PLEKHA7 (pleckstrin homology domain containing family A member 7) has been found in multiple studies as a candidate gene for human hypertension, yet functional data supporting this association are lacking. We investigated the contribution of this gene to the pathogenesis of salt-sensitive hypertension by mutating
Plekha7 in the Dahl salt-sensitive (SS/JrHsdMcwi) rat using zinc-finger nuclease technology. After four weeks on an 8% NaCl diet, homozygous mutant rats had lower mean arterial (149 ± 9 mmHg vs. 178 ± 7 mmHg;
P < 0.05) and systolic (180 ± 7 mmHg vs. 213 ± 8 mmHg;
P < 0.05) blood pressure compared with WT littermates. Albumin and protein excretion rates were also significantly lower in mutant rats, demonstrating a renoprotective effect of the mutation. Total peripheral resistance and perivascular fibrosis in the heart and kidney were significantly reduced in
Plekha7 mutant animals, suggesting a potential role of the vasculature in the attenuation of hypertension. Indeed, both flow-mediated dilation and endothelium-dependent vasodilation in response to acetylcholine were improved in isolated mesenteric resistance arteries of
Plekha7 mutant rats compared with WT. These vascular improvements were correlated with changes in intracellular calcium handling, resulting in increased nitric oxide bioavailability in mutant vessels. Collectively, these data provide the first functional evidence that
Plekha7 may contribute to blood pressure regulation and cardiovascular function through its effects on the vasculature.Hypertension is a complex disease that is characterized by increased blood pressure, renal damage, and vascular dysfunction which collectively increase risk of atherosclerosis, stroke, heart disease, and renal failure in one-quarter of all adults worldwide (
1–
3). Because there is strong evidence of heritability in hypertension (
2,
4,
5), considerable effort has been put toward identifying novel candidate genes and their molecular mechanisms. Genome-wide association studies (GWAS) have identified many potential hypertension loci, which shed light on the genetic complexity of this disease (
5–
8) but have provided little mechanistic insight. As such, validation and elucidation of the functional roles and disease mechanisms for these gene candidates are the next important challenges (
4).Because hypertension is a complex disease (i.e., multiple variants of small effect sizes contributing to disease risk), we hypothesized candidate gene targeting on a genetically sensitized background would reveal functional role(s) of genetic disease modifiers. The Dahl salt-sensitive (SS) rat is an inbred genetic model of salt-sensitive hypertension that displays hypertension-induced renal damage, cardiac hypertrophy and vascular dysfunction (
9–
11). These phenotypes are induced by exposing SS rats to a high-salt diet, which results in rapid induction of hypertensive phenotypes that closely resemble salt-induced hypertension seen in humans (
12–
15). Knockout of specific genes in this disease model using zinc-finger nuclease (ZFN) technology have revealed the importance of key mechanisms contributing to hypertension risk, such as the protection from salt-induced hypertension and renal injury by selective ablation of adaptive immune cells in the SS-
Rag1em1Mcwi and SS-
Cd247em1Mcwi knockout rats (
16,
17) and reduced hypertension and renal injury in the SS-
Ncf2em1Mcwi (p67phox) null model exhibiting reduced medullary oxidative stress (
18). Additionally, we have recently demonstrated multiple genes at a single hypertension GWAS-nominated locus (
Agtrap-Plod1 locus) can have additive or subtractive effects on blood pressure and renal function when mutated in the SS rat (
19). These previous studies highlight the utility of this model system for testing the roles of GWAS candidate human disease genes by disrupting their specific rat orthologs using ZFN technology (
20).A single-nucleotide polymorphism (SNP) (rs381815, minor allele frequency 0.26) in intron 1 of the pleckstrin homology domain containing family A member 7 (PLEKHA7) gene, was identified by five independent GWAS to be associated with elevated systolic blood pressure and hypertension in multiple populations (
5,
6,
8,
21,
22). The associated locus contains only the PLEKHA7 gene (
5); however, the genetic mechanism(s) underlying this locus have not yet been functionally characterized.
PLEKHA7 is highly expressed in the kidney and heart, where it may be involved in formation and maintenance of the apical junction complex of epithelial cells (
23). However, limited data on
PLEKHA7 function are available to extrapolate its potential role(s) in the pathogenesis of hypertension. Here we used ZFN mutagenesis to obtain the first evidence to our knowledge in any model system that
Plekha7 has a functional role in several hypertension-associated phenotypes in the rat. We found that mutation of
Plekha7 in the SS rat attenuated salt-induced hypertension, reduced renal damage, and improved cardiac function. We also show that
Plekha7 modulates calcium handling and nitric oxide (NO) bioavailability, both of which are required for normal vascular health. Collectively, these studies provide significant mechanistic insight to the role of
Plekha7 in salt-sensitive hypertension.
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