Single-cell total-RNA profiling unveils regulatory hubs of transcription factors

• Published in: Nature communications Vol. 15; no. 1; pp. 5941 - 18
• Main Authors: Niu, Yichi, Luo, Jiayi, Zong, Chenghang
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group 15.07.2024; Nature Publishing Group UK; Nature Portfolio
• Subjects: Cell cycle; Chemistry; Comparative analysis; Gene expression; Gene regulation; Hubs; Oncogenes; Post-transcription; Ribonucleic acid; RNA; Senescence; Transcription factors; Velocity
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-024-50291-3

Abstract Recent development of RNA velocity uses master equations to establish the kinetics of the life cycle of RNAs from unspliced RNA to spliced RNA (i.e., mature RNA) to degradation. To feed this kinetic analysis, simultaneous measurement of unspliced RNA and spliced RNA in single cells is greatly desired. However, the majority of single-cell RNA-seq chemistry primarily captures mature RNA species to measure gene expressions. Here, we develop a one-step total-RNA chemistry-based single-cell RNA-seq method: snapTotal-seq. We benchmark this method with multiple single-cell RNA-seq assays in their performance in kinetic analysis of cell cycle by RNA velocity. Next, with LASSO regression between transcription factors, we identify the critical regulatory hubs mediating the cell cycle dynamics. We also apply snapTotal-seq to profile the oncogene-induced senescence and identify the key regulatory hubs governing the entry of senescence. Furthermore, from the comparative analysis of unspliced RNA and spliced RNA, we identify a significant portion of genes whose expression changes occur in spliced RNA but not to the same degree in unspliced RNA, indicating these gene expression changes are mainly controlled by post-transcriptional regulation. Overall, we demonstrate that snapTotal-seq can provide enriched information about gene regulation, especially during the transition between cell states.