IBIMOL   23987
INSTITUTO DE BIOQUIMICA Y MEDICINA MOLECULAR PROFESOR ALBERTO BOVERIS
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
Role of VMP1-USP9x interaction in the initial steps of autophagosome formation
Autor/es:
FELIPE RENNA; DANIEL GRASSO; MARÍA INÉS VACCARO
Lugar:
CABA
Reunión:
Congreso; Pancreas 2017; 2017
Institución organizadora:
International Association of Pancreatology / European Pancreatic Club
Resumen:
Introduction: The intracellular activation of zymogens is involved in acute pancreatitis pathogenesis and the eventual pancreas self-digestion. Most of the cases of acute pancreatitis are self-limited, suggesting the importance of the adaptive response mechanism of pancreatic acinar cells. VMP1 is an autophagy-related protein which is able to induce autophagy even in non-starved condition. Moreover, VMP1 is induced by acute pancreatitis and mediates zymophagy, the selective autophagic degradation of zymogen granules. Here we demonstrated that the ubiquitin protease USP9x interacts with VMP1 during zymophagy in acute pancreatitis.Aim: The objective of this work is to know the role of VMP1-USP9x interaction in the initial steps of autophagosome formationResults: We demonstrate by immunofluorescence that USP9x responds as early as 10 min after autophagy induction, with a relocation pattern without changing its expression by western blot. At basal conditions, USP9x is distributed over the whole cytoplasm. However, upon autophagy induction, it quickly moves to a perinuclear area compatible with ERES/ERGIC compartment, considered as an autophagosome formation site. USP9x is esential for autophagosome formation since shRNA-mediated depletion of USP9x inhibits autophagy, evaluated by LC3 recruitment. USP9x interacts with VMP1 and colocalizes with VMP1 in the ERES/ERGIC compartment, suggesting its implication in the initiation mechanism of autophagosome formation. We also demonstrate that USP9x deubiquitinates BECN1, which is another VMP1 interactor essential for autophagosome formation. We show the ubiquitination of BECN1 during autophagy and its accumulation when it is co-expressed with a ubiquitin molecule without chain formation capability. Moreover, shRNA-mediated depletion of USP9x increases the general ubiquitin signal in the cytoplasm and inhibits autophagosome formation, confirming the critical role of VMP1, BECN1 and USP9x in the autophagosome formation.Conclusion: In conclusion, We demonstrated for the first time that the deubiquitin protease USP9x is a novel autophagy-related protein involved in initial steps of autophagosome formation. USP9x relocates to the ERES/ERGIC compartment interacting with VMP1 and BECN1 in response to autophagy induction. Hierarchically, USP9x acts downstream of VMP1, since VMP1 expression induces USP9x relocation during autophagosome formation. Finally, we suggest that USP9x is a new player in the molecular mechanism of zymophagy during acute pancreatitis, being part in the initial steps of VMP1-mediated autophagy.