Cleavage and polyadenylation specificity factor 1 (CPSF1) regulates alternative splicing of interleukin 7 receptor (IL7R) exon 6

Irina Evsyukova, Shelton Bradrick, Simon G. Gregory, Mariano Garcia-Blanco

Research output: Contribution to journalArticle

23 Citations (Scopus)

Abstract

Interleukin 7 receptor, IL7R, is expressed exclusively on cells of the lymphoid lineage, and its expression is crucial for the development and maintenance of T cells. Alternative splicing of IL7R exon 6 results in membrane-bound (exon 6 included) and soluble (exon 6 skipped) IL7R isoforms. Interestingly, the inclusion of exon 6 is affected by a single-nucleotide polymorphism associated with the risk of developing multiple sclerosis. Given the potential association of exon 6 inclusion with multiple sclerosis, we investigated the cis-acting elements and trans -acting factors that regulate exon 6 splicing. We identified multiple exonic and intronic cis -acting elements that impact inclusion of exon 6. Moreover, we utilized RNA affinity chromatography followed by mass spectrometry to identify trans -acting protein factors that bind exon 6 and regulate its splicing. These experiments identified cleavage and polyadenylation specificity factor 1 (CPSF1) among protein-binding candidates. A consensus polyadenylation signal AAUAAA is present in intron 6 of IL7R directly downstream from the 5′ splice site. Mutations to this site and CPSF1 knockdown both resulted in an increase in exon 6 inclusion. We found no evidence that this site is used to produce cleaved and polyadenylated mRNAs, suggesting that CPSF1 interaction with intronic IL7R pre-mRNA interferes with spliceosome binding to the exon 6 5′ splice site. Our results suggest that competing mRNA splicing and polyadenylation regulate exon 6 inclusion and consequently determine the ratios of soluble to membrane-bound IL7R . This may be relevant for both T cell ontogeny and function and development of multiple sclerosis.

Original languageEnglish (US)
Pages (from-to)103-115
Number of pages13
JournalRNA
Volume19
Issue number1
DOIs
StatePublished - Jan 2013
Externally publishedYes

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Cleavage And Polyadenylation Specificity Factor
Interleukin-7 Receptors
Alternative Splicing
Exons
Multiple Sclerosis
RNA Splice Sites
Polyadenylation
Trans-Activators
Spliceosomes
T-Lymphocytes
Messenger RNA
Membranes
RNA Precursors
Cell Lineage
Affinity Chromatography

Keywords

  • Alternative splicing
  • CPSF1
  • IL7R
  • Multiple sclerosis
  • Polyadenylation

ASJC Scopus subject areas

  • Molecular Biology

Cite this

Cleavage and polyadenylation specificity factor 1 (CPSF1) regulates alternative splicing of interleukin 7 receptor (IL7R) exon 6. / Evsyukova, Irina; Bradrick, Shelton; Gregory, Simon G.; Garcia-Blanco, Mariano.

In: RNA, Vol. 19, No. 1, 01.2013, p. 103-115.

Research output: Contribution to journalArticle

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abstract = "Interleukin 7 receptor, IL7R, is expressed exclusively on cells of the lymphoid lineage, and its expression is crucial for the development and maintenance of T cells. Alternative splicing of IL7R exon 6 results in membrane-bound (exon 6 included) and soluble (exon 6 skipped) IL7R isoforms. Interestingly, the inclusion of exon 6 is affected by a single-nucleotide polymorphism associated with the risk of developing multiple sclerosis. Given the potential association of exon 6 inclusion with multiple sclerosis, we investigated the cis-acting elements and trans -acting factors that regulate exon 6 splicing. We identified multiple exonic and intronic cis -acting elements that impact inclusion of exon 6. Moreover, we utilized RNA affinity chromatography followed by mass spectrometry to identify trans -acting protein factors that bind exon 6 and regulate its splicing. These experiments identified cleavage and polyadenylation specificity factor 1 (CPSF1) among protein-binding candidates. A consensus polyadenylation signal AAUAAA is present in intron 6 of IL7R directly downstream from the 5′ splice site. Mutations to this site and CPSF1 knockdown both resulted in an increase in exon 6 inclusion. We found no evidence that this site is used to produce cleaved and polyadenylated mRNAs, suggesting that CPSF1 interaction with intronic IL7R pre-mRNA interferes with spliceosome binding to the exon 6 5′ splice site. Our results suggest that competing mRNA splicing and polyadenylation regulate exon 6 inclusion and consequently determine the ratios of soluble to membrane-bound IL7R . This may be relevant for both T cell ontogeny and function and development of multiple sclerosis.",
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