Molecular Mechanisms for CFIm-Mediated Regulation of mRNA Alternative Polyadenylation

Zhu, Yong; Wang, Xiuye; Forouzmand, Elmira; Jeong, Joshua; Qiao, Feng; Sowd, Gregory A.; Engelman, Alan; Xie, Xiaohui; Hertel, Klemens J.; Shi, Yongsheng

doi:10.1016/j.molcel.2017.11.031

Cited by 172 publications

(242 citation statements)

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“…We found that 1134 individual PACs were differentially expressed (IHW-padj < 0.1, foldchange >1.5) (Supplemental Data 3). Volcano plots illustrating these changes in gene expression and PAC abundance are shown in The predominant shortening of 3'UTRs upon knock-down of CF25Im is the expected phenotype and is consistent with our and other previous analyses [3,14]. By virtue of measuring differential usage of each individual PAS independently, this pipeline revealed DE of PASs outside of annotated regions.…”

Section: Resultssupporting

confidence: 87%

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

Routh

2019

Preprint

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Poly(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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Section: Resultssupporting

confidence: 87%

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

Routh

2019

Preprint

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“…Some examples of APA changes triggered by Chtop knockdown are presented in Figures 4G and S3C. As a reference, knockdown of Cpsf6 in HEK293T has been reported to lead to 775 significant APA changes (718 distal to proximal and 57 proximal to distal) ( (Martin et al, 2012) data re-analyzed by (Zhu et al, 2018)).…”

Section: Chtop Participates In Apa Regulation In Vivomentioning

confidence: 99%

“…Various human APA trans-regulators have been reported, some directly involved in CPA (Cpsf5, Cpsf6, Cstf2) (Martin et al, 2012;Masamha et al, 2014;Yao et al, 2012;Zhu et al, 2018)1, others linked to splicing (hnRNPC, Tardbp, U2af2, U1 snRNP) (Gruber et al, 2016;…”

Section: Introductionmentioning

confidence: 99%

Co-transcriptional loading of RNA export factors shapes the human transcriptome

Viphakone

Sudbery

Heath

et al. 2018

Preprint

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During gene expression, RNA export factors are mainly known for driving nucleocytoplasmic transport. Whilst early studies suggested that the Exon Junction Complex provides a binding platform for them, subsequent work proposed that they are only recruited by the Cap-Binding Complex to the 5' end of RNAs, as part of TREX. Using iCLIP, we show that the export receptor Nxf1 and two TREX subunits, Alyref and Chtop, are actually recruited to the whole mRNA co-transcriptionally via splicing but before 3'-end processing. Consequently, Alyref alters splicing decisions and Chtop regulates alternative polyadenylation. Alyref is recruited to the 5'-end of RNAs by CBC and our data reveal subsequent binding to RNAs near EJCs. We demonstrate eIF4A3 stimulates Alyref deposition not only on spliced RNAs close to EJC sites, but also single exon transcripts. Our study reveals mechanistic insights into the cotranscriptional recruitment of mRNA export factors and how this shapes the human transcriptome. Kaida et al., 2010;Millevoi et al., 2006;Rot et al., 2017), and Thoc5, which plays a role in mRNA export (Katahira et al., 2013).A major pathway for nuclear mRNA export uses the TREX complex (Strässer et al., 2002) which contains a core THO sub-complex, the RNA helicase Ddx39b, and RNA export adaptors and co-adaptors that Ddx39b loads onto mRNAs (Heath et al., 2016). A major adaptor is Alyref (Stutz et al., 2000), though some shuttling SR proteins can also work as adaptors (Huang and Steitz, 2003). Several co-adaptors have been characterized: Chtop, Thoc5, Cpsf6 and Rbm15 (Chang et al., 2013;Katahira et al., 2009;Ruepp et al., 2009;Zolotukhin et al., 2009). These proteins together recruit the export receptor Nxf1 and stimulate its RNA binding activity, which promotes mRNA export Hautbergue et al., 2008;Viphakone et al., 2015;. The topology of TREX components on RNAs and the molecular mechanisms involved in their deposition are still unclear. It has been suggested that the EJC may serve as a binding platform for RNA export factors (Le Hir et al., 2001;Singh et al., 2012).Yet, these interactions were not seen in other studies which instead revealed a key role for the cap binding complex (CBC, containing Ncbp1 and Ncbp2) in recruiting TREX, consistent with the idea that mRNPs may be exported 5' end first (Cheng et al., 2006;Chi et al., 2013). Thus, the relative contribution of the EJC and CBC to TREX deposition on the RNA in vivo remains unresolved.Here, we address some of these outstanding questions by performing individual nucleotide resolution UV-crosslinking and immunoprecipitation (iCLIP) (Broughton and Pasquinelli, 2013;König et al., 2011;Lee and Ule, 2018) on the mRNA export factors Alyref, Chtop, and Nxf1. Our in vivo results suggest co-transcriptional recruitment of these proteins all along the RNA during splicing but before CPA, which grants them additional roles in gene expression. Alyref can bind inefficiently spliced introns and regulates their splicing, and Chtop binds last exons and participates in APA regulation. We est...

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“…Several studies in which key polyadenylation proteins were knocked down have revealed significant roles for each of them. For example, subunits of mammalian cleavage factor I (CFI m ) have been shown to play important roles in APA, favoring use of distal polyadenylation sites (Li et al, ; Martin, Gruber, Keller, & Zavolan, ; Y. Zhu et al, ). As such, CFI m influences neurite growth (Fukumitsu, Soumiya, & Furukawa, ) and may be involved in glioblastoma (Masamha et al, ).…”

Section: Polyadenylation and Alternative Polyadenylationmentioning

confidence: 99%

Tissue‐specific mechanisms of alternative polyadenylation: Testis, brain, and beyond (2018 update)

MacDonald

2019

WIREs RNA

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Alternative polyadenylation (APA) is how genes choose different sites for 3′ end formation for mRNAs during transcription. APA often occurs in a tissue‐ or developmental stage‐specific manner that can significantly affect gene activity by changing the protein product generated, the stability of the transcript, its localization within the cell, or its translatability. Despite the important regulatory effects that APA has on tissue‐specific gene expression, only a few examples have been characterized mechanistically. In this 2018 update to our 2010 review, we examine mechanisms for the control of APA and update our understanding of the older mechanisms since 2010. We once postulated the existence of tissue‐specific factors in APA. However, while a few tissue‐specific polyadenylation factors are known, the emerging conclusion is that the majority of APA is accomplished by altering levels of core polyadenylation proteins. Examples of those core proteins include CSTF2, CPSF1, and subunits of mammalian cleavage factor I. But despite support for these mechanisms, no one has yet documented any of these proteins changing in either a tissue‐specific or developmental manner. Given the profound effect that APA can have on gene expression and human health, improved understanding of tissue‐specific APA could lead to numerous advances in gene activity control. This article is categorized under: RNA Processing > 3′ End Processing RNA in Disease and Development > RNA in Development

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Molecular Mechanisms for CFIm-Mediated Regulation of mRNA Alternative Polyadenylation

Cited by 172 publications

References 50 publications

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

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

Co-transcriptional loading of RNA export factors shapes the human transcriptome

Tissue‐specific mechanisms of alternative polyadenylation: Testis, brain, and beyond (2018 update)

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