe-Carvajal, Salvador

2017-02-01

Oophorectomy in adult rats affected cardiac mitochondrial function. Progression of mitochondrial alterations was assessed at one, two and three months after surgery: at one month, very slight changes were observed, which increased at two and three months. Gradual effects included decrease in the rates of oxygen consumption and in respiratory uncoupling in the presence of complex I substrates, as well as compromised Ca 2+ buffering ability. Malondialdehyde concentration increased, whereas the ROS-detoxifying enzyme Mn 2+ superoxide dismutase (MnSOD) and aconitase lost activity. In the mitochondrial respiratory chain, the concentration and activity of complex I and complex IV decreased. Among other mitochondrial enzymes and transporters, adenine nucleotide carrier and glutaminase decreased. 2-Oxoglutarate dehydrogenase and pyruvate dehydrogenase also decreased. Data strongly suggest that in the female rat heart, estrogen depletion leads to progressive, severe mitochondrial dysfunction. © 2017 Society for Endocrinology.

Right ventricular exclusion and univentricular palliation for failed one and a half ventricle repair for Ebstein's anomaly.

PubMed

Sasikumar, Navaneetha; Krishna Manohar, Soman R; Philip, Saji; Cherian, Kottoorathu Mammen; Suresh Kumar, Raghavannair

2013-08-01

A 20 year-old male was diagnosed to have Ebstein's anomaly with severe right ventricular dysfunction. He was taken up for 1.5 ventricle repair. Post procedure, there was difficulty in weaning from cardiopulmonary bypass due to progressive right ventricular dilatation compromising the systemic output. An atrial septectomy did not help. Progressive right ventricular dilatation compressing the left ventricle, demonstrated on transoesophageal echocardiogram, prompted us to perform a right ventricular exclusion and univentricular palliation. The patient was successfully weaned off cardiopulmonary bypass and had a smooth postoperative recovery. Judicious use of right ventricular exclusion and univentricular palliation could be an effective bailout strategy in difficult surgical scenarios in Ebstein's anomaly. Copyright © 2012 Australian and New Zealand Society of Cardiac and Thoracic Surgeons (ANZSCTS) and the Cardiac Society of Australia and New Zealand (CSANZ). Published by Elsevier B.V. All rights reserved.

Targeting cancer’s weaknesses (not its strengths): Therapeutic strategies suggested by the atavistic model

PubMed Central

Lineweaver, Charles H.; Davies, Paul C.W.; Vincent, Mark D.

2014-01-01

In the atavistic model of cancer progression, tumor cell dedifferentiation is interpreted as a reversion to phylogenetically earlier capabilities. The more recently evolved capabilities are compromised first during cancer progression. This suggests a therapeutic strategy for targeting cancer: design challenges to cancer that can only be met by the recently evolved capabilities no longer functional in cancer cells. We describe several examples of this target-the-weakness strategy. Our most detailed example involves the immune system. The absence of adaptive immunity in immunosuppressed tumor environments is an irreversible weakness of cancer that can be exploited by creating a challenge that only the presence of adaptive immunity can meet. This leaves tumor cells more vulnerable than healthy tissue to pathogenic attack. Such a target-the-weakness therapeutic strategy has broad applications, and contrasts with current therapies that target the main strength of cancer: cell proliferation. PMID:25043755

Dry Eye Management: Targeting the Ocular Surface Microenvironment.

PubMed

Zhang, Xiaobo; M, Vimalin Jeyalatha; Qu, Yangluowa; He, Xin; Ou, Shangkun; Bu, Jinghua; Jia, Changkai; Wang, Junqi; Wu, Han; Liu, Zuguo; Li, Wei

2017-06-29

Dry eye can damage the ocular surface and result in mild corneal epithelial defect to blinding corneal pannus formation and squamous metaplasia. Significant progress in the treatment of dry eye has been made in the last two decades; progressing from lubricating and hydrating the ocular surface with artificial tear to stimulating tear secretion; anti-inflammation and immune regulation. With the increase in knowledge regarding the pathophysiology of dry eye, we propose in this review the concept of ocular surface microenvironment. Various components of the microenvironment contribute to the homeostasis of ocular surface. Compromise in one or more components can result in homeostasis disruption of ocular surface leading to dry eye disease. Complete evaluation of the microenvironment component changes in dry eye patients will not only lead to appropriate diagnosis, but also guide in timely and effective clinical management. Successful treatment of dry eye should be aimed to restore the homeostasis of the ocular surface microenvironment.

Clinical Manifestations and Overall Management Strategies for Duchenne Muscular Dystrophy.

PubMed

Tsuda, Takeshi

2018-01-01

Duchenne muscular dystrophy (DMD) is an X-linked genetic disorder that causes progressive weakness and wasting of skeletal muscular and myocardium in boys due to mutation of dystrophin. The structural integrity of each individual skeletal and cardiac myocyte is significantly compromised upon physical stress due to the absence of dystrophin. The progressive destruction of systemic musculature and myocardium causes affected patients to develop multiple organ disabilities, including loss of ambulation, physical immobility, neuromuscular scoliosis, joint contracture, restrictive lung disease, obstructive sleep apnea, and cardiomyopathy. There are some central nervous system-related medical problems, as dystrophin is also expressed in the neuronal tissues. Although principal management is to mainly delay the pathological process, an enhanced understanding of underlying pathological processes has significantly improved quality of life and longevity for DMD patients. Future research in novel molecular approach is warranted to answer unanswered questions.

Progress Toward Poliomyelitis Eradication - Afghanistan, January 2016-June 2017.

PubMed

Martinez, Maureen; Shukla, Hemant; Nikulin, Joanna; Wadood, Mufti Zubair; Hadler, Stephen; Mbaeyi, Chukwuma; Tangermann, Rudolph; Jorba, Jaume; Ehrhardt, Derek

2017-08-18

Afghanistan, Pakistan, and Nigeria remain the only countries where the transmission of endemic wild poliovirus type 1 (WPV1) continues (1). This report describes polio eradication activities, progress, and challenges in Afghanistan during January 2016-June 2017 and updates previous reports (2,3). Thirteen WPV1 cases were confirmed in Afghanistan in 2016, a decrease of seven from the 20 cases reported in 2015. From January to June 2017, five WPV1 cases were reported, compared with six during the same period in 2016. The number of affected districts declined from 23 (including WPV1-positive acute flaccid paralysis [AFP] cases and positive environmental sewage samples) in 2015 to six in 2016. To achieve WPV1 eradication, it is important that Afghanistan's polio program continue to collaborate with that of neighboring Pakistan to track and vaccinate groups of high-risk mobile populations and strengthen efforts to reach children in security-compromised areas.

Dry Eye Management: Targeting the Ocular Surface Microenvironment

PubMed Central

Zhang, Xiaobo; Jeyalatha M, Vimalin; Qu, Yangluowa; He, Xin; Ou, Shangkun; Bu, Jinghua; Jia, Changkai; Wang, Junqi; Wu, Han; Liu, Zuguo

2017-01-01

Dry eye can damage the ocular surface and result in mild corneal epithelial defect to blinding corneal pannus formation and squamous metaplasia. Significant progress in the treatment of dry eye has been made in the last two decades; progressing from lubricating and hydrating the ocular surface with artificial tear to stimulating tear secretion; anti-inflammation and immune regulation. With the increase in knowledge regarding the pathophysiology of dry eye, we propose in this review the concept of ocular surface microenvironment. Various components of the microenvironment contribute to the homeostasis of ocular surface. Compromise in one or more components can result in homeostasis disruption of ocular surface leading to dry eye disease. Complete evaluation of the microenvironment component changes in dry eye patients will not only lead to appropriate diagnosis, but also guide in timely and effective clinical management. Successful treatment of dry eye should be aimed to restore the homeostasis of the ocular surface microenvironment. PMID:28661456

The anti-tumor activity of a neutralizing nanobody targeting leptin receptor in a mouse model of melanoma.

PubMed

McMurphy, Travis; Xiao, Run; Magee, Daniel; Slater, Andrew; Zabeau, Lennart; Tavernier, Jan; Cao, Lei

2014-01-01

Environmental and genetic activation of a brain-adipocyte axis inhibits cancer progression. Leptin is the primary peripheral mediator of this anticancer effect in a mouse model of melanoma. In this study we assessed the effect of a leptin receptor antagonist on melanoma progression. Local administration of a neutralizing nanobody targeting the leptin receptor at low dose adjacent to tumor decreased tumor mass with no effects on body weight or food intake. In contrast, systemic administration of the nanobody failed to suppress tumor growth. Daily intraperitoneal injection of high-dose nanobody led to weight gain, hyperphagia, increased adiposity, hyperleptinemia, and hyperinsulinemia, and central effects mimicking leptin deficiency. The blockade of central actions of leptin by systemic delivery of nanobody may compromise its anticancer effect, underscoring the need to develop peripherally acting leptin antagonists coupled with efficient cancer-targeting delivery.

The Light and Shadow of Senescence and Inflammation in Cardiovascular Pathology and Regenerative Medicine

PubMed Central

Dal Sasso, Eleonora; Schirone, Leonardo; Forte, Maurizio; Palmerio, Silvia; Gerosa, Gino; Sciarretta, Sebastiano

2017-01-01

Recent epidemiologic studies evidence a dramatic increase of cardiovascular diseases, especially associated with the aging of the world population. During aging, the progressive impairment of the cardiovascular functions results from the compromised tissue abilities to protect the heart against stress. At the molecular level, in fact, a gradual weakening of the cellular processes regulating cardiovascular homeostasis occurs in aging cells. Atherosclerosis and heart failure are particularly correlated with aging-related cardiovascular senescence, that is, the inability of cells to progress in the mitotic program until completion of cytokinesis. In this review, we explore the intrinsic and extrinsic causes of cellular senescence and their role in the onset of these cardiovascular pathologies. Additionally, we dissect the effects of aging on the cardiac endogenous and exogenous reservoirs of stem cells. Finally, we offer an overview on the strategies of regenerative medicine that have been advanced in the quest for heart rejuvenation. PMID:29118467

Measuring facial expression of emotion.

PubMed

Wolf, Karsten

2015-12-01

Research into emotions has increased in recent decades, especially on the subject of recognition of emotions. However, studies of the facial expressions of emotion were compromised by technical problems with visible video analysis and electromyography in experimental settings. These have only recently been overcome. There have been new developments in the field of automated computerized facial recognition; allowing real-time identification of facial expression in social environments. This review addresses three approaches to measuring facial expression of emotion and describes their specific contributions to understanding emotion in the healthy population and in persons with mental illness. Despite recent progress, studies on human emotions have been hindered by the lack of consensus on an emotion theory suited to examining the dynamic aspects of emotion and its expression. Studying expression of emotion in patients with mental health conditions for diagnostic and therapeutic purposes will profit from theoretical and methodological progress.

A Human-in-the Loop Exploration of the Dynamic Airspace Configuration Concept

NASA Technical Reports Server (NTRS)

Homola, Jeffrey; Lee, Paul U.; Prevot, Thomas; Lee, Hwasoo; Kessell, Angela; Brasil, Connie; Smith, Nancy

2010-01-01

An exploratory human-in-the-loop study was conducted to better understand the impact of Dynamic Airspace Configuration (DAC) on air traffic controllers. To do so, a range of three progressively more aggressive algorithmic approaches to sectorizations were chosen. Sectorizations from these algorithms were used to test and quantify the range of impact on the controller and traffic. Results show that traffic count was more equitably distributed between the four test sectors and duration of counts over MAP were progressively lower as the magnitude of boundary change increased. However, taskload and workload were also shown to increase with the increase in aggressiveness and acceptability of the boundary changes decreased. Overall, simulated operations of the DAC concept did not appear to compromise safety. Feedback from the participants highlighted the importance of limiting some aspects of boundary changes such as amount of volume gained or lost and the extent of change relative to the initial airspace design.

Heterogeneous blood-tumor barrier permeability determines drug efficacy in experimental brain metastases of breast cancer.

PubMed

Lockman, Paul R; Mittapalli, Rajendar K; Taskar, Kunal S; Rudraraju, Vinay; Gril, Brunilde; Bohn, Kaci A; Adkins, Chris E; Roberts, Amanda; Thorsheim, Helen R; Gaasch, Julie A; Huang, Suyun; Palmieri, Diane; Steeg, Patricia S; Smith, Quentin R

2010-12-01

Brain metastases of breast cancer appear to be increasing in incidence, confer significant morbidity, and threaten to compromise gains made in systemic chemotherapy. The blood-tumor barrier (BTB) is compromised in many brain metastases; however, the extent to which this influences chemotherapeutic delivery and efficacy is unknown. Herein, we answer this question by measuring BTB passive integrity, chemotherapeutic drug uptake, and anticancer efficacy in vivo in two breast cancer models that metastasize preferentially to brain. Experimental brain metastasis drug uptake and BTB permeability were simultaneously measured using novel fluorescent and phosphorescent imaging techniques in immune-compromised mice. Drug-induced apoptosis and vascular characteristics were assessed using immunofluorescent microscopy. Analysis of over 2,000 brain metastases from two models (human 231-BR-Her2 and murine 4T1-BR5) showed partial BTB permeability compromise in greater than 89% of lesions, varying in magnitude within and between metastases. Brain metastasis uptake of ¹⁴C-paclitaxel and ¹⁴C-doxorubicin was generally greater than normal brain but less than 15% of that of other tissues or peripheral metastases, and only reached cytotoxic concentrations in a small subset (∼10%) of the most permeable metastases. Neither drug significantly decreased the experimental brain metastatic ability of 231-BR-Her2 tumor cells. BTB permeability was associated with vascular remodeling and correlated with overexpression of the pericyte protein desmin. This work shows that the BTB remains a significant impediment to standard chemotherapeutic delivery and efficacy in experimental brain metastases of breast cancer. New brain permeable drugs will be needed. Evidence is presented for vascular remodeling in BTB permeability alterations. ©2010 AACR.

Heterogeneous Blood-Tumor Barrier Permeability Determines Drug Efficacy in Experimental Brain Metastases of Breast Cancer

PubMed Central

Lockman, Paul R.; Mittapalli, Rajendar K.; Taskar, Kunal S.; Rudraraju, Vinay; Gril, Brunilde; Bohn, Kaci A.; Adkins, Chris E.; Roberts, Amanda; Thorsheim, Helen R.; Gaasch, Julie A.; Huang, Suyun; Palmieri, Diane; Steeg, Patricia S.; Smith, Quentin R.

2010-01-01

Purpose Brain metastases of breast cancer appear to be increasing in incidence, confer significant morbidity, and threaten to compromise gains made in systemic chemotherapy. The blood-tumor barrier (BTB) is compromised in many brain metastases, however, the extent to which this influences chemotherapeutic delivery and efficacy is unknown. Herein, we answer this question by measuring BTB passive integrity, chemotherapeutic drug uptake, and anticancer efficacy in vivo in two breast cancer models that metastasize preferentially to brain. Experimental Design Experimental brain metastasis drug uptake and BTB permeability were simultaneously measured using novel fluorescent and phosphorescent imaging techniques in immune compromised mice. Drug-induced apoptosis and vascular characteristics were assessed using immunofluorescent microscopy. Results Analysis of >2000 brain metastases from two models (human 231-BR-Her2 and murine 4T1-BR5) demonstrated partial BTB permeability compromise in >89% lesions, varying in magnitude within and between metastases. Brain metastasis uptake of 14C- paclitaxel and 14C- doxorubicin was generally greater than normal brain but <15% of that of other tissues or peripheral metastases, and only reached cytotoxic concentrations in a small subset (~10%) of the most permeable metastases. Neither drug significantly decreased the experimental brain metastatic ability of 231-BR-Her2 tumor cells. BTB permeability was associated with vascular remodeling and correlated with over expression of the pericyte protein, desmin. Conclusions This work demonstrates that the BTB remains a significant impediment to standard chemotherapeutic delivery and efficacy in experimental brain metastases of breast cancer. New brain permeable drugs will be needed. Evidence is presented for vascular remodeling in BTB permeability alterations. PMID:20829328

Mutant analysis of Cdt1's function in suppressing nascent strand elongation during DNA replication in Xenopus egg extracts.

PubMed

Nakazaki, Yuta; Tsuyama, Takashi; Azuma, Yutaro; Takahashi, Mikiko; Tada, Shusuke

2017-09-02

The initiation of DNA replication is strictly regulated by multiple mechanisms to ensure precise duplication of chromosomes. In higher eukaryotes, activity of the Cdt1 protein is temporally regulated during the cell cycle, and deregulation of Cdt1 induces DNA re-replication. In previous studies, we showed that excess Cdt1 inhibits DNA replication by suppressing progression of replication forks in Xenopus egg extracts. Here, we investigated the functional regions of Cdt1 that are required for the inhibition of DNA replication. We constructed a series of N-terminally or C-terminally deleted mutants of Cdt1 and examined their inhibitory effects on DNA replication in Xenopus egg extracts. Our results showed that the region spanning amino acids (a. a.) 255-620 is required for efficient inhibition of DNA replication, and that, within this region, a. a. 255-289 have a critical role in inhibition. Moreover, one of the Cdt1 mutants, Cdt1 R285A, was compromised with respect to the licensing activity but still inhibited DNA replication. This result suggests that Cdt1 has an unforeseen function in the negative regulation of DNA replication, and that this function is located within a molecular region that is distinct from those required for the licensing activity. Copyright © 2017 Elsevier Inc. All rights reserved.

Membrane pore formation in atomistic and coarse-grained simulations.

PubMed

Kirsch, Sonja A; Böckmann, Rainer A

2016-10-01

Biological cells and their organelles are protected by ultra thin membranes. These membranes accomplish a broad variety of important tasks like separating the cell content from the outer environment, they are the site for cell-cell interactions and many enzymatic reactions, and control the in- and efflux of metabolites. For certain physiological functions e.g. in the fusion of membranes and also in a number of biotechnological applications like gene transfection the membrane integrity needs to be compromised to allow for instance for the exchange of polar molecules across the membrane barrier. Mechanisms enabling the transport of molecules across the membrane involve membrane proteins that form specific pores or act as transporters, but also so-called lipid pores induced by external fields, stress, or peptides. Recent progress in the simulation field enabled to closely mimic pore formation as supposed to occur in vivo or in vitro. Here, we review different simulation-based approaches in the study of membrane pores with a focus on lipid pore properties such as their size and energetics, poration mechanisms based on the application of external fields, charge imbalances, or surface tension, and on pores that are induced by small molecules, peptides, and lipids. This article is part of a Special Issue entitled: Biosimulations edited by Ilpo Vattulainen and Tomasz Róg. Copyright © 2015 Elsevier B.V. All rights reserved.

Progress towards genetic and pharmacological therapies for keratin genodermatoses: current perspective and future promise

PubMed Central

Chamcheu, Jean Christopher; Wood, Gary S.; Siddiqui, Imtiaz A.; Syed, Deeba N.; Adhami, Vaqar M.; Teng, Joyce M.; Mukhtar, Hasan

2012-01-01

Hereditary keratin disorders of the skin and its appendages comprise a large group of clinically heterogeneous disfiguring blistering and ichthyotic diseases, primarily characterized by the loss of tissue integrity, blistering and hyperkeratosis in severely affected tissues. Pathogenic mutations in keratins cause these afflictions. Typically, these mutations in concert with characteristic features have formed the basis for improved disease diagnosis, prognosis and most recently therapy development. Examples include epidermolysis bullosa simplex, keratinopathic ichthyosis, pachyonychia congenita and several other tissue-specific hereditary keratinopathies. Understanding the molecular and genetic events underlying skin dysfunction has initiated alternative treatment approaches that may provide novel therapeutic opportunities for affected patients. Animal and in vitro disease modelling studies have shed more light on molecular pathogenesis, further defining the role of keratins in disease processes and promoting the translational development of new gene and pharmacological therapeutic strategies. Given that the molecular basis for these monogenic disorders is well established, gene therapy and drug discovery targeting pharmacological compounds with the ability to reinforce the compromised cytoskeleton may lead to promising new therapeutic strategies for treating hereditary keratinopathies. In this review, we will summarize and discuss recent advances in the preclinical and clinical modelling and development of gene, natural product, pharmacological and protein-based therapies for these disorders, highlighting the feasibility of new approaches for translational clinical therapy. PMID:22716242

DNA Methylation Program in Developing Hippocampus and Its Alteration by Alcohol

PubMed Central

Chen, Yuanyuan; Ozturk, Nail Can; Zhou, Feng C.

2013-01-01

During hippocampal development, the Cornus Ammonis (CA) and the dentate gyrus (DG) undergo waves of neurogenesis and neuronal migration and maturation independently. This stage is widely known to be vulnerable to environmental stresses, but its underlying mechanism is unclear. Alcohol exposure has been shown to alter the expression of genes that regulate the fate, survival, migration and differentiation of pyramidal and granule cells. Undermining this process might compromise hippocampal development underlying the learning and memory deficits known in Fetal Alcohol Spectrum Disorders (FASD). We have previously demonstrated that DNA methylation was programmed along with neural tube development. Here, we demonstrated that DNA methylation program (DMP) proceeded along with hippocampal neuronal differentiation and maturation, and how this DMP was affected by fetal alcohol exposure. C57BL/6 mice were treated with 4% v/v ethanol through a liquid diet along with pair-fed and chow-fed controls from gestation day (E) 7 to E16. We found that a characteristic DMP, including 5-methylcytidine (5mC), 5-hydroxylmethylcytidine (5hmC) and their binding proteins, led the hippocampal neuronal differentiation and maturation spatiotemporally as indicated by their phenotypic marks in the CA and DG pre- and post-natally. Alcohol hindered the acquisition and progression of methylation marks, and altered the chromatin translocation of these marks in the nucleus, which was correlated with developmental retardation. PMID:23544149

High doses of gamma radiation suppress allergic effect induced by food lectin

NASA Astrophysics Data System (ADS)

Vaz, Antônio F. M.; Souza, Marthyna P.; Vieira, Leucio D.; Aguiar, Jaciana S.; Silva, Teresinha G.; Medeiros, Paloma L.; Melo, Ana M. M. A.; Silva-Lucca, Rosemeire A.; Santana, Lucimeire A.; Oliva, Maria L. V.; Perez, Katia R.; Cuccovia, Iolanda M.; Coelho, Luana C. B. B.; Correia, Maria T. S.

2013-04-01

One of the most promising areas for the development of functional foods lies in the development of effective methods to reduce or eliminate food allergenicity, but few reports have summarized information concerning the progress made with food irradiation. In this study, we investigated the relationship between allergenicity and molecular structure of a food allergen after gamma irradiation and evaluate the profile of the allergic response to irradiated allergens. Cramoll, a lectin isolated from a bean and used as a food allergen, was irradiated and the possible structural changes were accompanied by spectrofluorimetry, circular dichroism and microcalorimetry. Subsequently, sensitized animals subjected to intragastric administration of non-irradiated and irradiated Cramoll were treated for 7 days. Then, body weight, leukocytes, cytokine profiles and histological parameters were also determined. Cramoll showed complete inhibition of intrinsic activity after high radiation doses. Changes in fluorescence and CD spectra with a simultaneous collapse of the tertiary structure followed by a pronounced decrease of native secondary structure were observed after irradiation. After oral challenge, sensitized mice demonstrate an association between Cramoll intake, body weight loss, eosinophilia, lymphocytic infiltrate in the gut and Eotaxin secretion. Irradiation significantly reduces, according to the dose, the effects observed by non-irradiated food allergens. We confirm that high-dose radiation may render protein food allergens innocuous by irreversibly compromising their molecular structure.

Sustained Endocannabinoid Signaling Compromises Decidual Function and Promotes Inflammation-induced Preterm Birth.

PubMed

Sun, Xiaofei; Deng, Wenbo; Li, Yingju; Tang, Shuang; Leishman, Emma; Bradshaw, Heather B; Dey, Sudhansu K

2016-04-08

Recent studies provide evidence that premature maternal decidual senescence resulting from heightened mTORC1 signaling is a cause of preterm birth (PTB). We show here that mice devoid of fatty acid amide hydrolase (FAAH) with elevated levels ofN-arachidonyl ethanolamide (anandamide), a major endocannabinoid lipid mediator, were more susceptible to PTB upon lipopolysaccharide (LPS) challenge. Anandamide is degraded by FAAH and primarily works by activating two G-protein-coupled receptors CB1 and CB2, encoded by Cnr1 and Cnr2, respectively. We found thatFaah(-/-)decidual cells progressively underwent premature senescence as marked by increased senescence-associated β-galactosidase (SA-β-Gal) staining and γH2AX-positive decidual cells. Interestingly, increased endocannabinoid signaling activated MAPK p38, but not p42/44 or mTORC1 signaling, inFaah(-/-)deciduae, and inhibition of p38 halted premature decidual senescence. We further showed that treatment of a long-acting anandamide in wild-type mice at midgestation triggered premature decidual senescence utilizing CB1, since administration of a CB1 antagonist greatly reduced the rate of PTB inFaah(-/-)females exposed to LPS. These results provide evidence that endocannabinoid signaling is critical in regulating decidual senescence and parturition timing. This study identifies a previously unidentified pathway in decidual senescence, which is independent of mTORC1 signaling. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Autophagy in human skin fibroblasts: Comparison between young and aged cells and evaluation of its cellular rhythm and response to Ultraviolet A radiation.

PubMed

Pernodet, Nadine; Dong, Kelly; Pelle, Edward

2016-01-01

Autophagic mechanisms play critical roles in cell maintenance. Damaged organelles that are not removed by autophagosomes, which act by engulfing and degrading these cellular components, have been linked to various pathologies. Recently, the progression of aging has also been correlated to a compromised autophagic response. Here, we report for the first time a significant reduction in autophagic levels in synchronized aged normal human skin fibroblasts as compared to young fibroblasts. We measured a 77.9% reduction in autophagy as determined by reverse transcription-polymerase chain reaction for LC3B expression, a microtubule-associated protein correlated to late stage autophagosome formation. In addition, we visualized these same changes by immunocytofluorescence with antibodies directed against LC3B. By harvesting synchronized, as well as unsynchronized cells over time, we were also able to measure for the first time a nighttime peak in autophagy that was present in young but absent in aged fibroblasts. Finally, since human skin is constantly subjected to environmentally induced oxidative stress from sunlight, we exposed fibroblasts to 10 J/cm2 ultraviolet A and found, in good agreement with current literature, not only that irradiation could partially reactivate autophagy in the aged cells, but also that this increase was phase shifted earlier from its endogenous temporal pattern because of its loss of synchronization with circadian rhythm.

Progress towards genetic and pharmacological therapies for keratin genodermatoses: current perspective and future promise.

PubMed

Chamcheu, Jean Christopher; Wood, Gary S; Siddiqui, Imtiaz A; Syed, Deeba N; Adhami, Vaqar M; Teng, Joyce M; Mukhtar, Hasan

2012-07-01

Hereditary keratin disorders of the skin and its appendages comprise a large group of clinically heterogeneous disfiguring blistering and ichthyotic diseases, primarily characterized by the loss of tissue integrity, blistering and hyperkeratosis in severely affected tissues. Pathogenic mutations in keratins cause these afflictions. Typically, these mutations in concert with characteristic features have formed the basis for improved disease diagnosis, prognosis and most recently therapy development. Examples include epidermolysis bullosa simplex, keratinopathic ichthyosis, pachyonychia congenita and several other tissue-specific hereditary keratinopathies. Understanding the molecular and genetic events underlying skin dysfunction has initiated alternative treatment approaches that may provide novel therapeutic opportunities for affected patients. Animal and in vitro disease modelling studies have shed more light on molecular pathogenesis, further defining the role of keratins in disease processes and promoting the translational development of new gene and pharmacological therapeutic strategies. Given that the molecular basis for these monogenic disorders is well established, gene therapy and drug discovery targeting pharmacological compounds with the ability to reinforce the compromised cytoskeleton may lead to promising new therapeutic strategies for treating hereditary keratinopathies. In this review, we will summarize and discuss recent advances in the preclinical and clinical modelling and development of gene, natural product, pharmacological and protein-based therapies for these disorders, highlighting the feasibility of new approaches for translational clinical therapy. © 2012 John Wiley & Sons A/S.

Differential expression proteomics of human colorectal cancer based on a syngeneic cellular model for the progression of adenoma to carcinoma.

PubMed

Roth, Udo; Razawi, Hanieh; Hommer, Julia; Engelmann, Katja; Schwientek, Tilo; Müller, Stefan; Baldus, Stephan E; Patsos, Georgios; Corfield, Anthony P; Paraskeva, Christos; Hanisch, Franz-Georg

2010-01-01

This is the first differential expression proteomics study on a human syngeneic cellular in vitro progression model of the colorectal adenoma-to-carcinoma sequence, the anchorage-dependent non-tumorigenic adenoma derived cell line AA/C1 and the derived anchorage-independent and tumorigenic carcinoma cell line AA/C1/SB10C. The study is based on quantitative 2-DE and is complemented by Western blot validation. Excluding redundancies due to proteolysis and post-translational modified isoforms of over 2000 protein spots, 13 proteins were revealed as regulated with statistical variance being within the 95th confidence level and were identified by peptide mass fingerprinting in MALDI MS. Progression-associated proteins belong to the functional complexes of anaerobic glycolysis/gluconeogenesis, steroid biosynthesis, prostaglandin biosynthesis, the regulation and maintenance of the cytoskeleton, protein biosynthesis and degradation, the regulation of apoptosis or other functions. Partial but significant overlap was revealed with previous proteomics and transcriptomics studies in colorectal carcinoma. Among upregulated proteins we identified 3-HMG-CoA synthase, protein phosphatase 1, prostaglandin E synthase 2, villin 1, annexin A1, triosephosphate isomerase, phosphoserine aminotransferase 1, fumarylacetoacetate hydrolase and pyrroline-5-carboxylate reductase 1 (PYCR1), while glucose-regulated protein 78, cathepsin D, lamin A/C and quinolate phosphoribosyltransferase were downregulated.

Dominant negative RPW8.2 fusion proteins reveal the importance of haustorium-oriented protein trafficking for resistance against powdery mildew in Arabidopsis.

PubMed

Zhang, Qiong; Berkey, Robert; Pan, Zhiyong; Wang, Wenming; Zhang, Yi; Ma, Xianfeng; King, Harlan; Xiao, Shunyuan

2015-01-01

Powdery mildew fungi form feeding structures called haustoria inside epidermal cells of host plants to extract photosynthates for their epiphytic growth and reproduction. The haustorium is encased by an interfacial membrane termed the extrahaustorial membrane (EHM). The atypical resistance protein RPW8.2 from Arabidopsis is specifically targeted to the EHM where RPW8.2 activates haustorium-targeted (thus broad-spectrum) resistance against powdery mildew fungi. EHM-specific localization of RPW8.2 suggests the existence of an EHM-oriented protein/membrane trafficking pathway during EHM biogenesis. However, the importance of this specific trafficking pathway for host defense has not been evaluated via a genetic approach without affecting other trafficking pathways. Here, we report that expression of EHM-oriented, nonfunctional RPW8.2 chimeric proteins exerts dominant negative effect over functional RPW8.2 and potentially over other EHM-localized defense proteins, thereby compromising both RPW8.2-mediated and basal resistance to powdery mildew. Thus, our results highlight the importance of the EHM-oriented protein/membrane trafficking pathway for host resistance against haustorium-forming pathogens such as powdery mildew fungi.

Clathrin and AP1 are required for apical sorting of glycosyl phosphatidyl inositol-anchored proteins in biosynthetic and recycling routes in Madin-Darby canine kidney cells.

PubMed

Castillon, Guillaume A; Burriat-Couleru, Patricia; Abegg, Daniel; Criado Santos, Nina; Watanabe, Reika

2018-03-01

Recently, studies in animal models demonstrate potential roles for clathrin and AP1 in apical protein sorting in epithelial tissue. However, the precise functions of these proteins in apical protein transport remain unclear. Here, we reveal mistargeting of endogenous glycosyl phosphatidyl inositol-anchored proteins (GPI-APs) and soluble secretory proteins in Madin-Darby canine kidney (MDCK) cells upon clathrin heavy chain or AP1 subunit knockdown (KD). Using a novel directional endocytosis and recycling assay, we found that these KD cells are not only affected for apical sorting of GPI-APs in biosynthetic pathway but also for their apical recycling and basal-to-apical transcytosis routes. The apical distribution of the t-SNARE syntaxin 3, which is known to be responsible for selective targeting of various apical-destined cargo proteins in both biosynthetic and endocytic routes, is compromised suggesting a molecular explanation for the phenotype in KD cells. Our results demonstrate the importance of biosynthetic and endocytic routes for establishment and maintenance of apical localization of GPI-APs in polarized MDCK cells. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Proteostasis: bad news and good news from the endoplasmic reticulum.

PubMed

Noack, Julia; Brambilla Pisoni, Giorgia; Molinari, Maurizio

2014-01-01

The endoplasmic reticulum (ER) is an intracellular compartment dedicated to the synthesis and maturation of secretory and membrane proteins, totalling about 30% of the total eukaryotic cells proteome. The capacity to produce correctly folded polypeptides and to transport them to their correct intra- or extracellular destinations relies on proteostasis networks that regulate and balance the activity of protein folding, quality control, transport and degradation machineries. Nutrient and environmental changes, pathogen infection aging and, more relevant for the topics discussed in this review, mutations that impair attainment of the correct 3D structure of nascent polypeptide chains may compromise the activity of the proteostasis networks with devastating consequences on cells, organs and organisms' homeostasis. Here we present a review of mechanisms regulating folding and quality control of proteins expressed in the ER, and we describe the protein degradation and the ER stress pathways activated by the expression of misfolded proteins in the ER lumen. Finally, we highlight select examples of proteopathies (also known as conformational disorders or protein misfolding diseases) caused by protein misfolding in the ER and/or affecting cellular proteostasis and therapeutic interventions that might alleviate or cure the disease symptoms.

A Catalogue of Altered Salivary Proteins Secondary to Invasive Ductal Carcinoma: A Novel In Vivo Paradigm to Assess Breast Cancer Progression

PubMed Central

Streckfus, Charles F.; Bigler, Lenora

2016-01-01

The objective of this manuscript is to introduce a catalogue of salivary proteins that are altered secondary to carcinoma of the breast. The catalogue of salivary proteins is a compilation of twenty years of research by the authors and consists of 233 high and low abundant proteins which have been identified by LC-MS/MS mass spectrometry, 2D-gel analysis and by enzyme-linked immunosorbent assay. The body of research suggests that saliva is a fluid suffused with solubilized by-products of oncogenic expression and that these proteins may be useful in the study of breast cancer progress, treatment efficacy and the tailoring of individualized patient care. PMID:27477923

Polycystic Kidney Disease: Pathogenesis and Potential Therapies

PubMed Central

Takiar, Vinita; Caplan, Michael J.

2011-01-01

Autosomal dominant polycystic kidney disease (ADPKD) is a prevalent, inherited condition for which there is currently no effective specific clinical therapy. The disease is characterized by the progressive development of fluid-filled cysts derived from renal tubular epithelial cells which gradually compress the parenchyma and compromise renal function. Current interests in the field focus on understanding and exploiting signaling mechanisms underlying disease pathogenesis as well as delineating the role of the primary cilium in cystogenesis. This review highlights the pathogenetic pathways underlying renal cyst formation as well as novel therapeutic targets for the treatment of PKD. PMID:21146605

Computer-Aided Design/Computer-Assisted Manufacture Monolithic Restorations for Severely Worn Dentition: A Case History Report.

PubMed

Abou-Ayash, Samir; Boldt, Johannes; Vuck, Alexander

Full-arch rehabilitation of patients with severe tooth wear due to parafunctional behavior is a challenge for dentists and dental technicians, especially when a highly esthetic outcome is desired. A variety of different treatment options and prosthetic materials are available for such a clinical undertaking. The ongoing progress of computer-aided design/computer-assisted manufacture technologies in combination with all-ceramic materials provides a predictable workflow for these complex cases. This case history report describes a comprehensive, step-by-step treatment protocol leading to an optimally predictable treatment outcome for an esthetically compromised patient.

A design for integration.

PubMed

Fenna, D

1977-09-01

For nearly two decades, the development of computerized information systems has struggled for acceptable compromises between the unattainable "total system" and the unacceptable separate applications. Integration of related applications is essential if the computer is to be exploited fully, yet relative simplicity is necessary for systems to be implemented in a reasonable time-scale. This paper discusses a system being progressively developed from minimal beginnings but which, from the outset, had a highly flexible and fully integrated system basis. The system is for batch processing, but can accommodate on-line data input; it is similar in its approach to many transaction-processing real-time systems.

Exploring the structure and organization of information within nursing clinical handovers.

PubMed

Johnson, Maree; Jefferies, Diana; Nicholls, Daniel

2012-10-01

Clinical handover is the primary source of patient information for nurses; however, inadequate information transfer compromises patient safety. We investigated the content and organization of information conveyed at 81 handovers. A structure that captures and presents the information transferred at handover emerged: identification of the patient and clinical risks, clinical history/presentation, clinical status, care plan and outcomes/goals of care (ICCCO). This approach covers essential information while allowing for prioritization of information when required. Further research into the impact of ICCCO on patient safety is in progress. © 2012 Wiley Publishing Asia Pty Ltd.

Aspergillus spondylodiscitis after multivisceral transplantation.

PubMed

Gerlach, Undine A; Kohler, Sven; Sauer, Igor M; Joerres, Dinah; Kandziora, Frank; Neuhaus, Peter; Pratschke, Johann; Pascher, Andreas

2009-01-01

Although spondylodiscitis is rare, it is increasingly described in patients with compromised immunity due to malignancy, chemotherapy or immunosuppression. Typical pathogens are staphylococcus aureus and enterobacteria; fungal spondylodiscitis is uncommon. We present a case of aspergillus spondylodiscitis following pulmonary aspergillosis in a patient with multivisceral and kidney transplantation. Due to irreversible disc destruction, surgical restoration by autologous iliac crest graft was required in addition to intravenous antifungal therapy, which consisted of voriconazole, caspofungin and liposomal amphotericin B. Aspergillus spondylodiscitis is a diagnostic and therapeutic challenge, a combination of surgical debridement and antifungal therapy is inevitable to prevent rapid progression of invasive aspergillosis and neurological damage.

Hazards to Early Development: The Biological Embedding of Early Life Adversity.

PubMed

Nelson, Charles A

2017-10-11

The number of children under 18 years of age has increased worldwide over the past decade. This growth spurt is due, in part, to remarkable progress in child survival. Alas, surviving early hazards, like prematurity or infectious disease, does not guarantee that children's development will not be compromised by other hazards as they grow older. Throughout the world, children continue to be confronted with a large number of biological and psychosocial challenges that greatly limit their developmental potential. The focus of this article is how such adverse experiences impact the developing brain. Copyright © 2017 Elsevier Inc. All rights reserved.

Interactions of Cryptococcus with Dendritic Cells

PubMed Central

Wozniak, Karen L.

2018-01-01

The fungal pathogens Cryptococcus neoformans and Cryptococcus gattii can cause life-threatening infections in immune compromised and immune competent hosts. These pathogens enter the host via inhalation, and respiratory tract innate immune cells such as dendritic cells (DCs) are one of the first host cells they encounter. The interactions between Cryptococcus and innate immune cells play a critical role in the progression of disease in the host. This review will focus specifically on the interactions between Cryptococcus and dendritic cells (DCs), including recognition/processing by DCs, effects of immune mediators on DC recruitment and activity, and the potential for DC vaccination against cryptococcosis. PMID:29543719

Interactions of Cryptococcus with Dendritic Cells.

PubMed

Wozniak, Karen L

2018-03-15

The fungal pathogens Cryptococcus neoformans and Cryptococcus gattii can cause life-threatening infections in immune compromised and immune competent hosts. These pathogens enter the host via inhalation, and respiratory tract innate immune cells such as dendritic cells (DCs) are one of the first host cells they encounter. The interactions between Cryptococcus and innate immune cells play a critical role in the progression of disease in the host. This review will focus specifically on the interactions between Cryptococcus and dendritic cells (DCs), including recognition/processing by DCs, effects of immune mediators on DC recruitment and activity, and the potential for DC vaccination against cryptococcosis.

Magnet safety and stability related coolant states: critical fluid dynamics at peak flux

NASA Astrophysics Data System (ADS)

Ravikumar, K. V.; Carandang, R. M.; Frederking, T. H. K.

The stability of superconducting magnets is endangered under certain distinct conditions of the fluid serving as magnet coolant. A severe compromising of safety takes place at the peak heat flux of nucleate boiling. Progress in analysing first order phase transitions for cryoliquids and room temperature liquids, in the presence of heat flow, has led to better understanding of the parameters related to vapour bubble phenomena. The present work addresses the consequences arising from bubble frequency results, including model calculations for the effective masses of the saturated fluids involved in the two-phase transport at the peak flux.

Surfing the big WAVE: Insights into the role of WAVE3 as a driving force in cancer progression and metastasis.

PubMed

Sossey-Alaoui, Khalid

2013-04-01

WAVE3 belongs to the WASP/WAVE family of actin cytoskeleton remodeling proteins. These proteins are known to be involved in several biological functions ranging from controlling cell shape and movement, to being closely associated with pathological conditions such as cancer progression and metastasis. Last decade has seen an explosion in the literature reporting significant scientific advances on the molecular mechanisms whereby the WASP/WAVE proteins are regulated both in normal physiological as well as pathological conditions. The purpose of this review is to present the major findings pertaining to how WAVE3 has become a critical player in the regulation of signaling pathways involved in cancer progression and metastasis. The review will conclude with suggesting options for the potential use of WAVE3 as a therapeutic target to prevent the progression of cancer to the lethal stage that is the metastatic disease. Copyright © 2012 Elsevier Ltd. All rights reserved.

Protein quality control system in neurodegeneration: a healing company hard to beat but failure is fatal.

PubMed

Chhangani, Deepak; Mishra, Amit

2013-08-01

A common feature in most neurodegenerative diseases and aging is the progressive accumulation of damaged proteins. Proteins are essential for all crucial biological functions. Under some notorious conditions, proteins loss their three dimensional native conformations and are converted into disordered aggregated structures. Such changes rise into pathological conditions and eventually cause serious protein conformation disorders. Protein aggregation and inclusion bodies formation mediated multifactorial proteotoxic stress has been reported in the progression of Parkinson's disease (PD), Huntington's disease (HD), Alzheimer's disease (AD), amyotrophic lateral sclerosis (ALS) and Prion disease. Ongoing studies have been remarkably informative in providing a systematic outlook for better understanding the concept and fundamentals of protein misfolding and aggregations. However, the precise role of protein quality control system and precursors of this mechanism remains elusive. In this review, we highlight recent insights and discuss emerging cytoprotective strategies of cellular protein quality control system implicated in protein deposition diseases. Our current review provides a clear, understandable framework of protein quality control system that may offer the more suitable therapeutic strategies for protein-associated diseases.

Antimicrobial preservatives induce aggregation of interferon alpha-2a: The order in which preservatives induce protein aggregation is independent of the protein

PubMed Central

Bis, Regina L.; Mallela, Krishna M.G.

2014-01-01

Antimicrobial preservatives (APs) are included in liquid multi-dose protein formulations to combat the growth of microbes and bacteria. These compounds have been shown to cause protein aggregation, which leads to serious immunogenic and toxic side-effects in patients. Our earlier work on a model protein cytochrome c (Cyt c) demonstrated that APs cause protein aggregation in a specific manner. The aim of this study is to validate the conclusions obtained from our model protein studies on a pharmaceutical protein. Interferon α-2a (IFNA2) is available as a therapeutic treatment for numerous immune-compromised disorders including leukemia and hepatitis c, and APs have been used in its multi-dose formulation. Similar to Cyt c, APs induced IFNA2 aggregation, demonstrated by the loss of soluble monomer and increase in solution turbidity. The extent of IFNA2 aggregation increased with the increase in AP concentration. IFNA2 aggregation also depended on the nature of AP, and followed the order m-cresol > phenol > benzyl alcohol > phenoxyethanol. This specific order exactly matched with that observed for the model protein Cyt c. These and previously published results on antibodies and other recombinant proteins suggest that the general mechanism by which APs induce protein aggregation may be independent of the protein. PMID:24974985

Antimicrobial preservatives induce aggregation of interferon alpha-2a: the order in which preservatives induce protein aggregation is independent of the protein.

PubMed

Bis, Regina L; Mallela, Krishna M G

2014-09-10

Antimicrobial preservatives (APs) are included in liquid multi-dose protein formulations to combat the growth of microbes and bacteria. These compounds have been shown to cause protein aggregation, which leads to serious immunogenic and toxic side-effects in patients. Our earlier work on a model protein cytochrome c (Cyt c) demonstrated that APs cause protein aggregation in a specific manner. The aim of this study is to validate the conclusions obtained from our model protein studies on a pharmaceutical protein. Interferon α-2a (IFNA2) is available as a therapeutic treatment for numerous immune-compromised disorders including leukemia and hepatitis C, and APs have been used in its multi-dose formulation. Similar to Cyt c, APs induced IFNA2 aggregation, demonstrated by the loss of soluble monomer and increase in solution turbidity. The extent of IFNA2 aggregation increased with the increase in AP concentration. IFNA2 aggregation also depended on the nature of AP, and followed the order m-cresol>phenol>benzyl alcohol>phenoxyethanol. This specific order exactly matched with that observed for the model protein Cyt c. These and previously published results on antibodies and other recombinant proteins suggest that the general mechanism by which APs induce protein aggregation may be independent of the protein. Copyright © 2014 Elsevier B.V. All rights reserved.

UCS protein Rng3p activates actin filament gliding by fission yeast myosin-II

PubMed Central

Lord, Matthew; Pollard, Thomas D.

2004-01-01

We purified native Myo2p/Cdc4p/Rlc1p (Myo2), the myosin-II motor required for cytokinesis by Schizosaccharomyces pombe. The Myo2p heavy chain associates with two light chains, Cdc4p and Rlc1p. Although crude Myo2 supported gliding motility of actin filaments in vitro, purified Myo2 lacked this activity in spite of retaining full Ca-ATPase activity and partial actin-activated Mg-ATPase activity. Unc45-/Cro1p-/She4p-related (UCS) protein Rng3p restored the full motility and actin-activated Mg-ATPase activity of purified Myo2. The COOH-terminal UCS domain of Rng3p alone restored motility to pure Myo2. Thus, Rng3p contributes directly to the motility activity of native Myo2. Consistent with a role in Myo2 activation, Rng3p colocalizes with Myo2p in the cytokinetic contractile ring. The absence of Rlc1p or mutations in the Myo2p head or Rng3p compromise the in vitro motility of Myo2 and explain the defects in cytokinesis associated with some of these mutations. In contrast, Myo2 with certain temperature-sensitive forms of Cdc4p has normal motility, so these mutations compromise other functions of Cdc4p required for cytokinesis. PMID:15504913

Altered Osteocyte-Specific Protein Expression in Bone after Childhood Solid Organ Transplantation

PubMed Central

Pereira, Renata C.; Valta, Helena; Tumber, Navdeep; Salusky, Isidro B.; Jalanko, Hannu

2015-01-01

Background Bone fragility is common post solid organ transplantation but little is known about bone pathology on a tissue level. Abnormal osteocytic protein expression has been linked to compromised bone health in chronic kidney disease (CKD) and immunosuppressant medications may impact osteocyte function. Methods Transiliac bone biopsies were obtained from 22 pediatric solid organ allograft recipients (average age 15.6 years) an average of 6.3 ± 1.2 years after transplantation and from 12 pediatric pre-dialysis CKD patients (average age 13.2 years). Histomorphometry and immunohistochemistry for FGF23, DMP1, sclerostin, and osteopontin were performed on all biopsies. Results FGF23 and sclerostin were increased in transplant recipients relative to non-transplant CKD, regardless of the type of allograft received and despite, in the case of liver and heart recipients, a higher GFR. Bone DMP1 expression was higher in liver or heart than in kidney recipients, concomitant with higher serum phosphate values. Osteopontin expression was higher in CKD than in transplant recipients (p<0.01). Bone FGF23 and sclerostin correlated directly (r = 0.38, p<0.05); bone FGF23 expression and osteoid thickness correlated inversely (r = - 0.46, p<0.01). Conclusions Solid-organ transplantation is associated with increased FGF23 and sclerostin expression. The contribution of these findings to compromised bone health post transplantation warrants further evaluation. PMID:26390291

Altered Osteocyte-Specific Protein Expression in Bone after Childhood Solid Organ Transplantation.

PubMed

Pereira, Renata C; Valta, Helena; Tumber, Navdeep; Salusky, Isidro B; Jalanko, Hannu; Mäkitie, Outi; Wesseling Perry, Katherine

2015-01-01

Bone fragility is common post solid organ transplantation but little is known about bone pathology on a tissue level. Abnormal osteocytic protein expression has been linked to compromised bone health in chronic kidney disease (CKD) and immunosuppressant medications may impact osteocyte function. Transiliac bone biopsies were obtained from 22 pediatric solid organ allograft recipients (average age 15.6 years) an average of 6.3 ± 1.2 years after transplantation and from 12 pediatric pre-dialysis CKD patients (average age 13.2 years). Histomorphometry and immunohistochemistry for FGF23, DMP1, sclerostin, and osteopontin were performed on all biopsies. FGF23 and sclerostin were increased in transplant recipients relative to non-transplant CKD, regardless of the type of allograft received and despite, in the case of liver and heart recipients, a higher GFR. Bone DMP1 expression was higher in liver or heart than in kidney recipients, concomitant with higher serum phosphate values. Osteopontin expression was higher in CKD than in transplant recipients (p<0.01). Bone FGF23 and sclerostin correlated directly (r = 0.38, p<0.05); bone FGF23 expression and osteoid thickness correlated inversely (r = - 0.46, p<0.01). Solid-organ transplantation is associated with increased FGF23 and sclerostin expression. The contribution of these findings to compromised bone health post transplantation warrants further evaluation.

Differential effects of male nutrient balance on pre- and post-copulatory traits, and consequences for female reproduction in Drosophila melanogaster

PubMed Central

Morimoto, Juliano; Wigby, Stuart

2016-01-01

Male fitness depends on the expression of costly traits involved in obtaining mates (pre-copulatory) and fertilization (post-copulatory). However, very little is known about the nutrient requirements for these traits and whether males compromise their diet to maximize one trait at the expense of another. Here we used Nutritional Geometry to investigate macronutrient requirements for pre- and post-copulatory traits in Drosophila, when males were the first or second to mate with females. We found no significant effects of male diet on sperm competitiveness. However, although males self-regulate their macronutrient intake at a protein-to-carbohydrate ratio (“P:C ratio”) of 1:1.5, this ratio does not coincide with their optima for several key reproductive traits: both the short-term (~24 hr) rate of offspring production after a female’s first mating, as well as the total offspring number sired when males were second to mate were maximized at a P:C ratio of 1:9, whereas male attractiveness (latency to mate), were maximised at a P:C ratio of 1:1. These results suggest a compromised optimum diet, and no single diet that simultaneously maximizes all male reproductive traits. The protein intake of first males also negatively affected female offspring production following remating, suggesting a long-term intersexual effect of male nutrition. PMID:27270223

Plant growth enhancement and associated physiological responses are coregulated by ethylene and gibberellin in response to harpin protein Hpa1.

PubMed

Li, Xiaojie; Han, Bing; Xu, Manyu; Han, Liping; Zhao, Yanying; Liu, Zhilan; Dong, Hansong; Zhang, Chunling

2014-04-01

The harpin protein Hpa1 produced by the bacterial blight pathogen of rice induces several growth-promoting responses in plants, activating the ethylene signaling pathway, increasing photosynthesis rates and EXPANSIN (EXP) gene expression levels, and thereby enhancing the vegetative growth. This study was attempted to analyze any mechanistic connections among the above and the role of gibberellin in these responses. Hpa1-induced growth enhancement was evaluated in Arabidopsis, tomato, and rice. And growth-promoting responses were determined mainly as an increase of chlorophyll a/b ratio, which indicates a potential elevation of photosynthesis rates, and enhancements of photosynthesis and EXP expression in the three plant species. In Arabidopsis, Hpa1-induced growth-promoting responses were partially compromised by a defect in ethylene perception or gibberellin biosynthesis. In tomato and rice, compromises of Hpa1-induced growth-promoting responses were caused by a pharmacological treatment with an ethylene perception inhibitor or a gibberellin biosynthesis inhibitor. In the three plant species, moreover, Hpa1-induced growth-promoting responses were significantly impaired, but not totally eliminated, by abolishing ethylene perception or gibberellin synthesis. However, simultaneous nullifications in both ethylene perception and gibberellin biosynthesis almost canceled the full effects of Hpa1 on plant growth, photosynthesis, and EXP2 expression. Theses results suggest that ethylene and gibberellin coregulate Hpa1-induced plant growth enhancement and associated physiological and molecular responses.

Oxaliplatin-induced Oxidative Stress Provokes Toxicity in Isolated Rat Liver Mitochondria.

PubMed

Tabassum, Heena; Waseem, Mohammad; Parvez, Suhel; Qureshi, M Irfan

2015-11-01

Oxaliplatin is a widely employed platinum-derived chemotherapeutic agent commonly used for the treatment of colorectal cancer. Unfortunately, the benefit of this important drug is compromised by severe side effects such as neuropathy, ototoxicity, gastrointestinal toxicity, and hematological toxicity. Recently, few studies have also suggested the occurrence of hepatotoxicity in oxaliplatin-treated patients. Mitochondria have emerged as targets for anticancer drugs in various kinds of toxicity including hepatotoxicity that can lead to neoplastic disease. Oxidative stress is a well-established biomarker of mitochondrial toxicity. The purpose of this study was to investigate the dose-dependent damage caused by oxaliplatin on isolated liver mitochondria under in vitro conditions. The study was conducted in mitochondria isolated from liver of Wistar rats. Oxaliplatin was incubated with mitochondria in a dose-dependent manner under in vitro conditions. Oxidative stress indexes, non-enzymatic and enzymatic antioxidants were evaluated, looking at the overall armamentarium against the toxicity induced by oxaliplatin. Oxaliplatin caused a significant rise in the mitochondrial oxidative stress indexes lipid peroxidation and protein carbonyl. Alterations in the levels of non-enzymatic antioxidants and activities of enzymatic antioxidants were also observed. Oxidative stress plays an important role in the mitochondrial toxicity of oxaliplatin. The integrity of the hepatic tissue is compromised by the reactive oxygen species-mediated lipid peroxidation and protein carbonyl formation. Copyright © 2015 IMSS. Published by Elsevier Inc. All rights reserved.

Cold ischemia contributes to the development of chronic rejection and mitochondrial injury after cardiac transplantation.

PubMed

Schneeberger, Stefan; Amberger, Albert; Mandl, Julia; Hautz, Theresa; Renz, Oliver; Obrist, Peter; Meusburger, Hugo; Brandacher, Gerald; Mark, Walter; Strobl, Daniela; Troppmair, Jakob; Pratschke, Johann; Margreiter, Raimund; Kuznetsov, Andrey V

2010-12-01

Chronic rejection (CR) remains an unsolved hurdle for long-term heart transplant survival. The effect of cold ischemia (CI) on progression of CR and the mechanisms resulting in functional deficit were investigated by studying gene expression, mitochondrial function, and enzymatic activity. Allogeneic (Lew→F344) and syngeneic (Lew→Lew) heart transplantations were performed with or without 10 h of CI. After evaluation of myocardial contraction, hearts were excised at 2, 10, 40, and 60 days for investigation of vasculopathy, gene expression, enzymatic activities, and mitochondrial respiration. Gene expression studies identified a gene cluster coding for subunits of the mitochondrial electron transport chain regulated in response to CI and CR. Myocardial performance, mitochondrial function, and mitochondrial marker enzyme activities declined in all allografts with time after transplantation. These declines were more rapid and severe in CI allografts (CR-CI) and correlated well with progression of vasculopathy and fibrosis. Mitochondria related gene expression and mitochondrial function are substantially compromised with the progression of CR and show that CI impacts on progression, gene profile, and mitochondrial function of CR. Monitoring mitochondrial function and enzyme activity might allow for earlier detection of CR and cardiac allograft dysfunction. © 2010 The Authors. Journal compilation © 2010 European Society for Organ Transplantation.

GSTM3 and GSTP1: novel players driving tumor progression in cervical cancer

PubMed Central

Checa-Rojas, Alberto; Delgadillo-Silva, Luis Fernando; Velasco-Herrera, Martín del Castillo; Andrade-Domínguez, Andrés; Gil, Jeovanis; Santillán, Orlando; Lozano, Luis; Toledo-Leyva, Alfredo; Ramírez-Torres, Alberto; Talamas-Rohana, Patricia; Encarnación-Guevara, Sergio

2018-01-01

The molecular processes and proteomic markers leading to tumor progression (TP) in cervical cancer (CC) are either unknown or only partially understood. TP affects metabolic and regulatory mechanisms that can be identified as proteomic changes. To identify which proteins are differentially expressed and to understand the mechanisms of cancer progression, we analyzed the dynamics of the tumor proteome in CC cell lines. This analysis revealed two proteins that are up-regulated during TP, GSTM3 and GSTP1. These proteins are involved in cell maintenance, cell survival and the cellular stress response via the NF-κB and MAP kinase pathways during TP. Furthermore, GSTM3 and GSTP1 knockdown showed that evasion of apoptosis was affected, and tumor proliferation was significantly reduced. Our data indicate the critical role of GST proteins in the regulation and progression of cervical cancer cells. Hence, we suggest GSTM3 and GSTP1 as novel biomarkers and potential therapeutic targets for treating cervical cancer. Significance CC is particularly hazardous in the advanced stages, and there are few therapeutic strategies specifically targeting these stages. We performed analyses on CC tumor proteome dynamics and identified GSTM3 and GSTP1 as novel potential therapeutic targets. Knockdown of these proteins showed that they are involved in cell survival, cell proliferation and cellular evasion of apoptosis. PMID:29774096

Conservative Tryptophan Mutants of the Protein Tyrosine Phosphatase YopH Exhibit Impaired WPD-Loop Function and Crystallize with Divanadate Esters in Their Active Sites

PubMed Central

Moise, Gwendolyn; Gallup, Nathan M.; Alexandrova, Anastassia N.; Hengge, Alvan C.; Johnson, Sean J.

2016-01-01

Catalysis in protein tyrosine phosphatases (PTPs) involves movement of a protein loop called the WPD loop that brings a conserved aspartic acid into the active site to function as a general acid. Mutation of the tryptophan in the WPD loop of the PTP YopH to any other residue with a planar, aromatic side chain (phenylalanine, tyrosine, or histidine) disables general acid catalysis. Crystal structures reveal these conservative mutations leave this critical loop in a catalytically unproductive, quasi-open position. Although the loop positions in crystal structures are similar for all three conservative mutants, the reasons inhibiting normal loop closure differ for each mutant. In the W354F and W354Y mutants, steric clashes result from six-membered rings occupying the position of the five-membered ring of the native indole side chain. The histidine mutant dysfunction results from new hydrogen bonds stabilizing the unproductive position. The results demonstrate how even modest modifications can disrupt catalytically important protein dynamics. Crystallization of all the catalytically compromised mutants in the presence of vanadate gave rise to vanadate dimers at the active site. In W354Y and W354H, a divanadate ester with glycerol is observed. Such species have precedence in solution and are known from the small molecule crystal database. Such species have not been observed in the active site of a phosphatase, as a functional phosphatase would rapidly catalyze their decomposition. The compromised functionality of the mutants allows the trapping of species that undoubtedly form in solution and are capable of binding at the active sites of PTPs, and, presumably, other phosphatases. In addition to monomeric vanadate, such higher-order vanadium-based molecules are likely involved in the interaction of vanadate with PTPs in solution. PMID:26445170

A Critical Role of Lyst-Interacting Protein5, a Positive Regulator of Multivesicular Body Biogenesis, in Plant Responses to Heat and Salt Stresses1

PubMed Central

Wang, Fei; Yang, Yan; Wang, Zhe; Zhou, Jie; Fan, Baofang; Chen, Zhixiang

2015-01-01

Multivesicular bodies (MVBs) are unique endosomes containing vesicles in the lumen and play critical roles in many cellular processes. We have recently shown that Arabidopsis (Arabidopsis thaliana) Lyst-Interacting Protein5 (LIP5), a positive regulator of the Suppressor of K+ Transport Growth Defect1 (SKD1) AAA ATPase in MVB biogenesis, is a critical target of the mitogen-activated protein kinases MPK3 and MPK6 and plays an important role in the plant immune system. In this study, we report that the LIP5-regulated MVB pathway also plays a critical role in plant responses to abiotic stresses. Disruption of LIP5 causes compromised tolerance to both heat and salt stresses. The critical role of LIP5 in plant tolerance to abiotic stresses is dependent on its ability to interact with Suppressor of K+ Transport Growth Defect1. When compared with wild-type plants, lip5 mutants accumulate increased levels of ubiquitinated protein aggregates and NaCl under heat and salt stresses, respectively. Further analysis using fluorescent dye and MVB markers reveals that abiotic stress increases the formation of endocytic vesicles and MVBs in a largely LIP5-dependent manner. LIP5 is also required for the salt-induced increase of intracellular reactive oxygen species, which have been implicated in signaling of salt stress responses. Basal levels of LIP5 phosphorylation by MPKs and the stability of LIP5 are elevated by salt stress, and mutation of MPK phosphorylation sites in LIP5 reduces the stability and compromises the ability to complement the lip5 salt-sensitive mutant phenotype. These results collectively indicate that the MVB pathway is positively regulated by pathogen/stress-responsive MPK3/6 through LIP5 phosphorylation and plays a critical role in broad plant responses to biotic and abiotic stresses. PMID:26229051

Current Status of the Congenital Myasthenic Syndromes

PubMed Central

Engel, Andrew G.

2011-01-01

Congenital myasthenic syndromes (CMS) are heterogeneous disorders in which the safety margin of neuromuscular transmission is compromised by one or more specific mechanisms. Clinical, electrophysiologic, and morphologic studies have paved the way for detecting CMS-related mutations in proteins residing in the nerve terminal, the synaptic basal lamina, and in the postsynaptic region of the motor endplate. The disease proteins identified to date include choline acetyltransferase (ChAT), the endplate species of acetylcholinesterase (AChE), β2-laminin, the acetylcholine receptor (AChR), rapsyn, plectin, Nav1.4, the muscle specific protein kinase (MuSK), agrin, downstream of tyrosine kinase 7 (Dok-7), and glutamine-fructose-6-phosphate transaminase 1 (GFPT1). Myasthenic syndromes associated with centronuclear myopathies were recently recognized. Analysis of properties of expressed mutant proteins contributed to finding improved therapy for most CMS. Despite these advances, the molecular basis of some phenotypically characterized CMS remains elusive. Moreover, other types of CMS and disease genes likely exist and await discovery. PMID:22104196

The advances and perspectives of recombinant protein production in the silk gland of silkworm Bombyx mori.

PubMed

Xu, Hanfu

2014-10-01

The silk gland of silkworm Bombyx mori, is one of the most important organs that has been fully studied and utilized so far. It contributes finest silk fibers to humankind. The silk gland has excellent ability of synthesizing silk proteins and is a kind tool to produce some useful recombinant proteins, which can be widely used in the biological, biotechnical and pharmaceutical application fields. It's a very active area to express recombinant proteins using the silk gland as a bioreactor, and great progress has been achieved recently. This review recapitulates the progress of producing recombinant proteins and silk-based biomaterials in the silk gland of silkworm in addition to the construction of expression systems. Current challenges and future trends in the production of valuable recombinant proteins using transgenic silkworms are also discussed.

Research Results Ultra-fast Energy Transfer from Monomer to Dimer within a Trimeric Molecule New Progress in Heterogeneous Catalysis Research Key Progress in Research on Terrestrial Carbon Cycle in China A New Progress in Research on the Mechanism of Bio-Invasion New Findings in Anti-viral infection and Control of Inflammation Major Headway in Avian Origin Research New Progress in Gold-Nanoparticle-Based Biochips Topological Insulator Research Made Important Progress Major Progress in Biodiversity Achieved New Developments of Direct Methods in Protein Crystallography Major Progress in China-UK Collaboration on the Causal Relationship between Volcanic Activity and Biological Distinction News in Brief: NSFC set up "Research Fund for Young Foreign Scholars" How Often Does Human DNA Mutate? Research Progress on Colossal Anisotropic Magneto Resistive Effect

NASA Astrophysics Data System (ADS)

2009-01-01

Ultra-fast Energy Transfer from Monomer to Dimer within a Trimeric Molecule New Progress in Heterogeneous Catalysis Research Key Progress in Research on Terrestrial Carbon Cycle in China A New Progress in Research on the Mechanism of Bio-Invasion New Findings in Anti-viral infection and Control of Inflammation Major Headway in Avian Origin Research New Progress in Gold-Nanoparticle-Based Biochips Topological Insulator Research Made Important Progress Major Progress in Biodiversity Achieved New Developments of Direct Methods in Protein Crystallography Major Progress in China-UK Collaboration on the Causal Relationship between Volcanic Activity and Biological Distinction News in Brief: NSFC set up "Research Fund for Young Foreign Scholars" How Often Does Human DNA Mutate? Research Progress on Colossal Anisotropic Magneto Resistive Effect

Antiretrovirals, Methamphetamine, and HIV-1 Envelope Protein gp120 Compromise Neuronal Energy Homeostasis in Association with Various Degrees of Synaptic and Neuritic Damage.

PubMed

Sanchez, Ana B; Varano, Giuseppe P; de Rozieres, Cyrus M; Maung, Ricky; Catalan, Irene C; Dowling, Cari C; Sejbuk, Natalia E; Hoefer, Melanie M; Kaul, Marcus

2016-01-01

HIV-1 infection frequently causes HIV-associated neurocognitive disorders (HAND) despite combination antiretroviral therapy (cART). Evidence is accumulating that components of cART can themselves be neurotoxic upon long-term exposure. In addition, abuse of psychostimulants, such as methamphetamine, seems to aggravate HAND and compromise antiretroviral therapy. However, the combined effect of virus and recreational and therapeutic drugs on the brain is poorly understood. Therefore, we exposed mixed neuronal-glial cerebrocortical cells to antiretrovirals (ARVs) (zidovudine [AZT], nevirapine [NVP], saquinavir [SQV], and 118-D-24) of four different pharmacological categories and to methamphetamine and, in some experiments, the HIV-1 gp120 protein for 24 h and 7 days. Subsequently, we assessed neuronal injury by fluorescence microscopy, using specific markers for neuronal dendrites and presynaptic terminals. We also analyzed the disturbance of neuronal ATP levels and assessed the involvement of autophagy by using immunofluorescence and Western blotting. ARVs caused alterations of neurites and presynaptic terminals primarily during the 7-day incubation and depending on the specific compounds and their combinations with and without methamphetamine. Similarly, the loss of neuronal ATP was context specific for each of the drugs or combinations thereof, with and without methamphetamine or viral gp120. Loss of ATP was associated with activation of AMP-activated protein kinase (AMPK) and autophagy, which, however, failed to restore normal levels of neuronal ATP. In contrast, boosting autophagy with rapamycin prevented the long-term drop of ATP during exposure to cART in combination with methamphetamine or gp120. Our findings indicate that the overall positive effect of cART on HIV infection is accompanied by detectable neurotoxicity, which in turn may be aggravated by methamphetamine. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

"Product on Stopper" in a Lyophilized Drug Product: Cosmetic Defect or a Product Quality Concern?

PubMed

Mehta, Shyam B; Roy, Shouvik; Yang, Han-Chang Cathy

2018-06-01

During manufacturing of a lyophilized drug product, operator errors in product handling during loading of product filled vials onto the lyophilizer can lead to a seemingly cosmetic defect which can impact certain critical quality attributes of finished product. In this study, filling of a formulated monoclonal antibody in vials was performed using a peristaltic pump filling unit, and subsequently, the product was lyophilized. After lyophilization, upon visual inspection, around 40% of vials had cosmetic defect with residual product around stopper of the vial and were categorized as "product on stopper" vials, whereas remaining 60% vials with no cosmetic defect were called "acceptable vials." Both groups of vials from 1 single batch were tested for critical quality attributes including protein concentration (ultraviolet absorbance at 280), residual moisture (Karl Fischer), sterility (membrane filtration), and container closure integrity (CCI) (blue dye ingress). Analysis of protein quality attributes such as aggregation, protein concentration, residual moisture showed no significant difference between vials with "product on stopper" and "acceptable vials." However, CCI of the "product on stopper" vials was compromised due to the presence of product around stopper of the vial. The results from this case study demonstrate the following 2 important findings: (1) that a seemingly cosmetic defect may impact product quality, compromising the integrity of the product and (2) that CCI test method can be used as an orthogonal method to sterility testing to evaluate sterility assurance of the product. The corrective action proposed to mitigate this defect is use of a larger sized vial that can potentially minimize this defect that arises because of product handling errors. Copyright © 2018 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

Type I J-Domain NbMIP1 Proteins Are Required for Both Tobacco Mosaic Virus Infection and Plant Innate Immunity

PubMed Central

Liu, Qi; Zhang, Haili; Wang, Yan; Hong, Yiguo; Xiao, Fangming; Zhang, Ling; Shen, Qianhua; Liu, Yule

2013-01-01

Tm-22 is a coiled coil-nucleotide binding-leucine rich repeat resistance protein that confers durable extreme resistance against Tomato mosaic virus (ToMV) and Tobacco mosaic virus (TMV) by recognizing the viral movement protein (MP). Here we report that the Nicotiana benthamiana J-domain MIP1 proteins (NbMIP1s) associate with tobamovirus MP, Tm-22 and SGT1. Silencing of NbMIP1s reduced TMV movement and compromised Tm-22-mediated resistance against TMV and ToMV. Furthermore, silencing of NbMIP1s reduced the steady-state protein levels of ToMV MP and Tm-22. Moreover, NbMIP1s are required for plant resistance induced by other R genes and the nonhost pathogen Pseudomonas syringae pv. tomato (Pst) DC3000. In addition, we found that SGT1 associates with Tm-22 and is required for Tm-22-mediated resistance against TMV. These results suggest that NbMIP1s function as co-chaperones during virus infection and plant immunity. PMID:24098120

Ubiquitin-like protein UBL5 promotes the functional integrity of the Fanconi anemia pathway.

PubMed

Oka, Yasuyoshi; Bekker-Jensen, Simon; Mailand, Niels

2015-05-12

Ubiquitin and ubiquitin-like proteins (UBLs) function in a wide array of cellular processes. UBL5 is an atypical UBL that does not form covalent conjugates with cellular proteins and which has a known role in modulating pre-mRNA splicing. Here, we report an unexpected involvement of human UBL5 in promoting the function of the Fanconi anemia (FA) pathway for repair of DNA interstrand crosslinks (ICLs), mediated by a specific interaction with the central FA pathway component FANCI. UBL5-deficient cells display spliceosome-independent reduction of FANCI protein stability, defective FANCI function in response to DNA damage and hypersensitivity to ICLs. By mapping the sequence determinants underlying UBL5-FANCI binding, we generated separation-of-function mutants to demonstrate that key aspects of FA pathway function, including FANCI-FANCD2 heterodimerization, FANCD2 and FANCI monoubiquitylation and maintenance of chromosome stability after ICLs, are compromised when the UBL5-FANCI interaction is selectively inhibited by mutations in either protein. Together, our findings establish UBL5 as a factor that promotes the functionality of the FA DNA repair pathway. © 2015 The Authors.

Tobacco BY-2 Media Component Optimization for a Cost-Efficient Recombinant Protein Production

PubMed Central

Häkkinen, Suvi T.; Reuter, Lauri; Nuorti, Ninni; Joensuu, Jussi J.; Rischer, Heiko; Ritala, Anneli

2018-01-01

Plant cells constitute an attractive platform for production of recombinant proteins as more and more animal-free products and processes are desired. One of the challenges in using plant cells as production hosts has been the costs deriving from expensive culture medium components. In this work, the aim was to optimize the levels of most expensive components in the nutrient medium without compromising the accumulation of biomass and recombinant protein yields. Wild-type BY-2 culture and transgenic tobacco BY-2 expressing green fluorescent protein–Hydrophobin I (GFP-HFBI) fusion protein were used to determine the most inexpensive medium composition. One particularly high-accumulating BY-2 clone, named ‘Hulk,’ produced 1.1 ± 0.2 g/l GFP-HFBI in suspension and kept its high performance during prolonged subculturing. In addition, both cultures were successfully cryopreserved enabling truly industrial application of this plant cell host. With the optimized culture medium, 43–55% cost reduction with regard to biomass and up to 69% reduction with regard to recombinant protein production was achieved. PMID:29434617

The Small Viral Membrane-Associated Protein P32 Is Involved in Bacteriophage PRD1 DNA Entry

PubMed Central

Grahn, A. Marika; Daugelavičius, Rimantas; Bamford, Dennis H.

2002-01-01

The lipid-containing bacteriophage PRD1 infects a variety of gram-negative cells by injecting its linear double-stranded DNA genome into the host cell cytoplasm, while the protein capsid is left outside. The virus membrane and several structural proteins are involved in phage DNA entry. In this work we identified a new infectivity protein of PRD1. Disruption of gene XXXII resulted in a mutant phenotype defective in phage reproduction. The absence of the protein P32 did not compromise the particle assembly but led to a defect in phage DNA injection. In P32-deficient particles the phage membrane is unable to undergo a structural transformation from a spherical to a tubular form. Since P32− particles are able to increase the permeability of the host cell envelope to a degree comparable to that found with wild-type particles, we suggest that the tail-tube formation is needed to eject the DNA from the phage particle rather than to reach the host cell interior. PMID:11967303

Emerging players in the initiation of eukaryotic DNA replication

PubMed Central

2012-01-01

Faithful duplication of the genome in eukaryotes requires ordered assembly of a multi-protein complex called the pre-replicative complex (pre-RC) prior to S phase; transition to the pre-initiation complex (pre-IC) at the beginning of DNA replication; coordinated progression of the replisome during S phase; and well-controlled regulation of replication licensing to prevent re-replication. These events are achieved by the formation of distinct protein complexes that form in a cell cycle-dependent manner. Several components of the pre-RC and pre-IC are highly conserved across all examined eukaryotic species. Many of these proteins, in addition to their bona fide roles in DNA replication are also required for other cell cycle events including heterochromatin organization, chromosome segregation and centrosome biology. As the complexity of the genome increases dramatically from yeast to human, additional proteins have been identified in higher eukaryotes that dictate replication initiation, progression and licensing. In this review, we discuss the newly discovered components and their roles in cell cycle progression. PMID:23075259

RNA metabolism in the regulation of protein synthesis in plants. Progress report, 1975-1979

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

Key, J L

1979-01-01

The major objectives of the research for the contract period covered by this report were (1) to gain an insight into the sequence organization of the DNA of soybean, emphasizing the arrangement of single copy or unique sequences and repetitive sequences of DNA throughout the genome, (2) to characterize soybean RNAs relative to nucleotide sequence complexity and kinetics of synthesis and turnover of poly A/sup +/ mRNA, and (3) to study ribosomal proteins directed to an analysis of possible changes in proteins which relate to the activation of 80S ribosomes and thus mRNA utilization and protein synthesis in response tomore » environmental stimuli. Even with greatly reduced funding compared to that requested, objectives 1 and 2 were substantially accomplished. Because of reduced funding and the 20-month no cost extension, relatively little progress was made on objective 3. Accordingly objectives 1 and 2 will be summarized in some detail; a brief account of progress is presented on objective 3.« less

TES inhibits colorectal cancer progression through activation of p38.

PubMed

Li, Huili; Huang, Kun; Gao, Lu; Wang, Lixia; Niu, Yanfeng; Liu, Hongli; Wang, Zheng; Wang, Lin; Wang, Guobin; Wang, Jiliang

2016-07-19

The human TESTIN (TES) gene has been identified as a candidate tumor suppressor based on its location at a common fragile site - a region where loss of heterozygosity has been detected in numerous types of tumors. To investigate its role in colorectal cancer (CRC), we examined TES protein levels in CRC tissue samples and cell lines. We observed that TES was markedly reduced in both CRC tissue and cell lines. Additionally, overexpression of TES significantly inhibited cell proliferation, migration, and invasion, while increasing cell apoptosis in colon cancer cells. By contrast, shRNA-mediated TES knockdown elicited the opposite effects. TES inhibited the progression of CRC by up-regulating pro-apoptotic proteins, down-regulating anti-apoptotic proteins, and simultaneously activating p38 mitogen-activated protein kinase (MAPK) signaling pathways. Collectively, these data indicate that TES functions as a necessary suppressor of CRC progression by activating p38-MAPK signaling pathways. This suggests that TES may have a potential application in CRC diagnosis and targeted gene therapy.

Evidence for the Influence of the Iron Regulatory MHC Class I Molecule HFE on Tumor Progression in Experimental Models and Clinical Populations

PubMed Central

Weston, Cody; Connor, James

2014-01-01

Proteins involved in iron regulation are modifiers of cancer risk and progression. Of these, the HFE protein (high iron gene and its protein product) is of particular interest because of its interaction with both iron handling and immune function and the high rate of genetic polymorphisms resulting in a mutant protein. Clinical studies suggest that HFE polymorphisms increase the risk of certain cancers, but the inconsistent outcomes suggest a more nuanced effect, possibly interacting with other genetic or environmental factors. Some basic science research has been conducted to begin to understand the implications of variant HFE genotype on cancer, but the story is far from complete. In particular, putative mechanisms exist for HFE to affect tumor progression through its role in iron handling and its major histocompatibility complex class I structural features. In this review, the current understanding of the role of HFE in cancer is described and models for future directions are identified. PMID:25520556

TES inhibits colorectal cancer progression through activation of p38

PubMed Central

Gao, Lu; Wang, Lixia; Niu, Yanfeng; Liu, Hongli; Wang, Zheng; Wang, Lin; Wang, Guobin; Wang, Jiliang

2016-01-01

The human TESTIN (TES) gene has been identified as a candidate tumor suppressor based on its location at a common fragile site – a region where loss of heterozygosity has been detected in numerous types of tumors. To investigate its role in colorectal cancer (CRC), we examined TES protein levels in CRC tissue samples and cell lines. We observed that TES was markedly reduced in both CRC tissue and cell lines. Additionally, overexpression of TES significantly inhibited cell proliferation, migration, and invasion, while increasing cell apoptosis in colon cancer cells. By contrast, shRNA-mediated TES knockdown elicited the opposite effects. TES inhibited the progression of CRC by up-regulating pro-apoptotic proteins, down-regulating anti-apoptotic proteins, and simultaneously activating p38 mitogen-activated protein kinase (MAPK) signaling pathways. Collectively, these data indicate that TES functions as a necessary suppressor of CRC progression by activating p38-MAPK signaling pathways. This suggests that TES may have a potential application in CRC diagnosis and targeted gene therapy. PMID:27323777

Measures of Urinary Protein and Albumin in the Prediction of Progression of IgA Nephropathy

PubMed Central

Zhao, Yan-feng; Liu, Li-jun; Shi, Su-fang; Lv, Ji-cheng; Zhang, Hong

2016-01-01

Background and objectives Proteinuria is an independent predictor for IgA nephropathy (IgAN) progression. Urine albumin-to-creatinine ratio (ACR), protein-to-creatinine ratio, and 24-hour urine protein excretion (UPE) are widely used for proteinuria evaluation in clinical practice. Here, we evaluated the association of these measurements with clinical and histologic findings of IgAN and explored which was the best predictor of IgAN prognosis. Design, setting, participants, & measurements Patients with IgAN were followed up for ≥12 months, were diagnosed between 2003 and 2012, and had urine samples available (438 patients). Spot urine ACR, protein-to-creatinine ratio, and 24-hour UPE at the time of renal biopsy were measured on a Hitachi Automatic Biochemical Analyzer 7180 (Hitachi, Yokohama, Japan). Results In our patients, ACR, protein-to-creatinine ratio, and 24-hour UPE were highly correlated (correlation coefficients: 0.71–0.87). They showed good relationships with acknowledged markers reflecting IgAN severity, including eGFR, hypertension, and the biopsy parameter (Oxford severity of tubular atrophy/interstitial fibrosis parameter). However, only ACR presented with positive association with the Oxford segmental glomerulosclerosis/adhesion parameter and extracapillary proliferation lesions. The follow-up time was 37.0 (22.0–58.0) months, with the last follow-up on April 18, 2014. In total, 124 patients reached the composite end point (30% eGFR decline, ESRD, or death). In univariate survival analysis, ACR consistently had better performance than protein-to-creatinine ratio and 24-hour UPE as represented by higher area under the curve using time–dependent survival analysis. When adjusted for well known risk factors for IgAN progression, ACR was most significantly associated with the composite end point (hazard ratio, 1.56 per 1-SD change of standard normalized square root–transformed ACR; 95% confidence interval, 1.29 to 1.89; P<0.001). Compared with protein-to-creatinine ratio and 24-hour UPE, addition of ACR to traditional risk factors resulted in more improvement in the predictive ability of IgAN progression (c statistic: ACR=0.70; protein-to-creatinine ratio =0.68; 24-hour UPE =0.69; Akaike information criterion: ACR=1217.85; protein-to-creatinine ratio =1229.28; 24-hour UPE =1234.96; P<0.001). Conclusions In IgAN, ACR, protein-to-creatinine ratio, and 24-hour UPE had comparable association with severe clinical and histologic findings. Compared with protein-to-creatinine ratio and 24-hour UPE, ACR showed slightly better performance in predicting IgAN progression. PMID:27026518

Tick-borne encephalitis virus infects human brain microvascular endothelial cells without compromising blood-brain barrier integrity.

PubMed

Palus, Martin; Vancova, Marie; Sirmarova, Jana; Elsterova, Jana; Perner, Jan; Ruzek, Daniel

2017-07-01

Alteration of the blood-brain barrier (BBB) is a hallmark of tick-borne encephalitis (TBE), a life-threating human viral neuroinfection. However, the mechanism of BBB breakdown during TBE, as well as TBE virus (TBEV) entry into the brain is unclear. Here, primary human microvascular endothelial cells (HBMECs) were infected with TBEV to study interactions with the BBB. Although the number of infected cells was relatively low in culture (<5%), the infection was persistent with high TBEV yields (>10 6 pfu/ml). Infection did not induce any significant changes in the expression of key tight junction proteins or upregulate the expression of cell adhesion molecules, and did not alter the highly organized intercellular junctions between HBMECs. In an in vitro BBB model, the virus crossed the BBB via a transcellular pathway without compromising the integrity of the cell monolayer. The results indicate that HBMECs may support TBEV entry into the brain without altering BBB integrity. Copyright © 2017 Elsevier Inc. All rights reserved.

Carcinoma-risk variant of EBNA1 deregulates Epstein-Barr Virus episomal latency.

PubMed

Dheekollu, Jayaraju; Malecka, Kimberly; Wiedmer, Andreas; Delecluse, Henri-Jacques; Chiang, Alan K S; Altieri, Dario C; Messick, Troy E; Lieberman, Paul M

2017-01-31

Epstein-Barr Virus (EBV) latent infection is a causative co-factor for endemic Nasopharyngeal Carcinoma (NPC). NPC-associated variants have been identified in EBV-encoded nuclear antigen EBNA1. Here, we solve the X-ray crystal structure of an NPC-derived EBNA1 DNA binding domain (DBD) and show that variant amino acids are found on the surface away from the DNA binding interface. We show that NPC-derived EBNA1 is compromised for DNA replication and episome maintenance functions. Recombinant virus containing the NPC EBNA1 DBD are impaired in their ability to immortalize primary B-lymphocytes and suppress lytic transcription during early stages of B-cell infection. We identify Survivin as a host protein deficiently bound by the NPC variant of EBNA1 and show that Survivin depletion compromises EBV episome maintenance in multiple cell types. We propose that endemic variants of EBNA1 play a significant role in EBV-driven carcinogenesis by altering key regulatory interactions that destabilize latent infection.

Ran-dependent TPX2 activation promotes acentrosomal microtubule nucleation in neurons

PubMed Central

Chen, Wen-Shin; Chen, Yi-Ju; Huang, Yung-An; Hsieh, Bing-Yuan; Chiu, Ho-Chieh; Kao, Pei-Ying; Chao, Chih-Yuan; Hwang, Eric

2017-01-01

The microtubule (MT) cytoskeleton is essential for the formation of morphologically appropriate neurons. The existence of the acentrosomal MT organizing center in neurons has been proposed but its identity remained elusive. Here we provide evidence showing that TPX2 is an important component of this acentrosomal MT organizing center. First, neurite elongation is compromised in TPX2-depleted neurons. In addition, TPX2 localizes to the centrosome and along the neurite shaft bound to MTs. Depleting TPX2 decreases MT formation frequency specifically at the tip and the base of the neurite, and these correlate precisely with the regions where active GTP-bound Ran proteins are enriched. Furthermore, overexpressing the downstream effector of Ran, importin, compromises MT formation and neuronal morphogenesis. Finally, applying a Ran-importin signaling interfering compound phenocopies the effect of TPX2 depletion on MT dynamics. Together, these data suggest a model in which Ran-dependent TPX2 activation promotes acentrosomal MT nucleation in neurons. PMID:28205572

Vegetarian low-protein diets supplemented with keto analogues: a niche for the few or an option for many?

PubMed

Piccoli, Giorgina B; Ferraresi, Martina; Deagostini, Maria C; Vigotti, Federica Neve; Consiglio, Valentina; Scognamiglio, Stefania; Moro, Irene; Clari, Roberta; Fassio, Federica; Biolcati, Marilisa; Porpiglia, Francesco

2013-09-01

Low-protein diets are often mentioned but seldom used to slow chronic kidney disease (CKD) progression. The aim of the study was to investigate the potential for implementation of a simplified low-protein diet supplemented with alpha-keto analogues (LPD-KA) as part of the routine work-up in CKD patients. In an implementation study (December 2007-November 2011), all patients with CKD Stages IV-V not on dialysis, rapidly progressive Stage III and/or refractory proteinuria, were offered either a simplified LPD-KA, or commercially available low-protein food. LPD-KA consisted of proteins 0.6 g/kg/day, supplementation with Ketosteril 1 pill/10 Kg, 1-3 free-choice meals/week and a simplified schema based on 'allowed' and 'forbidden' foods. 'Success' was defined as at least 6 months on LPD-KA. Progression was defined as reduction in glomerular filtration rate (GFR)[(Chronic Kidney Disease Epidemiology Collaboration) formula CKD-EPI] in patients with at least 6 months of follow-up. Of about 2500 patients referred (8% CKD Stages IV-V), 139 started LPD-KA; median age (70 years) and prevalence of comorbidity (79%) were in line with the dialysis population. Start of dialysis was the main reason for discontinuation (40 cases, unplanned in 7); clinical reasons were recorded in 7, personal preference in 14 and improvement and death in 8 each. The low gross mortality (4% per year) and the progression rate (from -8 to 0 mL/min/year at 6 months) are reassuring concerning safety. None of the baseline conditions, including age, educational level, comorbidity or kidney function, discriminated the patients who followed the diet for at least 6 months. Our data suggest a wider offer of LPD-KA to patients with severe and progressive CKD. The promising results in terms of mortality and progression need confirmation with different study designs.

Electrocatalyzed O2 response of myoglobin immobilized on multi-walled carbon nanotube forest electrodes.

PubMed

Pacios, M; del Valle, M; Bartroli, J; Esplandiu, M J

2009-10-01

Direct electrochemistry and activity of myoglobin (Mb) immobilized on carbon nanotube (CNT) forest electrodes were investigated by probing mainly its electrocatalytical response towards oxygen. The protein was anchored on the CNT electrodes through carbodiimide coupling, which was shown to provide long term stability. The electrochemical response was monitored as a function of oxygen concentration and pH. Conformational changes together with the consequent loss of oxygen affinity were recorded at low pH, which delimits the operative range of the Mb/CNT electrodes for sensing purposes. In general, it can be concluded that CNT forests constitute suitable platforms for Mb attachment without compromising the protein bioactivity and by keeping at the same time the direct electron exchange with the heme core. All these characteristics confer to the protein modified CNT system promising properties for the implementation of (bio)sensor devices with impact in the clinical and environmental field.

iBodies: Modular Synthetic Antibody Mimetics Based on Hydrophilic Polymers Decorated with Functional Moieties.

PubMed

Šácha, Pavel; Knedlík, Tomáš; Schimer, Jiří; Tykvart, Jan; Parolek, Jan; Navrátil, Václav; Dvořáková, Petra; Sedlák, František; Ulbrich, Karel; Strohalm, Jiří; Majer, Pavel; Šubr, Vladimír; Konvalinka, Jan

2016-02-12

Antibodies are indispensable tools for biomedicine and anticancer therapy. Nevertheless, their use is compromised by high production costs, limited stability, and difficulty of chemical modification. The design and preparation of synthetic polymer conjugates capable of replacing antibodies in biomedical applications such as ELISA, flow cytometry, immunocytochemistry, and immunoprecipitation is reported. The conjugates, named "iBodies", consist of an HPMA copolymer decorated with low-molecular-weight compounds that function as targeting ligands, affinity anchors, and imaging probes. We prepared specific conjugates targeting several proteins with known ligands and used these iBodies for enzyme inhibition, protein isolation, immobilization, quantification, and live-cell imaging. Our data indicate that this highly modular and versatile polymer system can be used to produce inexpensive and stable antibody substitutes directed toward virtually any protein of interest with a known ligand. © 2016 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA.

iBodies: Modular Synthetic Antibody Mimetics Based on Hydrophilic Polymers Decorated with Functional Moieties

PubMed Central

Šácha, Pavel; Knedlík, Tomáš; Schimer, Jiří; Tykvart, Jan; Parolek, Jan; Navrátil, Václav; Dvořáková, Petra; Sedlák, František; Ulbrich, Karel; Strohalm, Jiří; Majer, Pavel

2016-01-01

Abstract Antibodies are indispensable tools for biomedicine and anticancer therapy. Nevertheless, their use is compromised by high production costs, limited stability, and difficulty of chemical modification. The design and preparation of synthetic polymer conjugates capable of replacing antibodies in biomedical applications such as ELISA, flow cytometry, immunocytochemistry, and immunoprecipitation is reported. The conjugates, named “iBodies”, consist of an HPMA copolymer decorated with low‐molecular‐weight compounds that function as targeting ligands, affinity anchors, and imaging probes. We prepared specific conjugates targeting several proteins with known ligands and used these iBodies for enzyme inhibition, protein isolation, immobilization, quantification, and live‐cell imaging. Our data indicate that this highly modular and versatile polymer system can be used to produce inexpensive and stable antibody substitutes directed toward virtually any protein of interest with a known ligand. PMID:26749427

Tricking the guard: exploiting plant defense for disease susceptibility.

PubMed

Lorang, J; Kidarsa, T; Bradford, C S; Gilbert, B; Curtis, M; Tzeng, S-C; Maier, C S; Wolpert, T J

2012-11-02

Typically, pathogens deploy virulence effectors to disable defense. Plants defeat effectors with resistance proteins that guard effector targets. We found that a pathogen exploits a resistance protein by activating it to confer susceptibility in Arabidopsis. The guard mechanism of plant defense is recapitulated by interactions among victorin (an effector produced by the necrotrophic fungus Cochliobolus victoriae), TRX-h5 (a defense-associated thioredoxin), and LOV1 (an Arabidopsis susceptibility protein). In LOV1's absence, victorin inhibits TRX-h5, resulting in compromised defense but not disease by C. victoriae. In LOV1's presence, victorin binding to TRX-h5 activates LOV1 and elicits a resistance-like response that confers disease susceptibility. We propose that victorin is, or mimics, a conventional pathogen virulence effector that was defeated by LOV1 and confers virulence to C. victoriae solely because it incites defense.

Modeling virus capsids and their protein binding -- the search for weak regions within the HIV capsid

NASA Astrophysics Data System (ADS)

Sankey, Otto; Benson, Daryn

2010-10-01

Viruses remain a threat to the health of humans worldwide with 33 million infected with AIDS. Viruses are ubiquitous infecting animals, plants, and bacteria. Each virus infects in its own unique manner making the problem seem intractable. However, some general physical steps apply to many viruses and the application of basic physical modeling can potentially have great impact. The aim of this theoretical study is to investigate the stability of the HIV viral capsid (protein shell). The structural shell can be compromised by physical probes such as pulsed laser light. But what are the weakest regions of the capsid so that we can begin to understand vulnerabilities of these deadly materials? The atomic structure of HIV capsids is not precisely known and we begin by describing our work to model the capsid structure. Next we describe a course grained model to investigate protein interactions within the capsid.

S100-A9 protein in exosomes from chronic lymphocytic leukemia cells promotes NF-κB activity during disease progression.

PubMed

Prieto, Daniel; Sotelo, Natalia; Seija, Noé; Sernbo, Sandra; Abreu, Cecilia; Durán, Rosario; Gil, Magdalena; Sicco, Estefanía; Irigoin, Victoria; Oliver, Carolina; Landoni, Ana Inés; Gabus, Raúl; Dighiero, Guillermo; Oppezzo, Pablo

2017-08-10

Chronic lymphocytic leukemia (CLL) is an incurable disease characterized by accumulation of clonal B lymphocytes, resulting from a complex balance between cell proliferation and apoptotic death. Continuous crosstalk between cancer cells and local/distant host environment is required for effective tumor growth. Among the main actors of this dynamic interplay between tumoral cells and their microenvironment are the nano-sized vesicles called exosomes. Emerging evidence indicates that secretion, composition, and functional capacity of exosomes are altered as tumors progress to an aggressive phenotype. In CLL, no data exist exploring the specific changes in the proteomic profile of plasma-derived exosomes from patients during disease evolution. We hereby report for the first time different proteomic profiles of plasma exosomes, both between indolent and progressive CLLs as well as within the individual patients at the onset of disease and during its progression. Next, we focus on the changes of the exosome protein cargoes, which are found exclusively in patients with progressive CLL after disease progression. The alterations in the proteomic cargoes underline different networks specific for leukemia progression related to inflammation, oxidative stress, and NF-κB and phosphatidylinositol 3-kinase/AKT pathway activation. Finally, our results suggest a preponderant role for the protein S100-A9 as an activator of the NFκB pathway during CLL progression and suggest that the leukemic clone can generate an autoactivation loop through S100-A9 expression, NF-κB activation, and exosome secretion. Collectively, our data propose a new pathway for NF-κB activation in CLL and highlight the importance of exosomes as extracellular mediators promoting tumor progression in CLL. © 2017 by The American Society of Hematology.

Aberrant p53 protein expression is associated with an increased risk of neoplastic progression in patients with Barrett's oesophagus.

PubMed

Kastelein, Florine; Biermann, Katharina; Steyerberg, Ewout W; Verheij, Joanne; Kalisvaart, Marit; Looijenga, Leendert H J; Stoop, Hans A; Walter, Laurens; Kuipers, Ernst J; Spaander, Manon C W; Bruno, Marco J

2013-12-01

The value of surveillance for patients with Barrett's oesophagus (BO) is under discussion given the overall low incidence of neoplastic progression and lack of discriminative tests for risk stratification. Histological diagnosis of low-grade dysplasia (LGD) is the only accepted predictor for progression to date, but has a low predictive value. The aim of this study was therefore to evaluate the value of p53 immunohistochemistry for predicting neoplastic progression in patients with BO. We conducted a case-control study within a prospective cohort of 720 patients with BO. Patients who developed high-grade dysplasia (HGD) or oesophageal adenocarcinoma (OAC) were classified as cases and patients without neoplastic progression were classified as controls. P53 protein expression was determined by immunohistochemistry in more than 12 000 biopsies from 635 patients and was scored independently by two expert pathologists who were blinded to long-term outcome. During follow-up, 49 (8%) patients developed HGD or OAC. P53 overexpression was associated with an increased risk of neoplastic progression in patients with BO after adjusting for age, gender, Barrett length and oesophagitis (adjusted relative risks (RR(a)) 5.6; 95% CI 3.1 to 10.3), but the risk was even higher with loss of p53 expression (RR(a) 14.0; 95% CI 5.3 to 37.2). The positive predictive value for neoplastic progression increased from 15% with histological diagnosis of LGD to 33% with LGD and concurrent aberrant p53 expression. Aberrant p53 protein expression is associated with an increased risk of neoplastic progression in patients with BO and appears to be a more powerful predictor of neoplastic progression than histological diagnosis of LGD.

Recent Progress in CFTR Interactome Mapping and Its Importance for Cystic Fibrosis.

PubMed

Lim, Sang Hyun; Legere, Elizabeth-Ann; Snider, Jamie; Stagljar, Igor

2017-01-01

Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) is a chloride channel found in secretory epithelia with a plethora of known interacting proteins. Mutations in the CFTR gene cause cystic fibrosis (CF), a disease that leads to progressive respiratory illness and other complications of phenotypic variance resulting from perturbations of this protein interaction network. Studying the collection of CFTR interacting proteins and the differences between the interactomes of mutant and wild type CFTR provides insight into the molecular machinery of the disease and highlights possible therapeutic targets. This mini review focuses on functional genomics and proteomics approaches used for systematic, high-throughput identification of CFTR-interacting proteins to provide comprehensive insight into CFTR regulation and function.

45 CFR 30.22 - Bases for compromise.

Code of Federal Regulations, 2011 CFR

2011-10-01

... 45 Public Welfare 1 2011-10-01 2011-10-01 false Bases for compromise. 30.22 Section 30.22 Public Welfare DEPARTMENT OF HEALTH AND HUMAN SERVICES GENERAL ADMINISTRATION CLAIMS COLLECTION Debt Compromise § 30.22 Bases for compromise. (a) Compromise. The Secretary may compromise a debt if the full amount...

45 CFR 30.22 - Bases for compromise.

Code of Federal Regulations, 2012 CFR

2012-10-01

... 45 Public Welfare 1 2012-10-01 2012-10-01 false Bases for compromise. 30.22 Section 30.22 Public Welfare DEPARTMENT OF HEALTH AND HUMAN SERVICES GENERAL ADMINISTRATION CLAIMS COLLECTION Debt Compromise § 30.22 Bases for compromise. (a) Compromise. The Secretary may compromise a debt if the full amount...

45 CFR 30.22 - Bases for compromise.

Code of Federal Regulations, 2013 CFR

2013-10-01

... 45 Public Welfare 1 2013-10-01 2013-10-01 false Bases for compromise. 30.22 Section 30.22 Public Welfare DEPARTMENT OF HEALTH AND HUMAN SERVICES GENERAL ADMINISTRATION CLAIMS COLLECTION Debt Compromise § 30.22 Bases for compromise. (a) Compromise. The Secretary may compromise a debt if the full amount...

45 CFR 30.22 - Bases for compromise.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 45 Public Welfare 1 2010-10-01 2010-10-01 false Bases for compromise. 30.22 Section 30.22 Public Welfare DEPARTMENT OF HEALTH AND HUMAN SERVICES GENERAL ADMINISTRATION CLAIMS COLLECTION Debt Compromise § 30.22 Bases for compromise. (a) Compromise. The Secretary may compromise a debt if the full amount...

Characterising the muscle anabolic potential of dairy, meat and plant-based protein sources in older adults.

PubMed

Gorissen, Stefan H M; Witard, Oliver C

2018-02-01

The age-related loss of skeletal muscle mass and function is caused, at least in part, by a reduced muscle protein synthetic response to protein ingestion. The magnitude and duration of the postprandial muscle protein synthetic response to ingested protein is dependent on the quantity and quality of the protein consumed. This review characterises the anabolic properties of animal-derived and plant-based dietary protein sources in older adults. While approximately 60 % of dietary protein consumed worldwide is derived from plant sources, plant-based proteins generally exhibit lower digestibility, lower leucine content and deficiencies in certain essential amino acids such as lysine and methionine, which compromise the availability of a complete amino acid profile required for muscle protein synthesis. Based on currently available scientific evidence, animal-derived proteins may be considered more anabolic than plant-based protein sources. However, the production and consumption of animal-derived protein sources is associated with higher greenhouse gas emissions, while plant-based protein sources may be considered more environmentally sustainable. Theoretically, the lower anabolic capacity of plant-based proteins can be compensated for by ingesting a greater dose of protein or by combining various plant-based proteins to provide a more favourable amino acid profile. In addition, leucine co-ingestion can further augment the postprandial muscle protein synthetic response. Finally, prior exercise or n-3 fatty acid supplementation have been shown to sensitise skeletal muscle to the anabolic properties of dietary protein. Applying one or more of these strategies may support the maintenance of muscle mass with ageing when diets rich in plant-based protein are consumed.

A Novel, Non-Apoptotic Role for Scythe/BAT3: A Functional Switch between the Pro- and Anti-Proliferative Roles of p21 during the Cell Cycle

PubMed Central

Yong, Sheila T.; Wang, Xiao-Fan

2012-01-01

Background Scythe/BAT3 is a member of the BAG protein family whose role in apoptosis has been extensively studied. However, since the developmental defects observed in Bat3-null mouse embryos cannot be explained solely by defects in apoptosis, we investigated whether BAT3 is also involved in cell-cycle progression. Methods/Principal Findings Using a stable-inducible Bat3-knockdown cellular system, we demonstrated that reduced BAT3 protein level causes a delay in both G1/S transition and G2/M progression. Concurrent with these changes in cell-cycle progression, we observed a reduction in the turnover and phosphorylation of the CDK inhibitor p21, which is best known as an inhibitor of DNA replication; however, phosphorylated p21 has also been shown to promote G2/M progression. Our findings indicate that in Bat3-knockdown cells, p21 continues to be synthesized during cell-cycle phases that do not normally require p21, resulting in p21 protein accumulation and a subsequent delay in cell-cycle progression. Finally, we showed that BAT3 co-localizes with p21 during the cell cycle and is required for the translocation of p21 from the cytoplasm to the nucleus during the G1/S transition and G2/M progression. Conclusion: Our study reveals a novel, non-apoptotic role for BAT3 in cell-cycle regulation. By maintaining a low p21 protein level during the G1/S transition, BAT3 counteracts the inhibitory effect of p21 on DNA replication and thus enables the cells to progress from G1 to S phase. Conversely, during G2/M progression, BAT3 facilitates p21 phosphorylation by cyclin A/Cdk2, an event required for G2/M progression. BAT3 modulates these pro- and anti-proliferative roles of p21 at least in part by regulating cyclin A abundance, as well as p21 translocation between the cytoplasm and the nucleus to ensure that it functions in the appropriate intracellular compartment during each phase of the cell cycle. PMID:22761665

Protein-protein docking with binding site patch prediction and network-based terms enhanced combinatorial scoring.

PubMed

Gong, Xinqi; Wang, Panwen; Yang, Feng; Chang, Shan; Liu, Bin; He, Hongqiu; Cao, Libin; Xu, Xianjin; Li, Chunhua; Chen, Weizu; Wang, Cunxin

2010-11-15

Protein-protein docking has made much progress in recent years, but challenges still exist. Here we present the application of our docking approach HoDock in CAPRI. In this approach, a binding site prediction is implemented to reduce docking sampling space and filter out unreasonable docked structures, and a network-based enhanced combinatorial scoring function HPNCscore is used to evaluate the decoys. The experimental information was combined with the predicted binding site to pick out the most likely key binding site residues. We applied the HoDock method in the recent rounds of the CAPRI experiments, and got good results as predictors on targets 39, 40, and 41. We also got good results as scorers on targets 35, 37, 40, and 41. This indicates that our docking approach can contribute to the progress of protein-protein docking methods and to the understanding of the mechanism of protein-protein interactions. © 2010 Wiley-Liss, Inc.

The Effects of Lipoplex Formulation Variables on the Protein Corona and Comparisons with in vitro Transfection Efficiency

PubMed Central

Betker, Jamie L.; Gomez, Joe; Anchordoquy, Thomas J.

2013-01-01

The use of lipoplexes for the intracellular delivery of nucleic acids typically involves the optimization of several parameters that are known to affect delivery. Researchers commonly vary charge ratio, and often incorporate different amounts of helper lipids (e.g., cholesterol) to optimize formulations for transfection in cell culture and in vivo. The results of such experiments are often interpreted in the context of nuclease resistance and cell association, but effects on the protein corona are usually not considered. While many studies have demonstrated that lipoplex structure and function can be dramatically compromised in the presence of serum, little attention has been paid to the adsorption of specific proteins and how this might be affected by formulation parameters. In this study, we characterize changes in the protein corona that occur as DOTAP-based lipoplexes are formulated with different amounts of cholesterol and prepared at different charge ratios. Our results demonstrate a significant effect of lipid composition on both total protein adsorption as well as the individual proteins from fetal calf serum that are associated with lipoplexes. In addition, we show that PEGylation increases protein adsorption with our formulations; effects that depend on the type of PEG conjugate employed in the lipoplex. Attempts to identify a specific protein responsible for enhancing transfection were unsuccessful. PMID:23920037

Acute supplementation of amino acids increases net protein accretion in IUGR fetal sheep

PubMed Central

Rozance, Paul J.; Thorn, Stephanie R.; Friedman, Jacob E.; Hay, William W.

2012-01-01

Placental insufficiency decreases fetal amino acid uptake from the placenta, plasma insulin concentrations, and protein accretion, thus compromising normal fetal growth trajectory. We tested whether acute supplementation of amino acids or insulin into the fetus with intrauterine growth restriction (IUGR) would increase net fetal protein accretion rates. Late-gestation IUGR and control (CON) fetal sheep received acute, 3-h infusions of amino acids (with euinsulinemia), insulin (with euglycemia and euaminoacidemia), or saline. Fetal leucine metabolism was measured under steady-state conditions followed by a fetal muscle biopsy to quantify insulin signaling. In CON, increasing amino acid delivery rates to the fetus by 100% increased leucine oxidation rates by 100%. In IUGR, amino acid infusion completely suppressed fetal protein breakdown rates but increased leucine oxidation rate by only 25%, resulting in increased protein accretion rates by 150%. Acute insulin infusion, however, had very little effect on amino acid delivery rates, fetal leucine disposal rates, or fetal protein accretion rates in CON or IUGR fetuses despite robust signaling of the fetal skeletal muscle insulin-signaling cascade. These results indicate that, when amino acids are given directly into the fetal circulation independently of changes in insulin concentrations, IUGR fetal sheep have suppressed protein breakdown rates, thus increasing net fetal protein accretion. PMID:22649066

Photosensitized UVA-Induced Cross-Linking between Human DNA Repair and Replication Proteins and DNA Revealed by Proteomic Analysis

PubMed Central

2016-01-01

Long wavelength ultraviolet radiation (UVA, 320–400 nm) interacts with chromophores present in human cells to induce reactive oxygen species (ROS) that damage both DNA and proteins. ROS levels are amplified, and the damaging effects of UVA are exacerbated if the cells are irradiated in the presence of UVA photosensitizers such as 6-thioguanine (6-TG), a strong UVA chromophore that is extensively incorporated into the DNA of dividing cells, or the fluoroquinolone antibiotic ciprofloxacin. Both DNA-embedded 6-TG and ciprofloxacin combine synergistically with UVA to generate high levels of ROS. Importantly, the extensive protein damage induced by these photosensitizer+UVA combinations inhibits DNA repair. DNA is maintained in intimate contact with the proteins that effect its replication, transcription, and repair, and DNA–protein cross-links (DPCs) are a recognized reaction product of ROS. Cross-linking of DNA metabolizing proteins would compromise these processes by introducing physical blocks and by depleting active proteins. We describe a sensitive and statistically rigorous method to analyze DPCs in cultured human cells. Application of this proteomics-based analysis to cells treated with 6-TG+UVA and ciprofloxacin+UVA identified proteins involved in DNA repair, replication, and gene expression among those most vulnerable to cross-linking under oxidative conditions. PMID:27654267

Effects of Reusing Gel Electrophoresis and Electrotransfer Buffers on Western Blotting.

PubMed

Heda, Ghanshyam D; Omotola, Oluwabukola B; Heda, Rajiv P; Avery, Jamie

2016-09-01

SDS-PAGE and Western blotting are 2 of the most commonly used biochemical methods for protein analysis. Proteins are electrophoretically separated based on their MWs by SDS-PAGE and then electrotransferred to a solid membrane surface for subsequent protein-specific analysis by immunoblotting, a procedure commonly known as Western blotting. Both of these procedures use a salt-based buffer, with the latter procedure consisting of methanol as an additive known for its toxicity. Previous reports present a contradictory view in favor or against reusing electrotransfer buffer, also known as Towbin's transfer buffer (TTB), with an aim to reduce the toxic waste. In this report, we present a detailed analysis of not only reusing TTB but also gel electrophoresis buffer (EB) on proteins of low to high MW range. Our results suggest that EB can be reused for at least 5 times without compromising the electrophoretic separation of mixture of proteins in an MW standard, BSA, and crude cell lysates. Additionally, reuse of EB did not affect the quality of subsequent Western blots. Successive reuse of TTB, on the other hand, diminished the signal of proteins of different MWs in a protein standard and a high MW membrane protein cystic fibrosis transmembrane-conductance regulator (CFTR) in Western blotting.

The endosomal protein CHARGED MULTIVESICULAR BODY PROTEIN1 regulates the autophagic turnover of plastids in Arabidopsis.

PubMed

Spitzer, Christoph; Li, Faqiang; Buono, Rafael; Roschzttardtz, Hannetz; Chung, Taijoon; Zhang, Min; Osteryoung, Katherine W; Vierstra, Richard D; Otegui, Marisa S

2015-02-01

Endosomal Sorting Complex Required for Transport (ESCRT)-III proteins mediate membrane remodeling and the release of endosomal intraluminal vesicles into multivesicular bodies. Here, we show that the ESCRT-III subunit paralogs CHARGED MULTIVESICULAR BODY PROTEIN1 (CHMP1A) and CHMP1B are required for autophagic degradation of plastid proteins in Arabidopsis thaliana. Similar to autophagy mutants, chmp1a chmp1b (chmp1) plants hyperaccumulated plastid components, including proteins involved in plastid division. The autophagy machinery directed the release of bodies containing plastid material into the cytoplasm, whereas CHMP1A and B were required for delivery of these bodies to the vacuole. Autophagy was upregulated in chmp1 as indicated by an increase in vacuolar green fluorescent protein (GFP) cleavage from the autophagic reporter GFP-ATG8. However, autophagic degradation of the stromal cargo RECA-GFP was drastically reduced in the chmp1 plants upon starvation, suggesting that CHMP1 mediates the efficient delivery of autophagic plastid cargo to the vacuole. Consistent with the compromised degradation of plastid proteins, chmp1 plastids show severe morphological defects and aberrant division. We propose that CHMP1 plays a direct role in the autophagic turnover of plastid constituents. © 2015 American Society of Plant Biologists. All rights reserved.

Argonaute: The executor of small RNA function.

PubMed

Azlan, Azali; Dzaki, Najat; Azzam, Ghows

2016-08-20

The discovery of small non-coding RNAs - microRNA (miRNA), short interfering RNA (siRNA) and PIWI-interacting RNA (piRNA) - represents one of the most exciting frontiers in biology specifically on the mechanism of gene regulation. In order to execute their functions, these small RNAs require physical interactions with their protein partners, the Argonaute (AGO) family proteins. Over the years, numerous studies have made tremendous progress on understanding the roles of AGO in gene silencing in various organisms. In this review, we summarize recent progress of AGO-mediated gene silencing and other cellular processes in which AGO proteins have been implicated with a particular focus on progress made in flies, humans and other model organisms as compliment. Copyright © 2016 Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, and Genetics Society of China. Published by Elsevier Ltd. All rights reserved.

Integration of the ubiquitin-proteasome pathway with a cytosolic oligopeptidase activity

PubMed Central

Wang, Evelyn W.; Kessler, Benedikt M.; Borodovsky, Anna; Cravatt, Benjamin F.; Bogyo, Matthew; Ploegh, Hidde L.; Glas, Rickard

2000-01-01

Cytosolic proteolysis is carried out predominantly by the proteasome. We show that a large oligopeptidase, tripeptidylpeptidase II (TPPII), can compensate for compromised proteasome activity. Overexpression of TPPII is sufficient to prevent accumulation of polyubiquitinated proteins and allows survival of EL-4 cells at otherwise lethal concentrations of the covalent proteasome inhibitor NLVS (NIP-leu-leu-leu-vinylsulfone). Elevated TPPII activity also partially restores peptide loading of MHC molecules. Purified proteasomes from adapted cells lack the chymotryptic-like activity, but still degrade longer peptide substrates via residual activity of their Z subunits. However, growth of adapted cells depends on induction of other proteolytic activities. Therefore, cytosolic oligopeptidases such as TPPII normalize rates of intracellular protein breakdown required for normal cellular function and viability. PMID:10954757

Abfraction lesions: etiology, diagnosis, and treatment options

PubMed Central

Nascimento, Marcelle M; Dilbone, Deborah A; Pereira, Patricia NR; Duarte, Wagner R; Geraldeli, Saulo; Delgado, Alex J

2016-01-01

Abfraction is a type of noncarious cervical lesion (NCCL) characterized by loss of tooth tissues with different clinical appearances. Evidence supports that abfraction lesions, as any NCCLs, have a multifactorial etiology. Particularly, the cervical wear of abfraction can occur as a result of normal and abnormal tooth function and may also be accompanied by pathological wear, such as abrasion and erosion. The interaction between chemical, biological, and behavioral factors is critical and helps to explain why some individuals exhibit more than one type of cervical wear mechanism than others. In an era of personalized dentistry, patient risk factors for NCCLs must be identified and addressed before any treatment is performed. Marked variations exist in dental practice concerning the diagnosis and management of these lesions. The lack of understanding about the prognosis of these lesions with or without intervention may be a major contributor to variations in dentists’ management decisions. This review focuses on the current knowledge and available treatment strategies for abfraction lesions. By recognizing that progressive changes in the cervical area of the tooth are part of a physiologically dynamic process that occurs with aging, premature and unnecessary intervention can be avoided. In cases of asymptomatic teeth, where tooth vitality and function are not compromised, abfraction lesions should be monitored for at least 6 months before any invasive procedure is planned. In cases of abfraction associated with gingival recession, a combined restorative-surgical approach may be performed. Restorative intervention and occlusal adjustment are not indicated as treatment options to prevent further tooth loss or progression of abfraction. The clinical decision to restore abfraction lesions may be based on the need to replace form and function or to relieve hypersensitivity of severely compromised teeth or for esthetic reasons. PMID:27217799

Crown-of-thorns starfish predation and physical injuries promote brown band disease on corals

NASA Astrophysics Data System (ADS)

Katz, Sefano M.; Pollock, F. Joseph; Bourne, David G.; Willis, Bette L.

2014-09-01

Brown band (BrB) disease manifests on corals as a ciliate-dominated lesion that typically progresses rapidly causing extensive mortality, but it is unclear whether the dominant ciliate Porpostoma guamense is a primary or an opportunistic pathogen, the latter taking advantage of compromised coral tissue or depressed host resistance. In this study, manipulative aquarium-based experiments were used to investigate the role of P. guamense as a pathogen when inoculated onto fragments of the coral Acropora hyacinthus that were either healthy, preyed on by Acanthaster planci (crown-of-thorns starfish; COTS), or experimentally injured. Following ciliate inoculation, BrB lesions developed on all of COTS-predated fragments ( n = 9 fragments) and progressed up to 4.6 ± 0.3 cm d-1, resulting in ~70 % of coral tissue loss after 4 d. Similarly, BrB lesions developed rapidly on experimentally injured corals and ~38 % of coral tissue area was lost 60 h after inoculation. In contrast, no BrB lesions were observed on healthy corals following experimental inoculations. A choice experiment demonstrated that ciliates are strongly attracted to physically injured corals, with over 55 % of inoculated ciliates migrating to injured corals and forming distinct lesions, whereas ciliates did not migrate to healthy corals. Our results indicate that ciliates characteristic of BrB disease are opportunistic pathogens that rapidly migrate to and colonise compromised coral tissue, leading to rapid coral mortality, particularly following predation or injury. Predicted increases in tropical storms, cyclones, and COTS outbreaks are likely to increase the incidence of coral injury in the near future, promoting BrB disease and further contributing to declines in coral cover.

Identification of Unstable Network Modules Reveals Disease Modules Associated with the Progression of Alzheimer’s Disease

PubMed Central

Kikuchi, Masataka; Ogishima, Soichi; Miyamoto, Tadashi; Miyashita, Akinori; Kuwano, Ryozo; Nakaya, Jun; Tanaka, Hiroshi

2013-01-01

Alzheimer’s disease (AD), the most common cause of dementia, is associated with aging, and it leads to neuron death. Deposits of amyloid β and aberrantly phosphorylated tau protein are known as pathological hallmarks of AD, but the underlying mechanisms have not yet been revealed. A high-throughput gene expression analysis previously showed that differentially expressed genes accompanying the progression of AD were more down-regulated than up-regulated in the later stages of AD. This suggested that the molecular networks and their constituent modules collapsed along with AD progression. In this study, by using gene expression profiles and protein interaction networks (PINs), we identified the PINs expressed in three brain regions: the entorhinal cortex (EC), hippocampus (HIP) and superior frontal gyrus (SFG). Dividing the expressed PINs into modules, we examined the stability of the modules with AD progression and with normal aging. We found that in the AD modules, the constituent proteins, interactions and cellular functions were not maintained between consecutive stages through all brain regions. Interestingly, the modules were collapsed with AD progression, specifically in the EC region. By identifying the modules that were affected by AD pathology, we found the transcriptional regulation-associated modules that interact with the proteasome-associated module via UCHL5 hub protein, which is a deubiquitinating enzyme. Considering PINs as a system made of network modules, we found that the modules relevant to the transcriptional regulation are disrupted in the EC region, which affects the ubiquitin-proteasome system. PMID:24348898

24 CFR 17.73 - Standards for compromise of claims.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 24 Housing and Urban Development 1 2010-04-01 2010-04-01 false Standards for compromise of claims... General Provisions § 17.73 Standards for compromise of claims. (a) Compromise offer. An offer to...) Documentary evidence of compromise. No compromise of a claim shall be final or binding on the Department...

Two overlapping domains of a lyssavirus matrix protein that acts on different cell death pathways.

PubMed

Larrous, Florence; Gholami, Alireza; Mouhamad, Shahul; Estaquier, Jérôme; Bourhy, Hervé

2010-10-01

The lyssavirus matrix (M) protein induces apoptosis. The regions of the M protein that are essential for triggering cell death pathways are not yet clearly defined. We therefore compared the M proteins from two viruses that have contrasting characteristics in terms of cellular apoptosis: a genotype 3 lyssavirus, Mokola virus (MOK), and a genotype 1 rabies virus isolated from a dog from Thailand (THA). We identified a 20-amino-acid fragment (corresponding to positions 67 to 86) that retained the cell death activities of the full-length M protein from MOK via both the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and inhibition of cytochrome c oxidase (CcO) activity. We found that the amino acids at positions 77 and 81 have an essential role in triggering these two cell death pathways. Directed mutagenesis demonstrated that the amino acid at position 77 affects CcO activity, whereas the amino acid at position 81 affects TRAIL-dependent apoptosis. Mutations in the full-length M protein that compromised induction of either of these two pathways resulted in delayed apoptosis compared with the time to apoptosis for the nonmutated control.

Two Overlapping Domains of a Lyssavirus Matrix Protein That Acts on Different Cell Death Pathways ▿

PubMed Central

Larrous, Florence; Gholami, Alireza; Mouhamad, Shahul; Estaquier, Jérôme; Bourhy, Hervé

2010-01-01

The lyssavirus matrix (M) protein induces apoptosis. The regions of the M protein that are essential for triggering cell death pathways are not yet clearly defined. We therefore compared the M proteins from two viruses that have contrasting characteristics in terms of cellular apoptosis: a genotype 3 lyssavirus, Mokola virus (MOK), and a genotype 1 rabies virus isolated from a dog from Thailand (THA). We identified a 20-amino-acid fragment (corresponding to positions 67 to 86) that retained the cell death activities of the full-length M protein from MOK via both the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and inhibition of cytochrome c oxidase (CcO) activity. We found that the amino acids at positions 77 and 81 have an essential role in triggering these two cell death pathways. Directed mutagenesis demonstrated that the amino acid at position 77 affects CcO activity, whereas the amino acid at position 81 affects TRAIL-dependent apoptosis. Mutations in the full-length M protein that compromised induction of either of these two pathways resulted in delayed apoptosis compared with the time to apoptosis for the nonmutated control. PMID:20631119

Maternal protein-energy malnutrition during early pregnancy in sheep impacts the fetal ornithine cycle to reduce fetal kidney microvascular development.

PubMed

Dunford, Louise J; Sinclair, Kevin D; Kwong, Wing Y; Sturrock, Craig; Clifford, Bethan L; Giles, Tom C; Gardner, David S

2014-11-01

This paper identifies a common nutritional pathway relating maternal through to fetal protein-energy malnutrition (PEM) and compromised fetal kidney development. Thirty-one twin-bearing sheep were fed either a control (n=15) or low-protein diet (n=16, 17 vs. 8.7 g crude protein/MJ metabolizable energy) from d 0 to 65 gestation (term, ∼ 145 d). Effects on the maternal and fetal nutritional environment were characterized by sampling blood and amniotic fluid. Kidney development was characterized by histology, immunohistochemistry, vascular corrosion casts, and molecular biology. PEM had little measureable effect on maternal and fetal macronutrient balance (glucose, total protein, total amino acids, and lactate were unaffected) or on fetal growth. PEM decreased maternal and fetal urea concentration, which blunted fetal ornithine availability and affected fetal hepatic polyamine production. For the first time in a large animal model, we associated these nutritional effects with reduced micro- but not macrovascular development in the fetal kidney. Maternal PEM specifically impacts the fetal ornithine cycle, affecting cellular polyamine metabolism and microvascular development of the fetal kidney, effects that likely underpin programming of kidney development and function by a maternal low protein diet. © FASEB.

Oxidative Damage to RPA Limits the Nucleotide Excision Repair Capacity of Human Cells.

PubMed

Guven, Melisa; Brem, Reto; Macpherson, Peter; Peacock, Matthew; Karran, Peter

2015-11-01

Nucleotide excision repair (NER) protects against sunlight-induced skin cancer. Defective NER is associated with photosensitivity and a high skin cancer incidence. Some clinical treatments that cause photosensitivity can also increase skin cancer risk. Among these, the immunosuppressant azathioprine and the fluoroquinolone antibiotics ciprofloxacin and ofloxacin interact with UVA radiation to generate reactive oxygen species that diminish NER capacity by causing protein damage. The replication protein A (RPA) DNA-binding protein has a pivotal role in DNA metabolism and is an essential component of NER. The relationship between protein oxidation and NER inhibition was investigated in cultured human cells expressing different levels of RPA. We show here that RPA is limiting for NER and that oxidative damage to RPA compromises NER capability. Our findings reveal that cellular RPA is surprisingly vulnerable to oxidation, and we identify oxidized forms of RPA that are associated with impaired NER. The vulnerability of NER to inhibition by oxidation provides a connection between cutaneous photosensitivity, protein damage, and increased skin cancer risk. Our findings emphasize that damage to DNA repair proteins, as well as to DNA itself, is likely to be an important contributor to skin cancer risk.

Rho/ROCK signaling in regulation of corneal epithelial cell cycle progression.

PubMed

Chen, Jian; Guerriero, Emily; Lathrop, Kira; SundarRaj, Nirmala

2008-01-01

The authors' previous study showed that the expression of a Rho-associated serine/threonine kinase (ROCK) is regulated during cell cycle progression in corneal epithelial cells. The present study was conducted to determine whether and how Rho/ROCK signaling regulates cell cycle progression. Rabbit corneal epithelial cells (RCECs) in culture were arrested in the G(0) phase of the cell cycle by serum deprivation and then allowed to re-enter the cell cycle in the presence or absence of the ROCK inhibitor (Y27632) in serum-supplemented medium. The number of cells in the S phase, the relative levels of specific cyclins and CDKs and their intracellular distribution, and the relative levels of mRNAs were determined by BrdU labeling, Western blot and immunocytochemical analyses, and real-time RT-PCR, respectively. ROCK inhibition delayed the progression of G(1) to S phase and led to a decrease in the number of RCECs entering the S phase between 12 and 24 hours from 31.5% +/- 4.5% to 8.1% +/- 2.6%. During the cell cycle progression, protein and mRNA levels of cyclin-D1 and -D3 and cyclin-dependent kinases CDK4 and CDK6 were significantly lower, whereas the protein levels of the CDK inhibitor p27(Kip1) were higher in ROCK-inhibited cells. Intracellular mRNA or protein levels of cyclin-E and protein levels of CDK2 were not significantly affected, but their nuclear translocation was delayed by ROCK inhibition. ROCK signaling is involved in cell cycle progression in RCECs, possibly by upregulation of cyclin-D1 and -D3 and CDK4, -6, and -2; nuclear translocation of CDK2 and cyclin-E; and downregulation of p27(Kip1).

The Barley Powdery Mildew Candidate Secreted Effector Protein CSEP0105 Inhibits the Chaperone Activity of a Small Heat Shock Protein1[OPEN

PubMed Central

Ahmed, Ali Abdurehim; Pedersen, Carsten; Schultz-Larsen, Torsten; Kwaaitaal, Mark; Jørgensen, Hans Jørgen Lyngs; Thordal-Christensen, Hans

2015-01-01

Pathogens secrete effector proteins to establish a successful interaction with their host. Here, we describe two barley (Hordeum vulgare) powdery mildew candidate secreted effector proteins, CSEP0105 and CSEP0162, which contribute to pathogen success and appear to be required during or after haustorial formation. Silencing of either CSEP using host-induced gene silencing significantly reduced the fungal haustorial formation rate. Interestingly, both CSEPs interact with the barley small heat shock proteins, Hsp16.9 and Hsp17.5, in a yeast two-hybrid assay. Small heat shock proteins are known to stabilize several intracellular proteins, including defense-related signaling components, through their chaperone activity. CSEP0105 and CSEP0162 localized to the cytosol and the nucleus of barley epidermal cells, whereas Hsp16.9 and Hsp17.5 are cytosolic. Intriguingly, only those specific CSEPs changed localization and became restricted to the cytosol when coexpressed with Hsp16.9 and Hsp17.5, confirming the CSEP-small heat shock protein interaction. As predicted, Hsp16.9 showed chaperone activity, as it could prevent the aggregation of Escherichia coli proteins during thermal stress. Remarkably, CSEP0105 compromised this activity. These data suggest that CSEP0105 promotes virulence by interfering with the chaperone activity of a barley small heat shock protein essential for defense and stress responses. PMID:25770154

Complement factor H-related proteins in IgA nephropathy-sometimes a gentle nudge does the trick.

PubMed

Thurman, Joshua M; Laskowski, Jennifer

2017-10-01

Complement activation probably contributes to glomerular inflammation and damage in IgA nephropathy. In this issue, 2 groups report that levels of factor H-related protein 1 are elevated in patients with IgA nephropathy and correlate with disease progression. These studies provide new evidence that the complement cascade is important to the pathogenesis of this disease. These results also suggest that factor H-related protein 1 levels may be useful for identifying those patients at high risk of disease progression. Copyright © 2017 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

Protein Malnutrition Modifies Innate Immunity and Gene Expression by Intestinal Epithelial Cells and Human Rotavirus Infection in Neonatal Gnotobiotic Pigs

PubMed Central

Paim, Francine C.; Kandasamy, Sukumar; Alhamo, Moyasar A.; Fischer, David D.; Langel, Stephanie N.; Deblais, Loic; Kumar, Anand; Chepngeno, Juliet; Shao, Lulu; Huang, Huang-Chi; Candelero-Rueda, Rosario A.; Rajashekara, Gireesh

2017-01-01

ABSTRACT Malnutrition affects millions of children in developing countries, compromising immunity and contributing to increased rates of death from infectious diseases. Rotavirus is a major etiological agent of childhood diarrhea in developing countries, where malnutrition is prevalent. However, the interactions between the two and their combined effects on immune and intestinal functions are poorly understood. In this study, we used neonatal gnotobiotic (Gn) pigs transplanted with the fecal microbiota of a healthy 2-month-old infant (HIFM) and fed protein-deficient or -sufficient bovine milk diets. Protein deficiency induced hypoproteinemia, hypoalbuminemia, hypoglycemia, stunting, and generalized edema in Gn pigs, as observed in protein-malnourished children. Irrespective of the diet, human rotavirus (HRV) infection early, at HIFM posttransplantation day 3 (PTD3), resulted in adverse health effects and higher mortality rates (45 to 75%) than later HRV infection (PTD10). Protein malnutrition exacerbated HRV infection and affected the morphology and function of the small intestinal epithelial barrier. In pigs infected with HRV at PTD10, there was a uniform decrease in the function and/or frequencies of natural killer cells, plasmacytoid dendritic cells, and CD103+ and apoptotic mononuclear cells and altered gene expression profiles of intestinal epithelial cells (chromogranin A, mucin 2, proliferating cell nuclear antigen, SRY-Box 9, and villin). Thus, we have established the first HIFM-transplanted neonatal pig model that recapitulates major aspects of protein malnutrition in children and can be used to evaluate physiologically relevant interventions. Our findings provide an explanation of why nutrient-rich diets alone may lack efficacy in malnourished children. IMPORTANCE Malnutrition and rotavirus infection, prevalent in developing countries, individually and in combination, affect the health of millions of children, compromising their immunity and increasing the rates of death from infectious diseases. However, the interactions between the two and their combined effects on immune and intestinal functions are poorly understood. We have established the first human infant microbiota-transplanted neonatal pig model of childhood malnutrition that reproduced the impaired immune, intestinal, and other physiological functions seen in malnourished children. This model can be used to evaluate relevant dietary and other health-promoting interventions. Our findings provide an explanation of why adequate nutrition alone may lack efficacy in malnourished children. PMID:28261667

Genetic and physical interactions between factors involved in both cell cycle progression and pre-mRNA splicing in Saccharomyces cerevisiae.

PubMed Central

Ben-Yehuda, S; Dix, I; Russell, C S; McGarvey, M; Beggs, J D; Kupiec, M

2000-01-01

The PRP17/CDC40 gene of Saccharomyces cerevisiae functions in two different cellular processes: pre-mRNA splicing and cell cycle progression. The Prp17/Cdc40 protein participates in the second step of the splicing reaction and, in addition, prp17/cdc40 mutant cells held at the restrictive temperature arrest in the G2 phase of the cell cycle. Here we describe the identification of nine genes that, when mutated, show synthetic lethality with the prp17/cdc40Delta allele. Six of these encode known splicing factors: Prp8p, Slu7p, Prp16p, Prp22p, Slt11p, and U2 snRNA. The other three, SYF1, SYF2, and SYF3, represent genes also involved in cell cycle progression and in pre-mRNA splicing. Syf1p and Syf3p are highly conserved proteins containing several copies of a repeated motif, which we term RTPR. This newly defined motif is shared by proteins involved in RNA processing and represents a subfamily of the known TPR (tetratricopeptide repeat) motif. Using two-hybrid interaction screens and biochemical analysis, we show that the SYF gene products interact with each other and with four other proteins: Isy1p, Cef1p, Prp22p, and Ntc20p. We discuss the role played by these proteins in splicing and cell cycle progression. PMID:11102353

Genetic and physical interactions between factors involved in both cell cycle progression and pre-mRNA splicing in Saccharomyces cerevisiae.

PubMed

Ben-Yehuda, S; Dix, I; Russell, C S; McGarvey, M; Beggs, J D; Kupiec, M

2000-12-01

The PRP17/CDC40 gene of Saccharomyces cerevisiae functions in two different cellular processes: pre-mRNA splicing and cell cycle progression. The Prp17/Cdc40 protein participates in the second step of the splicing reaction and, in addition, prp17/cdc40 mutant cells held at the restrictive temperature arrest in the G2 phase of the cell cycle. Here we describe the identification of nine genes that, when mutated, show synthetic lethality with the prp17/cdc40Delta allele. Six of these encode known splicing factors: Prp8p, Slu7p, Prp16p, Prp22p, Slt11p, and U2 snRNA. The other three, SYF1, SYF2, and SYF3, represent genes also involved in cell cycle progression and in pre-mRNA splicing. Syf1p and Syf3p are highly conserved proteins containing several copies of a repeated motif, which we term RTPR. This newly defined motif is shared by proteins involved in RNA processing and represents a subfamily of the known TPR (tetratricopeptide repeat) motif. Using two-hybrid interaction screens and biochemical analysis, we show that the SYF gene products interact with each other and with four other proteins: Isy1p, Cef1p, Prp22p, and Ntc20p. We discuss the role played by these proteins in splicing and cell cycle progression.

Interactome Mapping Guided by Tissue-Specific Phosphorylation in Age-Related Macular Degeneration

PubMed Central

Sripathi, Srinivas R.; He, Weilue; Prigge, Cameron L.; Sylvester, O’Donnell; Um, Ji-Yeon; Powell, Folami L.; Neksumi, Musa; Bernstein, Paul S.; Choo, Dong-Won; Bartoli, Manuela; Gutsaeva, Diana R.; Jahng, Wan Jin

2017-01-01

The current study aims to determine the molecular mechanisms of age-related macular degeneration (AMD) using the phosphorylation network. Specifically, we examined novel biomarkers for oxidative stress by protein interaction mapping using in vitro and in vivo models that mimic the complex and progressive characteristics of AMD. We hypothesized that the early apoptotic reactions could be initiated by protein phosphorylation in region-dependent (peripheral retina vs. macular) and tissue-dependent (retinal pigment epithelium vs. retina) manner under chronic oxidative stress. The analysis of protein interactome and oxidative biomarkers showed the presence of tissue- and region-specific post-translational mechanisms that contribute to AMD progression and suggested new therapeutic targets that include ubiquitin, erythropoietin, vitronectin, MMP2, crystalline, nitric oxide, and prohibitin. Phosphorylation of specific target proteins in RPE cells is a central regulatory mechanism as a survival tool under chronic oxidative imbalance. The current interactome map demonstrates a positive correlation between oxidative stress-mediated phosphorylation and AMD progression and provides a basis for understanding oxidative stress-induced cytoskeletal changes and the mechanism of aggregate formation induced by protein phosphorylation. This information could provide an effective therapeutic approach to treat age-related neurodegeneration. PMID:28580316

Interactome Mapping Guided by Tissue-Specific Phosphorylation in Age-Related Macular Degeneration.

PubMed

Sripathi, Srinivas R; He, Weilue; Prigge, Cameron L; Sylvester, O'Donnell; Um, Ji-Yeon; Powell, Folami L; Neksumi, Musa; Bernstein, Paul S; Choo, Dong-Won; Bartoli, Manuela; Gutsaeva, Diana R; Jahng, Wan Jin

2017-02-01

The current study aims to determine the molecular mechanisms of age-related macular degeneration (AMD) using the phosphorylation network. Specifically, we examined novel biomarkers for oxidative stress by protein interaction mapping using in vitro and in vivo models that mimic the complex and progressive characteristics of AMD. We hypothesized that the early apoptotic reactions could be initiated by protein phosphorylation in region-dependent (peripheral retina vs. macular) and tissue-dependent (retinal pigment epithelium vs. retina) manner under chronic oxidative stress. The analysis of protein interactome and oxidative biomarkers showed the presence of tissue- and region-specific post-translational mechanisms that contribute to AMD progression and suggested new therapeutic targets that include ubiquitin, erythropoietin, vitronectin, MMP2, crystalline, nitric oxide, and prohibitin. Phosphorylation of specific target proteins in RPE cells is a central regulatory mechanism as a survival tool under chronic oxidative imbalance. The current interactome map demonstrates a positive correlation between oxidative stress-mediated phosphorylation and AMD progression and provides a basis for understanding oxidative stress-induced cytoskeletal changes and the mechanism of aggregate formation induced by protein phosphorylation. This information could provide an effective therapeutic approach to treat age-related neurodegeneration.

The phallic castration complex and primary femininity: paired developmental lines toward female gender identity.

PubMed

Mayer, E L

1995-01-01

I suggest that two developmental lines contribute to the achievement of female gender identity. One is rooted in the phallic castration complex, and the other in primary femininity. Far from being mutually exclusive, the two comprise necessary aspects of every girl's progress toward becoming a woman. To that extent, every woman's analysis will include the analysis of compromise formations that emerge from both. In distinguishing clinical manifestations of each developmental line, I suggest that it may be useful to conceptualize primary femininity and the phallic castration complex as affect-defense configurations which incorporate two fundamentally different ideas about danger. In conflicts of primary femininity, danger is anticipated: anxiety is the signal for compromise formation, since what is actually possessed (the female genital) is valued and is therefore imagined as subject to danger. In the phallic castration complex, danger is imagined already to have occurred. Depressive affect becomes the primary motive for defense, based on a fantasy that what is valued (the male genital) has already been lost. This distinction may facilitate our efforts to specify exactly how recent revisions in theories of female development have explicit implications for practice.

Induction of Oral Tolerance with Transgenic Plants Expressing Antigens for Prevention/Treatment of Autoimmune, Allergic and Inflammatory Diseases.

PubMed

Ma, Shengwu; Liao, Yu-Cai; Jevnikar, Anthony M

2015-01-01

The prevalence and incidence of autoimmune and allergic diseases have increased dramatically over the last several decades, especially in the developed world. The treatment of autoimmune and allergic diseases is typically with the use of non-specific immunosuppressive agents that compromise the integrity of the host immune system and therefore, increase the risk of infections. Antigenspecific immunotherapy by reinstating immunological tolerance towards self antigens without compromising immune functions is a much desired goal for the treatment of autoimmune and allergic diseases. Mucosal administration of antigen is a long-recognized method of inducing antigen-specific immune tolerance known as oral tolerance, which is viewed as having promising potential in the treatment of autoimmune and allergic diseases. Plant-based expression and delivery of recombinant antigens provide a promising new platform to induce oral tolerance, having considerable advantages including reduced cost and increased safety. Indeed, in recent years the use of tolerogenic plants for oral tolerance induction has attracted increasing attention, and considerable progress has been made. This review summarizes recent advances in using plants to deliver tolerogens for induction of oral tolerance in the treatment of autoimmune, allergic and inflammatory diseases.

ICPD beyond 2014: moving beyond missed opportunities and compromises in the fulfilment of sexual and reproductive health and rights.

PubMed

Sippel, Serra

2014-01-01

The 1994 International Conference on Population and Development (ICPD) in Cairo marked a paradigm shift that took family planning out of a population control context and into the broader context of sexual and reproductive health and rights (SRHR). While progress has been made with increased access to family planning and a decrease in maternal deaths, we have not seen practical results for the majority of women and girls worldwide, who still experience unacceptably high rates of maternal deaths, unmet contraceptive needs and HIV infections. Three of the compromises made by governments at Cairo - integration, reproductive rights and resource allocation - hindered the fulfilment of women's and girls' SRHR. The post-2015 agenda must ensure that economic development and global health interventions are linked at the national and global levels; family planning, HIV, maternal health and other reproductive health services are integrated and delivered through primary health settings; and access to safe and voluntary abortion services is recognised as a human right. Non-governmental organisations and donors must move beyond siloed issue areas to challenge governments, multilateral agencies, the financial sector and each other to ensure that the promise of SRHR is realised.

Advances in functional brain imaging technology and developmental neuro-psychology: their applications in the Jungian analytic domain.

PubMed

Petchkovsky, Leon

2017-06-01

Analytical psychology shares with many other psychotherapies the important task of repairing the consequences of developmental trauma. The majority of analytic patients come from compromised early developmental backgrounds: they may have experienced neglect, abuse, or failures of empathic resonance from their carers. Functional brain imagery techniques including Quantitative Electroencephalogram (QEEG), and functional Magnetic Resonance Imagery (fMRI), allow us to track mental processes in ways beyond verbal reportage and introspection. This independent perspective is useful for developing new psychodynamic hypotheses, testing current ones, providing diagnostic markers, and monitoring treatment progress. Jung, with the Word Association Test, grasped these principles 100 years ago. Brain imaging techniques have contributed to powerful recent advances in our understanding of neurodevelopmental processes in the first three years of life. If adequate nurturance is compromised, a range of difficulties may emerge. This has important implications for how we understand and treat our psychotherapy clients. The paper provides an overview of functional brain imaging and advances in developmental neuropsychology, and looks at applications of some of these findings (including neurofeedback) in the Jungian psychotherapy domain. © 2017, The Society of Analytical Psychology.

Genetics Home Reference: progressive pseudorheumatoid dysplasia

MedlinePlus

... caused by mutations in the WISP3 gene. The function of the protein produced from this gene is not well understood, ... protein that may not function. Loss of WISP3 protein function likely disrupts normal cartilage maintenance and bone growth, ...

Recent progress in elucidating the structure, function and evolution of disease resistance genes in plants.

PubMed

Liu, Jinling; Liu, Xionglun; Dai, Liangying; Wang, Guoliang

2007-09-01

Plants employ multifaceted mechanisms to fight with numerous pathogens in nature. Resistance (R) genes are the most effective weapons against pathogen invasion since they can specifically recognize the corresponding pathogen effectors or associated protein(s) to activate plant immune responses at the site of infection. Up to date, over 70 R genes have been isolated from various plant species. Most R proteins contain conserved motifs such as nucleotide-binding site (NBS), leucine-rich repeat (LRR), Toll-interleukin-1 receptor domain (TIR, homologous to cytoplasmic domains of the Drosophila Toll protein and the mammalian interleukin-1 receptor), coiled-coil (CC) or leucine zipper (LZ) structure and protein kinase domain (PK). Recent results indicate that these domains play significant roles in R protein interactions with effector proteins from pathogens and in activating signal transduction pathways involved in innate immunity. This review highlights an overview of the recent progress in elucidating the structure, function and evolution of the isolated R genes in different plant-pathogen interaction systems.

Targeting BET bromodomain proteins in solid tumors

PubMed Central

Sahai, Vaibhav; Redig, Amanda J.; Collier, Katharine A.; Eckerdt, Frank D.; Munshi, Hidayatullah G.

2016-01-01

There is increasing interest in inhibitors targeting BET (bromodomain and extra-terminal) proteins because of the association between this family of proteins and cancer progression. BET inhibitors were initially shown to have efficacy in hematologic malignancies; however, a number of studies have now shown that BET inhibitors can also block progression of non-hematologic malignancies. In this Review, we summarize the efficacy of BET inhibitors in select solid tumors; evaluate the role of BET proteins in mediating resistance to current targeted therapies; and consider potential toxicities of BET inhibitors. We also evaluate recently characterized mechanisms of resistance to BET inhibitors; summarize ongoing clinical trials with these inhibitors; and discuss potential future roles of BET inhibitors in patients with solid tumors. PMID:27283767

The Emerging Roles of Forkhead Box (FOX) Proteins in Osteosarcoma

PubMed Central

Zhang, Wentao; Duan, Ning; Song, Tao; Li, Zhong; Zhang, Caiguo; Chen, Xun

2017-01-01

Osteosarcoma is the most common bone cancer primarily occurring in children and young adults. Over the past few years, the deregulation of a superfamily transcription factors, known as forkhead box (FOX) proteins, has been demonstrated to contribute to the pathogenesis of osteosarcoma. Molecular mechanism studies have demonstrated that FOX family proteins participate in a variety of signaling pathways and that their expression can be regulated by multiple factors. The dysfunction of FOX genes can alter osteosarcoma cell differentiation, metastasis and progression. In this review, we summarized the evidence that FOX genes play direct or indirect roles in the development and progression of osteosarcoma, and evaluated the emerging role of FOX proteins as targets for therapeutic intervention. PMID:28775781

Direct enzymatic extraction of starch from corn as an energy-saving alternative to production of high-fructose syrup

NASA Astrophysics Data System (ADS)

The liquefaction of pre-gelatinized starch was studied with various analytical techniques to determine the effects of starch molecular weight, granule structure, granule size, and mechanical depolymerization. Also, improvements were made in the chromatographic system used to characterize starch hydrolysates. Progress is reported on protein removal. The effects of pH, temperature, and ionic strength were examined for the removal of protein from a syrup stream by adsorption on a phenolic resin. Buffered systems, which maintain more stable pH values, were also examined. Mathematical modeling of the results is in progress. The pilot plant facility is complete and in operation. Starch streams containing 1% protein are being produced by the protein extraction process.

Protein malnutrition promotes dysregulation of molecules involved in T cell migration in the thymus of mice infected with Leishmania infantum

PubMed Central

Losada-Barragán, Monica; Umaña-Pérez, Adriana; Cuervo-Escobar, Sergio; Berbert, Luiz Ricardo; Porrozzi, Renato; Morgado, Fernanda N.; Mendes-da-Cruz, Daniella Areas; Savino, Wilson; Sánchez-Gómez, Myriam; Cuervo, Patricia

2017-01-01

Protein malnutrition, the most deleterious cause of malnutrition in developing countries, has been considered a primary risk factor for the development of clinical visceral leishmaniasis (VL). Protein malnutrition and infection with Leishmania infantum leads to lymphoid tissue disorganization, including changes in cellularity and lymphocyte subpopulations in the thymus and spleen. Here we report that protein malnutrition modifies thymic chemotactic factors by diminishing the CCL5, CXCL12, IGF1, CXCL9 and CXCL10 protein levels in infected animals. Nevertheless, T cells preserve their migratory capability, as they were able to migrate ex vivo in response to chemotactic stimuli, indicating that malnutrition may compromise the thymic microenvironment and alter in vivo thymocyte migration. Decrease in chemotactic factors protein levels was accompanied by an early increase in the parasite load of the spleen. These results suggest that the precondition of malnutrition is affecting the cell-mediated immune response to L. infantum by altering T cell migration and interfering with the capacity of protein-deprived animals to control parasite spreading and proliferation. Our data provide evidence for a disturbance of T lymphocyte migration involving both central and peripheral T-cells, which likely contribute to the pathophysiology of VL that occurs in malnourished individuals. PMID:28397794

Effect of hyperthyroidism on circulating prolactin and hypothalamic expression of tyrosine hydroxylase, prolactin signaling cascade members and estrogen and progesterone receptors during late pregnancy and lactation in the rat.

PubMed

Pennacchio, Gisela E; Neira, Flavia J; Soaje, Marta; Jahn, Graciela A; Valdez, Susana R

2017-02-15

Hyperthyroidism (HyperT) compromises pregnancy and lactation, hindering suckling-induced PRL release. We studied the effect of HyperT on hypothalamic mRNA (RT-qPCR) and protein (Western blot) expression of tyrosine hydroxylase (TH), PRL receptor (PRLR) and signaling pathway members, estrogen-α (ERα) and progesterone (PR) receptors on late pregnancy (days G19, 20 and 21) and early lactation (L2) in rats. HyperT advanced pre-partum PRL release, reduced circulating PRL on L2 and increased TH mRNA (G21 and L2), p-TH, PRLR mRNA, STAT5 protein (G19 and L2), PRLR protein (G21) and CIS protein (G19). PRs mRNAs and protein decreased on G19 but afterwards PRA mRNA (G20), PRB mRNA (G21) and PRA mRNA and protein (L2) increased. ERα protein increased on G19 and decreased on G20. Thus, the altered hypothalamic PRLR, STAT5, PR and ERα expression in hyperthyroid rats may induce elevated TH expression and activation, that consequently, elevate dopaminergic tone during lactation, blunting suckling-induced PRL release and litter growth. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

A DIGE analysis of developing poplar leaves subjected to ozone reveals major changes in carbon metabolism.

PubMed

Bohler, Sacha; Bagard, Matthieu; Oufir, Mouhssin; Planchon, Sébastien; Hoffmann, Lucien; Jolivet, Yves; Hausman, Jean-François; Dizengremel, Pierre; Renaut, Jenny

2007-05-01

Tropospheric ozone pollution is described as having major negative effects on plants, compromising plant survival. Carbon metabolism is especially affected. In the present work, the effects of chronic ozone exposure were evaluated at the proteomic level in developing leaves of young poplar plants exposed to 120 ppb of ozone for 35 days. Soluble proteins (excluding intrinsic membrane proteins) were extracted from leaves after 3, 14 and 35 days of ozone exposure, as well as 10 days after a recovery period. Proteins (pI 4 to 7) were analyzed by 2-D DIGE experiments, followed by MALDI-TOF-TOF identification. Additional observations were obtained on growth, lesion formation, and leaf pigments analysis. Although treated plants showed large necrotic spots and chlorosis in mature leaves, growth decreased only slightly and plant height was not affected. The number of abscised leaves was higher in treated plants, but new leaf formation was not affected. A decrease in chlorophylls and lutein contents was recorded. A large number of proteins involved in carbon metabolism were identified. In particular, proteins associated with the Calvin cycle and electron transport in the chloroplast were down-regulated. In contrast, proteins associated with glucose catabolism increased in response to ozone exposure. Other identified enzymes are associated with protein folding, nitrogen metabolism and oxidoreductase activity.

Comparative systems analysis of the secretome of the opportunistic pathogen Aspergillus fumigatus and other Aspergillus species.

PubMed

Vivek-Ananth, R P; Mohanraj, Karthikeyan; Vandanashree, Muralidharan; Jhingran, Anupam; Craig, James P; Samal, Areejit

2018-04-26

Aspergillus fumigatus and multiple other Aspergillus species cause a wide range of lung infections, collectively termed aspergillosis. Aspergilli are ubiquitous in environment with healthy immune systems routinely eliminating inhaled conidia, however, Aspergilli can become an opportunistic pathogen in immune-compromised patients. The aspergillosis mortality rate and emergence of drug-resistance reveals an urgent need to identify novel targets. Secreted and cell membrane proteins play a critical role in fungal-host interactions and pathogenesis. Using a computational pipeline integrating data from high-throughput experiments and bioinformatic predictions, we have identified secreted and cell membrane proteins in ten Aspergillus species known to cause aspergillosis. Small secreted and effector-like proteins similar to agents of fungal-plant pathogenesis were also identified within each secretome. A comparison with humans revealed that at least 70% of Aspergillus secretomes have no sequence similarity with the human proteome. An analysis of antigenic qualities of Aspergillus proteins revealed that the secretome is significantly more antigenic than cell membrane proteins or the complete proteome. Finally, overlaying an expression dataset, four A. fumigatus proteins upregulated during infection and with available structures, were found to be structurally similar to known drug target proteins in other organisms, and were able to dock in silico with the respective drug.

A Cul-3-BTB ubiquitylation pathway regulates junctional levels and asymmetry of core planar polarity proteins

PubMed Central

Strutt, Helen; Searle, Elizabeth; Thomas-MacArthur, Victoria; Brookfield, Rosalind; Strutt, David

2013-01-01

The asymmetric localisation of core planar polarity proteins at apicolateral junctions is required to specify cell polarity in the plane of epithelia. This asymmetric distribution of the core proteins is proposed to require amplification of an initial asymmetry by feedback loops. In addition, generation of asymmetry appears to require the regulation of core protein levels, but the importance of such regulation and the underlying mechanisms is unknown. Here we show that ubiquitylation acts through more than one mechanism to control core protein levels in Drosophila, and that without this regulation cellular asymmetry is compromised. Levels of Dishevelled at junctions are regulated by a Cullin-3-Diablo/Kelch ubiquitin ligase complex, the activity of which is most likely controlled by neddylation. Furthermore, activity of the deubiquitylating enzyme Fat facets is required to maintain Flamingo levels at junctions. Notably, ubiquitylation does not alter the total cellular levels of Dishevelled or Flamingo, but only that of the junctional population. When junctional core protein levels are either increased or decreased by disruption of the ubiquitylation machinery, their asymmetric localisation is reduced and this leads to disruption of planar polarity at the tissue level. Loss of asymmetry by altered core protein levels can be explained by reference to feedback models for amplification of asymmetry. PMID:23487316

Echodense spinal subarachnoid space in neonates with progressive ventricular dilatation: a marker of noncommunicating hydrocephalus.

PubMed

Rudas, G; Almássy, Z; Papp, B; Varga, E; Méder, U; Taylor, G A

1998-10-01

Our purpose was to evaluate the frequency and clinical significance of echogenic debris in the spinal subarachnoid space of neonates at risk for progressive ventricular dilatation. Spinal sonography was performed on 15 neonates with severe intracranial hemorrhage (n = 10) or bacterial meningitis (n = 5). Spinal sonography also was performed on 16 control neonates. Images were analyzed for the presence and location of echogeric debris within the thoracolumbar subarachnoid space. Lumbar punctures were performed on all 31 neonates, and CSF was analyzed for cell count and protein content. Ten of 15 neonates required ventricular drainage procedures. Progressive ventricular dilatation occurred in 11 of 15 neonates with intracranial hemorrhage or meningitis. Echogenic debris was present in the thoracolumbar subarachnoid space on spinal sonography in every neonate with progressive ventricular dilatation compared with none of the 16 control neonates (p < .0001 by chi-square analysis). In addition, the 11 neonates with echogenic subarachnoid space had significantly higher protein and RBC contents in the lumbar CSF (p < .04). Echogenic subarachnoid space revealed by sonography is associated with progressive ventricular dilatation after severe intracranial hemorrhage or bacterial meningitis and is caused by high protein and RBC contents in the subarachnoid space. This finding may be helpful in identifying neonates who will not benefit from serial lumbar punctures for treatment of hydrocephalus.

Pathophysiology of osteoporosis: new mechanistic insights.

PubMed

Armas, Laura A G; Recker, Robert R

2012-09-01

Understanding of the pathophysiology of osteoporosis has evolved to include compromised bone strength and skeletal fragility caused by several factors: (1) defects in microarchitecture of trabeculae, (2) defective intrinsic material properties of bone tissue, (3) defective repair of microdamage from normal daily activities, and (4) excessive bone remodeling rates. These factors occur in the context of age-related bone loss. Clinical studies of estrogen deprivation, antiresorptives, mechanical loading, and disuse have helped further knowledge of the factors affecting bone quality and the mechanisms that underlie them. This progress has led to several new drug targets in the treatment of osteoporosis. Copyright © 2012 Elsevier Inc. All rights reserved.

Clinical applications of penetrating neural interfaces and Utah Electrode Array technologies

NASA Astrophysics Data System (ADS)

Normann, Richard A.; Fernandez, Eduardo

2016-12-01

This paper briefly describes some of the recent progress in the development of penetrating microelectrode arrays and highlights the use of two of these devices, Utah electrode arrays and Utah slanted electrode arrays, in two therapeutic interventions: recording volitional skeletal motor commands from the central nervous system, and recording motor commands and evoking somatosensory percepts in the peripheral nervous system (PNS). The paper also briefly explores other potential sites for microelectrode array interventions that could be profitably pursued and that could have important consequences in enhancing the quality of life of patients that has been compromised by disorders of the central and PNSs.

The role of bacteriophages in periodontal health and disease.

PubMed

Pinto, Graça; Silva, Maria Daniela; Peddey, Mark; Sillankorva, Sanna; Azeredo, Joana

2016-10-01

The human periodontium health is commonly compromised by chronic inflammatory conditions and has become a major public health concern. Dental plaque, the precursor of periodontal disease, is a complex biofilm consisting mainly of bacteria, but also archaea, protozoa, fungi and viruses. Viruses that specifically infect bacteria - bacteriophages - are most common in the oral cavity. Despite this, their role in the progression of periodontal disease remains poorly explored. This review aims to summarize how bacteriophages interact with the oral microbiota, their ability to increase bacterial virulence and mediate the transfer of resistance genes and suggests how bacteriophages can be used as an alternative to the current periodontal disease therapies.

Protein and amino acid nutrition

USDA-ARS?s Scientific Manuscript database

Dairy cow protein and amino acid nutrition have a significant role in sustainable dairying. Protein, amino acids, and nitrogen are inextricably linked through effects in the rumen, metabolism of the cow, and environmental nutrient management. Feeding systems have been making progress toward emphasiz...

Glycosylation patterns of kidney proteins differ in rat diabetic nephropathy.

PubMed

Ravidà, Alessandra; Musante, Luca; Kreivi, Marjut; Miinalainen, Ilkka; Byrne, Barry; Saraswat, Mayank; Henry, Michael; Meleady, Paula; Clynes, Martin; Holthofer, Harry

2015-05-01

Diabetic nephropathy often progresses to end-stage kidney disease and, ultimately, to renal replacement therapy. Hyperglycemia per se is expected to have a direct impact on the biosynthesis of N- and O-linked glycoproteins. This study aims to establish the link between protein glycosylation and progression of experimental diabetic kidney disease using orthogonal methods. Kidneys of streptozotocin-diabetic and control rats were harvested at three different time points post streptozotocin injection. A panel of 12 plant lectins was used in the screening of lectin blots. The lectins UEAI, PHA-E, GSI, PNA, and RCA identified remarkable disease-associated differences in glycoprotein expression. Lectin affinity chromatography followed by mass spectrometric analyses led to the identification of several glycoproteins involved in salt-handling, angiogenesis, and extracellular matrix degradation. Our data confirm a substantial link between glycosylation signature and diabetes progression. Furthermore, as suggested by our findings on dipeptidyl peptidase-IV, altered protein glycosylation may reflect changes in biochemical properties such as enzymatic activity. Thus, our study demonstrates the unexplored potential of protein glycosylation analysis in the discovery of molecules linked to diabetic kidney disease.

Activation of blood coagulation in cancer: implications for tumour progression

PubMed Central

Lima, Luize G.; Monteiro, Robson Q.

2013-01-01

Several studies have suggested a role for blood coagulation proteins in tumour progression. Herein, we discuss (1) the activation of the blood clotting cascade in the tumour microenvironment and its impact on primary tumour growth; (2) the intravascular activation of blood coagulation and its impact on tumour metastasis and cancer-associated thrombosis; and (3) antitumour therapies that target blood-coagulation-associated proteins. Expression levels of the clotting initiator protein TF (tissue factor) have been correlated with tumour cell aggressiveness. Simultaneous TF expression and PS (phosphatidylserine) exposure by tumour cells promote the extravascular activation of blood coagulation. The generation of blood coagulation enzymes in the tumour microenvironment may trigger the activation of PARs (protease-activated receptors). In particular, PAR1 and PAR2 have been associated with many aspects of tumour biology. The procoagulant activity of circulating tumour cells favours metastasis, whereas the release of TF-bearing MVs (microvesicles) into the circulation has been correlated with cancer-associated thrombosis. Given the role of coagulation proteins in tumour progression, it has been proposed that they could be targets for the development of new antitumour therapies. PMID:23889169

Fatty acid binding protein 4 enhances prostate cancer progression by upregulating matrix metalloproteinases and stromal cell cytokine production

PubMed Central

Huang, Mingguo; Narita, Shintaro; Inoue, Takamitsu; Koizumi, Atsushi; Saito, Mitsuru; Tsuruta, Hiroshi; Numakura, Kazuyuki; Satoh, Shigeru; Nanjo, Hiroshi; Sasaki, Takehiko; Habuchi, Tomonori

2017-01-01

Fatty acid binding protein 4 (FABP4) is an abundant protein in adipocytes, and its production is influenced by high-fat diet (HFD) or obesity. The prostate stromal microenvironment induces proinflammatory cytokine production, which is key for the development and progression of prostate cancer (PCa). Here, we show that high FABP4 expression and its secretion by PCa cells directly stimulated PCa cell invasiveness by upregulating matrix metalloproteinases through phosphatidylinositol 3-kinase and mitogen-activated protein kinase signaling pathways. In addition, prostate stromal cells augmented PCa cell invasiveness by secreting interleukin-8 and -6 in response to FABP4. This was abrogated by the FABP4 specific inhibitor, BMS309403. Furthermore, a mouse xenograft experiment showed HFD enhanced PCa metastasis and invasiveness by the upregulation of FABP4 and interleukin-8. Clinically, the serum level of FABP4 was significantly associated with an aggressive type of PCa rather than obesity. Taken together, FABP4 may enhance PCa progression and invasiveness by upregulating matrix metalloproteinases and cytokine production in the PCa stromal microenvironment, especially under HFD or obesity. PMID:29340091

The Blood-Testis Barrier and Its Implications for Male Contraception

PubMed Central

Mruk, Dolores D.

2012-01-01

The blood-testis barrier (BTB) is one of the tightest blood-tissue barriers in the mammalian body. It divides the seminiferous epithelium into the basal and the apical (adluminal) compartments. Meiosis I and II, spermiogenesis, and spermiation all take place in a specialized microenvironment behind the BTB in the apical compartment, but spermatogonial renewal and differentiation and cell cycle progression up to the preleptotene spermatocyte stage take place outside of the BTB in the basal compartment of the epithelium. However, the BTB is not a static ultrastructure. Instead, it undergoes extensive restructuring during the seminiferous epithelial cycle of spermatogenesis at stage VIII to allow the transit of preleptotene spermatocytes at the BTB. Yet the immunological barrier conferred by the BTB cannot be compromised, even transiently, during the epithelial cycle to avoid the production of antibodies against meiotic and postmeiotic germ cells. Studies have demonstrated that some unlikely partners, namely adhesion protein complexes (e.g., occludin-ZO-1, N-cadherin-β-catenin, claudin-5-ZO-1), steroids (e.g., testosterone, estradiol-17β), nonreceptor protein kinases (e.g., focal adhesion kinase, c-Src, c-Yes), polarity proteins (e.g., PAR6, Cdc42, 14-3-3), endocytic vesicle proteins (e.g., clathrin, caveolin, dynamin 2), and actin regulatory proteins (e.g., Eps8, Arp2/3 complex), are working together, apparently under the overall influence of cytokines (e.g., transforming growth factor-β3, tumor necrosis factor-α, interleukin-1α). In short, a “new” BTB is created behind spermatocytes in transit while the “old” BTB above transiting cells undergoes timely degeneration, so that the immunological barrier can be maintained while spermatocytes are traversing the BTB. We also discuss recent findings regarding the molecular mechanisms by which environmental toxicants (e.g., cadmium, bisphenol A) induce testicular injury via their initial actions at the BTB to elicit subsequent damage to germ-cell adhesion, thereby leading to germ-cell loss, reduced sperm count, and male infertility or subfertility. Moreover, we also critically evaluate findings in the field regarding studies on drug transporters in the testis and discuss how these influx and efflux pumps regulate the entry of potential nonhormonal male contraceptives to the apical compartment to exert their effects. Collectively, these findings illustrate multiple potential targets are present at the BTB for innovative contraceptive development and for better delivery of drugs to alleviate toxicant-induced reproductive dysfunction in men. PMID:22039149

Reducing the Dietary Acid Load: How a More Alkaline Diet Benefits Patients With Chronic Kidney Disease.

PubMed

Passey, Caroline

2017-05-01

It has been proposed that a low-protein diet will slow progression of chronic kidney disease although studies have not always supported this belief. The accepted practice is that 60% to 70% of protein comes from high biological value (HBV) protein, but this limits patient choice and patients struggle to follow the diet. When a diet with only 30% HBV protein was trialed, there was a significant increase in serum bicarbonate, and patients preferred the diet. The dietary advice given in predialysis clinics was changed. HBV protein was restricted to approximately 50% of total protein, bread and cereal foods were allowed freely, and fruits and vegetables (F&V) were encouraged. Patients who followed the diet have seen a slowing of progression and occasionally regression of their renal function. Both observations and scientific literature indicate that this is because of a reduction in the acid content of the diet. When foods are metabolized, most proteins produce acid, and most F&V produce alkali. A typical 21 st -century diet produces 50 to 100 mEq H + per day which the kidney is challenged to excrete. Acid is excreted with phosphate and is limited to about 45 mEq H + per day. With chronic kidney disease, this falls progressively to below 20 mEq H + per day. Historically, ammonium excretion was believed to be excretion of acid (NH 3 +  + H + → NH 4 + ), but it is now understood to be a by-product in the neutralization of acid by glutamine. The remaining acid is neutralized or stored within the body. Bone and muscle are lost in order to neutralize the acid. Acid also accumulates within cells, and serum bicarbonate falls. The author postulates that reducing the acid load through a low-protein diet with greater use of vegetable proteins and increased F&V intake will slow progression or occasionally improve renal function while maintaining the nutritional status of the individual. Copyright © 2016 National Kidney Foundation, Inc. Published by Elsevier Inc. All rights reserved.

Phosphorylation of a WRKY transcription factor by two pathogen-responsive MAPKs drives phytoalexin biosynthesis in Arabidopsis.

PubMed

Mao, Guohong; Meng, Xiangzong; Liu, Yidong; Zheng, Zuyu; Chen, Zhixiang; Zhang, Shuqun

2011-04-01

Plant sensing of invading pathogens triggers massive metabolic reprogramming, including the induction of secondary antimicrobial compounds known as phytoalexins. We recently reported that MPK3 and MPK6, two pathogen-responsive mitogen-activated protein kinases, play essential roles in the induction of camalexin, the major phytoalexin in Arabidopsis thaliana. In search of the transcription factors downstream of MPK3/MPK6, we found that WRKY33 is required for MPK3/MPK6-induced camalexin biosynthesis. In wrky33 mutants, both gain-of-function MPK3/MPK6- and pathogen-induced camalexin production are compromised, which is associated with the loss of camalexin biosynthetic gene activation. WRKY33 is a pathogen-inducible transcription factor, whose expression is regulated by the MPK3/MPK6 cascade. Chromatin immunoprecipitation assays reveal that WRKY33 binds to its own promoter in vivo, suggesting a potential positive feedback regulatory loop. Furthermore, WRKY33 is a substrate of MPK3/MPK6. Mutation of MPK3/MPK6 phosphorylation sites in WRKY33 compromises its ability to complement the camalexin induction in the wrky33 mutant. Using a phospho-protein mobility shift assay, we demonstrate that WRKY33 is phosphorylated by MPK3/MPK6 in vivo in response to Botrytis cinerea infection. Based on these data, we conclude that WRKY33 functions downstream of MPK3/MPK6 in reprogramming the expression of camalexin biosynthetic genes, which drives the metabolic flow to camalexin production in Arabidopsis challenged by pathogens.

Regulatory actions of 3',5'-cyclic adenosine monophosphate on osteoclast function: possible roles of Epac-mediated signaling.

PubMed

Jeevaratnam, Kamalan; Salvage, Samantha C; Li, Mengye; Huang, Christopher L-H

2018-05-30

Alterations in cellular levels of the second messenger 3',5'-cyclic adenosine monophosphate ([cAMP] i ) regulate a wide range of physiologically important cellular signaling processes in numerous cell types. Osteoclasts are terminally differentiated, multinucleated cells specialized for bone resorption. Their systemic regulator, calcitonin, triggers morphometrically and pharmacologically distinct retraction (R) and quiescence (Q) effects on cell-spread area and protrusion-retraction motility, respectively, paralleling its inhibition of bone resorption. Q effects were reproduced by cholera toxin-mediated G s -protein activation known to increase [cAMP] i , unaccompanied by the [Ca 2+ ] i changes contrastingly associated with R effects. We explore a hypothesis implicating cAMP signaling involving guanine nucleotide-exchange activation of the small GTPase Ras-proximate-1 (Rap1) by exchange proteins directly activated by cAMP (Epac). Rap1 activates integrin clustering, cell adhesion to bone matrix, associated cytoskeletal modifications and signaling processes, and transmembrane transduction functions. Epac activation enhanced, whereas Epac inhibition or shRNA-mediated knockdown compromised, the appearance of markers for osteoclast differentiation and motility following stimulation by receptor activator of nuclear factor kappa-Β ligand (RANKL). Deficiencies in talin and Rap1 compromised in vivo bone resorption, producing osteopetrotic phenotypes in genetically modified murine models. Translational implications of an Epac-Rap1 signaling hypothesis in relationship to N-bisphosphonate actions on prenylation and membrane localization of small GTPases are discussed. © 2018 New York Academy of Sciences.

Chromosome thripsis by DNA double strand break clusters causes enhanced cell lethality, chromosomal translocations and 53BP1-recruitment

PubMed Central

Schipler, Agnes; Mladenova, Veronika; Soni, Aashish; Nikolov, Vladimir; Saha, Janapriya; Mladenov, Emil; Iliakis, George

2016-01-01

Chromosome translocations are hallmark of cancer and of radiation-induced cell killing, reflecting joining of incongruent DNA-ends that alter the genome. Translocation-formation requires DNA end-joining mechanisms and incompletely characterized, permissive chromatin conditions. We show that chromatin destabilization by clusters of DNA double-strand-breaks (DSBs) generated by the I-SceI meganuclease at multiple, appropriately engineered genomic sites, compromises c-NHEJ and markedly increases cell killing and translocation-formation compared to single-DSBs. Translocation-formation from DSB-clusters utilizes Parp1 activity, implicating alt-EJ in their formation. Immunofluorescence experiments show that single-DSBs and DSB-clusters uniformly provoke the formation of single γ-H2AX foci, suggesting similar activation of early DNA damage response (DDR). Live-cell imaging also shows similar single-focus recruitment of the early-response protein MDC1, to single-DSBs and DSB-clusters. Notably, the late DDR protein, 53BP1 shows in live-cell imaging strikingly stronger recruitment to DSB-clusters as compared to single-DSBs. This is the first report that chromatin thripsis, in the form of engineered DSB-clusters, compromises first-line DSB-repair pathways, allowing alt-EJ to function as rescuing-backup. DSB-cluster-formation is indirectly linked to the increased biological effectiveness of high ionization-density radiations, such as the alpha-particles emitted by radon gas or the heavy-ions utilized in cancer therapy. Our observations provide the first direct mechanistic explanation for this long-known effect. PMID:27257076

Mycobacterium tuberculosis Ser/Thr protein kinase B mediates an oxygen-dependent replication switch

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

Ortega, Corrie; Liao, Reiling; Anderson, Lindsey N.

In the majority of cases, Mycobacterium tuberculosis (Mtb) infections are clinically latent, characterized by little or no bacterial replication and drug tolerance. Low oxygen tension is a major host factor inducing bacteriostasis, but the molecular mechanisms driving oxygen-dependent replication are poorly understood. Mtb encodes eleven serine/threonine protein kinases, a family of signaling molecules known to regulate similar replicative adaptations in other bacteria. Here, we tested the role of serine/threonine phosphorylation in the Mtb response to altered oxygen status, using an in vitro model of latency (hypoxia) and reactivation (reaeration). Broad kinase inhibition compromised survival of Mtb in hypoxia. Activity-based proteinmore » profiling and genetic mutation identified PknB as the kinase critical for surviving hypoxia. Mtb replication was highly sensitive to changes in PknB levels in aerated culture, and even more so in hypoxia. A mutant overexpressing PknB specifically in hypoxia showed a 10-fold loss in viability in low oxygen conditions. In contrast, chemically reducing PknB activity during hypoxia specifically compromised resumption of growth during reaeration. These data support a model in which PknB activity is reduced to achieve bacteriostasis, and elevated when replication resumes. Together, these data show that phosphosignaling controls replicative transitions associated with latency and reactivation, that PknB is a major regulator of these transitions, and that PknB could provide a highly vulnerable therapeutic target at every step of the Mtb life cycle - active disease, latency, and reactivation.« less

Altered GPM6A/M6 dosage impairs cognition and causes phenotypes responsive to cholesterol in human and Drosophila.

PubMed

Gregor, Anne; Kramer, Jamie M; van der Voet, Monique; Schanze, Ina; Uebe, Steffen; Donders, Rogier; Reis, André; Schenck, Annette; Zweier, Christiane

2014-12-01

Glycoprotein M6A (GPM6A) is a neuronal transmembrane protein of the PLP/DM20 (proteolipid protein) family that associates with cholesterol-rich lipid rafts and promotes filopodia formation. We identified a de novo duplication of the GPM6A gene in a patient with learning disability and behavioral anomalies. Expression analysis in blood lymphocytes showed increased GPM6A levels. An increase of patient-derived lymphoblastoid cells carrying membrane protrusions supports a functional effect of this duplication. To study the consequences of GPM6A dosage alterations in an intact nervous system, we employed Drosophila melanogaster as a model organism. We found that knockdown of Drosophila M6, the sole member of the PLP family in flies, in the wing, and whole organism causes malformation and lethality, respectively. These phenotypes as well as the protrusions of patient-derived lymphoblastoid cells with increased GPM6A levels can be alleviated by cholesterol supplementation. Notably, overexpression as well as loss of M6 in neurons specifically compromises long-term memory in the courtship conditioning paradigm. Our findings thus indicate a critical role of correct GPM6A/M6 levels for cognitive function and support a role of the GPM6A duplication for the patient's phenotype. Together with other recent findings, this study highlights compromised cholesterol homeostasis as a recurrent feature in cognitive phenotypes. © 2014 WILEY PERIODICALS, INC.

Target identification in Fusobacterium nucleatum by subtractive genomics approach and enrichment analysis of host-pathogen protein-protein interactions.

PubMed

Kumar, Amit; Thotakura, Pragna Lakshmi; Tiwary, Basant Kumar; Krishna, Ramadas

2016-05-12

Fusobacterium nucleatum, a well studied bacterium in periodontal diseases, appendicitis, gingivitis, osteomyelitis and pregnancy complications has recently gained attention due to its association with colorectal cancer (CRC) progression. Treatment with berberine was shown to reverse F. nucleatum-induced CRC progression in mice by balancing the growth of opportunistic pathogens in tumor microenvironment. Intestinal microbiota imbalance and the infections caused by F. nucleatum might be regulated by therapeutic intervention. Hence, we aimed to predict drug target proteins in F. nucleatum, through subtractive genomics approach and host-pathogen protein-protein interactions (HP-PPIs). We also carried out enrichment analysis of host interacting partners to hypothesize the possible mechanisms involved in CRC progression due to F. nucleatum. In subtractive genomics approach, the essential, virulence and resistance related proteins were retrieved from RefSeq proteome of F. nucleatum by searching against Database of Essential Genes (DEG), Virulence Factor Database (VFDB) and Antibiotic Resistance Gene-ANNOTation (ARG-ANNOT) tool respectively. A subsequent hierarchical screening to identify non-human homologous, metabolic pathway-independent/pathway-specific and druggable proteins resulted in eight pathway-independent and 27 pathway-specific druggable targets. Co-aggregation of F. nucleatum with host induces proinflammatory gene expression thereby potentiates tumorigenesis. Hence, proteins from IBDsite, a database for inflammatory bowel disease (IBD) research and those involved in colorectal adenocarcinoma as interpreted from The Cancer Genome Atlas (TCGA) were retrieved to predict drug targets based on HP-PPIs with F. nucleatum proteome. Prediction of HP-PPIs exhibited 186 interactions contributed by 103 host and 76 bacterial proteins. Bacterial interacting partners were accounted as putative targets. And enrichment analysis of host interacting partners showed statistically enriched terms that were in positive correlation with CRC, atherosclerosis, cardiovascular, osteoporosis, Alzheimer's and other diseases. Subtractive genomics analysis provided a set of target proteins suggested to be indispensable for survival and pathogenicity of F. nucleatum. These target proteins might be considered for designing potent inhibitors to abrogate F. nucleatum infections. From enrichment analysis, it was hypothesized that F. nucleatum infection might enhance CRC progression by simultaneously regulating multiple signaling cascades which could lead to up-regulation of proinflammatory responses, oncogenes, modulation of host immune defense mechanism and suppression of DNA repair system.

Short-term calorie and protein restriction provide partial protection from chemotoxicity but do not delay glioma progression

PubMed Central

Brandhorst, Sebastian; Wei, Min; Hwang, Saewon; Morgan, Todd E.; Longo, Valter D.

2013-01-01

Short-term starvation (STS) protects normal cells while simultaneously sensitizing malignant cells to high-dose chemotherapeutic drugs in mice and possibly patients. The fasting-dependent protection of normal cells and sensitization of malignant cells depends, in part, on reduced levels of insulin-like growth factor-1 (IGF-1) and glucose. Calorie restricted diets with defined macronutrient (carbohydrate, protein, fat) ratios were evaluated for the effects on stress sensitization markers and protection in mice treated with high-dose chemotherapy. We show that short-term CR significantly reduced both glucose and IGF-1 levels, but when specific macronutrient deficiencies were tested, only the complete lack of proteins reduced IGF-1 levels. Short-term 50% CR combined with either severe protein-deficiency or ketogenic diets improved chemotoxicity resistance similarly to the standard 50% CR, but did not result in the high protection caused by STS. Notably, a high protein diet reversed the beneficial effects of short-term CR. In a subcutaneous mouse model of glioma, feeding a low protein (4% calories from protein) diet for more than 20 days did not delay tumor progression once the tumor became palpable. Also, cycles of short-term (3 days) 50% CR did not augment the chemotherapy efficacy of cisplatin in a murine breast cancer model. These results indicate that the protection from chemotoxicity and retardation of the progression of certain tumors achieved with fasting is not obtained with short-term calorie and/or macronutrient restriction. PMID:23454633

Overexpression of protein kinase FA/GSK-3 alpha (a proline-directed protein kinase) correlates with human hepatoma dedifferentiation/progression.

PubMed

Yang, S D; Yu, J S; Yang, C C; Lee, S C; Lee, T T; Ni, M H; Kuan, C Y; Chen, H C

1996-05-01

Computer analysis of protein phosphorylation sites sequence revealed that transcriptional factors and viral oncoproteins are prime targets for regulation of proline-directed protein phosphorylation, suggesting an association of the proline-directed protein kinase (PDPK) family with neoplastic transformation and tumorigenesis. In this report, an immunoprecipitate activity assay of protein kinase FA/glycogen synthase kinase-3 alpha (kinase F(A)/GSK-3 alpha) (a member of PDPK family) has been optimized for human hepatoma and used to demonstrate for the first time significantly increased (P < 0.01) activity in poorly differentiated SK-Hep-1 hepatoma (24.2 +/- 2.8 units/mg) and moderately differentiated Mahlavu hepatoma (14.5 +/- 2.2 units/mg) when compared to well differentiated Hep 3B hepatoma (8.0 +/- 2.4 units/mg). Immunoblotting analysis revealed that increased activity of kinase FA/GSK-3 alpha is due to overexpression of the protein. Elevated kinase FA/GSK-3 alpha expression in human hepatoma biopsies relative to normal liver tissue was found to be even more profound. This kinase appeared to be fivefold overexpressed in well differentiated hepatoma and 13-fold overexpressed in poorly differentiated hepatoma when compared to normal liver tissue. Taken together, the results provide initial evidence that overexpression of kinase FA/GSK-3 alpha is involved in human hepatoma dedifferentiation/progression. Since kinase FA/GSK-3 alpha is a PDPK, the results further support a potential role of this kinase in human liver tumorigenesis, especially in its dedifferentiation/progression.

The BAX/BAK-like protein BOK is a prognostic marker in colorectal cancer.

PubMed

Carberry, Steven; D'Orsi, Beatrice; Monsefi, Naser; Salvucci, Manuela; Bacon, Orna; Fay, Joanna; Rehm, Markus; McNamara, Deborah; Kay, Elaine W; Prehn, Jochen H M

2018-01-26

The intrinsic or mitochondrial apoptosis pathway is controlled by the interaction of antiapoptotic and pro-apoptotic members of the BCL-2 protein family. Activation of this death pathway plays a crucial role in cancer progression and chemotherapy responses. The BCL-2-related ovarian killer (BOK) possesses three BCL-2 homology domains and has been proposed to act in a similar pro-apoptotic pathway as the pro-apoptotic proteins BAX and BAK. In this study, we showed that stage II and III colorectal cancer patients possessed decreased levels of BOK protein in their tumours compared to matched normal tissue. BOK protein levels in tumours were also prognostic of clinical outcome but increased BOK protein levels surprisingly associated with earlier disease recurrence and reduced overall survival. We found no significant association of BOK protein tumour levels with ER stress markers GRP78 or GRP94 or with cleaved caspase-3. In contrast, BOK protein levels correlated with Calreticulin. These data indicate BOK as a prognostic marker in colorectal cancer and suggest that different activities of BOK may contribute to cancer progression and prognosis.