Blood pressure–associated polymorphism controls ARHGAP42 expression via serum response factor DNA binding
Xue Bai, … , Joan M. Taylor, Christopher P. Mack
Xue Bai, … , Joan M. Taylor, Christopher P. Mack
Published January 23, 2017
Citation Information: J Clin Invest. 2017;127(2):670-680. https://doi.org/10.1172/JCI88899.
View: Text | PDF
Research Article Genetics Vascular biology

Blood pressure–associated polymorphism controls ARHGAP42 expression via serum response factor DNA binding

Abstract

We recently demonstrated that selective expression of the Rho GTPase-activating protein ARHGAP42 in smooth muscle cells (SMCs) controls blood pressure by inhibiting RhoA-dependent contractility, providing a mechanism for the blood pressure–associated locus within the ARHGAP42 gene. The goals of the current study were to identify polymorphisms that affect ARHGAP42 expression and to better assess ARHGAP42’s role in the development of hypertension. Using DNase I hypersensitivity methods and ENCODE data, we have identified a regulatory element encompassing the ARHGAP42 SNP rs604723 that exhibits strong SMC-selective, allele-specific activity. Importantly, CRISPR/Cas9–mediated deletion of this element in cultured human SMCs markedly reduced endogenous ARHGAP42 expression. DNA binding and transcription assays demonstrated that the minor T allele variation at rs604723 increased the activity of this fragment by promoting serum response transcription factor binding to a cryptic cis-element. ARHGAP42 expression was increased by cell stretch and sphingosine 1-phosphate in a RhoA-dependent manner, and deletion of ARHGAP42 enhanced the progression of hypertension in mice treated with DOCA-salt. Our analysis of a well-characterized cohort of untreated borderline hypertensive patients suggested that ARHGAP42 genotype has important implications in regard to hypertension risk. Taken together, our data add insight into the genetic mechanisms that control blood pressure and provide a potential target for individualized antihypertensive therapies.

Authors

Xue Bai, Kevin D. Mangum, Rachel A. Dee, George A. Stouffer, Craig R. Lee, Akinyemi Oni-Orisan, Cam Patterson, Jonathan C. Schisler, Anthony J. Viera, Joan M. Taylor, Christopher P. Mack

×

Figure 5

ARHGAP42 expression is activated by RhoA signaling and cell stretch.

Options: View larger image (or click on image) Download as PowerPoint
ARHGAP42 expression is activated by RhoA signaling and cell stretch. (A...
(A) Primary rat aortic SMCs were treated with 10 μM sphingosine 1-phosphate (S1P) with or without the ROCK inhibitor Y-27632. Arhgap42 expression was measured after 72 hours by semiquantitative PCR. Data represent mean ± SEM of 4 experiments; *P < 0.001 vs. control; **P < 0.001 vs. S1P-treated (ANOVA). n = 4. (B) Using the FX-4000T Flexcell system, primary rat aortic SMCs were subjected to 0 (Ctrl) or 20% equibiaxial elongation at 1 Hz (cyclic strain). Arhgap42 message was measured at 18 hours by quantitative PCR. Data represent mean ± SEM; n = 3 from 2 independent experiments; *P < 0.001 vs. no cyclic strain; **P < 0.05 vs. minus Y-27632 (ANOVA). (C) Rat portal veins placed in ex vivo culture were subjected to 0 or 600 mg of static stretching force. At 72 hours Arhgap42 message was measured by semiquantitative RT-PCR. Graph shows ImageJ-based quantification of 3 independent experiments. *P < 0.05 (Student’s t test). (D) Portal veins isolated from Arhgap42+/gt mice were cultured ex vivo and subjected to 0 or 300 mg of static stretching force for 5 days. After LacZ staining, tissues were processed for standard microscopy including H&E staining. Scale bar: 200 μm. Data are representative of 3 independent experiments.