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Characterization of serotonin-5-HTR1E signaling pathways and its role in cell survival.

ABSTRACT: 5-Hydroxy tryptamine receptor 1E (5-HTR1E) is reported to activate cAMP and ERK pathways via its ligands and binding partners, but the detailed mechanism underlying the serotonin induced 5-HTR1E signaling is still not known. In the present study, we determined the cellular regulators of ERK and cAMP signaling pathways in response to serotonin induced 5-HTR1E activation in 5-HTR1E overexpressing HEK293 cells. We found that Pertussis Toxin (PTX) treatment completely reversed the effect of serotonin-5-HTR1E mediated signaling on cAMP and ERK pathways, confirming the involvement of a Gαi-linked cascade. We also observed that Gβγ and Gq were not associated with 5-HTR1E activation, while blocking PKA inhibited ERK signaling only, and had no effect on cAMP. Additionally, serotonin-stimulated ERK1/2 phosphorylation was similar in 5-HTR1E overexpressing, β-arrestin-deficient HEK293 cells and is solely dependent on G protein signaling. siRNA mediated gene knockout studies in SH-SY5Y cells revealed that the inhibition of 5-HTR1E reduced the expression of cMyc, Cyclin D1, Cyclin E and BCL2 genes which are related to cell cycle regulation and survival. MTT assays showed that 5-HTR1E knockdown in SHSY-5Y and U118 cells inhibited cell survival significantly. In addition to the signaling mechanism, we also performed RNA-seq analysis in 5-HTR1E overexpressing HEK293 cells and found that 5-HTR1E can regulate the expression of Receptor activity modifying protein 1 ( RAMP1 ), Nuclear receptor 1 ( NR4A1 ) and other Cyclin genes. These findings indicate that serotonin interaction with 5-HTR1E receptor simultaneously activates cAMP and ERK pathway in HEK293 cells and its expression is important for cell survival.

SUBMITTER: Sharma VK

PROVIDER: S-EPMC9928056 | biostudies-literature | 2023 Jan

REPOSITORIES: biostudies-literature

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Characterization of serotonin-5-HTR1E signaling pathways and its role in cell survival.

Sharma Vinay Kumar VK Campbell Kiersten K Yang Xuyu X Dale Ryan R Loh Y Peng YP

Research square 20230127

5-Hydroxy tryptamine receptor 1E (5-HTR1E) is reported to activate cAMP and ERK pathways <i>via</i> its ligands and binding partners, but the detailed mechanism underlying the serotonin induced 5-HTR1E signaling is still not known. In the present study, we determined the cellular regulators of ERK and cAMP signaling pathways in response to serotonin induced 5-HTR1E activation in 5-HTR1E overexpressing HEK293 cells. We found that Pertussis Toxin (PTX) treatment completely reversed the effect of s ...[more]

PMID: 36789437

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Project description:Rationale: Psychological stress has been linked to cancer development and resistance to therapy by many epidemiological and clinical studies. Stress-induced immunosuppressive microenvironment by stress hormones, in particular glucocorticoids, has been extensively studied. However, the impacts of other stress-related neurotransmitters, such as serotonin (5-hydroxytryptamine, 5-HT), on cancer development just start to be revealed. Here, we aimed to identify novel neurotransmitters involved in stress-induced growth and dissemination of ovarian cancer (OC) and reveal the major underlying signaling pathway and the therapeutic significance. Methods: Through a genome-wide CRISPR/Cas9 knockout screen in the murine orthotopic model of ovarian carcinoma (OC), we identified candidate genes regulating the peritoneal dissemination of OC. Among them, we picked out HTR1E, one member of 5-HT receptor family specifically expressed in the ovary and endometrium in addition to brain. The correlation of HTR1E expression with OC progression was analyzed in OC patient specimen by quantitative reverse transcription polymerase chain reaction (qRT-PCR), western blot, and immunohistochemistry (IHC). Gain-of-function and loss-of-function analyses were performed to explore the functions of 5-HT/HTR1E signaling in OC growth and dissemination in vitro and in vivo. In addition, we investigated the therapeutic values of HTR1E specific agonist and small molecular inhibitors against HTR1E downstream factor SRC in a stressed murine OC xenograft model. Results: In OC patients, the HTR1E expression is dramatically decreased in peritoneal disseminated OC cells, which correlates with poor clinical outcome. Silence of HTR1E in OC cells greatly promotes cell proliferation and epithelial mesenchymal transition (EMT) by the activation of SRC-mediated downstream signaling pathways. Furthermore, chronic stress results in significantly decreased serotonin in the ovary and the enhanced OC growth and peritoneal dissemination in mice, which can be strongly inhibited by specific HTR1E agonist or the SRC inhibitor. Conclusions: We discovered the essential role of serotonin/HTR1E signaling in preventing the chronic psychological stress-promoted progression of OC, suggesting the potential therapeutic value of the HTR1E specific agonist and the SRC inhibitor for OC patients who are suffering from psychological stress.

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Characterization of serotonin-5-HTR1E signaling pathways and its role in cell survival.

Publications

Characterization of serotonin-5-HTR1E signaling pathways and its role in cell survival.

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