Class II phosphatidylinositol 3-kinase \u03b1 and \u03b2 isoforms are required for vascular smooth muscle Rho activation, contraction and blood pressure regulation in mice

Shahidul Islam,Noriko Takuwa,Sho Aki,Hiroki Yamada,Kazuaki Yoshioka,Yoh Takuwa,Kazuhiro Ishimaru

doi:10.1186/s12576-020-00745-2

Shahidul Islam, Noriko Takuwa + Show 5 more

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https://doi.org/10.1186/s12576-020-00745-2

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Abstract

Class II phosphatidylinositol 3-kinases (PI3K), PI3K-C2α and PI3K-C2β, are involved in cellular processes including endocytosis, cilia formation and autophagy. However, the role of PI3K-C2α and PI3K-C2β at the organismal level is not well understood. We found that double knockout (KO) mice with both smooth muscle-specific KO of PI3K-C2α and global PI3K-C2β KO, but not single KO mice of either PI3K-C2α or PI3K-C2β, exhibited reductions in arterial blood pressure and substantial attenuation of contractile responses of isolated aortic rings. In wild-type vascular smooth muscle cells, double knockdown of PI3K-C2α and PI3K-C2β but not single knockdown of either PI3K markedly inhibited contraction with reduced phosphorylation of 20-kDa myosin light chain and MYPT1 and Rho activation, but without inhibition of the intracellular Ca2+ mobilization. These data indicate that PI3K-C2α and PI3K-C2β play the redundant but essential role for vascular smooth muscle contraction and blood pressure regulation mainly through their involvement in Rho activation.

Highlights

Phosphatidylinositol 3-kinases (PI3K) are lipid kinases that catalyze the phosphorylation of phosphoinositides at the D3 position of their inositol ring and, thereby, control a diverse array of cellular processes including vesicular trafficking, cell migration, cell proliferation and cell metabolism [1]
Decreased blood pressure in C2α‐ and C2β‐double knockout mice To study the effects of genetic deletion of C2α and C2β on blood pressure and vascular smooth muscle contraction, we employed smooth muscle-specific C2α knockout (C2αKO) mice, global C2β-knockout (C2β−/− (C2βKO)) mice, and double knockout (DKO) mice with smooth muscle-specific C2αKO and global C2βKO, which we described previously [3, 16]
We found that green fluorescent protein (GFP)-C2α was distributed diffusely as fine puncta and as coarse puncta locally in the perinuclear region with some signals at or near the plasma membrane in Human aortic smooth muscle cells (HASM) (Fig. 8a, upper left and lower views). mCherryC2β was distributed in a fibrillary pattern in the peripheral cytoplasmic region close to the plasma membrane and in perinuclear coarse puncta with some co-localization with GFP-tagged C2α (GFP-C2α) (Fig. 8a, upper right and lower views)

Summary

Introduction

Phosphatidylinositol 3-kinases (PI3K) are lipid kinases that catalyze the phosphorylation of phosphoinositides at the D3 position of their inositol ring and, thereby, control a diverse array of cellular processes including vesicular trafficking, cell migration, cell proliferation and cell metabolism [1]. C2β is highly homologous in its amino acid sequence to C2α and exhibits similar activities including cell migration and growth to those of C2α. Our study suggested that C2α was involved in Rho activation and Rho kinase-dependent phosphorylation of the myosin-targeting subunit MYPT1 of myosin light chain phosphatase (MLCP) and, thereby, an increase in phosphorylation of 20 kDa regulatory myosin light chain (MLC20). We further demonstrated that in vascular smooth muscle of spontaneously hypertensive rats, enhanced activation of C2α contributed to increased Rho activity, elevated vascular tone and hypertension [15]. These observations suggested that C2α is an important regulatory molecule for Rho activation and contraction in vascular smooth muscle

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