Human BAG-1 Proteins Bind to the Cellular Stress Response Protein GADD34 and Interfere with GADD34 Functions

Abstract

3The cellular stress response protein GADD34 mediates growth arrest and apoptosis in response to DNA damage, negative growth signals, and protein malfolding. GADD34 binds to protein phosphatase PP1 and can attenuate the translational elongation of key transcriptional factors through dephosphorylation of eukaryotic initiation factor 2␣ (eIF2␣). Recently, we reported the involvement of human SNF5/INI1 (hSNF5/INI1) protein in the functions of GADD34 and showed that hSNF5/INI1 binds GADD34 and stimulates the bound PP1 phosphatase activity. To better understand the regulatory and functional mechanisms of GADD34, we undertook a yeast two-hybrid screen with full-length GADD34 as bait in order to identify additional protein partners of GADD34. We report here that human cochaperone protein BAG-1 interacts with GADD34 in vitro and in SW480 cells treated with the proteasome inhibitor z-LLL-B to induce apoptosis. Two other proteins, Hsp70/ Hsc70 and PP1, associate reversibly with the GADD34-BAG-1 complex, and their dissociation is promoted by ATP. BAG-1 negatively modulates GADD34-bound PP1 activity, and the expression of BAG-1 isoforms can also mask GADD34-mediated inhibition of colony formation and suppression of transcription. Our findings suggest that BAG-1 may function to suppress the GADD34-mediated cellular stress response and support a role for BAG-1 in the survival of cells undergoing stress.The eventual fate of cells following exposure to genotoxic stress is determined by signaling from competing pathways favoring either death or survival. The subsequent down-regulation of both p53-dependent and -independent apoptosis pathways may be key to both cell survival and oncogenesis. The growth arrest and DNA damage-inducible protein GADD34 mediates growth arrest and apoptosis in response to stress signals elicited by genotoxic stress, amino acid deprivation, and protein malfolding at the endoplasmic reticulum (21,22,35). The GADD34 transcript is stabilized under conditions of cellular stress, leading to the observed rise in its mRNA level independent of cellular p53 status (24).The GADD34 protein harbors a highly conserved carboxyterminal peptide domain homologous to herpes simplex virus type 1 (HSV-1) ICP34.5, a virulence factor that blocks the premature shutoff of protein synthesis in HSV-1-infected neuroblastoma cells and allows HSV-1-infected cells to circumvent apoptosis (6,10,11,19). Both ICP34.5 and mammalian GADD34 engage protein phosphatase-1 (PP1) and target dephosphorylation of the eukaryotic translation elongation factor eIF2␣ (13,20). In HSV-1-infected cells, this molecular event enables protein synthesis (19). In mammalian cells, the dephosphorylation attenuates eIF2␣-mediated induction of stress-responsive genes and has been proposed in feedback regulation of cellular stress response initiated by protein malfolding at the endoplasmic reticulum (35).The mechanism and regulation of GADD34-mediated growth suppression and apoptosis remain poorly understood.Recently we reported the association of the human SNF5...

AtBAG6, a novel calmodulin-binding protein, induces programmed cell death in yeast and plants

“…Comparative sequence analysis of AtBAG6 with other known proteins revealed the presence of a putative CaM-binding IQ motif [25][26][27][28] and a BAG domain [17][18][19] in the middle of the sequence ( Figure 1). A glutamic acid-rich region was identified near the carboxyl terminus of the protein sequence on a database using the ScanProsite program (http:// www.expasy.org/cgi-bin/scanprosite).…”

“…As a consequence, this family of cochaperones functionally regulates diverse cellular pathways, such as programmed cell death (PCD) and stress responses. 19 While information is available on the biological roles of BAG proteins in mammals, no plant homologs of BAG family members have been functionally characterized to date.…”

The BAG-1 cochaperone is a negative regulator of p73-dependent transcription

“…We have shown earlier that endogenous BAG-1 enhances the function of NF-kB in colorectal cancer cells (Clemo et al, 2008). BAG-1 also interacts with and interferes with the function of the stress-responsive, pro-apoptotic GADD34 protein (Hung et al, 2003). However, the requirements for cochaperone binding in these activities are not clear.…”

Time-resolved gene expression profiling of human squamous cell carcinoma cells during the apoptosis process induced by photodynamic treatment with hypericin

“…GADD34 can induce both, apoptosis as well as recovery from a shutoff of protein synthesis induced by ER stress and was also found highly upregulated by Buytaert et al (10). As an antagonist, BAG1 [BCL2-associated athanogene (GRAP)], moderately upregulated at 5 h (2.3-fold), suppresses GADD34 mediated stress response in contrast to ATF4, and promotes survival of cells undergoing stress (84). By interacting with BCL2 it regulates the activity of HSP70 and HSC70 and activates RAF-1, which induces MAP kinases (signalling pathway).…”

“…this prosurvival molecule suppresses GADD34 mediated cellular stress response and promotes survival of cells undergoing stress (84). BAG1 interacts with BCL2 (preventing release of cytochrome C), regulating the activity of HSP70 and HSC70 (controlling formation of the apoptosome) and activating RAF-1, which induces MAP kinases.…”