Cystathionine β-synthase regulates endothelial function via protein S-sulfhydration
01 Jan 2016-The FASEB Journal (Federation of American Societies for Experimental Biology)-Vol. 30, Iss: 1, pp 441-456
TL;DR: It is shown that the loss of CBS function in endothelial cells (ECs) leads to a significant down‐regulation of cellular hydrogen sulfide (H2S) by 50% and of glutathione (GSH) by 40%, highlighting the importance of CBS‐mediated protein S‐sulfhydration in maintaining vascular health and function.
Abstract: Deficiencies of the human cystathionine β-synthase (CBS) enzyme are characterized by a plethora of vascular disorders and hyperhomocysteinemia. However, several clinical trials demonstrated that despite reduction in homocysteine levels, disease outcome remained unaffected, thus the mechanism of endothelial dysfunction is poorly defined. Here, we show that the loss of CBS function in endothelial cells (ECs) leads to a significant down-regulation of cellular hydrogen sulfide (H2S) by 50% and of glutathione (GSH) by 40%. Silencing CBS in ECs compromised phenotypic and signaling responses to the VEGF that were potentiated by decreased transcription of VEGF receptor (VEGFR)-2 and neuropilin (NRP)-1, the primary receptors regulating endothelial function. Transcriptional down-regulation of VEGFR-2 and NRP-1 was mediated by a lack in stability of the transcription factor specificity protein 1 (Sp1), which is a sulfhydration target of H2S at residues Cys68 and Cys755. Reinstating H2S but not GSH in CBS-silenced ECs restored Sp1 levels and its binding to the VEGFR-2 promoter and VEGFR-2, NRP-1 expression, VEGF-dependent proliferation, and migration phenotypes. Thus, our study emphasizes the importance of CBS-mediated protein S-sulfhydration in maintaining vascular health and function.-Saha, S., Chakraborty, P. K., Xiong, X., Dwivedi, S. K. D., Mustafi, S. B., Leigh, N. R., Ramchandran, R., Mukherjee, P., Bhattacharya, R. Cystathionine β-synthase regulates endothelial function via protein S-sulfhydration.
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TL;DR: The biology of NO and ROS in the cardiovascular system, with special emphasis on their routes of formation and regulation, are presented, as well as the therapeutic challenges and opportunities for the management of no/ROS in cardiovascular disease.
Abstract: Nitric oxide (NO) is a small free radical with critical signaling roles in physiology and pathophysiology. The generation of sufficient NO levels to regulate the resistance of the blood vessels and...
282 citations
TL;DR: The biosynthesis and catabolism of Hcy is examined and recent findings linking disruption of this metabolism and endothelial dysfunction are critically revised, emphasizing the impact of HHcy on endothelial cell methylation status.
Abstract: Homocysteine (Hcy) is a sulfur-containing non-proteinogenic amino acid formed during the metabolism of the essential amino acid methionine. Hcy is considered a risk factor for atherosclerosis and cardiovascular disease (CVD), but the molecular basis of these associations remains elusive. The impairment of endothelial function, a key initial event in the setting of atherosclerosis and CVD, is recurrently observed in hyperhomocysteinemia (HHcy). Various observations may explain the vascular toxicity associated with HHcy. For instance, Hcy interferes with the production of nitric oxide (NO), a gaseous master regulator of endothelial homeostasis. Moreover, Hcy deregulates the signaling pathways associated with another essential endothelial gasotransmitter: hydrogen sulfide. Hcy also mediates the loss of critical endothelial antioxidant systems and increases the intracellular concentration of reactive oxygen species (ROS) yielding oxidative stress. ROS disturb lipoprotein metabolism, contributing to the growth of atherosclerotic vascular lesions. Moreover, excess Hcy maybe be indirectly incorporated into proteins, a process referred to as protein N-homocysteinylation, inducing vascular damage. Lastly, cellular hypomethylation caused by build-up of S-adenosylhomocysteine (AdoHcy) also contributes to the molecular basis of Hcy-induced vascular toxicity, a mechanism that has merited our attention in particular. AdoHcy is the metabolic precursor of Hcy, which accumulates in the setting of HHcy and is a negative regulator of most cell methyltransferases. In this review, we examine the biosynthesis and catabolism of Hcy and critically revise recent findings linking disruption of this metabolism and endothelial dysfunction, emphasizing the impact of HHcy on endothelial cell methylation status.
170 citations
Cites background from "Cystathionine β-synthase regulates ..."
...Taken together, these findings suggest that CBS deficiency will contribute to endothelial dysfunction by decreasing H2S-induced vascular relaxation [101,104]....
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TL;DR: This review has focused on the impact of H2S on vascular structure and function with an emphasis on angiogenesis, vascular tone, vascular permeability and atherosclerosis.
Abstract: Hydrogen sulfide (H2S) is a ubiquitous signaling molecule with important functions in many mammalian organs and systems. Observations in the 1990s ascribed physiological actions to H2S in the nervous system, proposing that this gasotransmitter acts as a neuromodulator. Soon after that, the vasodilating properties of H2S were demonstrated. In the past decade, H2S was shown to exert a multitude of physiological effects in the vessel wall. H2S is produced by vascular cells and exhibits antioxidant, antiapoptotic, anti-inflammatory, and vasoactive properties. In this concise review, we have focused on the impact of H2S on vascular structure and function with an emphasis on angiogenesis, vascular tone, vascular permeability and atherosclerosis. H2S reduces arterial blood pressure, limits atheromatous plaque formation, and promotes vascularization of ischemic tissues. Although the beneficial properties of H2S are well established, mechanistic insights into the molecular pathways implicated in disease prevention and treatment remain largely unexplored. Unraveling the targets and downstream effectors of H2S in the vessel wall in the context of disease will aid in translation of preclinical observations. In addition, acute regulation of H2S production is still poorly understood and additional work delineating the pathways regulating the enzymes that produce H2S will allow pharmacological manipulation of this pathway. As the field continues to grow, we expect that H2S-related compounds will find their way into clinical trials for diseases affecting the blood vessels.
148 citations
TL;DR: Mechanisms by which hyperhomocysteinemia affects cellular proteostasis are described, a comprehensive account of the biological chemistry of homocysteine-containing proteins is provided, and pathophysiological consequences and clinical implications of their formation are discussed.
Abstract: Epidemiological studies established that elevated homocysteine, an important intermediate in folate, vitamin B12, and one carbon metabolism, is associated with poor health, including heart and brai...
147 citations
TL;DR: This work will detail the known mechanisms of hydrogen sulfide in the mitochondria and the implications of its mitochondrial‐specific impacts in several pathologic conditions.
Abstract: Hydrogen sulfide can signal through 3 distinct mechanisms: 1) reduction and/or direct binding of metalloprotein heme centers, 2) serving as a potent antioxidant through reactive oxygen species/reactive nitrogen species scavenging, or 3) post-translational modification of proteins by addition of a thiol (-SH) group onto reactive cysteine residues: a process known as persulfidation. Below toxic levels, hydrogen sulfide promotes mitochondrial biogenesis and function, thereby conferring protection against cellular stress. For these reasons, increases in hydrogen sulfide and hydrogen sulfide-producing enzymes have been implicated in several human disease states. This review will first summarize our current understanding of hydrogen sulfide production and metabolism, as well as its signaling mechanisms; second, this work will detail the known mechanisms of hydrogen sulfide in the mitochondria and the implications of its mitochondrial-specific impacts in several pathologic conditions.-Murphy, B., Bhattacharya, R., Mukherjee, P. Hydrogen sulfide signaling in mitochondria and disease.
144 citations
Cites background from "Cystathionine β-synthase regulates ..."
...Persulfidationof these residues stabilizes SP1and regulates its transcription ofVEGF receptor 2 andneurolipin-1, 2major regulators of endothelial cell function (140)....
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...Moreover, our grouphas shown that persulfidation of SP1 by H2S can enhance its transcriptional activity, thereby indicating a potential mechanism by which HDmay be treated by H2S (140)....
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References
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TL;DR: The generation of mice deficient in Flk-1 by disruption of the gene using homologous recombination in embryonic stem (ES) cells is reported, indicating that FlK-1 is essential for yolk-sac blood-island formation and vasculogenesis in the mouse embryo.
Abstract: The receptor tyrosine kinase Flk-1 (ref. 1) is believed to play a pivotal role in endothelial development. Expression of the Flk-1 receptor is restricted to endothelial cells and their embryonic precursors, and is complementary to that of its ligand, vascular endothelial growth factor (VEGF), which is an endothelial-specific mitogen. Highest levels of flk-1 expression are observed during embryonic vasculogenesis and angiogenesis, and during pathological processes associated with neovascularization, such as tumour angiogenesis. Because flk-1 expression can be detected in presumptive mesodermal yolk-sac blood-island progenitors as early as 7.0 days postcoitum, Flk-1 may mark the putative common embryonic endothelial and haematopoietic precursor, the haemangioblast, and thus may also be involved in early haematopoiesis. Here we report the generation of mice deficient in Flk-1 by disruption of the gene using homologous recombination in embryonic stem (ES) cells. Embryos homozygous for this mutation die in utero between 8.5 and 9.5 days post-coitum, as a result of an early defect in the development of haematopoietic and endothelial cells. Yolk-sac blood islands were absent at 7.5 days, organized blood vessels could not be observed in the embryo or yolk sac at any stage, and haematopoietic progenitors were severely reduced. These results indicate that Flk-1 is essential for yolk-sac blood-island formation and vasculogenesis in the mouse embryo.
4,063 citations
TL;DR: Recent insights have shed light onto VEGFR signal transduction and the interplay between different V EGFRs and VEGF co-receptors in development, adult physiology and disease.
Abstract: Vascular endothelial growth-factor receptors (VEGFRs) regulate the cardiovascular system. VEGFR1 is required for the recruitment of haematopoietic precursors and migration of monocytes and macrophages, whereas VEGFR2 and VEGFR3 are essential for the functions of vascular endothelial and lymphendothelial cells, respectively. Recent insights have shed light onto VEGFR signal transduction and the interplay between different VEGFRs and VEGF co-receptors in development, adult physiology and disease.
2,894 citations
"Cystathionine β-synthase regulates ..." refers background in this paper
...To determine how CBS regulates EC function, we focused on the VEGF/VEGFR-2 signaling axis, arguably the predominant pathway responsible for EC proliferation, migration, and survival (28)....
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Journal Article•
TL;DR: Since the enzymatic abnormalities in both disorders share certain metabolic consequences, the conclusion has been reached that an elevated concentration of homocysteine, homocystine, or a derivative of hornocysteines is the common factor leading to arterial damage.
Abstract: NDiviDuAI with homocystinuria 12 have been found to lack normal activity of the enzyme cystathionine synthetase.3 In many of the patients progressive arterial disease develops in ildhood, frequently resulting in death from thrombosis in a vital organ45 In addition, congenital dislocation of the lenses, mental retardation, and skeletal abnormalitieseg, osteoporosis, arachnodactyly, and pectus excavatum or pectus carinatum-usually are foundL5' The vascular changes and other abnormalities encoumtered in homocystinuria have been attributed either to the metabolic effects of the elevated tissue concentrations of methionine, homocysteine, or homocystine, or to the metabolic consequences of decreased tissue concentrations of cystathionine found in the disease.7 In a child dying with homocystinuria, cystathioninuria, and methyl malonic aciduria, secondary to an abnormality of cobalamin (B12 ) metabolism, arterial lesions have been discovered that resemble in a striking way many of those found in cystathionine synthetase deficiency. The vascular findigs i this patient will be presented and compared with those in a patient with cystathionine synthetase deficiency. Since the enzymatic abnormalities in both disorders share certain metabolic consequences, the conclusion has been reached that an elevated concentration of homocysteine, homocystine, or a derivative of hornocysteine is the common factor leading to arterial damage. The possible role of elevated concentrations of homocysteine or its derivatives in the pathogenesis of arteriosclerosis in individls free of known enzyme deficiencies will be discussed and interpreted with particular reference to the findings in experimentally produced arteriosclerosis.
1,860 citations
TL;DR: Treatment with B vitamins did not lower the risk of recurrent cardiovascular disease after acute myocardial infarction and a harmful effect from combined B vitamin treatment was suggested.
Abstract: BACKGROUND Homocysteine is a risk factor for cardiovascular disease. We evaluated the efficacy of homocysteine-lowering treatment with B vitamins for secondary prevention in patients who had had an acute myocardial infarction. METHODS The trial included 3749 men and women who had had an acute myocardial infarction within seven days before randomization. Patients were randomly assigned, in a twoby-two factorial design, to receive one of the following four daily treatments: 0.8 mg of folic acid, 0.4 mg of vitamin B 12 , and 40 mg of vitamin B 6 ; 0.8 mg of folic acid and 0.4 mg of vitamin B 12 ; 40 mg of vitamin B 6 ; or placebo. The primary end point during a median follow-up of 40 months was a composite of recurrent myocardial infarction, stroke, and sudden death attributed to coronary artery disease. RESULTS The mean total homocysteine level was lowered by 27 percent among patients given folic acid plus vitamin B 12 , but such treatment had no significant effect on the primary end point (risk ratio, 1.08; 95 percent confidence interval, 0.93 to 1.25; P = 0.31). Also, treatment with vitamin B 6 was not associated with any significant benefit with regard to the primary end point (relative risk of the primary end point, 1.14; 95 percent confidence interval, 0.98 to 1.32; P = 0.09). In the group given folic acid, vitamin B 12 , and vitamin B 6 , there was a trend toward an increased risk (relative risk, 1.22; 95 percent confidence interval, 1.00 to 1.50; P = 0.05). CONCLUSIONS Treatment with B vitamins did not lower the risk of recurrent cardiovascular disease after acute myocardial infarction. A harmful effect from combined B vitamin treatment was suggested. Such treatment should therefore not be recommended. (ClinicalTrials.gov number, NCT00266487.)
1,371 citations
"Cystathionine β-synthase regulates ..." refers background in this paper
...Although theobservations reportedare suggestive of a causal role for Hcy in endothelial dysfunction, clinical trials using vitamin supplementation though lowered Hcy levels did not alter disease outcome (13, 14)....
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...However, clinical trials using vitamin supplements lowered Hcy but did not alter disease outcome, suggesting an intricate involvement of metabolites generated through the transsulfuration pathway in causing endothelial dysfunction (13, 14)....
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TL;DR: Ex vivo endogenous H2S physiologically modifies cysteine residues in many proteins, including glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and actin, converting Cysteine -SH groups to -SSH groups in a process the authors call S-sulfhydration.
Abstract: Hydrogen sulfide (H2S), a messenger molecule generated by cystathionine gamma-lyase, acts as a physiologic vasorelaxant. Mechanisms whereby H2S signals have been elusive. We now show that H2S physiologically modifies cysteines in a large number of proteins by S-sulfhydration. About 10 to 25% of many liver proteins, including actin, tubulin, and glyceraldehyde-3-phosphate dehydrogenase (GAPDH), are sulfhydrated under physiological conditions. Sulfhydration augments GAPDH activity and enhances actin polymerization. Sulfhydration thus appears to be a physiologic posttranslational modification for proteins.
1,027 citations
"Cystathionine β-synthase regulates ..." refers background or methods in this paper
...In conclusion, our work emphasizes the importance of CBS-mediated protein S-sulfhydration in maintaining vascular health and function and purports possible H2Sdonor based therapies for treatment of endothelial dysfunction in CBS deficiency....
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...Sulfhydration stabilizes and enhances binding of SP1 to the VEGFR-2 promoter Given thatH2S canmodify proteins through S-sulfhydration (24), we performed themaleimide assay in which the -SH and -SSH groups but not S-S in a protein react with fluorescentmaleimide (27) (Fig....
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...A few significant proteins have recently been identified to be targets of protein S-sulfhydration, including glyceraldehyde 3-phosphate dehydrogenase (GAPDH), actin, tubulin, NF-kB, Parkin, andCa2+-TRP channels (23, 25–27)....
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...Using the modified biotin-switch assay (23), we then determined if Sp1 could be sulfhydratedbyNaHS....
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...Recent studies have demonstrated that H2S functions by modifying free thiol groups (-SH) in a protein to form persulfides (-SSH), a process known as protein S-sulfhydration (23, 24)....
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