PCSK6 Is a Key Protease in the Control of Smooth Muscle Cell Function in Vascular Remodeling

Rationale: Proprotein convertase subtilisins/kexins (PCSKs) are a protease family with unknown functions in vasculature. Previously, we demonstrated PCSK6 upregulation in human atherosclerotic plaques associated with smooth muscle cells (SMCs), inflammation, extracellular matrix (ECM) remodeling and mitogens. Objective: Here, we applied a systems biology approach to gain deeper insights into the PCSK6 role in normal and diseased vessel wall. Methods and Results: Genetic analyses revealed association of intronic PCSK6 variant rs1531817 with maximum internal carotid intima-media thickness progression in high-cardiovascular risk subjects. This variant was linked with PCSK6 mRNA expression in healthy aortas and plaques, but also with overall plaque SMA+ cell content and pericyte fraction. Increased PCSK6 expression was found in several independent human cohorts comparing atherosclerotic lesions vs. healthy arteries, using transcriptomic and proteomic datasets. By immunohistochemistry, PCSK6 was localised to fibrous cap SMA+ cells and neovessels in plaques. In human, rat, and mouse intimal hyperplasia, PCSK6 was expressed by proliferating SMA+ cells and upregulated after 5 days in rat carotid balloon injury model, with positive correlation to PDGFB and MMP2/MMP14. Here, PCSK6 was shown to co-localise and co-interact with MMP2/MMP14 by in situ proximity ligation assay. Microarrays of carotid arteries from Pcsk6-/- vs. control mice revealed suppression of contractile SMC markers, ECM remodeling enzymes and cytokines/receptors. Pcsk6-/- mice showed reduced intimal hyperplasia response upon carotid ligation in vivo, accompanied by decreased MMP14 activation and impaired SMC outgrowth from aortic rings ex vivo. PCSK6 silencing in human SMCs in vitro lead to downregulation of contractile markers and increase in MMP2 expression. Conversely, PCSK6 overexpression increased PDGFBB-induced cell proliferation and particularly migration. Conclusions: PCSK6 is a novel protease that induces SMC migration in response to PDGFB, mechanistically via modulation of contractile markers and MMP14 activation. This study establishes PCSK6 as a key regulator of SMC function in vascular remodeling.