Afadin Promotes Vascular Smooth Muscle Cell Contraction by Interacting With Phospholipase C to Enhance Ca Signaling for Blood Pressure Regulation.

BACKGROUND

Vascular smooth muscle cells (VSMCs) regulate vascular tone and blood pressure. Stimulation of VSMCs with vasoconstrictors, such as AngII (angiotensin II) or norepinephrine, activates the G-protein-coupled receptor-mediated cascade, leading to a hypercontractile state and vascular remodeling. Afadin, an intracellular adaptor protein that mainly localizes at cell-cell junctions, regulates various biological phenomena. However, its role in VSMCs remains unclear.

METHODS

VSMCs were isolated from newly generated VSMC-specific afadin conditional knockout mice. A small peptide (7 amino acids) designed in silico to inhibit the afadin-PLC (phospholipase C) β association was administered to the mouse VSMCs and aortic media using adeno-associated virus.

RESULTS

Unlike control mice, afadin conditional knockout mice did not exhibit AngII- or norepinephrine-induced elevation in blood pressure. VSMCs isolated from afadin conditional knockout mice were less responsive to AngII- or norepinephrine-induced cell contractility compared with control VSMCs, as evidenced by reduced release of intracellular Ca resulting from lowered production of AngII- or norepinephrine-induced inositol 1,4,5-trisphosphate. Mechanistically, the PDZ (disk large tumor suppressor, zonula occludens-1) domain of afadin was shown to associate with the C terminus of PLCβ, providing support for the localization of PLCβ on the plasma membrane, where it generates inositol 1,4,5-trisphosphate. Furthermore, the newly designed small peptide, which inhibited the afadin-PLCβ association, attenuated AngII-induced cell contractility and intracellular Ca release in vitro and blocked AngII-stimulated blood pressure elevation in vivo.

CONCLUSIONS

Afadin expression in VSMCs promotes cell contraction by interacting with PLCβ to enhance Ca signaling and has potential as a novel molecular target for blood pressure regulation.