{"doi":"10.1093/nar/gkae209","title":"RNA polymerase SI3 domain modulates global transcriptional pausing and pause-site fluctuations","abstract":"Transcriptional pausing aids gene regulation by cellular RNA polymerases (RNAPs). A surface-exposed domain inserted into the catalytic trigger loop (TL) of Escherichia coli RNAP, called SI3, modulates pausing and is essential for growth. Here we describe a viable E. coli strain lacking SI3 enabled by a suppressor TL substitution (β'Ala941→Thr; ΔSI3*). ΔSI3* increased transcription rate in vitro relative to ΔSI3, possibly explaining its viability, but retained both positive and negative effects of ΔSI3 on pausing. ΔSI3* inhibited pauses stabilized by nascent RNA structures (pause hairpins; PHs) but enhanced other pauses. Using NET-seq, we found that ΔSI3*-enhanced pauses resemble the consensus elemental pause sequence whereas sequences at ΔSI3*-suppressed pauses, which exhibited greater association with PHs, were more divergent. ΔSI3*-suppressed pauses also were associated with apparent pausing one nucleotide upstream from the consensus sequence, often generating tandem pause sites. These '-2 pauses' were stimulated by pyrophosphate in vitro and by addition of apyrase to degrade residual NTPs during NET-seq sample processing. We propose that some pauses are readily reversible by pyrophosphorolysis or single-nucleotide cleavage. Our results document multiple ways that SI3 modulates pausing in vivo and may explain discrepancies in consensus pause sequences in some NET-seq studies.","journal":"Nucleic Acids Research","year":2024,"id":438115,"datarank":0.40390274789667496,"base_score":2.3978952727983707,"endowment":2.3978952727983707,"self_citation_contribution":0.3596842909197557,"citation_network_contribution":0.044218456976919264,"self_endowment_contribution":0.3596842909197557,"citer_contribution":0.044218456976919264,"corpus_percentile":null,"corpus_rank":null,"citation_count":10,"citer_count":7,"citers_with_citation_signal":3,"citers_with_endowment":3,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9503,"is_data_producer":true,"deposit_databanks":{"GEO":["GSE230757","GSE240163"]},"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2024-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":631129,"name":"Xinyun Cao","orcid":"0000-0002-7346-3909","position":1,"is_corresponding":false},{"id":461536,"name":"Robert Landick","orcid":"0000-0002-5042-0383","position":2,"is_corresponding":false},{"id":816120,"name":"Yu Bao","orcid":"0000-0002-7694-8880","position":0,"is_corresponding":true}],"reference_count":75,"raw_metadata":null,"created_at":"2026-07-19T02:00:38.888629Z","pmid":"38554114","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}