Vitamin A metabolite, all-trans-retinoic acid, mediates alternative splicing of protein kinase C δVIII (PKCδVIII) isoform via splicing factor SC35

Hercules Apostolatos, André Apostolatos, Timothy Vickers, James E. Watson, Shijie Song, Fernando Vale, Denise R. Cooper, Juan Sanchez-Ramos, Niketa A. Patel

Research output: Contribution to journalArticle

21 Scopus citations

Abstract

Vitamin A metabolite, all-trans-retinoic acid (RA), induces cell growth, differentiation, and apoptosis and has an emerging role in gene regulation and alternative splicing events. Protein kinase Cδ (PKCδ), a serine/threonine kinase, has a role in cell proliferation, differentiation, and apoptosis. We reported an alternatively spliced variant of human PKCδ, PKCδVIII that functions as a pro-survival protein (1). RA regulates the splicing and expression of PKCδVIII via utilization of a downstream 5′ splice site of exon 10 on PKCδ pre-mRNA. Here, we further elucidate the molecular mechanisms involved in RA regulation of alternative splicing of PKCδVIII mRNA. Overexpression and knockdown of the splicing factor SC35 (i.e. SRp30b) indicated that it is involved in PKCδVIII alternative splicing. To identify the cis-elements involved in 5′ splice site selection we cloned a minigene, which included PKCδ exon 10 and its flanking introns in the pSPL3 splicing vector. Alternative 5′ splice site utilization in the minigene was promoted by RA. Further, co-transfection of SC35 with PKCδ minigene promoted selection of 5′ splice site II. Mutation of the SC35 binding site in the PKCδ minigene abolished RA-mediated utilization of 5′ splice splice II. RNA binding assays demonstrated that the enhancer element downstream of PKCδ exon 10 is a SC35 cis-element. We conclude that SC35 is pivotal in RA-mediated PKCδ pre-mRNA alternative splicing. This study demonstrates how a nutrient, vitamin A, via its metabolite RA, regulates alternative splicing and thereby gene expression of the pro-survival protein PKCδVIII.

Original languageEnglish (US)
Pages (from-to)25987-25995
Number of pages9
JournalJournal of Biological Chemistry
Volume285
Issue number34
DOIs
StatePublished - Aug 20 2010

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Apostolatos, H., Apostolatos, A., Vickers, T., Watson, J. E., Song, S., Vale, F., Cooper, D. R., Sanchez-Ramos, J., & Patel, N. A. (2010). Vitamin A metabolite, all-trans-retinoic acid, mediates alternative splicing of protein kinase C δVIII (PKCδVIII) isoform via splicing factor SC35. Journal of Biological Chemistry, 285(34), 25987-25995. https://doi.org/10.1074/jbc.M110.100735