Biology subjects

Vlasenok, M.

Publications and source records attributed to Vlasenok, M..

6 recordsLinked to original sources

bioRxiv2026

Estimation of splicing metrics for NMD-sensitive transcripts

Alternative splicing is commonly quantified using the Percent-Spliced-In (PSI) metric, which measures the relative abundances of alternatively spliced isoforms. However, some transcript isoforms are targeted by the nonsense-mediated decay (NMD) pathway, introducing a strong bias that leads to underestimation of their true splicing rates. To correct for this bias, we developed an analytical framework and a set of statistical models employing a linear fractional transformation depending on a single parameter capturing the degradation rate of NMD-sensitive transcripts relative to normal mRNA decay. Using Gaussian mixture models, we demonstrated a clear separation of splicing events into two classes, responders and non-responders, with the former exhibiting strong upregulation upon NMD inhibition and the latter showing little or no response. Moreover, non-responders displayed higher coding potential and stronger translation signals both upstream and downstream of the stop codon, which are characteristic of NMD escape through translational readthrough. We further showed that incorporation of event-specific relative decay rates improves the interpretation of differential splicing patterns for NMD-sensitive transcripts. In sum, our results provide a solid framework for unbiased estimation of splicing metrics in NMD-sensitive transcripts from short-read RNA-seq data, without requiring NMD inhibition experiments.

bioinformatics↗

bioRxiv2025

Novel examples of NMD escape through alternative intronic polyadenylation

The nonsense-mediated mRNA decay (NMD) surveillance system detects and selectively eliminates transcripts with premature stop codons. A stop codon is considered premature if it is followed by an exon-exon junction more than 50 nucleotides downstream. Pruning of the 3-untranslated region containing such junctions through alternative polyadenylation may provide a mechanism of NMD escape. Here, we systematically examine a subclass of poison exons that carry a premature stop codon for the presence of polyadenylation sites in the downstream intron. Using data from the GTEx consortium, we observed that poison exons are more often followed by an active polyadenylation site compared with cassette exons. We also identified tissue-specific switches between NMD-targeted and NMD-escape isoforms in several human genes, including the vaccinia-related kinase VRK3, nuclear transcription factor NFX1, Notch pathway regulator TM2D3, and RNA helicase DDX31. Blocking the cleavage and polyadenylation sites in these genes using antisense oligonucleotides in human cells led to a switch from NMD-escape to NMD-target isoform, accompanied by a decrease in gene expression levels. This study reveals that NMD escape via alternative polyadenylation is a widespread, yet currently overlooked post-transcriptional mechanism of gene expression regulation.

bioinformatics↗

bioRxiv2022

Transcriptional Responses of Cancer Cells to Heat Shock-Inducing Stimuli Involve Amplification of Robust HSF1 Binding

Responses of cells to signals are increasingly discovered to involve the binding of sequence-specific transcription factors outside of known target genes. We wanted to determine to what extent the genome-wide binding and function of a transcription factor are shaped by the cell type versus the stimulus. To do so, we induced the Heat Shock Response pathway in two distant cell lines with two different stimuli and related the binding of its master regulator HSF1 to nascent RNA and chromatin accessibility. We show that HSF1 binding patterns robustly retain their identity under different magnitudes of activation so that common HSF1 binding is globally associated with stimulus-specific transcription outcomes. HSF1-induced increase in DNA accessibility was modest in scale but occurred predominantly at remote genomic sites. Apart from regulating transcription at existing elements including promoters and enhancers, responses to heat shock may directly engage inactive chromatin.

genomics↗

bioRxiv2022

RNAcontacts, a pipeline for predicting contacts from RNA proximity ligation assays

BackgroundHigh-throughput RNA proximity ligation assays are molecular methods that simultaneously analyze spatial proximity of many RNAs in living cells. Their principle is based on cross-linking, fragmentation, and consequent religation of RNAs followed by high-throughput sequencing. The generated fragments have two distinct types of splits, one resulting from pre-mRNA splicing, and the other resulting from ligating spatially close RNA strands. FindingsHere, we present RNAcontacts, a universal pipeline for detecting RNA-RNA contacts in high-throughput RNA proximity ligation assays. It circumvents the inherent problem of mapping sequences with two distinct split types using a two-pass alignment, in which splice junctions are inferred from a control RNA-seq experiment on the first pass and then provided to the aligner on the second pass as bona fide introns. This approach allows for a more sensitive detection of RNA contacts and has higher specificity with respect to splice junctions that are present in the biological sample in comparison to previously developed methods. RNAcontacts extracts contacts, clusters their ligation points, computes the read support, and generates tracks for the visualization through the UCSC Genome Browser. It is implemented in a reproducible and scalable workflow management system Snakemake that allows fast and uniform processing of multiple datasets. ConclusionsRNAcontacts represents a generic pipeline for the detection of RNA contacts that can be used with any proximity ligation method as long as one of the interacting partners is RNA. RNAcontacts is available via github at https://github.com/smargasyuk/RNAcontacts/

bioinformatics↗

bioRxiv2022

Tissue-specific regulation of gene expression via unproductive splicing

Eukaryotic gene expression is regulated post-transcriptionally by a mechanism called unproductive splicing, in which mRNA is triggered to degradation by the nonsense-mediated decay (NMD) pathway as a result of regulated alternative splicing (AS). Only a few dozen unproductive splicing events (USEs) are currently documented, and many more remain to be identified. Here, we analyzed RNA-seq experiments from the Genotype-Tissue Expression (GTEx) Consortium to identify USEs, in which an increase in the NMD isoform splicing rate is accompanied by tissue-specific downregulation of the host gene. Further, to characterize RBPs that regulate USEs, we superimposed these results with RNA-binding protein (RBP) footprinting data and experiments on the response of the transcriptome to the perturbation of expression of a large panel of RBPs. Concordant tissue-specific changes between the expression of RBP and USE splicing rate revealed a high-confidence regulatory network including 27 tissue-specific USEs with strong evidence of RBP binding. Among them, we found previously unknown PTBP1-controlled events in the DCLK2 and IQGAP1 genes, for which we confirmed the regulatory effect using siRNA-knockdown experiments in the A549 cell line. In sum, we present a transcriptomic pipeline that allows the identification of tissue-specific USEs, potentially many more than we have reported here using stringent filters.

bioinformatics↗

bioRxiv2022

Transcriptome sequencing suggests that pre-mRNA splicing counteracts premature intronic polyadenylation

Alternative splicing (AS) and alternative polyadenylation (APA) are two crucial steps in the post-transcriptional regulation of eukaryotic gene expression. Protocols capturing and sequencing RNA 3-ends have uncovered widespread intronic polyadenylation (IPA) in normal and disease conditions, where it is currently attributed to stochastic variations in the pre-mRNA processing. Here, we took advantage of the massive amount of RNA-seq data generated by the Genotype Tissue Expression project (GTEx) to simultaneously identify and match tissue-specific expression of intronic polyadenylation sites with tissue-specific splicing. A combination of computational methods including the analysis of short reads with non-templated adenines revealed that APA events are more abundant in introns than in exons. While the rate of IPA in composite terminal exons and skipped terminal exons expectedly correlates with splicing, we observed a considerable fraction of IPA events that lack AS support and attributed them to spliced polyadenylated introns (SPI). We hypothesize that SPIs represent transient byproducts of a dynamic coupling between APA and AS, in which the spliceosome removes an intron after cleavage and polyadenylation have already occurred in it. These findings indicate that cotranscriptional pre-mRNA splicing could serve as a rescue mechanism to suppress premature transcription termination at intronic polyadenylation sites.

bioinformatics↗