The Role of NPGPx in Stress Responses

• Main Authors: Wei,Pei-Chi, 魏珮琪
• Other Authors: Shew, Jin-Yuh
• Format: Others
• Language: en_US
• Published: 2012
• Online Access: http://ndltd.ncl.edu.tw/handle/55511598998300862097

博士 === 國防醫學院 === 生命科學研究所 === 101 === NPGPx is a member of the glutathione peroxidase (GPx) family; however, it lacks GPx enzymatic activity due to the absence of a critical selenocysteine residue, rendering its function an enigma. Here, we show that NPGPx is a newly identified stress sensor that transmits oxidative stress signals by forming the disulfide bond between its Cys57 and Cys86 residues. This oxidized form of NPGPx binds to glucose-regulated protein (GRP)78 and forms covalent bonding intermediates between Cys86 of NPGPx and Cys41/Cys420 of GRP78. Subsequently, the formation of the disulfide bond between Cys41 and Cys420 of GRP78 enhances its chaperone activity. NPGPx-deficient cells display increased reactive oxygen species, accumulated misfolded proteins, and impaired GRP78 chaperone activity. Complete loss of NPGPx in animals causes systemic oxidative stress, increases carcinogenesis, and shortens lifespan. These results suggest that NPGPx is essential for releasing excessive ER stress by enhancing GRP78 chaperone activity to maintain physiological homeostasis. On the other hand, we found that the presence of non-targeting (NT)-siRNA selectively induced the expression of NPGPx, but not other ER-stress proteins including GRP78, Calnexin, and XBP1. The proximal promoter of NPGPx, which contained a mixed G-quadruplex (G4) structure, was sensitive to NT-siRNA stress. Disrupting the G4 structure diminished NT-siRNA induced NPGPx promoter activity. Under the stress, nucleolin (NCL) specifically bonded to the G4 containing sequences to replace the originally bound Sp1 at the NPGPx promoter upon NT-siRNA stress. Consistently, overexpression of NCL further increased NPGPx promoter activity; while depletion of NCL desensitized NPGPx promoter to NT-siRNA stress. Upon NT-siRNA stress, NPGPx covalently bound to exoribonuclease XRN2, facilitating XRN2 to remove accumulated NT-siRNA. Cells suffering from constant NT-siRNA stress grew slower and prolonged G1 phase, while NPGPx-depleted cells accumulated mature NT-siRNA and underwent apoptosis. These results suggest that cis-element with mixed G4 structure at the NPGPx promoter plays an essential role for its transactivation mediated by NCL, and NPGPx serves as a novel responder to NT-siRNA-induced stress in facilitating XRN2 to release the NT-siRNA accumulation. In summary, we provided a new insight of how cells sense and response to ER/oxidative stress by using NPGPx as an example. NPGPx has multiple functions when encounter different stresses. It can sense the oxidative stress in ER lumen and activates GRP78 to chaperone misfolded protein. On the other hand, NPGPx can also interact and activate XRN2 to release NT-siRNA accumulation. Taken together, NPGPx is a novel protein peroxidase and it is important in sensing and also mediating stress response to protect cells from stress-induced damages.