Endoplasmic reticulum stress in the absence of calnexin.
Calnexin is a type I integral endoplasmic reticulum (ER) membrane chaperone involved in folding of newly synthesized (glycol)proteins. In this study, we used beta-galactosidase reporter gene knock-in and reverse transcriptase polymerase chain reaction (RT-PCR) to investigate activation of the calnexin gene during embryonic development. We showed that the calnexin gene was activated in neuronal tissue at the early stages of embryonic development but remained low in the heart, intestine, and smooth muscle. At early stages of embryonic development, large quantities of calnexin messenger RNA (mRNA) were also found in neuronal tissue and liver. There was no detectable calnexin mRNA in the heart, lung, and intestine. The absence of calnexin had no significant effect on ER stress response (unfolded protein response, UPR) at the tissue level as tested by IRE1-dependent splicing of Xbp1 mRNA. In contrast, non-stimulated calnexin-deficient cells showed increased activation of IRE1, as measured by RT-PCR and luciferase reporter gene analysis of splicing of Xbp1 mRNA and activation of the BiP promoter. This indicates that cnx (-/-) cells have increased constitutively active UPR. Importantly, cnx (-/-) cells have significantly increased proteasomal activity, which may play a role in the adaptive mechanisms addressing the acute ER stress observed in the absence of calnexin.
Department of Biochemistry, University of Alberta, Edmonton, Alberta, Canada.
Helen
H
Coe
Karen
K
Bedard
Jody
J
Groenendyk
Joanna
J
Jung
Marek
M
Michalak
D000818:Animals; D037281:Calnexin / Q000172:deficiency* / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D047108:Embryonic Development; D004721:Endoplasmic Reticulum / Q000378:metabolism / Q000473:pathology*; D000091342:Endoplasmic Reticulum Chaperone BiP; D005347:Fibroblasts / Q000166:cytology / Q000201:enzymology; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D006360:Heat-Shock Proteins / Q000235:genetics; D051379:Mice; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism; D009417:Nerve Tissue / Q000378:metabolism; D011401:Promoter Regions, Genetic; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D017510:Protein Folding; D012326:RNA Splicing; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D000071701:Regulatory Factor X Transcription Factors; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013312:Stress, Physiological; D014157:Transcription Factors / Q000378:metabolism; D000071717:X-Box Binding Protein 1
D004268:DNA-Binding Proteins; D000091342:Endoplasmic Reticulum Chaperone BiP; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D012333:RNA, Messenger; D000071701:Regulatory Factor X Transcription Factors; D014157:Transcription Factors; D000071717:X-Box Binding Protein 1; C000605544:Xbp1 protein, mouse; D037281:Calnexin; D046988:Proteasome Endopeptidase Complex