RGS6 interacts with DMAP1 and DNMT1 and inhibits DMAP1 transcriptional repressor activity

J Biol Chem. 2004 Apr 2;279(14):14120-8. doi: 10.1074/jbc.M309547200. Epub 2004 Jan 20.

Abstract

RGS6 is a member of a subfamily of mammalian RGS proteins that possess DEP (disheveled, Egl-10, pleckstrin) and GGL (G protein gamma subunit-like) domains in addition to the hallmark RGS domain. RGS proteins negatively regulate heterotrimeric G protein signaling by virtue of the GTPase-activating protein activity of their RGS domains. RGS6 exists in multiple splice forms with a long (6L) or short (6S) N terminus, a complete or incomplete GGL domain, in combination with various C-terminal domains. Green fluorescent protein-tagged RGS6L and RGS6S forms exhibit predominantly cytoplasmic and nuclear patterns of distribution in COS-7 cells, respectively, and traffic from these sites to nucleoli in response to stress signaling. We undertook a yeast two-hybrid screen for nuclear RGS6-binding proteins and here identify DMAP1 as an RGS6-interacting protein. DMAP1 is a component of the Dnmt1 complex involved in repression of newly replicated genes. The domains of interaction were mapped to the N-terminal region of the GGL domain of RGS6, a region distinct from its Gbeta5 binding region, and the C-terminal domain of DMAP1. Gbeta5 and DMAP1 did not compete for each other's interaction with RGS6. Co-immunoprecipitation studies in COS-7 cells showed that RGS6L and RGS6S, but not RGS6LDelta258-293 deletion mutant lacking a DMAP1-binding module, co-immunoprecipitate DMAP1 as well as Dnmt1 in a DMAP1-dependent manner. A recombinant GGL domain of RGS6 precipitated endogenous DMAP1 and Dnmt1 in neuroblastoma cell lysates and endogenous DMAP1 co-immunoprecipitated with RGS6L from mouse brain. Co-expression of DMAP1 with RGS6L promoted nuclear migration of RGS6L and its co-localization with DMAP1, a response not observed with RGS6LDelta258-293. RGS6 inhibited the transcriptional repressor activity of DMAP1. RGS6 is the first member of the RGS protein family shown to interact with proteins involved in transcriptional regulation.

Publication types

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

MeSH terms

  • Animals
  • COS Cells
  • Cell Line, Tumor
  • DNA (Cytosine-5-)-Methyltransferase 1
  • DNA (Cytosine-5-)-Methyltransferases / chemistry
  • DNA (Cytosine-5-)-Methyltransferases / genetics
  • DNA (Cytosine-5-)-Methyltransferases / metabolism*
  • Gene Expression Regulation / physiology
  • Humans
  • Protein Binding
  • Protein Structure, Tertiary
  • RGS Proteins / chemistry
  • RGS Proteins / genetics
  • RGS Proteins / metabolism*
  • Repressor Proteins / chemistry
  • Repressor Proteins / genetics
  • Repressor Proteins / metabolism*
  • Transcription, Genetic / physiology*

Substances

  • DMAP1 protein, human
  • Dmap1 protein, mouse
  • RGS Proteins
  • RGS6 protein, human
  • Repressor Proteins
  • Rgs6 protein, mouse
  • DNA (Cytosine-5-)-Methyltransferase 1
  • DNA (Cytosine-5-)-Methyltransferases
  • DNMT1 protein, human
  • Dnmt1 protein, mouse