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

Sooin Jang; Nicola J Cook; Valerie E Pye; Gregory J Bedwell; Amanda M Dudek; Parmit K Singh; Peter Cherepanov; Alan N Engelman

doi:10.1093/nar/gkz206

Back

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

Journal article

Peer reviewed

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

Sooin Jang, Nicola J Cook, Valerie E Pye, Gregory J Bedwell, Amanda M Dudek, Parmit K Singh, Peter Cherepanov and Alan N Engelman

Nucleic acids research, Vol.47(9), pp.4663-4683

05/21/2019

DOI: 10.1093/nar/gkz206

PMCID: PMC6511849

PMID: 30916345

View Online

Abstract

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 vitro, and we demonstrate here that while the TNPO1 binding site is dispensable for CPSF6 nuclear import, the arginine/serine (RS)-like domain (RSLD) that mediates TNPO3 binding is critical. The crystal structure of the RSLD-TNPO3 complex revealed potential CPSF6 interaction residues, which were confirmed to mediate TNPO3 binding and CPSF6 nuclear import. Both binding and nuclear import were independent of RSLD phosphorylation, though a hyperphosphorylated mimetic mutant failed to bind TNPO3 and mislocalized to the cell cytoplasm. Although hypophosphorylated CPSF6 largely supported normal polyadenylation site usage, a significant number of mRNAs harbored unnaturally extended 3' UTRs, similar to what is observed when other APA regulators, such as CFIIm component proteins, are depleted. Our results clarify the mechanism of CPSF6 nuclear import and highlight differential roles for RSLD phosphorylation in nuclear translocation versus regulation of APA.

Active Transport, Cell Nucleus - genetics

beta Karyopherins - chemistry

beta Karyopherins - genetics

Crystallography, X-Ray

Drosophila Proteins - chemistry

Drosophila Proteins - genetics

Humans

mRNA Cleavage and Polyadenylation Factors - chemistry

mRNA Cleavage and Polyadenylation Factors - genetics

Phosphorylation - genetics

Polyadenylation - genetics

Protein Binding - genetics

Protein Conformation

Protein Serine-Threonine Kinases - chemistry

Protein-Tyrosine Kinases - chemistry

RNA, Messenger

RNA-Binding Proteins - chemistry

RNA-Binding Proteins - genetics

Details

Title: Subtitle: Differential role for phosphorylation in alternative polyadenylation function versus nuclear import of SR-like protein CPSF6
Creators: Sooin Jang - Harvard Medical School
Nicola J Cook - The Francis Crick Institute
Valerie E Pye - The Francis Crick Institute
Gregory J Bedwell - Harvard Medical School
Amanda M Dudek - Harvard Medical School
Parmit K Singh - Harvard Medical School
Peter Cherepanov - Imperial College London
Alan N Engelman - Harvard Medical School
Resource Type: Journal article
Publication Details: Nucleic acids research, Vol.47(9), pp.4663-4683
DOI: 10.1093/nar/gkz206
PMID: 30916345
PMCID: PMC6511849
NLM abbreviation: Nucleic Acids Res
ISSN: 0305-1048
eISSN: 1362-4962
Grant note: P50 AI150481 / NIAID NIH HHS P30 AI060354 / NIAID NIH HHS T32 AI007386 / NIAID NIH HHS P50 GM082251 / NIGMS NIH HHS
Language: English
Date published: 05/21/2019
Academic Unit: Microbiology and Immunology
Record Identifier: 9984823121202771

Metrics

5 Record Views

See more details