We examined the role of the smoothelin-like 1 (SMTNL1) protein in mediating vascular smooth muscle contractile responses to pressure in resistance vessels. Arterioles from wild type (WT) and SMTNL1 global knock-out (KO) mice were examined with pressure myography, and tissues were analyzed by western blotting. Mesenteric arterioles (3rd order) from Smtnl1 KO mice exhibited strikingly enhanced myogenic tone potential compared to WT littermates. Vessels from KO animals are more responsive to acute pressure changes, (i.e., exhibit greater contractile response to increasing luminal pressure). Specifically, vessels from Smtnl1 KO animals were able to generate pressure induced constriction down to 75% of their passive diameter over pressures ranging from 40-120mmHg. Conversely, wild type vessels generate only modest tone. SMTNL1 was previously shown to regulate the activity of smooth muscle myosin phosphatase by altering MYPT1 expression levels. In this regard, WT and KO mesenteric arterioles showed no differences in total MYPT1 expression; however, a 4- fold increase in CPI-17 expression (transcript and protein) was associated with SMTNL1 KO. Based on these findings, we conclude that SMTNL1 contributes to the regulation of pressure induced contractility through modulation of the activity of myosin light chain phosphatase, both directly and indirectly (via CPI-17).