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The thrombin receptor (PAR1) is associated with microtubules, mitosis and process formation in glioma cells.

ABSTRACT: The cell surface protease-activated receptor 1 (PAR1) is overexpressed in glioblastoma multiforme (GBM). We studied the function and structure of intracellular microtubule (MT) and PAR1 in a tubulin-mediated process. We found that exposure to thrombin increased the percentage of proliferative, S, and M phases cells, affected morphology, and increased process elongation. PAR1 antagonist inversely affects these measures, increases tubulin end-binding protein 3 (EB3) mRNA expression in C6 cells, and reduces EB3 comet length, track length, and duration in neuroblastoma cells. In addition, immunofluorescence staining suggests that PAR1 is in close association with the MT α-tubulin and with coagulation cascade proteins during cell division stages. Our findings support PAR1 involvement in MT dynamics.

SUBMITTER: Golderman V

PROVIDER: S-EPMC11254606 | biostudies-literature | 2024 Jun

REPOSITORIES: biostudies-literature

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The thrombin receptor (PAR1) is associated with microtubules, mitosis and process formation in glioma cells.

Golderman Valery V Gofrit Shany Guly SG Ivashko-Pachima Yanina Y Gozes Illana I Chapman Joab J Shavit-Stein Efrat E

Heliyon 20240619 12

The cell surface protease-activated receptor 1 (PAR1) is overexpressed in glioblastoma multiforme (GBM). We studied the function and structure of intracellular microtubule (MT) and PAR1 in a tubulin-mediated process. We found that exposure to thrombin increased the percentage of proliferative, S, and M phases cells, affected morphology, and increased process elongation. PAR1 antagonist inversely affects these measures, increases tubulin end-binding protein 3 (EB3) mRNA expression in C6 cells, an ...[more]

PMID: 39027436

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Project description:Essentials Elimination of PDAC tumor cell PAR1 increased cytotoxic T cells and reduced tumor macrophages. PAR1KO PDAC cells are preferentially eliminated from growing tumors. Thrombin-PAR1 signaling in PDAC tumor cells drives an immunosuppressive gene signature. Csf2 and Ptgs2 are thrombin-PAR1 downstream immune suppressor genes in PDAC tumor cells. ABSTRACT: Background Pancreatic ductal adenocarcinoma (PDAC) is characterized by a prothrombotic state and a lack of host antitumor immune responsiveness. Linking these two key features, we previously demonstrated that tumor-derived coagulation activity promotes immune evasion. Specifically, thrombin-protease-activated receptor-1 (PAR1) signaling in mouse PDAC cells drives tumor growth by evading cytotoxic CD8a+ cells. Methods Syngeneic mixed cell tumor growth, transcriptional analyses, and functional tests of immunosuppressive response genes were used to identify cellular and molecular immune evasion mechanisms mediated by thrombin-PAR-1 signaling in mouse PDAC tumor cells. Results Elimination of tumor cell PAR1 in syngeneic graft studies increased cytotoxic T lymphocyte (CTL) infiltration and decreased tumor-associated macrophages in the tumor microenvironment. Co-injection of PAR1-expressing and PAR1-knockout (PAR-1KO ) tumor cells into immunocompetent mice resulted in preferential elimination of PAR-1KO cells from developing tumors, suggesting that PAR1-dependent immune evasion is not reliant on CTL exclusion. Transcriptomics analyses revealed no PAR1-dependent changes in the expression of immune checkpoint proteins and no difference in major histocompatibility complex-I cell surface expression. Importantly, thrombin-PAR1 signaling in PDAC cells upregulated genes linked to immunosuppression, including Csf2 and Ptgs2. Functional analyses confirmed that both Csf2 and Ptgs2 are critical for PDAC syngeneic graft tumor growth and overexpression of each factor partially restored tumor growth of PAR1KO cells in immunocompetent mice. Conclusions Our results provide novel insight into the mechanisms of a previously unrecognized pathway coupling coagulation to PDAC immune evasion by identifying PAR1-dependent changes in the tumor microenvironment, a PAR1-driven immunosuppressive gene signature, and Csf2 and Ptgs2 as critical PAR1 downstream targets.

Project description:ATP-binding cassette transporters A1 (ABCA1) and G1 (ABCG1) play a vital role in promoting cholesterol efflux. Although, the dysregulation of these transporters was attributed as one of the mechanisms of atherogenesis, what renders their dysfunction is not well explored. Previously, we have reported that thrombin without having any effect on ABCG1 levels depletes ABCA1 levels affecting cholesterol efflux. In this study, we explored the mechanisms underlying thrombin-induced depletion of ABCA1 levels both in macrophages and smooth muscle cells. Under normal physiological conditions, COP9 signalosome subunit 3 (CSN3) was found to exist in complex with ABCA1 and in the presence of proatherogenic stimulants such as thrombin, ABCA1 was phosphorylated and dissociated from CSN3, leading to its degradation. Forced expression of CSN3 inhibited thrombin-induced ABCA1 ubiquitination and degradation, restored cholesterol efflux and suppressed foam cell formation. In Western diet (WD)-fed ApoE-/- mice, CSN3 was also disassociated from ABCA1 otherwise remained as a complex in Chow diet (CD)-fed ApoE-/- mice. Interestingly, depletion of CSN3 levels in WD-fed ApoE-/- mice significantly lowered ABCA1 levels, inhibited cholesterol efflux and intensified foam cell formation exacerbating the lipid laden atherosclerotic plaque formation. Mechanistic studies have revealed the involvement of Par1-Gα12-Pyk2-Gab1-PKCθ signaling in triggering phosphorylation of ABCA1 and its disassociation from CSN3 curtailing cholesterol efflux and amplifying foam cell formation. In addition, although both CSN3 and ABCA1 were found to be colocalized in human non-lesion coronary arteries, their levels were decreased as well as dissociated from each other in advanced atherosclerotic lesions. Together, these observations reveal for the first time an anti-atherogenic role of CSN3 and hence, designing therapeutic drugs protecting its interactions with ABCA1 could be beneficial against atherosclerosis.

Dataset Information

The thrombin receptor (PAR1) is associated with microtubules, mitosis and process formation in glioma cells.

Publications

The thrombin receptor (PAR1) is associated with microtubules, mitosis and process formation in glioma cells.

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