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Molecular Cell Mar 2022The nucleus is highly compartmentalized through the formation of distinct classes of membraneless domains. However, the composition and function of many of these...
The nucleus is highly compartmentalized through the formation of distinct classes of membraneless domains. However, the composition and function of many of these structures are not well understood. Using APEX2-mediated proximity labeling and RNA sequencing, we surveyed human transcripts associated with nuclear speckles, several additional domains, and the lamina. Remarkably, speckles and lamina are associated with distinct classes of retained introns enriched in genes that function in RNA processing, translation, and the cell cycle, among other processes. In contrast to the lamina-proximal introns, retained introns associated with speckles are relatively short, GC-rich, and enriched for functional sites of RNA-binding proteins that are concentrated in these domains. They are also highly differentially regulated across diverse cellular contexts, including the cell cycle. Thus, our study provides a resource of nuclear domain-associated transcripts and further reveals speckles and lamina as hubs of distinct populations of retained introns linked to gene regulation and cell cycle progression.
Topics: Cell Nucleus; Gene Expression Regulation; Humans; Introns; RNA Splicing; RNA-Binding Proteins
PubMed: 35182477
DOI: 10.1016/j.molcel.2021.12.010 -
Human Mutation Apr 2022DMD pathogenic variants for Duchenne and Becker muscular dystrophy are detectable with high sensitivity by standard clinical exome analyses of genomic DNA. However, up...
DMD pathogenic variants for Duchenne and Becker muscular dystrophy are detectable with high sensitivity by standard clinical exome analyses of genomic DNA. However, up to 7% of DMD mutations are deep intronic and analysis of muscle-derived RNA is an important diagnostic step for patients who have negative genomic testing but abnormal dystrophin expression in muscle. In this study, muscle biopsies were evaluated from 19 patients with clinical features of a dystrophinopathy, but negative clinical DMD mutation analysis. Reverse transcription-polymerase chain reaction or high-throughput RNA sequencing methods identified 19 mutations with one of three pathogenic pseudoexon types: deep intronic point mutations, deletions or insertions, and translocations. In association with point mutations creating intronic splice acceptor sites, we observed the first examples of DMD pseudo 3'-terminal exon mutations causing high efficiency transcription termination within introns. This connection between splicing and premature transcription termination is reminiscent of U1 snRNP-mediating telescripting in sustaining RNA polymerase II elongation across large genes, such as DMD. We propose a novel classification of three distinct types of mutations identifiable by muscle RNA analysis, each of which differ in potential treatment approaches. Recognition and appropriate characterization may lead to therapies directed toward full-length dystrophin expression for some patients.
Topics: Dystrophin; Humans; Introns; Muscular Dystrophy, Duchenne; Mutation; RNA Splice Sites
PubMed: 35165973
DOI: 10.1002/humu.24343 -
Memorias Do Instituto Oswaldo Cruz 2018In Cryptococcus neoformans, nearly all genes are interrupted by small introns. In recent years, genome annotation and genetic analysis have illuminated the major roles... (Review)
Review
In Cryptococcus neoformans, nearly all genes are interrupted by small introns. In recent years, genome annotation and genetic analysis have illuminated the major roles these introns play in the biology of this pathogenic yeast. Introns are necessary for gene expression and alternative splicing can regulate gene expression in response to environmental cues. In addition, recent studies have revealed that C. neoformans introns help to prevent transposon dissemination and protect genome integrity. These characteristics of cryptococcal introns are probably not unique to Cryptococcus, and this yeast likely can be considered as a model for intron-related studies in fungi.
Topics: Cryptococcus; DNA, Fungal; Genome, Fungal; Introns; Sequence Analysis, DNA
PubMed: 29513783
DOI: 10.1590/0074-02760170519 -
Cancer Science Sep 2022Pre-mRNA splicing is now widely recognized as a cotranscriptional and post-transcriptional mechanism essential for regulating gene expression and modifying gene product... (Review)
Review
Pre-mRNA splicing is now widely recognized as a cotranscriptional and post-transcriptional mechanism essential for regulating gene expression and modifying gene product function. Mutations in genes encoding core spliceosomal proteins and accessory regulatory splicing factors are now considered among the most recurrent genetic abnormalities in patients with cancer, particularly hematologic malignancies. These include mutations in the major (U2-type) and minor (U12-type) spliceosomes, which remove >99% and ~0.35% of introns, respectively. Growing evidence indicates that aberrant splicing of evolutionarily conserved U12-type minor introns plays a crucial role in cancer as the minor spliceosome component, ZRSR2, is subject to recurrent, leukemia-associated mutations, and intronic mutations have been shown to disrupt the splicing of minor introns. Here, we review the importance of minor intron regulation, the molecular effects of the minor (U12-type) spliceosomal mutations and cis-regulatory regions, and the development of minor intron studies for better understanding of cancer biology.
Topics: Humans; Introns; Mutation; Neoplasms; RNA Splicing; Spliceosomes
PubMed: 35766428
DOI: 10.1111/cas.15476 -
Journal of Neuromuscular Diseases 2020The DMD gene is the largest in the human genome, with a total intron content exceeding 2.2Mb. In the decades since DMD was discovered there have been numerous reported... (Review)
Review
The DMD gene is the largest in the human genome, with a total intron content exceeding 2.2Mb. In the decades since DMD was discovered there have been numerous reported cases of pseudoexons (PEs) arising in the mature DMD transcripts of some individuals, either as the result of mutations or as low-frequency errors of the spliceosome. In this review, I collate from the literature 58 examples of DMD PEs and examine the diversity and commonalities of their features. In particular, I note the high frequency of PEs that arise from deep intronic SNVs and discuss a possible link between PEs induced by distal mutations and the regulation of recursive splicing.
Topics: Dystrophin; Exons; Humans; Introns; Muscular Dystrophy, Duchenne; RNA Splicing
PubMed: 32176650
DOI: 10.3233/JND-190431 -
BMC Genomics Sep 2022Spliceosomal introns are parts of primary transcripts that are removed by RNA splicing. Although introns apparently do not contribute to the function of the mature...
BACKGROUND
Spliceosomal introns are parts of primary transcripts that are removed by RNA splicing. Although introns apparently do not contribute to the function of the mature transcript, in vertebrates they comprise the majority of the transcribed region increasing the metabolic cost of transcription. The persistence of long introns across evolutionary time suggests functional roles that can offset this metabolic cost. The teleosts comprise one of the largest vertebrate clades. They have unusually compact and variable genome sizes and provide a suitable system for analysing intron evolution.
RESULTS
We have analysed intron lengths in 172 vertebrate genomes and show that teleost intron lengths are relatively short, highly variable and bimodally distributed. Introns that were long in teleosts were also found to be long in mammals and were more likely to be found in regulatory genes and to contain conserved sequences. Our results argue that intron length has decreased in parallel in a non-random manner throughout teleost evolution and represent a deviation from the ancestral state.
CONCLUSION
Our observations indicate an accelerated rate of intron size evolution in the teleosts and that teleost introns can be divided into two classes by their length. Teleost intron sizes have evolved primarily as a side-effect of genome size evolution and small genomes are dominated by short introns (<256 base pairs). However, a non-random subset of introns has resisted this process across the teleosts and these are more likely have functional roles in all vertebrate clades.
Topics: Animals; Evolution, Molecular; Exons; Genome; Introns; Mammals; Vertebrates
PubMed: 36050638
DOI: 10.1186/s12864-022-08760-w -
Human Genetics Sep 2017Intron retention has long been an exemplar of regulated splicing with case studies of individual events serving as models that provided key mechanistic insights into the... (Review)
Review
Intron retention has long been an exemplar of regulated splicing with case studies of individual events serving as models that provided key mechanistic insights into the process of splicing control. In organisms such as plants and budding yeast, intron retention is well understood as a major mechanism of gene expression regulation. In contrast, in mammalian systems, the extent and functional significance of intron retention have, until recently, remained greatly underappreciated. Technical challenges to the global detection and quantitation of transcripts with retained introns have often led to intron retention being overlooked or dismissed as "noise". Now, however, with the wealth of information available from high-throughput deep sequencing, combined with focused computational and statistical analyses, we are able to distinguish clear intron retention patterns in various physiological and pathological contexts. Several recent studies have demonstrated intron retention as a central component of gene expression programs during normal development as well as in response to stress and disease. Furthermore, these studies revealed various ways in which intron retention regulates protein isoform production, RNA stability and translation efficiency, and rapid induction of expression via post-transcriptional splicing of retained introns. In this review, we highlight critical findings from these transcriptomic studies and discuss commonalties in the patterns prevalent in intron retention networks at the functional and regulatory levels.
Topics: Animals; Gene Expression Regulation; Humans; Introns; Models, Genetic; Protein Biosynthesis; Protein Isoforms; RNA Stability
PubMed: 28391524
DOI: 10.1007/s00439-017-1791-x -
FEMS Microbiology Letters Oct 2021Spliceosomal introns are noncoding sequences that are spliced from pre-mRNA. They are ubiquitous in eukaryotic genomes, although the average number of introns per gene... (Review)
Review
Spliceosomal introns are noncoding sequences that are spliced from pre-mRNA. They are ubiquitous in eukaryotic genomes, although the average number of introns per gene varies considerably between different eukaryotic species. Fungi are diverse in terms of intron numbers ranging from 4% to 99% genes with introns. Alternative splicing is one of the most common modes of posttranscriptional regulation in eukaryotes, giving rise to multiple transcripts from a single pre-mRNA and is widespread in metazoans and drives extensive proteome diversity. Earlier, alternative splicing was considered to be rare in fungi, but recently, increasing numbers of studies have revealed that alternative splicing is also widespread in fungi and has been implicated in the regulation of fungal growth and development, protein localization and the improvement of survivability, likely underlying their unique capacity to adapt to changing environmental conditions. However, the role of alternative splicing in pathogenicity and development of drug resistance is only recently gaining attention. In this review, we describe the intronic landscape in fungi. We also present in detail the newly discovered functions of alternative splicing in various cellular processes and outline areas particularly in pathogenesis and clinical drug resistance for future studies that could lead to the development of much needed new therapeutics.
Topics: Alternative Splicing; Fungi; Introns; RNA Precursors; Spliceosomes
PubMed: 34718529
DOI: 10.1093/femsle/fnab135 -
BMC Genomics Nov 2021Alternative splicing is the process of selecting different combinations of splice sites to produce variably spliced mRNAs. However, the relationships between alternative...
BACKGROUND
Alternative splicing is the process of selecting different combinations of splice sites to produce variably spliced mRNAs. However, the relationships between alternative splicing prevalence and level (ASP/L) and variations of intron size and organism complexity (OC) remain vague. Here, we developed a robust protocol to analyze the relationships between ASP/L and variations of intron size and OC. Approximately 8 Tb raw RNA-Seq data from 37 eumetazoan species were divided into three sets of species based on variations in intron size and OC.
RESULTS
We found a strong positive correlation between ASP/L and OC, but no correlation between ASP/L and intron size across species. Surprisingly, ASP/L displayed a positive correlation with mean intron size of genes within individual genomes. Moreover, our results revealed that four ASP/L-related pathways contributed to the differences in ASP/L that were associated with OC. In particular, the spliceosome pathway displayed distinct genomic features, such as the highest gene expression level, conservation level, and fraction of disordered regions. Interestingly, lower or no obvious correlations were observed among these genomic features.
CONCLUSIONS
The positive correlation between ASP/L and OC ubiquitously exists in eukaryotes, and this correlation is not affected by the mean intron size of these species. ASP/L-related splicing factors may play an important role in the evolution of OC.
Topics: Alternative Splicing; Eukaryota; Genome; Introns; RNA Splicing; RNA, Messenger
PubMed: 34819032
DOI: 10.1186/s12864-021-08172-2 -
Proceedings of the National Academy of... Nov 2022Pre-messenger RNA splicing is initiated with the recognition of a single-nucleotide intronic branchpoint (BP) within a BP motif by spliceosome elements. Forty-eight rare...
Pre-messenger RNA splicing is initiated with the recognition of a single-nucleotide intronic branchpoint (BP) within a BP motif by spliceosome elements. Forty-eight rare variants in 43 human genes have been reported to alter splicing and cause disease by disrupting BP. However, until now, no computational approach was available to efficiently detect such variants in massively parallel sequencing data. We established a comprehensive human genome-wide BP database by integrating existing BP data and generating new BP data from RNA sequencing of lariat debranching enzyme DBR1-mutated patients and from machine-learning predictions. We characterized multiple features of BP in major and minor introns and found that BP and BP-2 (two nucleotides upstream of BP) positions exhibit a lower rate of variation in human populations and higher evolutionary conservation than the intronic background, while being comparable to the exonic background. We developed BPHunter as a genome-wide computational approach to systematically and efficiently detect intronic variants that may disrupt BP recognition. BPHunter retrospectively identified 40 of the 48 known pathogenic BP variants, in which we summarized a strategy for prioritizing BP variant candidates. The remaining eight variants all create AG-dinucleotides between the BP and acceptor site, which is the likely reason for missplicing. We demonstrated the practical utility of BPHunter prospectively by using it to identify a novel germline heterozygous BP variant of in a patient with critical COVID-19 pneumonia and a novel somatic intronic 59-nucleotide deletion of in a lymphoma patient, both of which were validated experimentally. BPHunter is publicly available from https://hgidsoft.rockefeller.edu/BPHunter and https://github.com/casanova-lab/BPHunter.
Topics: Humans; Introns; Retrospective Studies; COVID-19; RNA Splicing; Nucleotides
PubMed: 36306325
DOI: 10.1073/pnas.2211194119