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Phosphorylation of CPSF6 RS-like domain

ABSTRACT: Cleavage factor I mammalian (CFIm) complex, composed of cleavage and polyadenylation specificity factor CPSF6, regulates alternative polyadenylation (APA). CPSF6 has a RS-like domain which plays role in protein -protein interactions. This interaction might have role in alternative polyadenylation site selection. The phosphorylation of RS- like domain might play role in protein -protein interaction and thus might have a role in alternative polyadenylation site selection. So we did mass specteroanalysis to analyse phosphorylation sites of RS-like domain of CPSF6.

INSTRUMENT(S): LTQ Orbitrap Velos

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Kidney

SUBMITTER: Parmit Singh

LAB HEAD: Alan N. Engelman

PROVIDER: PXD012713 | Pride | 2019-04-01

REPOSITORIES: Pride

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			Action	DRS
	293T_rep1.raw	Raw
	293T_rep1.tgz	Other
	293T_rep2.raw	Raw
	293T_rep2_GFY_52380_out.tgz	Other
	CKO_551_rep1.raw	Raw

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Publications

Differential role for phosphorylation in alternative polyadenylation function versus nuclear import of SR-like protein CPSF6.

Jang Sooin S Cook Nicola J NJ Pye Valerie E VE Bedwell Gregory J GJ Dudek Amanda M AM Singh Parmit K PK Cherepanov Peter P Engelman Alan N AN

Nucleic acids research 20190501 9

Cleavage factor I mammalian (CFIm) complex, composed of cleavage and polyadenylation specificity factor 5 (CPSF5) and serine/arginine-like protein CPSF6, regulates alternative polyadenylation (APA). Loss of CFIm function results in proximal polyadenylation site usage, shortening mRNA 3' untranslated regions (UTRs). Although CPSF6 plays additional roles in human disease, its nuclear translocation mechanism remains unresolved. Two β-karyopherins, transportin (TNPO) 1 and TNPO3, can bind CPSF6 in v ...[more]

PMID: 30916345

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Integrative genome-wide analysis reveals HLP1, a novel RNA-binding protein? regulates plant flowering by targeted alternative polyadenylation

Project description:Alternative polyadenylation (APA) is a widespread mechanism for gene regulation and has been implicated in flowering, but the molecular basis governing the choice of a specific poly(A) site during the vegetative-to-reproductive growth transition remains unclear. Here we characterize HLP1, an hnRNP A/B protein as a novel regulator for pre-mRNA 3’ end processing in Arabidopsis. Genetic analysis reveals that HLP1 suppresses Flowering Locus C (FLC), a key repressor of flowering in Arabidopsis. Genome-wide mapping of HLP1-RNA interactions indicates that HLP1 binds preferentially to A-rich and U-rich elements around cleavage and polyadenylation sites, implicating a role in 3’-end formation. We show significantly enriched binding of HLP1 to transcripts involved in RNA metabolism and flowering. Comprehensive profiling of the poly(A) site usage reveals that HLP1 mutations cause thousands of poly(A) site shifts. A distal-to-proximal poly(A) site shift in the flowering regulator FCA, a direct target of HLP1, further leading to up-regulated FLC and delayed flowering. Our results elucidate that HLP1 is a novel factor involved in 3’ end processing and controls reproductive timing via targeted APA. Investigate the role of HLP1 in Alternative Polyadenylation Contributed by: The Institute of Genetics and Developmental Biology (IGDB) of the Chinese Academy of Sciences

2015-06-30 | E-GEOD-39051 | biostudies-arrayexpress

Regulation of alternative cleavage and polyadenylation by 3’ end processing and splicing factors

Project description:Alternative cleavage and polyadenylation (APA) results in mRNA isoforms containing different 3’ untranslated regions (3’UTRs) and/or coding sequences. How core cleavage and polyadenylation (C/P) factors regulate APA is not well understood. Using siRNA knockdown coupled with deep sequencing, we found that several C/P factors can play significant roles in 3’UTR-APA. Whereas Pcf11 and Fip1 enhance usage of proximal poly(A) sites (pAs), CFI-25/68, PABPN1, and PABPC1 promote usage of distal pAs. Strong cis element biases were found for pAs regulated by CFI or Fip1, and the distance between pAs plays an important role in APA regulation. In addition, intronic pAs are substantially regulated by splicing factors, with U1 mostly influencing C/P events in 5’ introns and U2 impacting those in efficiently spliced introns. Furthermore, PABPN1 regulates expression of transcripts with pAs near the transcription start site, a property possibly related to its role in RNA degradation. Finally, we found that groups of APA events regulated by C/P factors are also modulated in cell differentiation and development with distinct trends. Together, our results indicate that the abundance of different C/P factors and splicing factors plays diverse roles in APA, and is relevant to APA regulation in biological conditions. knockdown experiments of 23 C/P factors, 3 splicing factors and U1D in mouse C2C12 myoblast cells

2015-04-01 | E-GEOD-62001 | biostudies-arrayexpress

Alternative cleavage and polyadenylation analysis in an isoform-resolved way by Second- and Third-Generation Sequencing technologies

Project description:Alternative cleavage and polyadenylation (APA) is emerging as an important mechanism of gene regulation in eukaryotes and plays important regulatory roles in human development and diseases. Despite the widespread application of Second Generation Sequencing (SGS) technology for polyadenylation site identification, matching each identified polyadenylation site within a gene to its derived isoform remains a major challenge. To achieve the isoform-resolved APA analysis, we developed a tool termed “IDP-APA” that constructs truly expressed isoforms and identifies polyadenylation sites by integrating the respective strengths of Third Generation Sequencing (TGS) long reads and SGS short reads. Compared to existing tools, IDP-APA demonstrated superior performance in both isoform reconstruction and polyadenylation site identification. Applications to human embryonic stem cells, breast cancer cells and brain tissue from a patient with Alzheimer’s disease revealed prevalent APA events and cell-/tissue-specific APA patterns, especially in an isoform-resolved way.

2017-06-01 | GSE97688 | GEO

A developmental mechanism to Regulate Alternative Polyadenylation in an Adult Stem Cell Lineage [3'seq_OEs]

Project description:Alternative Cleavage and Polyadenylation (APA) often results in production of mRNA isoforms with either longer or shorter 3’UTRs from the same genetic locus, potentially impacting mRNA translation, localization and stability. Developmentally regulated APA can thus make major contributions to cell-type-specific gene expression programs as cells differentiate. During Drosophila spermatogenesis, approximately 500 genes undergo APA when proliferating spermatogonia differentiate into spermatocytes, producing transcripts with shortened 3’ UTRs, leading to profound stage-specific changes in the proteins expressed. The molecular mechanisms that specify usage of upstream polyadenylation sites in spermatocytes are thus key to understanding the changes in cell state. Here, we show that upregulation of PCF11 and Cbc, the two components of Cleavage Factor II (CFII), orchestrates APA during Drosophila spermatogenesis. Knock down of PCF11 or cbc in spermatocytes caused dysregulation of APA, with many transcripts normally cleaved at a proximal site in spermatocytes now cleaved at their distal site, as in spermatogonia. Forced overexpression of CFII components in spermatogonia switched cleavage of some transcripts to the proximal site normally used in spermatocytes. Our findings reveal a developmental mechanism where changes in expression of specific cleavage factors can direct cell-type-specific APA at selected genes.

2024-07-22 | GSE272580 | GEO

A developmental mechanism to Regulate Alternative Polyadenylation in an Adult Stem Cell Lineage [3'seq_KDs]

2024-07-22 | GSE272579 | GEO

A developmental mechanism to Regulate Alternative Polyadenylation in an Adult Stem Cell Lineage [RNA-seq]

2024-07-22 | GSE272581 | GEO

Genome-wide analysis of polyadenylation events in Schmidtea mediterranea (I)

Project description:This study is the first to report genome-scale polyadenylation events in S.mediterranea using poly-A position profiling (3P-Seq).Various cis-acting elements such as hexameric PAS & U/GU enrichment after cleavage site were observed to be conserved in planaria.The cleavage site derived from 3P-Seq could be successfully associated with ~38-60% of transcripts (Makers -38%, oxford- 60% and Dresden- 44%).We also investigated the functional consequences of altered 3’UTRs arising from ApA. Around 97 transcripts were observed to undergo coding region alternate polyadenylation (CR-ApA) that resulted in loss of specific domains from proteins (as inferred from pfam domain search). In this study, we also demonstrated that microRNA-mediated regulation might be one of the key factors playing an important role in selection/evolution of alternate 3’UTRs. The 3’ UTR is one of the key regulatory element that decides the fate of mRNA inside a cell. Switching isoforms according to the need of cell and environmental cues could help the cell to adapt. In this study, we also attempted to study the tissue-specific role of ApA pattern in planaria. Due to the limitations associated with isolation of different tissue-specific cells from planaria, we performed a high-throughput microarray analysis across X1, X2 and Xins cell populations. We were clearly able to identify the differential expression pattern of the ApA events across cell population.

2022-10-01 | GSE78161 | GEO

Genome-wide analysis of polyadenylation events in Schmidtea mediterranea (II)

2022-10-01 | GSE78261 | GEO

A compendium of conserved cleavage and polyadenylation events in mammalian genes

Project description:Cleavage and polyadenylation is essential for 3’ end processing of almost all eukaryotic mRNAs. Recent studies have shown widespread alternative cleavage and polyadenylation (APA) events leading to mRNA isoforms with different 3’UTRs and/or coding sequences. Here we present a compendium of conserved cleavage and polyadenylation sites (PASs) in mammalian genes, based on ~1.2 billion 3’ end sequencing reads from over 360 human, mouse and rat samples. We show that ~80% of mammalian mRNA genes contain at least one conserved PAS, and ~50% have conserved APA events. PAS conservation generally reduces promiscuous 3’ end processing, stabling gene expression levels across species. Conservation of APA correlates with gene age, gene expression features, and gene functions. Genes with certain functions, such as cell morphology, cell proliferation, and mRNA metabolism, are particularly enriched with APA events. While tissue-specific genes typically have a low APA rate, brain-specific genes tend to evolve APA. We show enrichment of mRNA destabilizing motifs in alternative 3’UTR sequences, leading to substantial differences in mRNA stability between 3’UTR APA isoforms. Using conserved PASs, we reveal sequence motifs surrounding APA sites and a preference of adenosine at the cleavage site. Mutations of the U-rich motif around the PAS often accompany APA profile changes between species. Analysis of lncRNA PASs indicates a mechanism of PAS fixation involving evolution of A-rich motifs. Taken together, our results present a comprehensive view of PAS evolution in mammals, and a phylogenic understanding of APA functions.

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hnRNPC regulates cancer-specific alternative cleavage and polyadenylation profiles

Project description:Alternative cleavage and polyadenylation (APA) can occur at more than half of all human genes, greatly enhancing the cellular repertoire of mRNA isoforms. As these isoforms can have altered stability, localisation and coding potential, deregulation of APA can disrupt gene expression and this has been linked to many diseases including cancer progression. How APA generates cancer-specific isoform profiles and what their physiological consequences are, however, is largely unclear. Here we use a subcellular fractionation approach to determine the nuclear and cytoplasmic APA profiles of successive stages of colon cancer using a cell line-based model. Using this approach, we show that during cancer progression specific APA profiles are established. We identify that overexpression of hnRNPC has a critical role in the establishment of APA profiles characteristic for metastatic colon cancer cells, by regulating poly(A) site selection in a subset of genes that have been implicated in cancer progression including MTHFD1L.

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