scholarly journals Systematic identification of functional SNPs interrupting 3’ UTR polyadenylation signals

2019 ◽  
Author(s):  
Eldad David Shulman ◽  
Ran Elkon
Keyword(s):  
Gene Expression ◽  
Nuclear Export ◽  
Cellular Localization ◽  
Alternative Polyadenylation ◽  
Regulatory Elements ◽  
Translation Efficiency ◽  
Protein Coding ◽  
A Site ◽  
The Impact ◽  
Main Variant

AbstractAlternative polyadenylation (APA) is emerging as a widespread regulatory layer as the majority of human protein-coding genes contain several polyadenylation (p(A)) sites in their 3’ UTRs. By generating isoforms with different 3’ UTR length, APA potentially affects mRNA stability, translation efficiency, nuclear export, and cellular localization. Polyadenylation sites are regulated by adjacent RNA cis-regulatory elements, the principals among them are the polyadenylation signal (PAS) AAUAAA and its main variant AUUAAA, typically located ~20- nt upstream of the p(A) site. Mutations in PAS and other auxiliary poly(A) cis-elements in the 3’ UTR of several genes have been shown to cause human Mendelian diseases, and to date, only a few common SNPs that regulate APA were associated with complex diseases. Here, we systematically searched for SNPs that affect gene expression and human traits by modulation of 3’ UTR APA. Focusing on the variants most likely to exert the strongest effect, we identified 2,305 SNPs that interrupt the canonical PAS or its main variant. Implementing pA-QTL tests using GTEx RNA-seq data, we identified 139 PAS SNPs significantly associated with the usage of their p(A) site. As expected, PAS-interrupting alleles were significantly linked with decreased cleavage at their p(A) site and the consequential 3’ UTR lengthening. As an indication for a functional effect of these PAS SNPs on gene expression, 65 of the pA-QTLs were also detected as eQTLs of the same gene in the same tissue. Furthermore, we observed that PAS-interrupting alleles linked with 3’ UTR lengthening were also strongly associated with decreased gene expression, pointing that shorter isoforms generated by APA are generally more stable than longer ones. Last, indicative of the impact of PAS SNPs on human phenotypes, 53 pA-QTLs overlapped GWAS SNPs that are significantly linked with human traits.

scholarly journals FIP1L1 Regulates Alternative Polyadenylation of Leukemia-Associated Genes in Acute Myeloid Leukemia

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 3882-3882
Author(s):  
Amanda G Davis ◽  
Takahiro Shima ◽  
Ruijia Wang ◽  
Dinghai Zheng ◽  
Bin Tian ◽  
...  
Keyword(s):  
Gene Expression ◽  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Leukemia Cell ◽  
Alternative Polyadenylation ◽  
Translation Efficiency ◽  
Hematopoietic Stem ◽  
Altered Expression ◽  
The Impact ◽  
Acute Myeloid

Abstract Alternative polyadenylation (APA) is a prevalent mechanism of post-transcriptional gene regulation. APA can impact the protein coding portion of a gene or the length of the 3' untranslated region (UTR), a crucial mRNA region mediating stability, localization, and translation efficiency. Global shifts in APA usage have been reported during development and in cancer transformation; however, the impact of APA in acute myeloid leukemia (AML) is currently unexplored. Therefore, we sought to address whether global shifts in APA patterns can be seen in leukemic blasts and if APA changes contribute to leukemogenesis. Here we report that leukemic blasts exhibit global transcript shortening that contributes to the upregulation of leukemia-associated genes: NRAS, BAALC, and MAPKAPK3. We also implicate FIP1L1 as an important mediator of APA alterations in AML. To identify whether global trends of APA are seen in leukemia patients, we performed a 3'RNA sequencing experiment comparing healthy CD34+ hematopoietic stem and progenitor cells (HSPCs) with CD34+ enriched leukemic blasts from primary patient samples. We observed a trend in whole transcript shortening, with an emphasis on 3'UTR shortening of affected genes. Importantly, genes with altered APA were among pathways associated with oncogenic transformation and specifically AML. We selected three genes for further functional validation (NRAS, BAALC, and MAPKAPK3) that exhibited a correlation of shortened 3'UTRs with higher mRNA expression. Using mRNA stability assays and luciferase assays, we confirmed that 3'UTR shortening contributes to the upregulation of these leukemia-associated genes in blood cells. To probe upstream regulators of APA alterations in leukemia, we performed overexpression and knockdown studies of APA machinery members that have altered expression in leukemia patients. We identified FIP1L1 as a novel regulator of APA in AML. Overexpression of FIP1L1 resulted in 3'UTR lengthening and downregulation of our three selected genes. Conversely, knockdown led to 3' UTR shortening and upregulation. Interestingly, altering FIP1L1 expression (both up and downregulation) is detrimental to leukemia cell lines, suggesting that modest changes in gene expression can have a dramatic impact on hematopoietic cell fitness. We conclude that APA changes represent another dysregulated layer of gene expression that can contribute to AML development. Current studies aimed at identifying the role of FIP1L1 in healthy HSPCs will elucidate the feasibility of FIP1L1 as a novel therapeutic target in AML. Disclosures No relevant conflicts of interest to declare.

scholarly journals RNA polyadenylation and its consequences in prokaryotes

2018 ◽  
Vol 373 (1762) ◽  
pp. 20180166 ◽  
Author(s):  
Eliane Hajnsdorf ◽  
Vladimir R. Kaberdin
Keyword(s):  
Gene Expression ◽  
State Level ◽  
Rna Degradation ◽  
Plasmid Copy Number ◽  
Plasmid Copy ◽  
Protein Coding ◽  
Mrna Metabolism ◽  
Isolation And Characterization ◽  
Polymerase I ◽  
The Impact

Post-transcriptional addition of poly(A) tails to the 3′ end of RNA is one of the fundamental events controlling the functionality and fate of RNA in all kingdoms of life. Although an enzyme with poly(A)-adding activity was discovered in Escherichia coli more than 50 years ago, its existence and role in prokaryotic RNA metabolism were neglected for many years. As a result, it was not until 1992 that E. coli poly(A) polymerase I was purified to homogeneity and its gene was finally identified. Further work revealed that, similar to its role in surveillance of aberrant nuclear RNAs of eukaryotes, the addition of poly(A) tails often destabilizes prokaryotic RNAs and their decay intermediates, thus facilitating RNA turnover. Moreover, numerous studies carried out over the last three decades have shown that polyadenylation greatly contributes to the control of prokaryotic gene expression by affecting the steady-state level of diverse protein-coding and non-coding transcripts including antisense RNAs involved in plasmid copy number control, expression of toxin–antitoxin systems and bacteriophage development. Here, we review the main findings related to the discovery of polyadenylation in prokaryotes, isolation, and characterization and regulation of bacterial poly(A)-adding activities, and discuss the impact of polyadenylation on prokaryotic mRNA metabolism and gene expression. This article is part of the theme issue ‘5′ and 3′ modifications controlling RNA degradation’.

scholarly journals Characterization of the nuclear and cytosolic transcriptomes in human brain tissue reveals new insights into the subcellular distribution of RNA transcripts

2020 ◽  
Author(s):  
Ammar Zaghlool ◽  
Adnan Niazi ◽  
Åsa K. Björklund ◽  
Jakub Orzechowski Westholm ◽  
Adam Ameur ◽  
...  
Keyword(s):  
Gene Expression ◽  
Subcellular Localization ◽  
Human Brain ◽  
Expression Analysis ◽  
Differential Expression Analysis ◽  
Adult Brain ◽  
Sequencing Data ◽  
Human Brain Tissue ◽  
Protein Coding ◽  
The Impact

AbstractTranscriptome analysis has mainly relied on analyzing RNA sequencing data from whole cells, overlooking the impact of subcellular RNA localization and its influence on our understanding of gene function, and interpretation of gene expression signatures in cells. Here, we performed a comprehensive analysis of cytosolic and nuclear transcriptomes in human fetal and adult brain samples. We show significant differences in RNA expression for protein-coding and lncRNA genes between cytosol and nucleus. Transcripts displaying differential subcellular localization belong to particular functional categories and display tissue-specific localization patterns. We also show that transcripts encoding the nuclear-encoded mitochondrial proteins are significantly enriched in the cytosol compared to the rest of protein-coding genes. Further investigation of the use of the cytosolic or the nuclear transcriptome for differential gene expression analysis indicates important differences in results depending on the cellular compartment. These differences were manifested at the level of transcript types and the number of differentially expressed genes. Our data provide a resource of RNA subcellular localization in the human brain and highlight differences in using the cytosolic or the nuclear transcriptomes for differential expression analysis.

scholarly journals Actively translated uORFs reduce translation and mRNA stability independent of NMD in Arabidopsis

2021 ◽  
Author(s):  
Hsin-Yen Larry Wu ◽  
Polly Yingshan Hsu
Keyword(s):  
Gene Expression ◽  
Mrna Stability ◽  
Gene Expression Regulation ◽  
Regulatory Elements ◽  
Expression Regulation ◽  
Ribosome Profiling ◽  
Translation Efficiency ◽  
Nonsense Mediated Decay ◽  
Protein Levels ◽  
Eukaryotic Genes

ABSTRACTUpstream ORFs (uORFs) are widespread cis-regulatory elements in the 5’ untranslated regions of eukaryotic genes. Translation of uORFs could negatively regulate protein synthesis by repressing main ORF (mORF) translation and by reducing mRNA stability presumably through nonsense-mediated decay (NMD). While the above expectations were supported in animals, they have not been extensively tested in plants. Using ribosome profiling, we systematically identified 2093 Actively Translated uORFs (ATuORFs) in Arabidopsis seedlings and examined their roles in gene expression regulation by integrating multiple genome-wide datasets. Compared with genes without uORFs, we found ATuORFs result in 38%, 14%, and 43% reductions in translation efficiency, mRNA stability, and protein levels, respectively. The effects of predicted but not actively translated uORFs are much weaker than those of ATuORFs. Interestingly, ATuORF-containing genes are also expressed at higher levels and encode longer proteins with conserved domains, features that are common in evolutionarily older genes. Moreover, we provide evidence that uORF translation in plants, unlike in vertebrates, generally does not trigger NMD. We found ATuORF-containing transcripts are degraded through 5’ to 3’ decay, while NMD targets are degraded through both 5’ to 3’ and 3’ to 5’ decay, suggesting uORF-associated mRNA decay and NMD have distinct genetic requirements. Furthermore, we showed ATuORFs and NMD repress translation through separate mechanisms. Our results reveal that the potent inhibition of uORFs on mORF translation and mRNA stability in plants are independent of NMD, highlighting a fundamental difference in gene expression regulation by uORFs in the plant and animal kingdoms.

scholarly journals Codon usage is an important determinant of gene expression levels largely through its effects on transcription

2016 ◽  
Vol 113 (41) ◽  
pp. E6117-E6125 ◽  
Author(s):  
Zhipeng Zhou ◽  
Yunkun Dang ◽  
Mian Zhou ◽  
Lin Li ◽  
Chien-hung Yu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Codon Usage ◽  
Important Determinant ◽  
Mrna Translation ◽  
Mrna Levels ◽  
Protein Coding ◽  
Expression Levels ◽  
Highly Expressed Genes ◽  
The Impact ◽  
Gene Expression Levels

Codon usage biases are found in all eukaryotic and prokaryotic genomes, and preferred codons are more frequently used in highly expressed genes. The effects of codon usage on gene expression were previously thought to be mainly mediated by its impacts on translation. Here, we show that codon usage strongly correlates with both protein and mRNA levels genome-wide in the filamentous fungus Neurospora. Gene codon optimization also results in strong up-regulation of protein and RNA levels, suggesting that codon usage is an important determinant of gene expression. Surprisingly, we found that the impact of codon usage on gene expression results mainly from effects on transcription and is largely independent of mRNA translation and mRNA stability. Furthermore, we show that histone H3 lysine 9 trimethylation is one of the mechanisms responsible for the codon usage-mediated transcriptional silencing of some genes with nonoptimal codons. Together, these results uncovered an unexpected important role of codon usage in ORF sequences in determining transcription levels and suggest that codon biases are an adaptation of protein coding sequences to both transcription and translation machineries. Therefore, synonymous codons not only specify protein sequences and translation dynamics, but also help determine gene expression levels.

scholarly journals The Impact of Leadered and Leaderless Gene Structures on Translation Efficiency, Transcript Stability, and Predicted Transcription Rates in Mycobacterium smegmatis

2020 ◽  
Vol 202 (9) ◽  
Author(s):  
Tien G. Nguyen ◽  
Diego A. Vargas-Blanco ◽  
Louis A. Roberts ◽  
Scarlet S. Shell
Keyword(s):  
Gene Expression ◽  
Mycobacterium Tuberculosis ◽  
Mycobacterium Smegmatis ◽  
Half Life ◽  
Untranslated Regions ◽  
Translation Efficiency ◽  
Production Rates ◽  
Content Type ◽  
The Impact ◽  
Mrna Half Life

ABSTRACT Regulation of gene expression is critical for Mycobacterium tuberculosis to tolerate stressors encountered during infection and for nonpathogenic mycobacteria such as Mycobacterium smegmatis to survive environmental stressors. Unlike better-studied models, mycobacteria express ∼14% of their genes as leaderless transcripts. However, the impacts of leaderless transcript structures on mRNA half-life and translation efficiency in mycobacteria have not been directly tested. For leadered transcripts, the contributions of 5′ untranslated regions (UTRs) to mRNA half-life and translation efficiency are similarly unknown. In M. tuberculosis and M. smegmatis, the essential sigma factor, SigA, is encoded by a transcript with a relatively short half-life. We hypothesized that the long 5′ UTR of sigA causes this instability. To test this, we constructed fluorescence reporters and measured protein abundance, mRNA abundance, and mRNA half-life and calculated relative transcript production rates. The sigA 5′ UTR conferred an increased transcript production rate, shorter mRNA half-life, and decreased apparent translation rate compared to a synthetic 5′ UTR commonly used in mycobacterial expression plasmids. Leaderless transcripts appeared to be translated with similar efficiency as those with the sigA 5′ UTR but had lower predicted transcript production rates. A global comparison of M. tuberculosis mRNA and protein abundances failed to reveal systematic differences in protein/mRNA ratios for leadered and leaderless transcripts, suggesting that variability in translation efficiency is largely driven by factors other than leader status. Our data are also discussed in light of an alternative model that leads to different conclusions and suggests leaderless transcripts may indeed be translated less efficiently. IMPORTANCE Tuberculosis, caused by Mycobacterium tuberculosis, is a major public health problem killing 1.5 million people globally each year. During infection, M. tuberculosis must alter its gene expression patterns to adapt to the stress conditions it encounters. Understanding how M. tuberculosis regulates gene expression may provide clues for ways to interfere with the bacterium’s survival. Gene expression encompasses transcription, mRNA degradation, and translation. Here, we used Mycobacterium smegmatis as a model organism to study how 5′ untranslated regions affect these three facets of gene expression in multiple ways. We furthermore provide insight into the expression of leaderless mRNAs, which lack 5′ untranslated regions and are unusually prevalent in mycobacteria.

scholarly journals Regulation of mRNA cap methylation

2009 ◽  
Vol 425 (2) ◽  
pp. 295-302 ◽  
Author(s):  
Victoria H. Cowling
Keyword(s):  
Gene Expression ◽  
Cell Viability ◽  
Nuclear Export ◽  
Present Review ◽  
Biological Processes ◽  
Experimental Systems ◽  
Efficient Gene ◽  
Cellular Mrnas ◽  
Eukaryotic Organisms ◽  
The Impact

The 7-methylguanosine cap added to the 5′ end of mRNA is essential for efficient gene expression and cell viability. Methylation of the guanosine cap is necessary for the translation of most cellular mRNAs in all eukaryotic organisms in which it has been investigated. In some experimental systems, cap methylation has also been demonstrated to promote transcription, splicing, polyadenylation and nuclear export of mRNA. The present review discusses how the 7-methylguanosine cap is synthesized by cellular enzymes, the impact that the 7-methylguanosine cap has on biological processes, and how the mRNA cap methylation reaction is regulated.

scholarly journals Impact and evolutionary determinants of Neanderthal introgression on transcriptional and post-transcriptional regulation

2019 ◽  
Author(s):  
Martin Silvert ◽  
Lluis Quintana-Murci ◽  
Maxime Rotival
Keyword(s):  
Gene Expression ◽  
Gene Regulation ◽  
Transcriptional Regulation ◽  
Regulatory Elements ◽  
Strong Impact ◽  
Modern Humans ◽  
Archaic Admixture ◽  
Archaic Introgression ◽  
Post Transcriptional Regulation ◽  
The Impact

AbstractArchaic admixture is increasingly recognized as an important source of diversity in modern humans, with Neanderthal haplotypes covering 1-3% of the genome of present-day Eurasians. Recent work has shown that archaic introgression has contributed to human phenotypic diversity, mostly through the regulation of gene expression. Yet, the mechanisms through which archaic variants alter gene expression, and the forces driving the introgression landscape at regulatory regions remain elusive. Here, we explored the impact of archaic introgression on transcriptional and post-transcriptional regulation, focusing on promoters and enhancers across 127 different tissues as well as microRNA-mediated regulation. Although miRNAs themselves harbor few archaic variants, we found that some of these variants may have a strong impact on miRNA-mediated gene regulation. Enhancers were by far the regulatory elements most affected by archaic introgression, with one third of the tissues tested presenting significant enrichments. Specifically, we found strong enrichments of archaic variants in adipose-related tissues and primary T cells, even after accounting for various genomic and evolutionary confounders such as recombination rate and background selection. Interestingly, we identified signatures of adaptive introgression at enhancers of some key regulators of adipogenesis, raising the interesting hypothesis of a possible adaptation of early Eurasians to colder climates. Collectively, this study sheds new light onto the mechanisms through which archaic admixture have impacted gene regulation in Eurasians and, more generally, increases our understanding of the contribution of Neanderthals to the regulation of acquired immunity and adipose homeostasis in modern humans.

scholarly journals Candidate cancer driver mutations in superenhancers and long-range chromatin interaction networks

2017 ◽  
Author(s):  
Lina Wadi ◽  
Liis Uusküla-Reimand ◽  
Keren Isaev ◽  
Shimin Shuai ◽  
Vincent Huang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Long Range ◽  
Target Genes ◽  
Tumor Biology ◽  
Regulatory Elements ◽  
Chromatin Interaction ◽  
Driver Mutations ◽  
Regulatory Regions ◽  
Protein Coding ◽  
Primary Tumors

AbstractA comprehensive catalogue of the mutations that drive tumorigenesis and progression is essential to understanding tumor biology and developing therapies. Protein-coding driver mutations have been well-characterized by large exome-sequencing studies, however many tumors have no mutations in protein-coding driver genes. Non-coding mutations are thought to explain many of these cases, however few non-coding drivers besides TERT promoter are known. To fill this gap, we analyzed 150,000 cis-regulatory regions in 1,844 whole cancer genomes from the ICGC-TCGA PCAWG project. Using our new method, ActiveDriverWGS, we found 41 frequently mutated regulatory elements (FMREs) enriched in non-coding SNVs and indels (FDR<0.05) characterized by aging-associated mutation signatures and frequent structural variants. Most FMREs are distal from genes, reported here for the first time and also recovered by additional driver discovery methods. FMREs were enriched in super-enhancers, H3K27ac enhancer marks of primary tumors and long-range chromatin interactions, suggesting that the mutations drive cancer by distally controlling gene expression through threedimensional genome organization. In support of this hypothesis, the chromatin interaction network of FMREs and target genes revealed associations of mutations and differential gene expression of known and novel cancer genes (e.g., CNNB1IP1, RCC1), activation of immune response pathways and altered enhancer marks. Thus distal genomic regions may include additional, infrequently mutated drivers that act on target genes via chromatin loops. Our study is an important step towards finding such regulatory regions and deciphering the somatic mutation landscape of the non-coding genome.

scholarly journals DPAC: a tool for Differential Poly(A) Site usage from poly(A)–targeted RNAseq data

2019 ◽  
Author(s):  
Andrew Routh
Keyword(s):  
Gene Expression ◽  
Alternative Polyadenylation ◽  
Data Mapping ◽  
Library Synthesis ◽  
Sequencing Data ◽  
Synthesis Strategy ◽  
Rnaseq Data ◽  
Differential Gene ◽  
A Site

AbstractPoly(A)-tail targeted RNAseq approaches, such as 3’READS, PAS-seq and Poly(A)-ClickSeq, are becoming popular alternatives to random-primed RNAseq for simplified gene expression analyses as well as to measure changes in poly(A) site usage. We and others have recently demonstrated that these approaches perform similarly to other RNAseq strategies, while saving on the volume of sequencing data required and providing a simpler library synthesis strategy. Here, we present DPAC; a streamlined pipeline for the preprocessing of poly(A)-tail targeted RNAseq data, mapping of poly(A)-sites and poly(A) clustering, and determination of differential poly(A) site usage using DESeq2. Changes in poly(A) site usage is simultaneously used to report differential gene expression, differential terminal exon usage and alternative polyadenylation (APA).

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