Transcriptional Induction of Cystathionine γ-Lyase, a Reactive Sulfur-Producing Enzyme, by Copper Diethyldithiocarbamate in Cultured Vascular Endothelial Cells

Abstract

As toxic substances can enter the circulating blood and cross endothelial monolayers to reach parenchymal cells in organs, vascular endothelial cells are an important target compartment for such substances. Reactive sulfur species protect cells against oxidative stress and toxic substances, including heavy metals. Reactive sulfur species are produced by enzymes, such as cystathionine γ-lyase (CSE), cystathionine β-synthase, 3-mercaptopyruvate sulfurtransferase, and cysteinyl-tRNA synthetase. However, little is known about the regulatory mechanisms underlying the expression of these enzymes in vascular endothelial cells. Bio-organometallics is a research field that analyzes biological systems using organic-inorganic hybrid molecules (organometallic compounds and metal coordinating compounds) as molecular probes. In the present study, we analyzed intracellular signaling pathways that mediate the expression of reactive sulfur species-producing enzymes in cultured bovine aortic endothelial cells, using copper diethyldithiocarbamate (Cu10). Cu10 selectively upregulated CSE gene expression in vascular endothelial cells independent of cell density. This transcriptional induction of endothelial CSE required both the diethyldithiocarbamate scaffold and the coordinated copper ion. Additionally, the present study revealed that ERK1/2, p38 MAPK, and hypoxia-inducible factor (HIF)-1α/HIF-1β pathways mediate transcriptional induction of endothelial CSE by Cu10. The transcription factors NF-κB, Sp1, and ATF4 were suggested to act in constitutive CSE expression, although the possibility that they are involved in the CSE induction by Cu10 cannot be excluded. The present study used a copper complex as a molecular probe to reveal that the transcription of CSE is regulated by multiple pathways in vascular endothelial cells, including ERK1/2, p38 MAPK, and HIF-1α/HIF-1β. Bio-organometallics appears to be an effective strategy for analyzing the functions of intracellular signaling pathways in vascular endothelial cells.

Synthesis of Reactive Sulfur Species in Cultured Vascular Endothelial Cells after Exposure to TGF-β1: Induction of Cystathionine γ-Lyase and Cystathionine β-Synthase Expression Mediated by the ALK5-Smad2/3/4 and ALK5-Smad2/3-ATF4 Pathways

“…Induction of CSE expression by a calcium-sensing receptor agonist is mediated by the calmodulin signaling pathway [38]. We previously found that transcriptional induction of CSE by Cu10 was mediated by the ERK1/2, p38 MAPK, and HIF-1α/β pathways in vascular endothelial cells [17]. Moreover, we observed that heavy metals can induce endothelial CSE expression through pathways other than those mentioned above (unpublished data).…”

“…Recently, we reported that among RSS-producing enzymes, the expression of CSE is selectively induced by FGF-2 [19]. Additionally, copper diethyldithiocarbamate (Cu10), a copper complex, also selectively enhances the transcriptional induction of CSE without changing the levels of other RSS-producing enzymes [17]. These results suggest that intracellular RSS levels are regulated particularly by the activity of CSE, and other RSS-producing enzymes such as CBS, 3-MST, and CARS2 contribute to the constitutive levels of RSS in vascular endothelial cells.…”

“…Reactive sulfur species (RSS), such as cysteine persulfide, glutathione persulfide, and hydrogen persulfide, can act as cytoprotective factors against electrophiles, including reactive oxygen species (ROS) and heavy metals [14][15][16]. Previously, we found that transcription of the gene encoding cystathionine γ-lyase (CSE), an RSS-producing enzyme, is induced by copper diethyldithiocarbamate through activation of the ERK1/2, p38 MAPK, and hypoxia-inducible factor-1α/β (HIF-1α/β) pathways in vascular endothelial cells [17]. Additionally, we revealed that fibroblast growth factor-2 (FGF-2) increases intracellular RSS levels by inducing CSE expression through the ERK1/2 pathway in vascular endothelial cells [18].…”

Sodium trisulfide, a sulfane sulfur donor, stimulates bovine aortic endothelial cell proliferation in culture

“…RSS production is catalyzed by enzymes such as cystathionine γ-lyase (Ida et al, 2014), cystathionine β-synthase (Ida et al, 2014), 3-mercaptopyruvate sulfurtransferase (Módis et al, 2013), and cysteinyl-tRNA synthetase (Akaike et al, 2017). We report a copper complex, copper diethyldithiocarbamate, that induces transcription of cystathionine γ-lyase in vascular endothelial cells (Fujie et al, 2020), suggesting that RSS production is regulated in response to exposure to chemical compounds, and may protect cells from toxic compound exposure. RSS are known to reduce cadmium cytotoxicity in vascular endothelial cells (Shinkai et al, 2017).…”

Fibroblast Growth Factor-2 Upregulates Reactive Sulfur Species Production <i>via</i> ERK1/2 Signal-Mediated Cystathionine γ-Lyase Induction in Cultured Bovine Aortic Endothelial Cells

“…9) We have reported that a copper complex, copper diethyldithiocarbamate, causes transcriptional induction of cystathionine γ-lyase (CSE), an RSS-producing enzyme, in the cells. 10) Additionally, RSS may modulate vascular endothelial cell functions such as proliferation. We recently revealed that sodium trisulfide, a sulfane sulfur donor, stimulates the proliferation of vascular endothelial cells.…”