Analysis of the steroid receptor coactivator 1 (SRC1)-CREB binding protein interaction interface and its importance for the function of SRC1.
The transcriptional activity of nuclear receptors is mediated by coactivator proteins, including steroid receptor coactivator 1 (SRC1) and its homologues and the general coactivators CREB binding protein (CBP) and p300. SRC1 contains an activation domain (AD1) which functions via recruitment of CBP and and p300. In this study, we have used yeast two-hybrid and in vitro interaction-peptide inhibition experiments to map the AD1 domain of SRC1 to a 35-residue sequence potentially containing two alpha-helices. We also define a 72-amino-acid sequence in CBP necessary for SRC1 binding, designated the SRC1 interaction domain (SID). We show that in contrast to SRC1, direct binding of CBP to the estrogen receptor is weak, suggesting that SRC1 functions primarily as an adaptor to recruit CBP and p300. In support of this, we show that the ability of SRC1 to enhance ligand-dependent nuclear receptor activity in transiently transfected cells is dependent upon the integrity of the AD1 region. In contrast, the putative histone acetyltransferase domain, the Per-Arnt-Sim basic helix-loop-helix domain, the glutamine-rich domain, and AD2 can each be removed without loss of ligand-induced activity. Remarkably, a construct corresponding to residues 631 to 970, which contains only the LXXLL motifs and the AD1 region of SRC1, retained strong coactivator activity in our assays.
Sheppard
H M
HM
Department of Biochemistry, University of Leicester, Leicester, LE1 7RH, United Kingdom.
Harries
J C
JC
Hussain
S
S
Bevan
C
C
Heery
D M
DM
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D050882:CREB-Binding Protein; D002460:Cell Line; D017930:Genes, Reporter; D051548:Histone Acetyltransferases; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D056920:Nuclear Receptor Coactivator 1; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011960:Receptors, Estrogen / Q000378:metabolism; D020218:Response Elements / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics; D013816:Thermodynamics; D015534:Trans-Activators / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D020798:Two-Hybrid System Techniques
D009687:Nuclear Proteins; D010446:Peptide Fragments; D011960:Receptors, Estrogen; D015534:Trans-Activators; D014157:Transcription Factors; D050882:CREB-Binding Protein; C494363:CREBBP protein, human; D051548:Histone Acetyltransferases; C538859:NCOA1 protein, human; D056920:Nuclear Receptor Coactivator 1