title: S-Nitrosylation of Bcl-2 Negatively Affects Autophagy in Lung Epithelial Cells.
abstract: Autophagy is a catabolic cellular mechanism involving lysosomal degradation of unwanted cellular components. Interaction between Beclin-1 and Bcl-2 proteins is known to play a critical role in the initiation of autophagy. We report that malignantly transformed lung epithelial cells are resistant to autophagy and express lower basal levels of autophagic proteins, Beclin-1 and LC3-II as compared to non-tumorigenic cells. Additionally, increased levels of nitric oxide (NO) and Bcl-2 were observed in transformed cells. Nitric oxide was found to negatively regulate autophagy initiation and autophagic flux by nitrosylating Bcl-2 and stabilizing its interaction with Beclin-1, resulting in inhibition of Beclin-1 activity. An increase in the apoptotic initiator caspase-9 and the apoptosis and autophagy-associated kinase p38/MAPK in both cell types indicated possible autophagy-apoptosis crosstalk. Pre-treatments with ABT-737 (Bcl-2 inhibitor) and aminoguanidine (NO inhibitor), and transfection with a non-nitrosylable Bcl-2 cysteine double-mutant plasmid resulted in increased autophagic flux (LC3-II/p62 upregulation) corresponding with decreased S-nitrocysteine expression, thus corroborating the regulatory role of Bcl-2 S-nitrosylation in autophagy. In conclusion, our study reveals a novel mechanism of autophagy resistance via post-translational modification of Bcl-2 protein by NO, which may be critical in driving cellular tumorigenesis.
authors.affiliation: Department of Pharmaceutical Sciences, Hampton University, Hampton, Virginia.
authors.forename: Clayton
authors.initials: C
authors.lastname: Wright
authors.affiliation: Department of Pharmaceutical Sciences, Hampton University, Hampton, Virginia.
authors.forename: Anand Krishnan V
authors.initials: AK
authors.lastname: Iyer
authors.affiliation: Department of Pharmaceutical Sciences, Hampton University, Hampton, Virginia.
authors.forename: Yogesh
authors.initials: Y
authors.lastname: Kulkarni
authors.affiliation: Department of Pharmaceutical Sciences, Hampton University, Hampton, Virginia.
authors.forename: Neelam
authors.initials: N
authors.lastname: Azad
mesh_terms: D056809:Alveolar Epithelial Cells / Q000378:metabolism*; D001343:Autophagy; D002460:Cell Line; D003545:Cysteine / Q000031:analogs & derivatives / Q000378:metabolism; D006801:Humans; D009569:Nitric Oxide / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D026403:S-Nitrosothiols / Q000378:metabolism; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
chemical_list: C000595707:BCL2 protein, human; D019253:Proto-Oncogene Proteins c-bcl-2; D026403:S-Nitrosothiols; D009569:Nitric Oxide; C021315:S-nitrosocysteine; D048051:p38 Mitogen-Activated Protein Kinases; D003545:Cysteine
keywords: AUTOPHAGY; BCL-2; BECLIN-1; NITRIC OXIDE; S-NITROSYLATION