TMEM158-mediated TGF-β signaling pathway modulates the sensitivity of TP53-deficient osteosarcoma to USP14 inhibitors

Background Previous studies have demonstrated that the USP14 inhibitor IU1 and USP14/UCHL5 inhibitor b-AP15 can extend the survival period of TP53-deficient mice with spontaneous osteosarcoma (OS). However, the underlying molecular mechanisms remain to be fully elucidated. The transmembrane protein TMEM158 has been identified as a key regulator in the progression of various cancers. Nevertheless, its functional role in OS remains largely unknown. Methods In this study, we conducted comprehensive bioinformatics analyses—including cluster analysis, differential expression analysis, and functional enrichment analysis—on clinical OS databases to assess the correlation between TMEM158 expression and the proteasome-associated USP14 and UCHL5. Primary tumor cells (TP53-deficient OS cells) and U2OS cells were treated with IU1 or b-AP15, respectively. The expression levels of TMEM158 were quantified using qPCR. Subsequently, TMEM158 was knocked down in both cell lines, and changes in cell viability and TGF-β signaling activity were evaluated. Additionally, single-cell RNA sequencing data were analyzed to identify cell types with high TMEM158 expression and their potential roles in intercellular communication. Results Both IU1 and b-AP15 significantly prolonged the survival of TP53-deficient OS mice and exhibited enhanced cytotoxic effects on TP53-deficient OS cells. These compounds selectively suppressed TMEM158 expression in TP53-deficient OS cells. Bioinformatics analysis revealed that TMEM158 is positively correlated with USP14 and UCHL5 expression and serves as an independent prognostic marker for poor clinical outcomes in OS patients. Experimental validation showed that TMEM158 knockdown significantly reduced the viability of TP53-deficient OS cells and inhibited TGF-β pathway activation. Osteoblastic OS cells displayed concurrent suppression of the P53 pathway and activation of the TGF-β pathway, with a strong covariant relationship between TMEM158 and TGF-β activity. Meanwhile, osteoblastic OS cells with high expression levels of TMEM158 demonstrate enhanced intercellular TGF-β signaling communication with macrophages. Conclusion Our findings demonstrated that the TMEM158-TGF-β pathway plays a central role in mediating the heightened sensitivity of TP53-deficient OS to USP14 inhibition. Targeting this pathway may offer a promising therapeutic strategy for precision treatment of osteosarcoma.