Role of PTP-1B in aortic smooth muscle cell motility  and tyrosine phosphorylation of focal adhesion proteins

Recent studies have focused attention on the role of protein tyrosine kinases in vascular smooth muscle cell biology, but similar information regarding protein tyrosine phosphatases (PTP) is sparse. PTP-1B is a ubiquitous nonreceptor phosphatase with uncertain function and substrates that are mostly unidentified. We used antisense oligodeoxynucleotides (ODN) against PTP-1B to investigate the role of endogenous PTP-1B in motility of primary cultures of rat aortic smooth muscle cells (RASMC). Antisense ODN decreased PTP-1B protein levels and activity in a concentration-dependent fashion, whereas sense, scrambled, or three-base mismatch antisense ODN had little or no effect. Treatment of cells with antisense ODN, but not sense, scrambled, or three-base mismatch antisense ODN, enhanced cell motility and increased tyrosine phosphorylation levels of focal adhesion proteins paxillin, p130cas, and focal adhesion kinase. Our findings indicate that PTP-1B is a negative regulator of RASMC motility via modulation of phosphotyrosine levels in several focal adhesion proteins and suggest the involvement of PTP-1B in events such as atherosclerosis and restenosis, which are associated with increased vascular smooth muscle cell motility.

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Abstract 480: Non-muscle Myosin II Regulates Aortic Stiffness via Tension-dependent Phosphorylation of Specific Focal Adhesion Proteins

Circulation Research ◽

10.1161/res.127.suppl_1.480 ◽

2020 ◽

Vol 127 (Suppl_1) ◽

Author(s):

Kuldeep Singh ◽

Anne B Kim ◽

Kathleen G Morgan

Keyword(s):

Smooth Muscle ◽

Muscle Cell ◽

Focal Adhesion ◽

Aortic Stiffness ◽

Myosin Ii ◽

Cell Cortex ◽

Dependent Manner ◽

Adhesion Proteins ◽

Focal Adhesion Proteins ◽

Muscle Myosin

Non-muscle myosin II plays a role in many fundamental cellular processes including cell adhesion, migration, and cytokinesis. However, its role in vascular function is not well understood. Here, we investigated the function of non-muscle myosin II in the biomechanical properties of mouse proximal aorta. We found that blebbistatin, a specific inhibitor of non-muscle myosin II decreases agonist-induced aortic stress and stiffness in a dose-dependent manner. We also specifically demonstrate, in freshly isolated contractile aortic smooth muscle cells, using deconvolution microscopy that the NM myosin IIA isoform co-localizes with contractile filaments in the core of the cell as well as in the non-muscle cell cortex. However, the NM myosin IIB isoform is only colocalized with contractile filaments, and is excluded from the cell cortex. Furthermore, both the siRNA knockdown of NMIIA and NMIIB isoforms in a differentiated smooth muscle cell line A7r5 and blebbistatin-mediated inhibition of NM myosin II suppresses agonist-activated increases in phosphorylation of FAK Y925 and paxillin Y118. Thus, in the present study, we show, for the first time, that NM myosin II regulates aortic stiffness and that this regulation is mediated at least in part through the tension-dependent phosphorylation of focal adhesion proteins FAK and paxillin.

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