title: Familial CJD associated PrP mutants within transmembrane region induced Ctm-PrP retention in ER and triggered apoptosis by ER stress in SH-SY5Y cells.
abstract: BACKGROUND
  Genetic prion diseases are linked to point and inserted mutations in the prion protein (PrP) gene that are presumed to favor conversion of the cellular isoform of PrP (PrP(C)) to the pathogenic one (PrP(Sc)). The pathogenic mechanisms and the subcellular sites of the conversion are not completely understood. Here we introduce several PRNP gene mutations (such as, PrP-KDEL, PrP-3AV, PrP-A117V, PrP-G114V, PrP-P102L and PrP-E200K) into the cultured cells in order to explore the pathogenic mechanism of familial prion disease.
  
  
  RESULTS
  To address the roles of aberrant retention of PrP in endoplasmic reticulum (ER), the recombinant plasmids expressing full-length human PrP tailed with an ER signal peptide at the COOH-terminal (PrP-KDEL) and PrP with three amino acids exchange in transmembrane region (PrP-3AV) were constructed. In the preparations of transient transfections, 18-kD COOH-terminal proteolytic resistant fragments (Ctm-PrP) were detected in the cells expressing PrP-KDEL and PrP-3AV. Analyses of the cell viabilities in the presences of tunicamycin and brefeldin A revealed that expressions of PrP-KDEL and PrP-3AV sensitized the transfected cells to ER stress stimuli. Western blots and RT-PCR identified the clear alternations of ER stress associated events in the cells expressing PrP-KDEL and PrP-3AV that induced ER mediated apoptosis by CHOP and caspase-12 apoptosis pathway. Moreover, several familial CJD related PrP mutants were transiently introduced into the cultured cells. Only the mutants within the transmembrane region (G114V and A117V) induced the formation of Ctm-PrP and caused the ER stress, while the mutants outside the transmembrane region (P102L and E200K) failed.
  
  
  CONCLUSIONS
  The data indicate that the retention of PrP in ER through formation of Ctm-PrP results in ER stress and cell apoptosis. The cytopathic activities caused by different familial CJD associated PrP mutants may vary, among them the mutants within the transmembrane region undergo an ER-stress mediated cell apoptosis.
authors.affiliation: State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, People's Republic of China.
authors.forename: Xin
authors.initials: X
authors.lastname: Wang
authors.forename: Qi
authors.initials: Q
authors.lastname: Shi
authors.forename: Kun
authors.initials: K
authors.lastname: Xu
authors.forename: Chen
authors.initials: C
authors.lastname: Gao
authors.forename: Cao
authors.initials: C
authors.lastname: Chen
authors.forename: Xiao-Li
authors.initials: XL
authors.lastname: Li
authors.forename: Gui-Rong
authors.initials: GR
authors.lastname: Wang
authors.forename: Chan
authors.initials: C
authors.lastname: Tian
authors.forename: Jun
authors.initials: J
authors.lastname: Han
authors.forename: Xiao-Ping
authors.initials: XP
authors.lastname: Dong
mesh_terms: D017209:Apoptosis; D045744:Cell Line, Tumor; D007562:Creutzfeldt-Jakob Syndrome / Q000235:genetics* / Q000473:pathology; D004721:Endoplasmic Reticulum / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D050505:Mutant Proteins / Q000235:genetics*; D018621:PrPC Proteins / Q000235:genetics; D018620:PrPSc Proteins / Q000235:genetics; D011328:Prions / Q000378:metabolism*; D011506:Proteins; D014162:Transfection
chemical_list: C110396:(ctm)PrP; D050505:Mutant Proteins; D018621:PrPC Proteins; D018620:PrPSc Proteins; D011328:Prions; D011506:Proteins