Dataset Information

Endothelial GATA6 deficiency suppresses intracellular TLR3-interferon signaling in HPAECs and promotes interferon response in HPASMCs

ABSTRACT: GATA6 is a key transcription factor crucial for maintaining endothelial cell (EC) homeostasis. The dysregulation of endothelial immune function is a central feature in diseases such as pulmonary arterial hypertension (PAH). In this study, we explored the consequences of GATA6 deficiency in human pulmonary arterial endothelial cells (HPAECs) and its impact on immune response pathways. We report that siRNA-induced GATA6 deficiency or the GATA inhibitor led to significant downregulation of interferon response genes and a marked reduction in toll-like receptor 3 (TLR3) expression. GATA6 overexpression enhanced the expression of these genes, and TLR3 inhibition abrogated this response. Furthermore, conditioned medium from GATA6-deficient HPAECs upregulated interferon genes in pulmonary artery smooth muscle cells (HPASMCs), indicating a paracrine effect. Overall, these findings highlight the critical role of GATA6 in modulating TLR3 signaling and immune responses in endothelial cells and suggest its involvement in endothelial-smooth muscle cell interactions in vascular inflammation.

ORGANISM(S): Homo sapiens

PROVIDER: GSE293379 | GEO | 2026/03/23

REPOSITORIES: GEO

ACCESS DATA

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

Project description:Oxidants induce cell cycle arrest to halt cell proliferation; however, little is known about the redox-regulated effector proteins that mediate these processes. Here, we report a novel kinase-inhibitory disulfide bond in cyclin D-CDK4 and investigate its role in cell proliferation and PH. Oxidative modifications of cyclin D-CDK4 were detected in human pulmonary arterial smooth muscle cells (HPASMCs) and human pulmonary arterial endothelial cells (HPAECs). Cysteine to alanine mutants were generated, and cell cycle experiments were employed to characterize the nature of this reversible intermolecular disulfide bond. The functional role of the disulfide was delineated using in vitro kinase activity assays, HPASMCs and knock-in cells. Finally, the cyclin D-CDK4 disulfide was assessed in vivo in the pulmonary arteries and isolated HPASMCs of PAH patients, and in three preclinical models of PH. Cyclin D-CDK4 forms an oxidant-induced heterodimeric disulfide dimer between C7/8 and C135, respectively. This reversible modification forms in cells in vitro and in pulmonary arteries in vivo to inhibit cyclin D-CDK4 kinase activity and decrease retinoblastoma protein (Rb) phosphorylation. Correspondingly, treatment of HPASMCs with H2O2 or auranofin induces cell cycle arrest. Notably, mutation of CDK4 C135 causes a kinase-impaired phenotype, which decreases the proliferation rate of cells, suggesting this cysteine is indispensable for cyclin D-CDK4 kinase activity. Pulmonary arteries and HPASMCs from patients with pulmonary arterial hypertension (PAH) display a decreased level of CDK4 disulfide, consistent with CDK4 being hyperactive in PAH. Furthermore, auranofin treatment, which induces the cyclin D-CDK4 disulfide, attenuates disease severity in experimental models of PH, by mitigating pulmonary vascular remodeling. A novel disulfide bond in cyclin D-CDK4 acts as a rapid switch to inhibit kinase activity and halt cell proliferation. This oxidative modification forms at a critical cysteine residue, which is unique to CDK4, offering the potential for design of a selective covalent inhibitor that may prove beneficial in pulmonary hypertension

Project description:Introduction. Pulmonary arterial hypertension (PAH) is a severe cardiopulmonary disease that may be triggered by exposure to drugs such as dasatinib or facilitated by genetic predispositions. The incidence of dasatinib-associated PAH is estimated at 0.45%, suggesting individual predispositions. The mechanisms of dasatinib-associated PAH are still incomplete. This study studied the link between dasatinib and KCNK3 and the consequences of dasatinib exposure or KCNK3 knockdown in pulmonary endothelial cells (PECs) and pulmonary arterial smooth muscle cells (PASMC). Methods. We discovered a KCNK3 gene variant in a patient with dasatinib-associated PAH and investigated the impact of this variant on KCNK3 function using patch-clamp analysis. Additionally, we assessed the effects of dasatinib exposure on KCNK3 expression and function in human and rat pulmonary arteries. In control-human PASMCs and PECs (hPASMCs and hPECs), we evaluated the consequence of KCNK3 knockdown on cell migration, mitochondrial membrane potential, ATP production, and in vitro tube formation. Using mass spectrometry, we determined the KCNK3 interactome. Results. Patch-clamp revealed that the identified KCNK3 variant represents a loss-of-function variant. Dasatinib contributed to pulmonary artery constriction by decreasing KCNK3 function and expression. In control-hPASMCs, KCNK3 knockdown promotes mitochondrial membrane depolarization and glycolytic shift. Additionally, dasatinib exposure or KCNK3 knockdown reduced the number of caveolae in PECs. Moreover, KCNK3 knockdown in control-hPECs reduced migration, proliferation, and in vitro tubulogenesis. Lastly, using proximity ligation assay and mass spectrometry, we identified the KCNK3 interactome revealing that KCNK3 interaction with various proteins across different cellular compartments could impact these cellular functions. Conclusion. We identified a novel pathogenic variant in the KCNK3 gene, suggesting that KCNK3 gene variations could also influence the development of dasatinib-associated PAH. Our results support that one of the mechanisms of action of dasatinib-associated PAH results from the downregulation of KCNK3.

Project description:Despite recent improvements in management of idiopathic pulmonary arterial hypertension, mortality remains high. Understanding the alterations in the transcriptome–phenotype of the key lung cells involved could provide insight into the drivers of pathogenesis. In this study, we examined differential gene expression of cell types implicated in idiopathic pulmonary arterial hypertension from lung explants of patients with idiopathic pulmonary arterial hypertension compared to control lungs. After tissue digestion, we analyzed all cells from three idiopathic pulmonary arterial hypertension and six control lungs using droplet-based single cell RNA-sequencing. After dimensional reduction by t-stochastic neighbor embedding, we compared the transcriptomes of endothelial cells, pericyte/smooth muscle cells, fibroblasts, and macrophage clusters, examining differential gene expression and pathways implicated by analysis of Gene Ontology Enrichment. We found that endothelial cells and pericyte/smooth muscle cells had the most differentially expressed gene profile compared to other cell types. Top differentially upregulated genes in endothelial cells included novel genes: ROBO4, APCDD1, NDST1, MMRN2, NOTCH4, and DOCK6, as well as previously reported genes: ENG, ORAI2, TFDP1, KDR, AMOTL2, PDGFB, FGFR1, EDN1, and NOTCH1. Several transcription factors were also found to be upregulated in idiopathic pulmonary arterial hypertension endothelial cells including SOX18, STRA13, LYL1, and ELK, which have known roles in regulating endothelial cell phenotype. In particular, SOX18 was implicated through bioinformatics analyses in regulating the idiopathic pulmonary arterial hypertension endothelial cell transcriptome. Furthermore, idiopathic pulmonary arterial hypertension endothelial cells upregulated expression of FAM60A and HDAC7, potentially affecting epigenetic changes in idiopathic pulmonary arterial hypertension endothelial cells. Pericyte/smooth muscle cells expressed genes implicated in regulation of cellular apoptosis and extracellular matrix organization, and several ligands for genes showing increased expression in endothelial cells. In conclusion, our study represents the first detailed look at the transcriptomic landscape across idiopathic pulmonary arterial hypertension lung cells and provides robust insight into alterations that occur in vivo in idiopathic pulmonary arterial hypertension lungs.

Dataset Information

Endothelial GATA6 deficiency suppresses intracellular TLR3-interferon signaling in HPAECs and promotes interferon response in HPASMCs

Dataset's files

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets