P53 Impairs Endothelium-Dependent Vasomotor Function Through Transcriptional Upregulation of P66shc

Abstract

Abstract-The transcription factor, p53, and the adaptor protein, p66shc, both play essential roles in promoting oxidative stress in the vascular system. However, the relationship between the two in the context of endothelium-dependent vascular tone is unknown. Here, we report a novel, evolutionarily conserved, p53-mediated transcriptional mechanism that regulates p66shc expression and identify p53 as an important determinant of endothelium-dependent vasomotor function. We provide evidence of a p53 response element in the promoter of p66shc and show that angiotensin II-induced upregulation of p66shc in endothelial cells is dependent on p53.In addition, we demonstrate that downregulation of p66shc expression, as well as inhibition of p53 function in mice, mitigates angiotensin II-induced impairment of endothelium-dependent vasorelaxation, decrease in bioavailable nitric oxide, and hypertension. These findings reveal a novel p53-dependent transcriptional mechanism for the regulation of p66shc expression that is operative in the vascular endothelium and suggest that this mechanism is important in impairing endothelium-dependent vascular relaxation. Key Words: tumor suppressor p53 Ⅲ p66shc angiotensin II Ⅲ endothelial dysfunction P 66shc belongs to the shcA family of adaptor proteins. P66shc is structurally and functionally distinct from p52shc and p46shc, the other 2 members of this family. It has a unique N-terminal collagen homology (CH2) domain that is important in governing its activity, and it functions as a protein that promotes oxidative stress within cells and tissues. Cells lacking p66shc have reduced levels of oxidants, and mice deficient for p66shc are resistant to oxidative stresses, and have an extended lifespan. 1,2 In addition to living longer, p66shc-deficient mice are protected against age-associated and hyperglycemia-induced endothelial dysfunction and oxidative stress. 3,4 Similarly, downregulation of p66shc in normal blood vessels and cells increases endothelial NO synthase activity and improves endothelium-dependent vasorelaxation. 5 P66shc also promotes atherogenesis. P66shc-deficient mice have reduced susceptibility to atheroma formation when fed a high-fat diet. 6 Despite this growing body of evidence implicating p66shc in many pathophysiological states of the cardiovascular system, little information exists about the mechanisms that govern p66shc expression in the cardiovascular system and the relevance of these mechanisms to human cardiovascular disease states.The role that the tumor suppressor protein p53 plays in different cell types comprising the vascular wall, and in the overall pathogenesis of atherosclerosis, as well as in other vascular disorders that are not necessarily associated with atherosclerosis, is highly controversial. P53 is undetectable in normal vascular specimens, but is expressed in endothelial cells, smooth muscle cells, and macrophages of advanced human atherosclerosis. 7 In animal models of high-fat dietinduced atheroma formation, knockout of p53 results in an incre...

The p66Shc redox adaptor protein is induced by saturated fatty acids and mediates lipotoxicity-induced apoptosis in pancreatic beta cells

“…Moreover, pancreatic islets isolated from HFD-fed mice displayed robust increases in p66 Shc mRNA expression, establishing a link between in vivo lipotoxicity and p66 Shc regulation. Both the use of pifithrin-α, an inhibitor of p53 function, and siRNA-mediated p53 knockdown allowed us to prove that the ability of saturated FAs to promote p66 Shc expression is mediated by p53, in line with similar results in other cell systems [23]. We found that p53 expression was increased in response to palmitate in both mouse islets and INS-1E beta cells, as well as in human islets from overweight/obese compared with normal-weight donors.…”

“…Prolonged exposure of beta cells to elevated palmitate concentrations has been shown to cause downstream JNK activation and increased p53 expression, events which are in part mediated by enhanced oxidative stress and typically linked to cellular damage [7, 24, 33, this study]. The identification of a p53 response/binding element in the promoter region of p66 Shc indicated that p66 Shc is a p53 target gene and that p66 Shc is indispensable for p53-induced apoptosis [22,23,34]. In this study, murine and human islets, as well as INS-1E cells, showed increased levels of p66 Shc when chronically exposed to elevated palmitate concentrations.…”

“…Levels of p66 Shc mRNA and p66 Shc protein were found to be increased in the kidney cortex of diabetic mice [19] and in circulating leucocytes from diabetic patients [21], suggesting that p66 Shc could 'sense' the impaired metabolic milieu in diabetes and promote cellular dysfunction. In support of this concept, the p66 Shc protein was found to act as a downstream effector of the tumour suppressor p53 gene in oxidative stress-induced apoptosis [18,22], and a p53 response element in the gene promoter of p66 Shc has also been identified [23]. On the other hand, FAs increase the level of p53 acetylation in human monocytes [24].…”

p66Shc mediates high-glucose and angiotensin II-induced oxidative stress renal tubular injury via mitochondrial-dependent apoptotic pathway

“…The HG has been shown to increase the synthesis of ANG II (8) and expression of p53, a transcription factor. Interestingly, the consensus sequences for the binding of this transcription factor have been localized in the promoter region of p66Shc (19). Thus the observed increase of p66Shc mRNA expression in DN may be via the ANG II-induced modulation of p53.…”

Nicotine Exposure Augments Renal Toxicity of 5-azacytidine Through p66shc: Prevention by Resveratrol

“…NRK52E cells were transfected with the following reporter luciferase plasmids: p66shc promoter (14), antioxidant response element (ARE; Qiagen, Germantown, MD, USA) and a HO-1-promoter (15) together with a Renilla luciferase (Promega, Madison, WI, USA) using Lipofectamine 3000 reagent (Life Technologies, Grand Island, NY, USA) as recommended by the manufacturer. 24 h after treatment firefly and renilla luciferase activities were determined using the Dual Luciferase assay kit (Promega, Madison, WI, USA) as recommended by the manufacturer.…”