HIF‐dependent regulation of AKAP12 (gravin) in the control of human vascular endothelial function

Abstract

Hypoxia has been widely implicated in many pathological conditions, including those associated with inflammation and tumorigenesis. A number of recent studies have implicated hypoxia in the control of vasculogenesis and permeability, the basis for which is not fully understood. Here we examine the transcriptional regulation of angiogenesis and permeability by hypoxia in endothelial cells. Guided by a global profiling approach in cultured endothelial cells, these studies revealed the selective induction of human gravin (protein kinase A anchoring protein 12) by hypoxia. Analysis of the cloned gravin promoter identified a functional hypoxia-responsive region including 2 binding sites for hypoxia-inducible factor (HIF). Site-directed mutagenesis identified the most distal HIF-binding site as essential for the induction of gravin by hypoxia. Further studies examining gravin gain and loss of function confirmed strong dependence of gravin in control of microvascular endothelial tube formation, wherein gravin functions as a "braking" system for angiogenesis. Additional studies in confluent endothelia revealed that gravin functionally couples to control endothelial barrier function in response to protein kinase A (PKA) agonists. Taken together, these results demonstrate transcriptional coordination of gravin by HIF-1α and amplified PKA-dependent endothelial responses. These findings provide an important link between hypoxia and metabolic conditions associated with inflammation and angiogenesis.

AKAP12 deficiency impairs VEGF‐induced endothelial cell migration and sprouting

“…At the molecular level, this effect was attributed to the downregulation of matrix metalloproteinase 9 and hypoxia‐inducible factor‐1α (HIF‐1α), by enhancing the interaction of HIF‐1α with von Hippel‐Lindau tumour suppressor protein and prolyl hydroxylase 2 . Also, in cultured endothelial cells, exposure to hypoxia was reported to increase AKAP12 expression and the protein was linked to vascular stability and the inhibition rather than stimulation of angiogenesis . Such observations fit with reports that AKAP12 has been reported to act as a tumour suppressor that blocks the cell cycle and inhibits oncogenic proliferation, invasion, chemotaxis and neovascularization .…”

FRET biosensors reveal AKAP-mediated shaping of subcellular PKA activity and a novel mode of Ca2+/PKA crosstalk

“…This is distinct from other known crosstalk mechanisms [reviewed in 47] and lays groundwork for future studies to investigate the impact of gravin-mediated Ca 2+ /PKA crosstalk in vivo . For example, while agonists such as thrombin, bradykinin, and histamine stimulate G protein pathways linked to [Ca 2+ ] i elevation and regulate endothelial permeability, gravin is also known to support endothelial barrier strength through cAMP-dependent pathways [17, 44]. Our finding that ATP treatment impacts PKA activity suggests that receptor-mediated Ca 2+ agonists that stimulate endothelial permeability may accomplish this in part by redirecting gravin/PKA away from the cell periphery.…”

“…The specificity with which gravin directs PKA to target substrates is currently unknown, although some AKAPs require very high submolecular specificity in the range of 10–20nm, where both PKA and its substrate are tethered to the same AKAP [36]. Our observation that gravin increased the phosphorylation of AKAR3, a non-endogenous protein, suggests that colocalization of gravin and PKA substrates at the PM may be sufficient for phosphorylation to occur at nearby substrates such as ion channels [37–39], cytoskeletal regulators [40, 41], and junctional proteins [17, 42–44]. Understanding the mechanisms underlying specificity in gravin-PKA signaling will require further identification of additional substrates beyond those already known, as well as studies of gravin-substrate binding, gravin structural dynamics, and mobility in the membrane under normal physiologic conditions.…”

“…However, gravin’s impact on subcellular PKA activity, both basally and following Ca 2+ mediated redistribution, is poorly understood. This could have important implications for disease contexts that utilize crosstalk between Ca 2+ /PKC-dependent and PKA-dependent signaling pathways, such as cellular migration [10–12], cancer [reviewed in 13], learning and memory [14], cardiac function [15], and vascular biology [16, 17]. …”

Peripheral blood mononuclear cells as a source to detect markers of homeostatic alterations caused by the intake of diets with an unbalanced macronutrient composition

“…Their activity may be linked to insulin resistance since mice lacking Msra are prone to the development of HF diet-induced insulin resistance and present a reduced insulin response [70]. Further indication for increased oxidative stress comes from the observed induction of Akap12, coding for protein kinase A anchoring protein 12 (also known as gravin), which is responsive to hypoxia [71], a condition associated with increased levels of reactive oxygen species. Ethe1 codes for a dioxygenase located in the mitochondria involved in hydrogen sulphide detoxification [72].…”