biostudies-literatureNie JJiangxi Provincial Natural Science FoundationNational Natural Science Foundation of ChinaJiangxi Province Postgraduate Innovation Special Fund Project159https://www.ebi.ac.uk/biostudies/studies/S-EPMC12016182biostudies-literatureUnknown25(1)Dysregulation of R-loops has been implicated in tumor development, progression, and the regulation of tumor immune microenvironment (TME). However, their roles in osteosarcoma (OS) remain underexplored. In this study, we firstly constructed a novel R-loop Gene Prognostic Score Model (RGPSM) based on the RNA-sequencing (RNA-seq) datasets and evaluated the relationships between the RGPSM scores and the TME. Additionally, we identified key R-loop-related genes involved in OS progression using single-cell RNA sequencing (scRNA-seq) dataset, and validated these findings through experiments. We found that patients with high-RGPSM scores exhibited poorer prognosis, lower Huvos grades and a more suppressive TME. Moreover, the proportion of malignant cells was significantly higher in the high-RGPSMCancer cell internationalIdentifying PSIP1 as a critical R-loop regulator in osteosarcoma via machine-learning and multi-omics analysis.PMC120161828236054320232ACB206043YC2023-B067Liu JLiu ZWang SNie JZhong YYang FfalseIdentifying PSIP1 as a critical R-loop regulator in osteosarcoma via machine-learning and multi-omics analysis.Dysregulation of R-loops has been implicated in tumor development, progression, and the regulation of tumor immune microenvironment (TME). However, their roles in osteosarcoma (OS) remain underexplored. In this study, we firstly constructed a novel R-loop Gene Prognostic Score Model (RGPSM) based on the RNA-sequencing (RNA-seq) datasets and evaluated the relationships between the RGPSM scores and the TME. Additionally, we identified key R-loop-related genes involved in OS progression using single-cell RNA sequencing (scRNA-seq) dataset, and validated these findings through experiments. We found that patients with high-RGPSM scores exhibited poorer prognosis, lower Huvos grades and a more suppressive TME. Moreover, the proportion of malignant cells was significantly higher in the high-RGPSM2025-01-01T00:00:00Z2025 Apr2026-06-02T04:12:53.844Z2025-07-06T03:04:31.003ZS-EPMC120161824026416410.1186/s12935-025-03775-1