RHOJ enhances adhesion and proliferation capabilities and suppresses apoptosis of melanoma cells by activating the Rap1 signaling pathway.

BACKGROUND

Melanoma is an aggressive skin cancer derived from melanocytes, known for its high metastatic potential and poor prognosis. Understanding the molecular mechanisms underlying melanoma progression could provide novel therapeutic targets for improving treatment outcomes. Our study aims to investigate the role of the RHO family GTPase RHOJ in melanoma progression and its regulation of cell adhesion, proliferation, and apoptosis through the Rap1 signaling pathway.

METHODS

The Gene Expression Omnibus (GEO) dataset GSE122907 and the Gene Expression Profiling Interactive Analysis (GEPIA) database were used to analyze differentially expressed genes related to melanoma. A375 cells were employed as the in vitro melanoma model. The STRING database was utilized to identify RHOJ-associated genes, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis was performed based on these genes. A375 cells were transfected with si-RHOJ, with or without the addition of a Rap1 signaling pathway activator. Cell viability was assessed using the Cell Counting Kit-8 (CCK-8) assay, while cell proliferation was measured using the 5-ethynyl-2'-deoxyuridine (EdU) assay. Apoptosis was evaluated by flow cytometry, and cell adhesion was determined using a cell adhesion detection kit. The expression of relevant genes was analyzed by real-time polymerase chain reaction (PCR), Western blot, and immunofluorescence techniques.

RESULTS

RHOJ, a key differentially expressed gene associated with melanoma, was significantly upregulated in melanoma cells, particularly in A375 cells. Knockdown of RHOJ reduced cell viability and proliferation, increased cell apoptosis, upregulated Bax, and downregulated Bcl-2. Additionally, cell adhesion was diminished, accompanied by the upregulation of E-cadherin and the downregulation of vinculin. The Rap1 signaling pathway was identified as a key pathway regulated by RHOJ. The levels of RAP1, RAP1GAP, and RasGRP3 were decreased in A375 cells transfected with si-RHOJ; however, these changes were reversed by activation of the Rap1 signaling pathway. Moreover, we found that the Rap1 signaling pathway activator could reverse the reduction in cell viability, proliferation, and adhesion, as well as the increase in apoptosis induced by si-RHOJ.

CONCLUSIONS

In conclusion, RHOJ promotes melanoma cell adhesion and proliferation while inhibiting apoptosis through the activation of the Rap1 signaling pathway, highlighting the potential clinical implications of targeting RHOJ in melanoma treatment.