title: Effect of HSP47 expression levels on heterotrimer formation among type IV collagen α3, α4 and α5 chains.
abstract: We previously established stable transformants of the human embryonic kidney 293 (HEK293) cell line that express type IV collagen α3, α4 and wild-type or mutant-type α5 chains. Using these cell lines, we confirmed that these three chains form a heterotrimer and that α5 chains containing mutations seen in Alport syndrome are defective in heterotrimerization. In these studies, the amount of heterotrimer that formed was much less than expected relative to the amount of α(IV) chains expressed. The aim of the present study was to determine the effect of the collagen-specific molecular chaperone heat shock protein 47 (HSP47), whose expression is low in HEK293 cells, on the heterotrimerization of α3(IV), α4(IV) and α5(IV) chains. Reduction of HSP47 levels by siRNA resulted in defects of heterotrimerization among the three chains, indicating that HSP47 plays a critical role in the heterotrimerization. On the other hand, overexpression of HSP47 did not influence heterotrimerization. Since many enzymes and molecular chaperons assist correct folding and trimerization of collagens, one or more enzymes and/or molecular chaperones, other than HSP47, might be deficient in HEK293 cells. Overexpression of HSP47 decreased the secretion of heterotrimers containing the mutant α5(IV) chain, suggesting that HSP47 overexpression might enhance the quality control mechanisms of collagen synthesis by inhibiting the secretion of incorrectly structured heterotrimers.
authors.affiliation: Division of Pediatrics, Department of Homeostatic Regulation and Development, Niigata University Graduate School of Medical and Dental Sciences, 1-757 Asahimachi-dori, Niigata, Japan. takekoba@med.niigata-u.ac.jp
authors.forename: Takehiro
authors.initials: T
authors.lastname: Kobayashi
authors.forename: Makoto
authors.initials: M
authors.lastname: Uchiyama
mesh_terms: D024141:Collagen Type IV / Q000235:genetics* / Q000378:metabolism*; D057809:HEK293 Cells; D050958:HSP47 Heat-Shock Proteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D007678:Kidney Glomerulus / Q000378:metabolism; D009154:Mutation; D009394:Nephritis, Hereditary / Q000235:genetics / Q000378:metabolism; D055503:Protein Multimerization
chemical_list: D024141:Collagen Type IV; D050958:HSP47 Heat-Shock Proteins; C494660:SERPINH1 protein, human