The Pancreatitis-induced Vacuole Membrane Protein 1 Triggers Autophagy in Mammalian Cells

Department of Physiology, School of Medicine, University of Buenos Aires, and CEMIC University Hospital, 4102 Av. E. Galvan, Buenos Aires C1431FWO, Argentina.
Journal of Biological Chemistry (Impact Factor: 4.57). 01/2008; 282(51):37124-33. DOI: 10.1074/jbc.M706956200
Source: PubMed


Autophagy is a degradation process of cytoplasmic cellular constituents, which serves as a survival mechanism in starving cells, and it is characterized by sequestration of bulk cytoplasm and organelles in double-membrane vesicles called autophagosomes. Autophagy has been linked to a variety of pathological processes such as neurodegenerative diseases and tumorigenesis, which highlights its biological and medical importance. We have previously characterized the vacuole membrane protein 1 (VMP1) gene, which is highly activated in acute pancreatitis, a disease associated with morphological changes resembling autophagy. Here we show that VMP1 expression triggers autophagy in mammalian cells. VMP1 expression induces the formation of ultrastructural features of autophagy and recruitment of the microtubule-associated protein 1 light-chain 3 (LC3), which is inhibited after treatment with the autophagy inhibitor 3-methiladenine. VMP1 is induced by starvation and rapamycin treatments. Its expression is necessary for autophagy, because VMP1 small interfering RNA inhibits autophagosome formation under both autophagic stimuli. VMP1 is a transmembrane protein that co-localizes with LC3, a marker of the autophagosomes. It interacts with Beclin 1, a mammalian autophagy initiator, through the VMP1-Atg domain, which is essential for autophagosome formation. VMP1 endogenous expression co-localizes with LC3 in pancreas tissue undergoing pancreatitis-induced autophagy. Finally, VMP1 stable expression targeted to pancreas acinar cell in transgenic mice induces autophagosome formation. Our results identify VMP1 as a novel autophagy-related membrane protein involved in the initial steps of the mammalian cell autophagic process.

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Available from: Daniel Grasso, Oct 04, 2015
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    • "Beclin 1 enhanced SDT-induced apoptosis 475 2007, Ropolo et al. 2007 "
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    ABSTRACT: To prove the occurrence of autophagy after treatment by protoporphyrin IX (PpIX)- mediated sonodynamic therapy (SDT) of human chronic myelogenous leukemia K562 cells as well as its relationship with apoptosis. The 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenylter-trazolium bromide tetrazolium (MTT) assay was adopted to examine cytotoxicity of different treatments. Nuclear morphology changes were observed under a fluorescence microscopy with 4'-6-Diamidino-2-Phenylindole (DAPI) staining. Western blotting was used to analyze the expression of caspase-3, Beclin 1 (BECN 1) and the conversion of LC3-phosphatidylethanolamine conjugate / a cytosolic form of LC3 (LC3 II / I). Fluorescence microscope was used to identify the formation of autophagic vacuoles (AVO) during autophagy. Under optimal conditions, SDT was shown to induce autophagy in K562 cells, which caused the up-regulation of Beclin-1 and the formation of AVO. In addition, pre-treatment of cancer cells with beclin 1-targeted short hairpin RNA (Beclin 1 shRNA) was shown to reduce the level of LC3-II accumulation and staining with punctate spots of monodansylcadaverine (MDC) staining. Besides, the cytotoxic effect of SDT was significantly increased by Beclin 1 shRNA. Furthermore, studies showed a marked effect on the apoptosis of cells by Beclin 1 shRNA to sonodamage with increased DAPI staining and caspase-3 cleavage. These results demonstrated that SDT significantly induced autophagy of K562 cells, probably to protect the K562 cells from sonodamage.
    International Journal of Radiation Biology 03/2015; 91(6):1-32. DOI:10.3109/09553002.2015.1021961 · 1.69 Impact Factor
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    • "Interestingly, an accumulation of huge ubiquitinpositive protein aggregates containing the autophagy marker ATG8/LC3 was seen and p62 homolog [45] in Dictyostelium cells lacking Vmp1 gene showed. Moreover, the knockdown of VMP1 expression abolishes starvation and rapamycininduced autophagosome formation [40]. It also abolishes autophagy induced by hyperstimulation of Gq-coupled CCK receptor in pancreatic acinar cells [41] or by chemotherapy in pancreatic tumor cells [46]. "
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    ABSTRACT: Autophagy is a highly regulated-cell pathway for degrading long-lived proteins as well as for clearing cytoplasmic organelles. Autophagy is a key contributor to cellular homeostasis and metabolism. Warburg hypothesized that cancer growth is frequently associated with a deviation of a set of energy generation mechanisms to a nonoxidative breakdown of glucose. This cellular phenomenon seems to rely on a respiratory impairment, linked to mitochondrial dysfunction. This mitochondrial dysfunction results in a switch to anaerobic glycolysis. It has been recently suggested that epithelial cancer cells may induce the Warburg effect in neighboring stromal fibroblasts in which autophagy was activated. These series of observations drove to the proposal of a putative reverse Warburg effect of pathophysiological relevance for, at least, some tumor phenotypes. In this review we introduce the autophagy process and its regulation and its selective pathways and role in cancer cell metabolism. We define and describe the Warburg effect and the newly suggested "reverse" hypothesis. We also discuss the potential value of modulating autophagy with several pharmacological agents able to modify the Warburg effect. The association of the Warburg effect in cancer and stromal cells to tumor-related autophagy may be of relevance for further development of experimental therapeutics as well as for cancer prevention.
    BioMed Research International 08/2014; 2014:926729. DOI:10.1155/2014/926729 · 3.17 Impact Factor
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    • "VMP1 (Vacuole Membrane Protein 1) is a transmembrane protein with homologues in all eukaryotic kingdoms, with fungi being a notable exception. It has been shown to be located in several cellular compartments: the Golgi apparatus [1], the endoplasmic reticulum [1,2], autophagosomes [3], and the plasma membrane [4]. It has been implicated in an array of cellular processes: organellar biogenesis, endo-, exo-, phagocytosis and protein secretion [2]; apoptosis [1]; adhesion [4]; and, perhaps most notably, macroautophagy (henceforth: "autophagy"; [3]), as well as a special type of autophagy, namely zymophagy, in which zymogen granules—subcellular structures that contain inactive digestive enzyme—are selectively degraded [5]. "
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    ABSTRACT: Background The versatile Vacuole Membrane Protein 1 (VMP1) has been previously investigated in six species. It has been shown to be essential in macroautophagy, where it takes part in autophagy initiation. In addition, VMP1 has been implicated in organellar biogenesis; endo-, exo- and phagocytosis, and protein secretion; apoptosis; and cell adhesion. These roles underly its proven involvement in pancreatitis, diabetes and cancer in humans. Results In this study we analyzed a VMP1 homologue from the green alga Chlamydomonas reinhardtii. CrVMP1 knockdown lines showed severe phenotypes, mainly affecting cell division as well as the morphology of cells and organelles. We also provide several pieces of evidence for its involvement in macroautophagy. Conclusion Our study adds a novel role to VMP1's repertoire, namely the regulation of cytokinesis. Though the directness of the observed effects and the mechanisms underlying them remain to be defined, the protein's involvement in macroautophagy in Chlamydomonas, as found by us, suggests that CrVMP1 shares molecular characteristics with its animal and protist counterparts.
    BMC Plant Biology 05/2014; 14(1):121. DOI:10.1186/1471-2229-14-121 · 3.81 Impact Factor
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