scholarly journals Regulation, diversity and function of MECP2 exon and 3′UTR isoforms

2020 ◽  
Vol 29 (R1) ◽  
pp. R89-R99
Author(s):  
Deivid Carvalho Rodrigues ◽  
Marat Mufteev ◽  
James Ellis
Keyword(s):  
Dna Binding ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Alternative Polyadenylation ◽  
Cell Types ◽  
Protein Isoforms ◽  
Translation Efficiency ◽  
Mrna Isoforms ◽  
Exon 2 ◽  
Messenger Ribonucleic Acid

Abstract The methyl-CpG-binding protein 2 (MECP2) is a critical global regulator of gene expression. Mutations in MECP2 cause neurodevelopmental disorders including Rett syndrome (RTT). MECP2 exon 2 is spliced into two alternative messenger ribonucleic acid (mRNA) isoforms encoding MECP2-E1 or MECP2-E2 protein isoforms that differ in their N-termini. MECP2-E2, isolated first, was used to define the general roles of MECP2 in methyl-deoxyribonucleic acid (DNA) binding, targeting of transcriptional regulatory complexes, and its disease-causing impact in RTT. It was later found that MECP2-E1 is the most abundant isoform in the brain and its exon 1 is also mutated in RTT. MECP2 transcripts undergo alternative polyadenylation generating mRNAs with four possible 3′untranslated region (UTR) lengths ranging from 130 to 8600 nt. Together, the exon and 3′UTR isoforms display remarkable abundance disparity across cell types and tissues during development. These findings indicate discrete means of regulation and suggest that protein isoforms perform non-overlapping roles. Multiple regulatory programs have been explored to explain these disparities. DNA methylation patterns of the MECP2 promoter and first intron impact MECP2-E1 and E2 isoform levels. Networks of microRNAs and RNA-binding proteins also post-transcriptionally regulate the stability and translation efficiency of MECP2 3′UTR isoforms. Finally, distinctions in biophysical properties in the N-termini between MECP2-E1 and E2 lead to variable protein stabilities and DNA binding dynamics. This review describes the steps taken from the discovery of MECP2, the description of its key functions, and its association with RTT, to the emergence of evidence revealing how MECP2 isoforms are differentially regulated at the transcriptional, post-transcriptional and post-translational levels.

scholarly journals Crosstalk Between mRNA 3'-End Processing and Epigenetics

2021 ◽  
Vol 12 ◽  
Author(s):  
Lindsey V. Soles ◽  
Yongsheng Shi
Keyword(s):  
Histone Modifications ◽  
Dynamic Process ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Alternative Polyadenylation ◽  
Regulatory Mechanisms ◽  
Epigenetic Mechanisms ◽  
Cis Elements ◽  
Mrna Isoforms ◽  
Eukaryotic Genes

The majority of eukaryotic genes produce multiple mRNA isoforms by using alternative poly(A) sites in a process called alternative polyadenylation (APA). APA is a dynamic process that is highly regulated in development and in response to extrinsic or intrinsic stimuli. Mis-regulation of APA has been linked to a wide variety of diseases, including cancer, neurological and immunological disorders. Since the first example of APA was described 40 years ago, the regulatory mechanisms of APA have been actively investigated. Conventionally, research in this area has focused primarily on the roles of regulatory cis-elements and trans-acting RNA-binding proteins. Recent studies, however, have revealed important functions for epigenetic mechanisms, including DNA and histone modifications and higher-order chromatin structures, in APA regulation. Here we will discuss these recent findings and their implications for our understanding of the crosstalk between epigenetics and mRNA 3'-end processing.

Stage-Specific Switches in Alternative Pre-mRNA Splicing during Late Erythropoiesis Are Conserved from Mouse to Human

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 531-531
Author(s):  
Sherry Gee ◽  
Jonathan Villalobos ◽  
Miki Yamamoto ◽  
Tyson A. Clark ◽  
Jeong-Ah Kang ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Mrna Splicing ◽  
Protein Isoforms ◽  
Comparative Genomic ◽  
Erythroid Progenitors ◽  
Mrna Isoforms ◽  
Stop Codons ◽  
Primary Mouse

Abstract Spatial and temporal regulation of alternative pre-mRNA splicing determines which exons are incorporated into mature mRNA, modulating mRNA coding capacity to ensure synthesis of appropriate protein isoforms throughout normal differentiation and development. During erythropoiesis, a stage-specific switch in pre-mRNA splicing activates incorporation of protein 4.1R exon 16, thereby increasing 4.1R affinity for spectrin and actin and mechanically strengthening red blood cell membranes. We are exploring the hypothesis that stage-specific changes in pre-mRNA splicing regulate expression of other critical genes during terminal erythropoiesis. Last year we described exon microarray and RT-PCR studies that revealed several novel pre-mRNA splicing switches in terminally differentiating human erythroid progenitors. These alternative splicing events involved well-annotated exons with consensus exon-intron boundaries, supporting a model in which these events represent a regulated alternative splicing program rather than a breakdown of splicing integrity in late erythropoiesis. Here we report additional evidence for this model by showing that several erythroid stage-specific switches in alternative pre-mRNA splicing are conserved between human and mouse. Primary mouse splenic erythroblasts from FVA-infected mice were cultured in vitro under differentiation conditions and used as the source of RNA for analysis of murine erythroid splicing events. From a total of seven internal cassette exons whose splicing was activated in late human erythroblasts, five exhibited an analogous splicing switch in murine erythroblasts. Comparative genomic analysis showed that these alternative exons are embedded in regions of unusually high sequence conservation among vertebrate species, suggesting that important regulatory signals are contained within the adjacent introns. Indeed, the flanking introns for several of these exons contain binding motifs for Fox2, an RNA binding protein and known splicing regulator for many tissue-specific splicing events. Further analysis of the conserved erythroid splicing events revealed the following: three splicing switches occur in transcripts encoding RNA binding proteins (MBNL2, HNRPLL, and SNRP70), suggesting significant changes in the RNA processing machinery of late erythroblasts; and three of these alternative exons encode premature stop codons that could induce nonsense mediated decay (NMD) and contribute to down-regulation of these genes during terminal erythropoiesis. Consistent with the latter hypothesis, inhibition of NMD in murine erythroblast cultures led to increased accumulation of mRNA isoforms containing the premature stop codons. Together these results suggest the existence of a highly regulated alternative splicing program that is critical for late erythroid differentiation.

scholarly journals Emerging Roles for 3′ UTRs in Neurons

2020 ◽  
Vol 21 (10) ◽  
pp. 3413
Author(s):  
Bongmin Bae ◽  
Pedro Miura
Keyword(s):  
Translational Control ◽  
Transcriptional Control ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Alternative Polyadenylation ◽  
Mrna Localization ◽  
Structural Features ◽  
Mrna Isoforms ◽  
Rna Sequences ◽  
Neuronal Functions

The 3′ untranslated regions (3′ UTRs) of mRNAs serve as hubs for post-transcriptional control as the targets of microRNAs (miRNAs) and RNA-binding proteins (RBPs). Sequences in 3′ UTRs confer alterations in mRNA stability, direct mRNA localization to subcellular regions, and impart translational control. Thousands of mRNAs are localized to subcellular compartments in neurons—including axons, dendrites, and synapses—where they are thought to undergo local translation. Despite an established role for 3′ UTR sequences in imparting mRNA localization in neurons, the specific RNA sequences and structural features at play remain poorly understood. The nervous system selectively expresses longer 3′ UTR isoforms via alternative polyadenylation (APA). The regulation of APA in neurons and the neuronal functions of longer 3′ UTR mRNA isoforms are starting to be uncovered. Surprising roles for 3′ UTRs are emerging beyond the regulation of protein synthesis and include roles as RBP delivery scaffolds and regulators of alternative splicing. Evidence is also emerging that 3′ UTRs can be cleaved, leading to stable, isolated 3′ UTR fragments which are of unknown function. Mutations in 3′ UTRs are implicated in several neurological disorders—more studies are needed to uncover how these mutations impact gene regulation and what is their relationship to disease severity.

scholarly journals A CRISPR RNA-binding protein screen reveals regulators of RUNX1 isoform generation

2021 ◽  
Vol 5 (5) ◽  
pp. 1310-1323
Author(s):  
Amanda G. Davis ◽  
Jaclyn M. Einstein ◽  
Dinghai Zheng ◽  
Nathan D. Jayne ◽  
Xiang-Dong Fu ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Rna Processing ◽  
Posttranscriptional Regulation ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Hematologic Malignancy ◽  
Alternative Polyadenylation ◽  
Protein Isoforms ◽  
Self Renewal ◽  
Hematopoietic Stem

Abstract The proper balance of hematopoietic stem cell (HSC) self-renewal and differentiation is critical for normal hematopoiesis and is disrupted in hematologic malignancy. Among regulators of HSC fate, transcription factors have a well-defined central role, and mutations promote malignant transformation. More recently, studies have illuminated the importance of posttranscriptional regulation by RNA-binding proteins (RBPs) in hematopoiesis and leukemia development. However, the RBPs involved and the breadth of regulation are only beginning to be elucidated. Furthermore, the intersection between posttranscriptional regulation and hematopoietic transcription factor function is poorly understood. Here, we studied the posttranscriptional regulation of RUNX1, a key hematopoietic transcription factor. Alternative polyadenylation (APA) of RUNX1 produces functionally antagonistic protein isoforms (RUNX1a vs RUNX1b/c) that mediate HSC self-renewal vs differentiation, an RNA-processing event that is dysregulated in malignancy. Consequently, RBPs that regulate this event directly contribute to healthy and aberrant hematopoiesis. We modeled RUNX1 APA using a split GFP minigene reporter and confirmed the sensitivity of our model to detect changes in RNA processing. We used this reporter in a clustered regularly interspaced short palindromic repeats (CRISPR) screen consisting of single guide RNAs exclusively targeting RBPs and uncovered HNRNPA1 and KHDRBS1 as antagonistic regulators of RUNX1a isoform generation. Overall, our study provides mechanistic insight into the posttranscriptional regulation of a key hematopoietic transcription factor and identifies RBPs that may have widespread and important functions in hematopoiesis.

scholarly journals PTBP1 mRNA isoforms and regulation of their translation

2018 ◽  
Author(s):  
Luisa M Arake de Tacca ◽  
Mia C Pulos ◽  
Stephen N Floor ◽  
Jamie Cate
Keyword(s):  
Cell Cycle ◽  
Alternative Splicing ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Translation Initiation Factor ◽  
Protein Isoforms ◽  
Initiation Factor ◽  
Dependent Manner ◽  
Mrna Isoforms

Polypyrimidine tract-binding proteins (PTBPs) are RNA binding proteins that regulate a number of post-transcriptional events. Human PTBP1 transits between the nucleus and cytoplasm and is thought to regulate RNA processes in both. However, information about PTBP1 mRNA isoforms and regulation of PTPB1 expression remain incomplete. Here we mapped the major PTBP1 mRNA isoforms in HEK293T cells, and identified alternative 5' and 3' untranslated regions (5' UTRs, 3' UTRs) as well as alternative splicing patterns in the protein coding region. We also assessed how the observed PTBP1 mRNA isoforms contribute to PTBP1 expression in different phases of the cell cycle. Previously, PTBP1 mRNAs were shown to crosslink to eukaryotic translation initiation factor 3 (eIF3). We find that eIF3 binds differently to each PTBP1 mRNA isoform in a cell cycle-dependent manner. We also observe a strong correlation between eIF3 binding to PTBP1 mRNAs and repression of PTBP1 levels during the S phase of the cell cycle. Our results provide evidence of translational regulation of PTBP1 protein levels during the cell cycle, which may affect downstream regulation of alternative splicing and translation mediated by PTBP1 protein isoforms.

scholarly journals LABRAT reveals association of alternative polyadenylation with transcript localization, RNA binding protein expression, transcription speed, and cancer survival

2020 ◽  
Author(s):  
Raeann Goering ◽  
Krysta L. Engel ◽  
Austin E. Gillen ◽  
Nova Fong ◽  
David L. Bentley ◽  
...  
Keyword(s):  
Cancer Survival ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Alternative Polyadenylation ◽  
Cell Types ◽  
Strongly Correlated ◽  
Multiple Cell ◽  
Patient Prognosis ◽  
Transcriptome Quantification

ABSTRACTThe sequence content of the 3′ UTRs of many mRNA transcripts is regulated through alternative polyadenylation (APA). The study of this process using RNAseq data, though, has been historically challenging. To combat this problem, we developed LABRAT, an APA quantification method. LABRAT takes advantage of newly developed transcriptome quantification techniques to accurately determine relative APA site usage and how it varies across conditions. Using LABRAT, we found consistent relationships between gene-distal APA and subcellular RNA localization in multiple cell types. We also observed connections between transcription speed and APA site choice as well as tumor-specific transcriptome-wide shifts in APA in hundreds of patient-derived tumor samples that were associated with patient prognosis. We investigated the effects of APA on transcript expression and found a weak overall relationship, although many individual genes showed strong correlations between APA and expression. We interrogated the roles of 191 RNA-binding proteins in the regulation of APA, finding that dozens promote broad, directional shifts in relative APA isoform abundance both in vitro and in patient-derived samples. Finally, we find that APA site shifts in the two classes of APA, tandem UTRs and alternative last exons, are strongly correlated across many contexts, suggesting that they are coregulated.

scholarly journals Alternative polyadenylation and RNA-binding proteins

2016 ◽  
Vol 57 (2) ◽  
pp. F29-F34 ◽  
Author(s):  
Ayse Elif Erson-Bensan
Keyword(s):  
Rna Binding ◽  
Biomarker Discovery ◽  
Rna Binding Proteins ◽  
Alternative Polyadenylation ◽  
Mrna Isoforms ◽  
New Therapeutic Approaches ◽  
Complex Interactions ◽  
Action Mechanisms ◽  
The Stability

Our understanding of the extent of microRNA-based gene regulation has expanded in an impressive pace over the past decade. Now, we are beginning to better appreciate the role of 3′-UTR (untranslated region) cis-elements which harbor not only microRNA but also RNA-binding protein (RBP) binding sites that have significant effect on the stability and translational rate of mRNAs. To add further complexity, alternative polyadenylation (APA) emerges as a widespread mechanism to regulate gene expression by producing shorter or longer mRNA isoforms that differ in the length of their 3′-UTRs or even coding sequences. Resulting shorter mRNA isoforms generally lack cis-elements where trans-acting factors bind, and hence are differentially regulated compared with the longer isoforms. This review focuses on the RBPs involved in APA regulation and their action mechanisms on APA-generated isoforms. A better understanding of the complex interactions between APA and RBPs is promising for mechanistic and clinical implications including biomarker discovery and new therapeutic approaches.

scholarly journals LABRAT reveals association of alternative polyadenylation with transcript localization, RNA binding protein expression, transcription speed, and cancer survival

BMC Genomics ◽  
2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Raeann Goering ◽  
Krysta L. Engel ◽  
Austin E. Gillen ◽  
Nova Fong ◽  
David L. Bentley ◽  
...  
Keyword(s):  
Cancer Survival ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Alternative Polyadenylation ◽  
Cell Types ◽  
Rnaseq Data ◽  
Multiple Cell ◽  
Patient Prognosis ◽  
Molecular Phenotypes

Abstract Background The sequence content of the 3′ UTRs of many mRNA transcripts is regulated through alternative polyadenylation (APA). The study of this process using RNAseq data, though, has been historically challenging. Results To combat this problem, we developed LABRAT, an APA isoform quantification method. LABRAT takes advantage of newly developed transcriptome quantification techniques to accurately determine relative APA site usage and how it varies across conditions. Using LABRAT, we found consistent relationships between gene-distal APA and subcellular RNA localization in multiple cell types. We also observed connections between transcription speed and APA site choice as well as tumor-specific transcriptome-wide shifts in APA isoform abundance in hundreds of patient-derived tumor samples that were associated with patient prognosis. We investigated the effects of APA on transcript expression and found a weak overall relationship, although many individual genes showed strong correlations between relative APA isoform abundance and overall gene expression. We interrogated the roles of 191 RNA-binding proteins in the regulation of APA isoforms, finding that dozens promote broad, directional shifts in relative APA isoform abundance both in vitro and in patient-derived samples. Finally, we find that APA site shifts in the two classes of APA, tandem UTRs and alternative last exons, are strongly correlated across many contexts, suggesting that they are coregulated. Conclusions We conclude that LABRAT has the ability to accurately quantify APA isoform ratios from RNAseq data across a variety of sample types. Further, LABRAT is able to derive biologically meaningful insights that connect APA isoform regulation to cellular and molecular phenotypes.

scholarly journals Alternative polyadenylation: methods, mechanism, function, and role in cancer

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
Yi Zhang ◽  
Lian Liu ◽  
Qiongzi Qiu ◽  
Qing Zhou ◽  
Jinwang Ding ◽  
...  
Keyword(s):  
Gene Regulation ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Single Gene ◽  
Alternative Polyadenylation ◽  
Length Change ◽  
Suppressor Gene ◽  
Gene Expressions ◽  
Related Factors ◽  
Mrna Maturation

AbstractOccurring in over 60% of human genes, alternative polyadenylation (APA) results in numerous transcripts with differing 3’ends, thus greatly expanding the diversity of mRNAs and of proteins derived from a single gene. As a key molecular mechanism, APA is involved in various gene regulation steps including mRNA maturation, mRNA stability, cellular RNA decay, and protein diversification. APA is frequently dysregulated in cancers leading to changes in oncogenes and tumor suppressor gene expressions. Recent studies have revealed various APA regulatory mechanisms that promote the development and progression of a number of human diseases, including cancer. Here, we provide an overview of four types of APA and their impacts on gene regulation. We focus particularly on the interaction of APA with microRNAs, RNA binding proteins and other related factors, the core pre-mRNA 3’end processing complex, and 3’UTR length change. We also describe next-generation sequencing methods and computational tools for use in poly(A) signal detection and APA repositories and databases. Finally, we summarize the current understanding of APA in cancer and provide our vision for future APA related research.

scholarly journals The landscape of alternative polyadenylation in single cells of the developing mouse embryo

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Vikram Agarwal ◽  
Sereno Lopez-Darwin ◽  
David R. Kelley ◽  
Jay Shendure
Keyword(s):  
Rna Binding ◽  
Developmental Stages ◽  
Single Cells ◽  
Neuronal Cell ◽  
Alternative Polyadenylation ◽  
Cell Types ◽  
Untranslated Regions ◽  
Mammalian Development ◽  
Translation Rate ◽  
Dynamic Landscape

Abstract3′ untranslated regions (3′ UTRs) post-transcriptionally regulate mRNA stability, localization, and translation rate. While 3′-UTR isoforms have been globally quantified in limited cell types using bulk measurements, their differential usage among cell types during mammalian development remains poorly characterized. In this study, we examine a dataset comprising ~2 million nuclei spanning E9.5–E13.5 of mouse embryonic development to quantify transcriptome-wide changes in alternative polyadenylation (APA). We observe a global lengthening of 3′ UTRs across embryonic stages in all cell types, although we detect shorter 3′ UTRs in hematopoietic lineages and longer 3′ UTRs in neuronal cell types within each stage. An analysis of RNA-binding protein (RBP) dynamics identifies ELAV-like family members, which are concomitantly induced in neuronal lineages and developmental stages experiencing 3′-UTR lengthening, as putative regulators of APA. By measuring 3′-UTR isoforms in an expansive single cell dataset, our work provides a transcriptome-wide and organism-wide map of the dynamic landscape of alternative polyadenylation during mammalian organogenesis.

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