Introduction

Liver cancer ranks as the sixth most common malignant tumor worldwide and the third leading cause of cancer-related deaths. Hepatocellular carcinoma (HCC), accounting for approximately 90% of primary liver cancers, is among the most lethal malignancies, with a mortality-to-incidence ratio as high as 0.92. Metastasis remains a pivotal determinant of HCC patient prognosis and therapeutic efficacy, making in-depth exploration of its underlying mechanisms a critical foundation for improving clinical outcomes. Ribosomes, composed of ribosomal RNA (rRNA) and ribosomal proteins (RPs), are central to protein synthesis. Emerging evidence highlights that ribosomal heterogeneity influences tumor biology through differentially regulating protein translation. However, the specific mechanisms by which ribosomal proteins contribute to HCC metastasis remain poorly characterized, underscoring a significant gap in oncological research.

Aims & Methods

We performed a screen for ribosomal genes associated with survival in HCC patients using multiple databases through a comprehensively bioinformatics analysis. We estimated the correlation between RPS8 expression and clinicopathological features of patients in HCC. We explored the biological function of RPS8 in HCC cells via lentiviral in vitro. We further assessed the tumorigenicity of RPS8 using both subcutaneous tumor formation in nude mouse and orthotopic liver tumorigenesis in NCG mouse. multi-omics were applied for exploring the mechanism of RPS8 in HCC.

Results

Comprehensive pathological evaluation of HCC clinical specimens revealed overexpression of RPS8 in tumor tissues, which exhibited significant associations with higher tumor stage, poorly differentiated histology , microvascular invasion,and reduced overall survival.Functional studies demonstrated that knockdown of RPS8 suppressed proliferation, migration, and invasion in HCC cell lines . Overexpression of RPS8 could enhance the malignant behavior of HCC cells. Moreover, RPS8 depletion reduced subcutaneous tumor volume and orthotopic liver tumor metastasis to lungs. Mechanistically, proteomic analysis and RIP-seq revealed interferon-stimulated gene 15 (ISG15) exhibited the most significant downregulation at the protein level upon RPS8 knockdown and RPS8 directly binds to ISG15 mRNA. GO and KEGG analysis indicate that RPS8 promotes HCC invasion and metastasis by modulating cell motility and migration. Among the top five differentially expressed proteins, CCDC25, a neutrophil extracellular trap (NET) repceptor, exhibited the most significant downregulation upon RPS8 knockdown. ISG15 can bind with target proteins to form ISGlation, thus preventing the ubiquitinlation. Bioinformatic predictions have identified six potential ubiquitination modification sites in CCDC25. Additionally, molecular docking analyses suggest the presence of potential ubiquitination modification sites within the binding interface between ISG15 and CCDC25. We then performed CCDC25 overexpression in RPS8-knockdown HCC cells and CCDC25 knockout in RPS8-overexpressing HCC cells. Downregulation or upregulation of CCDC25 partially reversed the oncogenic effects of RPS8 respectively, supporting CCDC25 as a critical downstream mediator of RPS8-driven HCC progression. Finally, we found that NETs from HCC patients could promote in vivo metastasis of control cells, but showed less efficacy in RPS8-knockdown cells.

Conclusion

Ribosomal protein RPS8 promotes HCC metastasis through upregulating ISG15 to stabilize NETs receptor CCDC25 protein. These findings provide a promising therapeutic strategy targeting .

References

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