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Nature Structural & Molecular Biology Aug 2023Combinatorially, intron excision within a given nascent transcript could proceed down any of thousands of paths, each of which would expose different dynamic landscapes...
Combinatorially, intron excision within a given nascent transcript could proceed down any of thousands of paths, each of which would expose different dynamic landscapes of cis-elements and contribute to alternative splicing. In this study, we found that post-transcriptional multi-intron splicing order in human cells is largely predetermined, with most genes spliced in one or a few predominant orders. Strikingly, these orders were conserved across cell types and stages of motor neuron differentiation. Introns flanking alternatively spliced exons were frequently excised last, after their neighboring introns. Perturbations to the spliceosomal U2 snRNA altered the preferred splicing order of many genes, and these alterations were associated with the retention of other introns in the same transcript. In one gene, early removal of specific introns was sufficient to induce delayed excision of three proximal introns, and this delay was caused by two distinct cis-regulatory mechanisms. Together, our results demonstrate that multi-intron splicing order in human cells is predetermined, is influenced by a component of the spliceosome and ensures splicing fidelity across long pre-mRNAs.
Topics: Humans; Introns; RNA Precursors; RNA Splicing; Alternative Splicing; Spliceosomes
PubMed: 37443198
DOI: 10.1038/s41594-023-01035-2 -
Biochimica Et Biophysica Acta. Gene... 2019RNA splicing, the process through which intervening segments of noncoding RNA (introns) are excised from pre-mRNAs to allow for the formation of a mature mRNA product,... (Review)
Review
RNA splicing, the process through which intervening segments of noncoding RNA (introns) are excised from pre-mRNAs to allow for the formation of a mature mRNA product, has long been appreciated for its capacity to add complexity to eukaryotic proteomes. However, evidence suggests that the utility of this process extends beyond protein output and provides cells with a dynamic tool for gene regulation. In this review, we aim to highlight the role that intronic RNA plays in mediating specific splicing outcomes in pre-mRNA processing, as well as explore an emerging class of stable intronic sequences that have been observed to act in gene expression control. Building from underlying flexibility in both sequence and structure, intronic RNA provides mechanisms for post-transcriptional gene regulation that are amenable to the tissue and condition specific needs of eukaryotic cells. This article is part of a Special Issue entitled: RNA structure and splicing regulation edited by Francisco Baralle, Ravindra Singh and Stefan Stamm.
Topics: Alternative Splicing; Animals; Gene Expression Regulation; Humans; Introns; RNA Precursors; RNA Processing, Post-Transcriptional; RNA, Messenger
PubMed: 31682938
DOI: 10.1016/j.bbagrm.2019.194439 -
Cell Death & Disease Feb 2023Dysregulation of pre-mRNA splicing is a common hallmark of cancer cells and it is associated with altered expression, localization, and mutations of the components of... (Review)
Review
Dysregulation of pre-mRNA splicing is a common hallmark of cancer cells and it is associated with altered expression, localization, and mutations of the components of the splicing machinery. In the last few years, it has been elucidated that spliceosome components can also influence cellular processes in a splicing-independent manner. Here, we analyze open source data to understand the effect of the knockdown of splicing factors in human cells on the expression and splicing of genes relevant to cell proliferation, migration, cell cycle regulation, DNA repair, and cell death. We supplement this information with a comprehensive literature review of non-canonical functions of splicing factors linked to cancer progression. We also specifically discuss the involvement of splicing factors in intercellular communication and known autoregulatory mechanisms in restoring their levels in cells. Finally, we discuss strategies to target components of the spliceosome machinery that are promising for anticancer therapy. Altogether, this review greatly expands understanding of the role of spliceosome proteins in cancer progression.
Topics: Humans; Spliceosomes; Neoplasms; RNA Splicing; RNA Splicing Factors; Cell Cycle Checkpoints; RNA Precursors
PubMed: 36732501
DOI: 10.1038/s41419-022-05470-9 -
Proceedings of the National Academy of... Oct 2022Telomerase is a eukaryotic ribonucleoprotein (RNP) enzyme that adds DNA repeats onto chromosome ends to maintain genomic stability and confer cellular immortality in...
Telomerase is a eukaryotic ribonucleoprotein (RNP) enzyme that adds DNA repeats onto chromosome ends to maintain genomic stability and confer cellular immortality in cancer and stem cells. The telomerase RNA (TER) component is essential for telomerase catalytic activity and provides the template for telomeric DNA synthesis. The biogenesis of TERs is extremely divergent across eukaryotic kingdoms, employing distinct types of transcription machinery and processing pathways. In ciliates and plants, TERs are transcribed by RNA polymerase III (Pol III), while animal and ascomycete fungal TERs are transcribed by RNA Pol II and share biogenesis pathways with small nucleolar RNA (snoRNA) and small nuclear RNA (snRNA), respectively. Here, we report an unprecedented messenger RNA (mRNA)-derived biogenesis pathway for the 1,291 nucleotide TER from the basidiomycete fungus . The TER (TER) contains a 5'-monophosphate, distinct from the 5' 2,2,7-trimethylguanosine (TMG) cap common to animal and ascomycete fungal TERs. The mature TER is processed from the 3'-untranslated region (3'-UTR) of a larger RNA precursor that possesses characteristics of mRNA including a 5' 7-methyl-guanosine (mG) cap, alternative splicing of introns, and a poly(A) tail. Moreover, this mRNA transcript encodes a protein called Early meiotic induction protein 1 (Emi1) that is conserved across dikaryotic fungi. A recombinant TER precursor expressed from an mRNA promoter is processed correctly to yield mature TER, confirming an mRNA-processing pathway for producing TER. Our findings expand the plethora of TER biogenesis mechanisms and demonstrate a pathway for producing a functional long noncoding RNA from a protein-coding mRNA precursor.
Topics: Animals; Guanosine; Nucleotides; RNA; RNA Polymerase II; RNA Polymerase III; RNA Precursors; RNA, Long Noncoding; RNA, Messenger; RNA, Small Nucleolar; Ribonucleoproteins; Telomerase; Untranslated Regions
PubMed: 36197996
DOI: 10.1073/pnas.2204636119 -
Nucleic Acids Research Feb 2017RNA G-quadruplexes (G4s) are formed by G-rich RNA sequences in protein-coding (mRNA) and non-coding (ncRNA) transcripts that fold into a four-stranded conformation....
RNA G-quadruplexes (G4s) are formed by G-rich RNA sequences in protein-coding (mRNA) and non-coding (ncRNA) transcripts that fold into a four-stranded conformation. Experimental studies and bioinformatic predictions support the view that these structures are involved in different cellular functions associated to both DNA processes (telomere elongation, recombination and transcription) and RNA post-transcriptional mechanisms (including pre-mRNA processing, mRNA turnover, targeting and translation). An increasing number of different diseases have been associated with the inappropriate regulation of RNA G4s exemplifying the potential importance of these structures on human health. Here, we review the different molecular mechanisms underlying the link between RNA G4s and human diseases by proposing several overlapping models of deregulation emerging from recent research, including (i) sequestration of RNA-binding proteins, (ii) aberrant expression or localization of RNA G4-binding proteins, (iii) repeat associated non-AUG (RAN) translation, (iv) mRNA translational blockade and (v) disabling of protein-RNA G4 complexes. This review also provides a comprehensive survey of the functional RNA G4 and their mechanisms of action. Finally, we highlight future directions for research aimed at improving our understanding on RNA G4-mediated regulatory mechanisms linked to diseases.
Topics: Computational Biology; G-Quadruplexes; Genetic Predisposition to Disease; Genetic Variation; Humans; Models, Biological; Polymorphism, Single Nucleotide; RNA; RNA Precursors; RNA, Messenger; RNA-Binding Proteins
PubMed: 28013268
DOI: 10.1093/nar/gkw1280 -
Expert Opinion on Drug Discovery Oct 2022RNA splicing is a pivotal step of eukaryotic gene expression during which the introns are excised from the precursor (pre-)RNA and the exons are joined together to form... (Review)
Review
INTRODUCTION
RNA splicing is a pivotal step of eukaryotic gene expression during which the introns are excised from the precursor (pre-)RNA and the exons are joined together to form mature RNA products (i.e a protein-coding mRNA or long non-coding (lnc)RNAs). The spliceosome, a complex ribonucleoprotein machine, performs pre-RNA splicing with extreme precision. Deregulated splicing is linked to cancer, genetic, and neurodegenerative diseases. Hence, the discovery of small-molecules targeting core spliceosome components represents an appealing therapeutic opportunity.
AREA COVERED
Several atomic-level structures of the spliceosome and distinct splicing-modulators bound to its protein/RNA components have been solved. Here, we review recent advances in the discovery of small-molecule splicing-modulators, discuss opportunities and challenges for their therapeutic applicability, and showcase how structural data and/or all-atom simulations can illuminate key facets of their mechanism, thus contributing to future drug-discovery campaigns.
EXPERT OPINION
This review highlights the potential of modulating pre-RNA splicing with small-molecules, and anticipates how the synergy of computer and wet-lab experiments will enrich our understanding of splicing regulation/deregulation mechanisms. This information will aid future structure-based drug-discovery efforts aimed to expand the currently limited portfolio of selective splicing-modulators.
Topics: Humans; Introns; RNA Precursors; RNA Splicing; Spliceosomes
PubMed: 35983696
DOI: 10.1080/17460441.2022.2114452 -
Journal of Biomedical Science Jul 2015Splicing of precursor mRNA takes place via two consecutive steps of transesterification catalyzed by a large ribonucleoprotein complex called the spliceosome. The... (Review)
Review
Splicing of precursor mRNA takes place via two consecutive steps of transesterification catalyzed by a large ribonucleoprotein complex called the spliceosome. The spliceosome is assembled through ordered binding to the pre-mRNA of five small nuclear RNAs and numerous protein factors, and is disassembled after completion of the reaction to recycle all components. Throughout the splicing cycle, the spliceosome changes its structure, rearranging RNA-RNA, RNA-protein and protein-protein interactions, for positioning and repositioning of splice sites. DExD/H-box RNA helicases play important roles in mediating structural changes of the spliceosome by unwinding of RNA duplexes or disrupting RNA-protein interactions. DExD/H-box proteins are also implicated in the fidelity control of the splicing process at various steps. This review summarizes the functional roles of DExD/H-box proteins in pre-mRNA splicing according to studies conducted mostly in yeast and will discuss the concept of the complicated splicing reaction based on recent findings.
Topics: Humans; RNA Helicases; RNA Precursors; RNA Splicing; Saccharomyces cerevisiae; Spliceosomes
PubMed: 26173448
DOI: 10.1186/s12929-015-0161-z -
Molekuliarnaia Biologiia 2015RNA-binding proteins (RBPs) play an important role in regulating gene expression at the posttranscriptional level, including the steps of pre-mRNA splicing,... (Review)
Review
RNA-binding proteins (RBPs) play an important role in regulating gene expression at the posttranscriptional level, including the steps of pre-mRNA splicing, polyadenylation, mRNA stabilization, mRNA export from the nucleus to the cytoplasm, mRNA localization, and translation. RBPs regulate these processes primarily by binding to specific sequence elements in newly synthesized or mature transcripts. While many RPBs are known to recognize certain nucleotide sequences in RNA, information is insufficient for others. In particular, RBPs often compete for RNA binding or interact with RNA cooperatively. Hence, it is of importance to study the RNA-protein interactions in vivo. Numerous methods have been developed to identify the target nucleotide sequences of RBPs. The methods include the electrophoretic mobility shift assay (EMSA), systematic evolution of ligands by exponential enrichment (SELEX), RNA pull-down assay, RNA footprinting, RNA immunoprecipitation (RIP), UV-induced crosslinking immunoprecipitation (CLIP) and its variants, and measurement of the level for newly synthesized transcripts. Each of the methods has its limitation, and several methods supplementing each other should be employed in order to detect the RNA sequence to which a protein binds.
Topics: Animals; Binding Sites; Electrophoretic Mobility Shift Assay; Gene Expression Regulation; Humans; Immunoprecipitation; Protein Binding; RNA Precursors; RNA, Messenger; RNA-Binding Proteins; SELEX Aptamer Technique; Sequence Analysis, RNA
PubMed: 26107901
DOI: 10.7868/S0026898415020111 -
The New Phytologist Mar 2023Pathogenic fungi are subject to DNA damage stress derived from host immune responses during infection. Small ubiquitin-like modifier (SUMO) modification and precursor...
Pathogenic fungi are subject to DNA damage stress derived from host immune responses during infection. Small ubiquitin-like modifier (SUMO) modification and precursor (pre)-mRNA splicing are both involved in DNA damage response (DDR). However, the mechanisms of how SUMOylation and splicing coordinated in DDR remain largely unknown. Combining with biochemical analysis, RNA-Seq method, and biological analysis, we report that SUMO pathway participates in DDR and virulence in Fusarium graminearum, a causal agent of Fusarium head blight of cereal crops world-wide. Interestingly, a key transcription factor FgSR is SUMOylated upon DNA damage stress. SUMOylation regulates FgSR nuclear-cytoplasmic partitioning and its phosphorylation by FgMec1, and promotes its interaction with chromatin remodeling complex SWI/SNF for activating the expression of DDR-related genes. Moreover, the SWI/SNF complex was found to further recruit splicing-related NineTeen Complex, subsequently modulates pre-mRNA splicing during DDR. Our findings reveal a novel function of SUMOylation in DDR by regulating a transcription factor to orchestrate gene expression and pre-mRNA splicing to overcome DNA damage during the infection of F. graminearum, which advances the understanding of the delicate regulation of DDR by SUMOylation in pathogenic fungi, and extends the knowledge of cooperation of SUMOylation and pre-mRNA splicing in DDR in eukaryotes.
Topics: Sumoylation; RNA Precursors; Small Ubiquitin-Related Modifier Proteins; Transcription Factors; DNA Damage
PubMed: 36539920
DOI: 10.1111/nph.18692 -
Genes Apr 2021The human immune response is a complex process that responds to numerous exogenous antigens in preventing infection by microorganisms, as well as to endogenous... (Review)
Review
The human immune response is a complex process that responds to numerous exogenous antigens in preventing infection by microorganisms, as well as to endogenous components in the surveillance of tumors and autoimmune diseases, and a great number of molecules are necessary to carry the functional complexity of immune activity. Alternative splicing of pre-mRNA plays an important role in immune cell development and regulation of immune activity through yielding diverse transcriptional isoforms to supplement the function of limited genes associated with the immune reaction. In addition, multiple factors have been identified as being involved in the control of alternative splicing at the , , or co-transcriptional level, and the aberrant splicing of RNA leads to the abnormal modulation of immune activity in infections, immune diseases, and tumors. In this review, we summarize the recent discoveries on the generation of immune-associated alternative splice variants, clinical disorders, and possible regulatory mechanisms. We also discuss the immune responses to the neoantigens produced by alternative splicing, and finally, we issue some alternative splicing and immunity correlated questions based on our knowledge.
Topics: Alternative Splicing; Gene Expression Regulation; Humans; Immunity; RNA Precursors; RNA, Untranslated
PubMed: 33921058
DOI: 10.3390/genes12040574