eNRSA: a faster and more powerful approach for nascent transcriptome analysis.

Nascent RNA sequencing tracks primary transcriptional events, making it crucial for studying the immediate regulatory changes of genes and enhancers in response to both endogenous and exogenous stimuli. NRSA is a widely used tool for analyzing nascent transcriptomic data, enabling quantification of transcriptional changes at proximal promoters and gene bodies, estimation of pausing indices, identifying active enhancers, and establishing enhancer-target gene relationships. To improve its functionality and broaden its applicability to diverse organisms and complex study designs, we have developed an enhanced version, eNRSA. Key advancements include adaptive selection of major transcripts, support for any organism with known gene structures, compatibility with complex study designs, and identification of alternative transcription start and termination sites, as well as transcription readthrough events. Additionally, eNRSA achieves a ∼20-fold increase in analysis speed while significantly reducing memory usage. These enhancements make eNRSA a faster, more versatile, and more powerful tool for nascent transcriptome analysis. eNRSA is freely available at https://bioinfo.vanderbilt.edu/eNRSA/.