{"doi":"10.1101/2025.10.27.684876","title":"An intron-based timer for circadian rhythms","abstract":"Abstract: Circadian clocks regulate daily rhythms in all eukaryotes through ∼24-hour transcriptional-translational feedback loops driven by clock proteins. However, the molecular mechanisms that set the 24-hour period remain poorly defined. Here, using single-molecule imaging and nascent RNA sequencing, we uncover an unexpected RNA-based molecular timer: a single intron in the Drosophila timeless ( tim ) gene that regulates circadian period length by controlling mRNA localization. Strikingly, we find that ∼50% of tim mRNAs are localized to the nucleus due to inefficient post-transcriptional splicing of a single intron (which we named intron P ), in contrast to other core clock transcripts that localize predominantly to the cytoplasm. CRISPR-mediated removal of intron P abolishes nuclear retention of tim transcripts, leading to accelerated TIM protein accumulation and a shortened ∼22-hour period with reduced rhythmic robustness. Remarkably, insertion of intron P alone into heterologous reporters is sufficient to promote nuclear retention in both Drosophila and human cells, acting as a conserved checkpoint that withholds transcripts in the nucleus until splicing is complete. Finally, we identify three RNA-binding proteins, two repressors (Hrb27C and Squid) and one activator (Qkr58E-2, a Sam68 homolog), that modulate intron P splicing in a rheostat-like manner. Together, these findings establish tim intron P as the first intron-based molecular timer in circadian clocks and reveal splicing kinetics as a critical regulatory layer in temporal gene expression programs, with broad implications for other processes such as development and immunity.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2025,"id":559381,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":1,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9367,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2025-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":822803,"name":"Amanda Linskens","orcid":"0000-0001-8678-3293","position":1,"is_corresponding":false},{"id":1461194,"name":"Rafael De Gouvea","orcid":null,"position":2,"is_corresponding":false},{"id":1461195,"name":"Haokai Liu","orcid":null,"position":3,"is_corresponding":false},{"id":572061,"name":"Yangbo Xiao","orcid":null,"position":4,"is_corresponding":false},{"id":1461196,"name":"Smriti Suresh","orcid":null,"position":5,"is_corresponding":false},{"id":1461197,"name":"Irene Hu","orcid":null,"position":6,"is_corresponding":false},{"id":334787,"name":"Swathi Yadlapalli","orcid":"0000-0001-5870-341X","position":7,"is_corresponding":false},{"id":571509,"name":"Ye Yuan","orcid":"0000-0001-9641-9102","position":0,"is_corresponding":true}],"reference_count":61,"raw_metadata":null,"created_at":"2026-07-19T02:55:34.849815Z","pmid":"41279210","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":[]}