Phosphorylation at Tyrosine 262 Promotes GADD34 Protein Turnover

Background: Proteasomal degradation of GADD34 dictates cell sensitivity to ER stress. Results: Phosphorylation at tyrosine 262 controls GADD34 protein turnover and cell fate following ER stress. Conclusion: Tyrosine 262 phosphorylation controls the cellular levels of GADD34, a key component of the stress-activated eIF2 phosphatase. Significance: Stress-induced GADD34 gene transcription and subsequent regulated GADD34 protein turnover control the feedback loop that ensures cell survival following ER stress. In mammalian cells, metabolic and environmental stress increases the phosphorylation of the eukaryotic translational initiation factor, eIF2, and attenuates global protein synthesis. Subsequent transcriptional activation of GADD34 assembles an eIF2 phosphatase that feeds back to restore mRNA translation. Active proteasomal degradation of GADD34 protein then reestablishes the sensitivity of cells to subsequent bouts of stress. Mass spectrometry established GADD34 phosphorylation on multiple serines, threonines, and tyrosines. Phosphorylation at tyrosine 262 enhanced the rate of the GADD34 protein turnover. Substrate-trapping studies identified TC-PTP (PTPN2) as a potential GADD34 phosphatase, recognizing phosphotyrosine 262. Reduced GADD34 protein levels in TC-PTP-null MEFs following ER stress emphasized the importance of TC-PTP in determining the cellular levels of GADD34 protein. The susceptibility of TC-PTP-null MEFs to ER stress-induced apoptosis was significantly ameliorated by ectopic expression of GADD34. The data suggested that GADD34 phosphorylation on tyrosine 262 modulates endoplasmic reticulum stress signaling and cell fate.