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Abstract
Background m6A RNA methylation modification is closely associated with tumor progression, but the key m6A regulatory network and molecular mechanisms leading to hepatocellular carcinoma (HCC) metastasis remain unclear. This study investigates the role of RBMX, a novel m6A reader, in HCC metastasis.
Methods HCC tissues and adjacent normal liver tissues were used to study the correlation between RBMX expression level and tumor metastasis. The role of RBMX in HCC metastasis was studied using HCC cell lines. Multi-omics sequencing including meRIP-seq, RIP-seq and RNA-seq were employed to unravel the molecular mechanisms of RBMX. The activity of RhoA was assessed by a Rho activation assay. Liquid-liquid phase separation (LLPS) was investigated through confocal immunofluorescence.
Results We discovered that RBMX was abnormally overexpressed in HCC tissues and correlated with vascular invasion and poor prognosis (IDDF2024-ABS-0240 Figure 1. The expression level of RBMX is correlated with the advanced clinical stage, metastasis and poor prognosis of HCC). Functional assays indicated that RBMX enhanced the migration and invasion of HCC cells (IDDF2024-ABS-0240 Figure 2. RBMX promotes HCC cell migration and invasion). Integrative analysis including meRIP-seq, RIP-seq and RNA-seq revealed that RBMX facilitates HCC metastasis by regulating small GTPase-mediated signal transduction (IDDF2024-ABS-0240 Figure 3. Multiomic analyses identify ARHGAP5 as the key target of RBMX). Specifically, RBMX could upregulate ARHGAP5 mRNA stability by directly recognizing the ARHGAP5 m6A modification. Both RBMX and ARHGAP5 knockdown increased RhoA-GTP levels, with subsequent functional assays verifying the role of ARHGAP5 in RBMX-driven metastasis (IDDF2024-ABS-0240 Figure 3. Multiomic analyses identify ARHGAP5 as the key target of RBMX, IDDF2024-ABS-0240 Figure 4. ARHGAP5 contributes to RBMX-driven metastasis). Furthermore, we found that RBMX binds to m6A-RNA and forms phase-separated nuclear condensates, which might protect ARHGAP5 mRNA from degradation (IDDF2024-ABS-0240 Figure 5. RBMX proteins undergo LLPS with m6A-modified ARHGAP5 mRNA). Finally, lactate accumulated in the tumor microenvironment was found to potently induce RBMX upregulation in HCC cells via H3K18 lactylation (IDDF2024-ABS-0240 Figure 6. H3K18 lactylation upregulates RBMX expression in HCC cells).
The expression level of RBMX is correlated with the advanced clinical stage, metastasis and poor prognosis of HCC
RBMX promotes HCC cell migration and invasion
Multiomic analyses identify ARHGAP5 as the key target of RBMX
ARHGAP5 contributes to RBMX-driven metastasis
RBMX proteins undergo LLPS with m6A-modified ARHGAP5 mRNA
H3K18 lactylation upregulates RBMX expression in HCC cells
Conclusions Our data demonstrated that RBMX promotes HCC metastasis by binding and stabilizing m6A-modified ARHGAP5 through LLPS. Targeting the RBMX-ARHGAP5 interaction presents a promising therapeutic strategy for the treatment of HCC metastasis.