Novel noncanonical regulation of soluble VEGF/VEGFR2 signaling by mechanosensitive ion channel TRPV4

Abstract

VEGF signaling via VEGF receptor-2 (VEGFR2) is a major regulator of endothelial cell (EC) functions, including angiogenesis. Although most studies of angiogenesis focus on soluble VEGF signaling, mechanical signaling also plays a critical role. Here, we examined the consequence of disruption of mechanical signaling on soluble signaling pathways. Specifically, we observed that small interfering RNA (siRNA) knockdown of a mechanosensitive ion channel, transient receptor potential vanilloid 4 (TRPV4), significantly reduced perinuclear (Golgi) VEGFR2 in human ECs with a concomitant increase in phosphorylation at Y1175 and membrane translocation. TRPV4 knockout (KO) ECs exhibited increased plasma membrane localization of phospho-VEGFR2 comparedwith normal ECs. The knockdown also increased phospho-VEGFR2 inwhole cell lysates andmembrane fractions comparedwith control siRNA-treatedcells. siRNAknockdown ofTRPV4 enhancednuclear localizationof mechanosensitive transcription factors, yes-associated protein/transcriptional coactivator with PDZ-binding motif via rho kinase, which were shown to increase VEGFR2 trafficking to the plasma membrane. Furthermore, TRPV4 deletion/knockdown enhanced VEGF-mediatedmigration in vitro and increased expression of VEGFR2 in vivo in the vasculature of TRPV4KOtumors comparedwith wild-type tumors.Our results thus show that TRPV4 channels regulate VEGFR2 trafficking and activation to identify novel cross-talk between mechanical (TRPV4) and soluble (VEGF) signaling that controls EC migration and angiogenesis.