Effect of HSP47 expression levels on hetero trimmer formation among type iv collagen α3, α4 and α5 chains

Abstract

We previously established stable transform ants 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 hetero trimmer and that α5 chains containing mutations seen in Apart syndrome are defective in hetero trimerization. In these studies, the amount of hetero trimmer 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 hetero trimerization of α3(IV), α4(IV) and α5(IV) chains. Reduction of HSP47 levels by SIRNA resulted in defects of hetero trimerization among the three chains, indicating that HSP47 plays a critical role in the hetero trimerization. On the other hand, over expression of HSP47 did not infuence 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. Over expression of HSP47 decreased the secretion of hetero trimmers containing the mutant α5(IV) chain, suggesting that HSP47 over expression might enhance the quality control mechanisms of collagen synthesis by inhibiting the secretion of incorrectly structured hetero trimmers.