Role of heat shock protein 47 in platelet glycoprotein VI dimerization and signaling

Abstract

BackgroundHeat shock protein 47 (HSP47) is an intracellular chaperone protein with an indispensable role in collagen biosynthesis in collagen-secreting cells. This chaperone has also been shown to be released and present on the surface of platelets. The inhibition of HSP47 in human platelets or its ablation in mouse platelets reduces platelet function in response to collagen and the glycoprotein (GP) VI collagen receptor agonist CRP-XL. ObjectivesIn this study, we sought, through experiments, to explore cellular distribution, trafficking, and influence on GPVI interactions to understand how HSP47 modulates collagen receptor signaling. MethodsHSP47-deficient mouse platelets and SMIH- treated human platelets were used to study the role of HSP47 in collagen mediated responses and signaling. ResultsUsing subcellular fractionation analysis and immunofluorescence microscopy, HSP47 was found to be localized to the platelet-dense tubular system. Following platelet stimulation, HSP47 mobilization to the cell surface was shown to be dependent on actin polymerization, a feature common to other dense tubular system resident platelet proteins that are released to the cell surface during activation. In this location, HSP47 was found to contribute to platelet adhesion to collagen or CRP-XL but not to GFOGER peptide (an integrin α2β1-binding sequence within collagens), indicating selective effects of HSP47 on GPVI function. Dimerization of GPVI on the platelet surface increases its affinity for collagen. GPVI dimerization was reduced following HSP47 inhibition, as was collagen and CRP-XL-mediated signaling. ConclusionThe present study identifies a role for cell surface-localized HSP47 in modulating platelet responses to collagen through dimerization of GPVI, thereby enhancing platelet signaling and activation.