SPLICE-q: a Python tool for genome-wide quantification of splicing efficiency

Integrator is a multi-subunit complex that directly interacts with the C-terminal domain (CTD) of RNA polymerase II (RNAPII). Through its RNA endonuclease activity, Integrator is required for 3′-end processing of both non-coding and coding transcripts. Here we demonstrate that depleting Integrator subunit 11 (INTS11), the main catalytic subunit of the Integrator complex, leads to a global elongation defect as a result of decreased polymerase processivity. We observe this defect in the region approximately 12 to 35 kb downstream of the transcription start site (TSS), where RNAPII normally transitions to its maximum processivity. We also identify an important role for INTS11, possibly in association with RNAPII CTD phospho-Tyr1, in repressing antisense transcription upstream of active promoters, as well as repressing transcription of genic regions near AsiSI-induced double-strand breaks. Altogether, this study points toward a novel function of Integrator in promoting termination of incompetent RNAPII molecules while facilitating the transition to fully processive polymerase in order to enable efficient elongation.

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“…Splicing efficiency was quantified using SPLICE-q with default parameters(de Melo Costa et al 2021).…”

Section: Methodsmentioning

confidence: 99%

Human Integrator provides a quality checkpoint during elongation to facilitate RNA polymerase II processivity

Rohban

Rafiee

Ule

et al. 2023

Preprint

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“…To quantify splicing efficiency of introns, we used SPLICE-q 81 version 1.0.0 to calculate the reverse intron expression ratio (IER) in protein coding genes using the Gencode v34 chromosomal annotation. By default, SPLICE-q uses the highest filtering settings for IER quantification, which selects only introns that do not overlap any exons of the same gene or of any other gene.…”

Section: Methodsmentioning

confidence: 99%

Global impact of aberrant splicing on human gene expression levels

Fair,

Abad Najar,

Zhao

et al. 2023

Preprint

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Nonsense-mediated decay (NMD) is an RNA surveillance mechanism that degrades transcripts containing premature termination codons. Alternative splicing (AS) is known to affect the expression levels of some human genes by producing transcripts rapidly degraded by NMD. However, the importance of AS in determining expression levels of most genes is unclear because rapidly degraded mRNA isoforms are difficult to quantify. To assess the impact of AS on gene expression levels, we analyzed population-scale data across eight molecular assays that capture gene regulation before, during, and after transcription and mRNA decay. Sequencing nascent RNA revealed that ~15% of all mRNA molecules are NMD targets, of which most result from low-usage cryptic splicing. Leveraging genetic variation across cell lines, we find that trait-associated loci explained by AS are similarly likely to associate with NMD-induced expression level differences as with differences in protein isoform usage, suggesting that much of the impact of AS is mediated by NMD. Finally, we used the splice-switching drug risdiplam to perturb AS at hundreds of genes, finding that ~3/4 of the splicing perturbations induce NMD. The remarkable prevalence of cryptic splice sites across the genome presents a rich opportunity for natural selection and therapeutics to shape the expression levels of nearly all human genes.

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“…The correlation between biological duplicates for all RNA-seq couples according to Pearson coefficient was > 0.98. SPLICE-q (de Melo Costa et al 2021) was used for splicing efficiency quantification.…”

Section: Methodsmentioning

confidence: 99%

A systematic screen identifies Saf5 as a link between splicing and transcription in fission yeast

Borao,

Vega,

Boronat

et al. 2023

Preprint

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Splicing is an important step of gene expression regulation in eukaryotes, as there are many mRNA precursors that can be alternatively spliced in different tissues, at different cell cycle phases or under different external stimuli. We have developed several life constructs that allow the quantification of fission yeast splicingin vivoon intact cells, and we have compared their splicing efficiency in a wild type strain and in aprp2-1(U2AF65) genetic background, showing a clear dependency between Prp2 and a consensus signal at 5’ splicing site (5’SS). To isolate novel genes involved in regulated splicing, we have crossed the reporter containing the weakest splicing rate with theSchizosaccharomyces pombeknock out collection. Among the candidate genes involved in the regulation of splicing, we have detected strong splicing defects in two of the mutants (Δcwf12andΔsaf5), both of them related to the NineTeen Complex (NTC). We have identified that strains with mutations incwf12have inefficient splicing, mainly when the 5’SS differs from the consensus. However, althoughΔsaf5cells also have some dependency on 5’SS sequence, we noticed that when one intron of a given pre-mRNA was affected, the rest of the introns of the same pre-mRNA had high probabilities of being also affected. This observation points Saf5 as a link between transcription rate and splicing.

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SPLICE-q: a Python tool for genome-wide quantification of splicing efficiency

Cited by 14 publications

References 38 publications

Human Integrator provides a quality checkpoint during elongation to facilitate RNA polymerase II processivity

Human Integrator provides a quality checkpoint during elongation to facilitate RNA polymerase II processivity

Global impact of aberrant splicing on human gene expression levels

A systematic screen identifies Saf5 as a link between splicing and transcription in fission yeast

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