title: The small GTPase Rac1 is a novel binding partner of Bcl-2 and stabilizes its antiapoptotic activity.
abstract: The small GTPase Rac1 is involved in the activation of the reduced NAD phosphate oxidase complex resulting in superoxide production. We recently showed that Bcl-2 overexpression inhibited apoptosis in leukemia cells by creating a pro-oxidant intracellular milieu, and that inhibiting intracellular superoxide production sensitized Bcl-2-overexpressing cells to apoptotic stimuli. We report here that silencing and functional inhibition of Rac1 block Bcl-2-mediated increase in intracellular superoxide levels in tumor cells. Using confocal, electron microscopy and coimmunoprecipitation, as well as glutathione S-transferase-fusion proteins, we provide evidence for a colocalization and physical interaction between the 2 proteins. This interaction is blocked in vitro and in vivo by the BH3 mimetics as well as by synthetic Bcl-2 BH3 domain peptides. That this interaction is functionally relevant is supported by the ability of the Bcl-2 BH3 peptide as well as the silencing and functional inhibition of Rac1 to inhibit intracellular superoxide production as well as overcome Bcl-2-mediated drug resistance in human leukemia cells and cervical cancer cells. Notably, the interaction was observed in primary cells derived from patients with B-cell lymphoma overexpressing Bcl-2 but not in noncancerous tissue. These data provide a novel facet in the biology of Bcl-2 with potential implications for targeted anticancer drug design.
authors.affiliation: Department of Physiology, Yong Loo Lin School of Medicine, Singapore.
authors.forename: Rathiga
authors.initials: R
authors.lastname: Velaithan
authors.forename: Jia
authors.initials: J
authors.lastname: Kang
authors.forename: Jayshree L
authors.initials: JL
authors.lastname: Hirpara
authors.forename: Thomas
authors.initials: T
authors.lastname: Loh
authors.forename: Boon Cher
authors.initials: BC
authors.lastname: Goh
authors.forename: Morgane
authors.initials: M
authors.lastname: Le Bras
authors.forename: Catherine
authors.initials: C
authors.lastname: Brenner
authors.forename: Marie-Veronique
authors.initials: MV
authors.lastname: Clement
authors.forename: Shazib
authors.initials: S
authors.lastname: Pervaiz
mesh_terms: D000818:Animals; D017209:Apoptosis; D020868:Gene Silencing; D006367:HeLa Cells; D006801:Humans; D019169:Jurkat Cells; D051379:Mice; D041681:NIH 3T3 Cells; D009369:Neoplasms / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism; D009479:Neuropeptides / Q000235:genetics / Q000378:metabolism*; D010446:Peptide Fragments / Q000494:pharmacology; D057786:Peptidomimetics / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects / Q000235:genetics; D011518:Proto-Oncogene Proteins / Q000494:pharmacology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D013481:Superoxides / Q000378:metabolism*; D020744:rac GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*
chemical_list: C474007:Bax protein (53-86); D009479:Neuropeptides; D010446:Peptide Fragments; D057786:Peptidomimetics; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; C107953:RAC1 protein, human; C484602:Rac1 protein, mouse; D013481:Superoxides; C499134:Rac1 protein, rat; D020744:rac GTP-Binding Proteins; D020830:rac1 GTP-Binding Protein