L'ANNEXINE 3 DANS LE NEUTROPHILE (EXPRESSION AU COURS DE LA DIFFERENCIATION ET CARACTERISATION DE MUTANTS FONCTIONNELS)

PARIS-AgroParisTech Centre Paris (751052302) / SudocSudocFranceF

Propriétés d'interactions aux lipides membranaires de sous-domaines de la dystrophine (effet de l'hétérogénéité moléculaire)

Les interactions entre les protéines et les lipides sont impliquées dans plusieurs processus biologiques au cours desquels leurs propriétés physicochimiques influencent leur comportement. Dans cette étude nous avons caractérisés des interactions des lipides membranaires avec des fragments du domaine central de la dystrophine humaine. Il s'agit d'une longue protéine musculaire qui joue un rôle déterminant dans la stabilité membranaire et cellulaire au cours de l'effort musculaire. Son absence ou son dysfonctionnement induisent des dystrophies musculaires. Le domaine central est constitué de répétitions hélicales et de charnières flexibles, nous avons étudié plus particulièrement les répétitions 16 à 21 (R16-21). En utilisant différents modèles membranaires et en combinant des méthodes biochimiques et biophysiques, nous avons mis en évidence le rôle de la charnière n 3 dans l'augmentation de l'affinité des répétitions R16-21 pour les lipides anioniques, ce qui favoriserait la stabilité membranaire et les interactions avec d'autres partenaires. Nous avons montré que le comportement de ce fragment avec les lipides est sensible à la présence de cholestérol et de lipides saturés. Des délétions de R16-21 retrouvés chez des patients, induisent une diminution de la stabilité structurale et un comportement différent avec les lipides par rapport à la protéine native. En conclusion, nous montrons que l'organisation à la membrane du fragment R16-21 de la dystrophine, ou de certains dérivés présents chez des patients myopathes, est modulée par l'environnement lipidique. Les modifications structurales qui en découlent pourraient influencer leur comportement dans la cellule musculaire.Interactions between protein and lipids are involved in several biological processes during which physicochemical properties of both partners influence each other behavior. We characterized here the nteractions of fragments of the central rod domain of dystrophin with membrane lipids. Dystrophin is a long muscular protein which plays an important role in the membrane and cellular stability during muscular effort. Its absence or dysfunction due to deletions causes muscular dystrophies. In particular, we studied fragment R16-21 of the central rod domain constituted by helical repeats and by flexible hinges. Using membrane models and combining biochemical and biophysical methods, we highlighted the role of hinge 3 in the increase of the affinity of R16-21 for anionic lipids, which would favor the membrane stability and the interactions with other partners. We showed that the behavior of this fragment with lipids is sensitive to the presence of cholesterol or saturated lipids. Deletions found in patients in this R16-21 fragment were shown to decrease the structural stability and to modify the behavior with lipids compared to the native protein. In conclusion, we show that the organization to the membrane of the R16-21 fragment of dystrophin, or mutated fragments, is modulated by the lipid environment and by their structure and conformation, which confer them different functions in the muscular cell.RENNES1-BU Sciences Philo (352382102) / SudocSudocFranceF

Aquaglyceroporins, one channel for two molecules

In the light of the recently published structure of GlpF and AQP1, we have analysed the nature of the residues which could be involved in the formation of the selectivity filter of aquaporins, glycerol facilitators and aquaglyceroporins. We demonstrate that the functional specificity for major intrinsic protein (MIP) channels can be explained on one side by analysing the polar environment of the residues that form the selective filter. On the other side, we show that the channel selectivity could be associated with the oligomeric state of the membrane protein. We conclude that a non-polar environment in the vicinity of the top of helix 5 could allow aquaglyceroporins and GlpF to exist as monomers within the hydrophobic environment of the membrane

Aquaglyceroporins, one channel for two molecules

In the light of the recently published structure of GlpF and AQP1, we have analysed the nature of the residues which could be involved in the formation of the selectivity filter of aquaporins, glycerol facilitators and aquaglyceroporins. We demonstrate that the functional specificity for major intrinsic protein (MIP) channels can be explained on one side by analysing the polar environment of the residues that form the selective filter. On the other side, we show that the channel selectivity could be associated with the oligomeric state of the membrane protein. We conclude that a non-polar environment in the vicinity of the top of helix 5 could allow aquaglyceroporins and GlpF to exist as monomers within the hydrophobic environment of the membrane

Non protective function of CXCL14 chemokine in acute viral hepatitis in coronavirus MHV infected mice and in human

International audienc

Depletion of RIPK1 in hepatocytes exacerbates PolyIC and Murine Hepatitis Virus induced liver damage

International audienc

Depletion of RIPK1 in hepatocytes exacerbates liver damage in fulminant viral hepatitis

Abstract The protein kinase RIPK1 plays a crucial role at the crossroad of stress-induced signaling pathways that affects cell’s decision to live or die. The present study aimed to define the role of RIPK1 in hepatocytes during fulminant viral hepatitis, a worldwide syndrome mainly observed in hepatitis B virus (HBV) infected patients. Mice deficient for RIPK1, specifically in liver parenchymal cells (Ripk1 LPC-KO) and their wild-type littermates (Ripk1 fl/fl), were challenged by either the murine hepatitis virus type 3 (MHV3) or poly I:C, a synthetic analog of double-stranded RNA mimicking viral pathogen-associated molecular pattern. Ripk1 LPC-KO mice developed more severe symptoms at early stage of the MHV3-induced fulminant hepatitis. Similarly, administration of poly I:C only triggered increase of systemic transaminases in Ripk1 LPC-KO mice, reflecting liver damage through induced apoptosis as illustrated by cleaved-caspase 3 labeling of liver tissue sections. Neutralization of TNF-α or prior depletion of macrophages were able to prevent the appearance of apoptosis of hepatocytes in poly I:C-challenged Ripk1 LPC-KO mice. Moreover, poly I:C never induced direct hepatocyte death in primary culture whatever the murine genotype, while it always stimulated an anti-viral response. Our investigations demonstrated that RIPK1 protects hepatocytes from TNF-α secreted from macrophages during viral induced fulminant hepatitis. These data emphasize the potential worsening risks of an HBV infection in people with polymorphism or homozygous amorphic mutations already described for the RIPK1 gene

Receptor-interacting protein kinase-1 ablation in liver parenchymal cells promotes liver fibrosis in murine NASH without affecting other symptoms

International audienceNon-alcoholic steatohepatitis (NASH), a chronic liver disease that emerged in industrialized countries, can further progress into liver fibrosis, cirrhosis, and hepatocellular carcinoma. In the next decade, NASH is predicted to become the leading cause of liver transplantation, the only current interventional therapeutic option. Hepatocyte death, triggered by different death ligands, plays key role in its progression. Previously, we showed that the receptor-interacting protein kinase-1 (RIPK1) in hepatocytes exhibits a protective role in ligand-induced death. Now, to decipher the role of RIPK1 in NASH, Ripk1(LPC-KO) mice, deficient for RIPK1 only in liver parenchymal cells, and their wild-type littermates (Ripk1(fl/fl)) were fed for 3, 5, or 12 weeks with high-fat high-cholesterol diet (HFHCD). The main clinical signs of NASH were analyzed to compare the pathophysiological state established in mice. Most of the symptoms evolved similarly whatever the genotype, whether it was the increase in liver to body weight ratio, the steatosis grade or the worsening of liver damage revealed by serum transaminase levels. In parallel, inflammation markers followed the same kinetics with significant equivalent inductions of cytokines (hepatic mRNA levels and blood cytokine concentrations) and a main peak of hepatic infiltration of immune cells at 3 weeks of HFHCD. Despite this identical inflammatory response, more hepatic fibrosis was significantly evidenced at week 12 in Ripk1(LPC-KO) mice. This coincided with over-induced rates of transcripts of genes implied in fibrosis development (Tgfb1, Tgfbi, Timp1, and Timp2) in Ripk1(LPC-KO) animals. In conclusion, our results show that RIPK1 in hepatocyte limits the progression of liver fibrosis during NASH