Nuclear coactivator-62 kDa/Ski-interacting protein is a nuclear matrix-associated coactivator that may couple vitamin D receptor-mediated transcription and RNA splicing

J Biol Chem. 2003 Sep 12;278(37):35325-36. doi: 10.1074/jbc.M305191200. Epub 2003 Jul 2.

Abstract

Nuclear coactivator-62 kDa/Ski-interacting protein (NCoA62/SKIP) is a putative vitamin D receptor (VDR) and nuclear receptor coactivator protein that is unrelated to other VDR coactivators such as those in the steroid receptor coactivator (SRC) family. The mechanism through which NCoA62/SKIP functions in VDR-activated transcription is unknown. In the present study, we identified a nuclear localization sequence in the COOH terminus of NCoA62/SKIP and showed that NCoA62/SKIP was targeted to nuclear matrix subdomains. Chromatin immunoprecipitation studies revealed that endogenous NCoA62/SKIP associated in a 1,25-dihydroxyvitamin D3-dependent manner with VDR target genes in ROS17/2.8 osteosarcoma cells. A cyclic pattern of promoter occupancy by VDR, SRC-1, and NCoA62/SKIP was observed, with NCoA62/SKIP entering these promoter complexes after SRC-1. These studies provide strong support for the proposed role of NCoA62/SKIP as a VDR transcriptional coactivator, and they indicate that key mechanistic differences probably exist between NCoA62/SKIP and SRC coactivators. To explore potential mechanisms, NCoA62/SKIP-interacting proteins were purified from HeLa cell nuclear extracts and identified by mass spectrometry. The identified proteins represent components of the spliceosome as well as other nuclear matrix-associated proteins. Here, we show that a dominant negative inhibitor of NCoA62/SKIP (dnNCoA62/SKIP) interfered with appropriate splicing of transcripts derived from 1,25-dihydroxyvitamin D3-induced expression of a growth hormone minigene cassette. Taken together, these data show that NCoA62/SKIP has properties that are consistent with those of nuclear receptor coactivators and with RNA spliceosome components, thus suggesting a potential role for NCoA62/SKIP in coupling VDR-mediated transcription to RNA splicing.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Base Sequence
  • COS Cells
  • Calcitriol / pharmacology
  • Cell Nucleus / metabolism
  • Chlorocebus aethiops
  • DNA Primers
  • HeLa Cells
  • Humans
  • Nuclear Proteins / metabolism*
  • Nuclear Receptor Coactivators
  • Plasmids
  • RNA Splicing* / drug effects
  • RNA, Messenger / drug effects
  • RNA, Messenger / genetics
  • Receptors, Calcitriol / metabolism*
  • Recombinant Fusion Proteins / metabolism
  • Reverse Transcriptase Polymerase Chain Reaction
  • Trans-Activators / metabolism
  • Transcription Factors / metabolism*
  • Transcription, Genetic* / drug effects
  • Transfection

Substances

  • DNA Primers
  • Nuclear Proteins
  • Nuclear Receptor Coactivators
  • RNA, Messenger
  • Receptors, Calcitriol
  • Recombinant Fusion Proteins
  • SNW1 protein, human
  • Trans-Activators
  • Transcription Factors
  • Calcitriol