Angiotensin II Promotes KV7.4 Channels Degradation Through Reduced Interaction With HSP90 (Heat Shock Protein 90)

Voltage-gated Kv7.4 channels have been implicated in vascular smooth muscle cells’ activity because they modulate basal arterial contractility, mediate responses to endogenous vasorelaxants, and are downregulated in several arterial beds in different models of hypertension. Angiotensin II (Ang II) is a key player in hypertension that affects the expression of several classes of ion channels. In this study, we evaluated the effects of Ang II on the expression and function of vascular Kv7.4. Western blot and quantitative polymerase chain reaction revealed that in whole rat mesenteric artery, Ang II incubation for 1 to 7 hours decreased Kv7.4 protein expression without reducing transcript levels. Moreover, Ang II decreased XE991 (Kv7)–sensitive currents and attenuated membrane potential hyperpolarization and relaxation induced by the Kv7 activator ML213. Ang II also reduced Kv7.4 staining at the plasma membrane of vascular smooth muscle cells. Proteasome inhibition with MG132 prevented Ang II–induced decrease of Kv7.4 levels and counteracted the functional impairment of ML213-induced relaxation in myography experiments. Proximity ligation assays showed that Ang II impaired the interaction of Kv7.4 with the molecular chaperone HSP90 (heat shock protein 90), enhanced the interaction of Kv7.4 with the E3 ubiquitin ligase CHIP (C terminus of Hsp70-interacting protein), and increased Kv7.4 ubiquitination. Similar alterations were found in mesenteric vascular smooth muscle cells isolated from Ang II–infused mice. The effect of Ang II was emulated by 17-AAG (17-demethoxy-17-(2-propenylamino) geldanamycin) that inhibits HSP90 interactions with client proteins. These results show that Ang II downregulates Kv7.4 by altering protein stability through a decrease of its interaction with HSP90. This leads to the recruitment of CHIP and Kv7.4 ubiquitination and degradation via the proteasome

Kcne4 Deletion Sex-Dependently Alters Vascular Reactivity

Voltage-gated potassium (Kv) channels formed by Kv7 (KCNQ) α-subunits are recognized as crucial for vascular smooth muscle function, in addition to their established roles in the heart (Kv7.1) and the brain (Kv7.2-5). In vivo, Kv7 α-subunits are often regulated by KCNE subfamily ancillary (β) subunits. We investigated the effects of targeted germline Kcne4 deletion on mesenteric artery reactivity in adult male and female mice. Kcne4 deletion increased mesenteric artery contractility in response to α-adrenoceptor agonist methoxamine, and decreased responses to Kv7.2-7.5 channel activator ML213, in male but not female mice. In contrast, Kcne4 deletion markedly decreased vasorelaxation in response to isoprenaline in both male and female mice. Kcne4 expression was two-fold lower in female versus male mouse mesenteric artery, and Kcne4 deletion elicited only moderate changes of other Kcne transcripts, with no striking sex-specific differences. However, Kv7.4 protein expression in females was twice that in males, and reduced in both sexes by Kcne4 deletion. Our findings confirm a crucial role for KCNE4 in the vasculature, and provide the first known molecular mechanism for sex-specificity of this modulation that has important implications on vascular reactivity and may underlie sex-differences in susceptibility to cardiovascular diseases

<b><i>Kcne4</i></b> Deletion Sex-Dependently Alters Vascular Reactivity

Voltage-gated potassium (Kv) channels formed by Kv7 (KCNQ) α-subunits are recognized as crucial for vascular smooth muscle function, in addition to their established roles in the heart (Kv7.1) and the brain (Kv7.2-5). In vivo, Kv7 α-subunits are often regulated by KCNE subfamily ancillary (β) subunits. We investigated the effects of targeted germline Kcne4 deletion on mesenteric artery reactivity in adult male and female mice. Kcne4 deletion increased mesenteric artery contractility in response to α-adrenoceptor agonist methoxamine, and decreased responses to Kv7.2-7.5 channel activator ML213, in male but not female mice. In contrast, Kcne4 deletion markedly decreased vasorelaxation in response to isoprenaline in both male and female mice. Kcne4 expression was two-fold lower in female versus male mouse mesenteric artery, and Kcne4 deletion elicited only moderate changes of other Kcne transcripts, with no striking sex-specific differences. However, Kv7.4 protein expression in females was twice that in males, and reduced in both sexes by Kcne4 deletion. Our findings confirm a crucial role for KCNE4 in the vasculature, and provide the first known molecular mechanism for sex-specificity of this modulation that has important implications on vascular reactivity and may underlie sex-differences in susceptibility to cardiovascular diseases

Identification of novel proteins and mechanistic pathways associated with early-onset hypertension by deep proteomic mapping of resistance arteries

Resistance arteries are small blood vessels that create resistance to blood flow. In hypertension, resistance arteries undergo remodeling, affecting their ability to contract and relax appropriately. To date, no study has mapped the hypertension-related proteomic changes in resistance arteries. Using a novel data-independent acquisition–mass spectrometry (DIA-MS) approach, we determined the proteomic changes in small mesenteric and renal arteries in pre- and early-onset hypertension from the spontaneously hypertensive rat (SHR) model, which represents human primary hypertension. Compared with normotensive controls, mesenteric arteries from 12-week-old SHRs had 286 proteins that were significantly up- or downregulated, whereas 52 proteins were identified as up- or downregulated in mesenteric arteries from 6-week-old SHRs. Of these proteins, 18 were also similarly regulated in SHR renal arteries. Our pathway analyses reveal several novel pathways in the pathogenesis of hypertension. Finally, using a matrisome database, we identified 38 altered extracellular-matrix-associated proteins, many of which have never previously been associated with hypertension. Taken together, this study reveals novel proteins and mechanisms that are associated with early-onset hypertension, thereby providing novel insights into disease progression

Cycling matters: Sex hormone regulation of vascular potassium channels

ABSTRACTSex hormones and the reproductive cycle (estrus in rodents and menstrual in humans) have a known impact on arterial function. In spite of this, sex hormones and the estrus/menstrual cycle are often neglected experimental factors in vascular basic preclinical scientific research. Recent research by our own laboratory indicates that cyclical changes in serum concentrations of sex -hormones across the rat estrus cycle, primarily estradiol, have significant consequences for the subcellular trafficking and function of KV7. Vascular potassium channels, including KV7, are essential components of vascular reactivity. Our study represents a small part of a growing field of literature aimed at determining the role of sex hormones in regulating arterial ion channel function. This review covers key findings describing the current understanding of sex hormone regulation of vascular potassium channels, with a focus on KV7 channels. Further, we highlight areas of research where the estrus cycle should be considered in future studies to determine the consequences of physiological oscillations in concentrations of sex hormones on vascular potassium channel function