He, K. L. | bioRxiv Science

bioRxiv2025

Transcription termination promotes splicing efficiency and fidelity in a compact genome

Splicing of terminal introns is coupled to 3'-end processing by cleavage and polyadenylation (CPA) of mRNAs in mammalian genes. Whether this functional coupling is universally conserved across eukaryotes is unclear. Here we show using long read RNA sequencing in S.cerevisiae that splicing inactivation does not result in widespread CPA impairment. We also show that inactivation of CPA has limited impact on splicing efficiency. The negative impact of CPA inactivation on splicing is mainly due to transcription termination defects that promote readthrough transcription, leading to splicing inhibition for downstream intron-containing genes. Splicing impairment due to 5' extensions is length-dependent and can be detected independently from CPA inactivation for endogenous or synthetic genes, and is likely due to an increased distance of splicing signals to the 5' cap. Finally, we found that deficient termination can promote novel intragenic and long-range intergenic splicing events. These results argue against a broad coupling between splicing and CPA in S.cerevisiae but show that efficient CPA-mediated transcription termination is critical for splicing fidelity and efficiency in a compact genome. Significance StatementAccurate gene expression requires that the enzyme that polymerizes RNA stops at the proper site (termination). In addition multiple RNA processing reactions, including removal of intervening sequences are necessary to produce mature mRNAs. How these different steps in the RNA biogenesis pathways influence each other is not well understood. In this study, we show that inactivation of termination induces mature RNA formation defects, including long RNAs that retain intervening sequences, or chimeric RNAs containing sequences from genes located next to each other on the genome. This study underscores the importance of proper termination to ensure accurate and efficient splicing of adjacent genes, which is particularly critical for compact genomes in which genes are located close to each other.

molecular biology↗

bioRxiv2020

Natural Antisense Transcript of Period2, Per2AS, regulates the amplitude of the mouse circadian clock

Circadian transcriptome studies identified a novel transcript at the Period2 (Per2) locus, which we named Per2AS. Per2AS is a long non-coding RNA transcribed from the antisense strand of Per2, and is expressed rhythmically and anti-phasic to Per2 mRNA. Previously, we mathematically tested the hypothesis that Per2AS and Per2 mutually inhibit each others expression by forming a double negative feedback loop, and found that Per2AS expands the oscillatory domain. In this study, we have experimentally tested this prediction by perturbing the expression of Per2AS in mouse fibroblasts. We found that Per2AS represses Per2 pre-transcriptionally in cis and regulates the amplitude of the circadian clock, but not period or phase. Unexpectedly, we also found that Per2 positively regulates Per2AS post-transcriptionally, indicating that Per2AS and Per2 form a single negative feedback loop. Because knock-down of Per2 does not recapitulate the phenotypes of Per2AS perturbation and Per2AS also activates Bmal1 in trans, we propose that Per2AS regulates the amplitude of the circadian clock without producing a protein by rewiring the molecular clock circuit.

molecular biology↗