Phosphorylation of the human retinoid X receptor alpha at serine 260 impairs coactivator(s) recruitment and induces hormone resistance to multiple ligands.
The retinoid X receptor alpha (RXRalpha) is a member of the nuclear receptor superfamily that regulates transcription of target genes through heterodimerization with several partners, including peroxisome proliferator-activated receptor, retinoic acid receptor, thyroid receptor, and vitamin D receptor (VDR). We have shown previously that signaling through VDR.RXRalpha heterodimers was attenuated in ras-transformed keratinocytes due to phosphorylation of serine 260 of the RXRalpha via the activated Ras-Raf-MAPK cascade in these cells. In this study we demonstrate that phosphorylation at serine 260, a site located in the omega loop-AF-2 interacting domain of RXRalpha, inhibits signaling through several heterodimeric partners of the RXRalpha. The inhibition of signaling results in reduced transactivational response to ligand presentation and the reduced physiological response of growth inhibition not only of 1,25-dihydroxyvitamin D3 but also of retinoic acid receptor alpha ligands and LG1069 (an RXRalpha ligand). This partial resistance to ligands could be reversed by inhibition of MAPK activity or by overexpression of a non-phosphorylable RXRalpha mutant at serine 260 (RXRalpha Ser-260-->Ala). Importantly, phosphorylation of RXRalpha at serine 260 impaired the recruitment of DRIP205 and other coactivators to the VDR.RXRalpha complex. Chromatin immunoprecipitation and pulldown assays further demonstrated that coactivator recruitment to the VDR.RXR complex could be restored by treatment with a MAPK inhibitor. Our data suggest that phosphorylation at serine 260 plays a critical role in inducing hormone resistance of RXRalpha-mediated signaling likely through structural changes in the H1-H3 omega loop-AF2 coactivator(s) interacting domain.
Macoritto
Michael
M
McGill University Health Centre, Montreal, Quebec H3A 1A1, Canada.
Nguyen-Yamamoto
Loan
L
Huang
Dao Chao
DC
Samuel
Sara
S
Yang
Xian Fang
XF
Wang
Tian Tian
TT
White
John H
JH
Kremer
Richard
R
D002117:Calcitriol / Q000378:metabolism / Q000494:pharmacology; D002461:Cell Line, Transformed; D004351:Drug Resistance / Q000502:physiology; D048049:Extracellular Signal-Regulated MAP Kinases; D006728:Hormones / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008024:Ligands; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000502:physiology; D056892:Mediator Complex; D046912:Multiprotein Complexes / Q000235:genetics / Q000378:metabolism*; D015689:Oncogene Protein p21(ras) / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D017434:Protein Structure, Tertiary / Q000502:physiology; D018167:Receptors, Calcitriol / Q000235:genetics / Q000378:metabolism*; D047490:Retinoid X Receptor alpha / Q000235:genetics / Q000378:metabolism*; D012694:Serine / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014815:Vitamins / Q000378:metabolism / Q000494:pharmacology; D048490:raf Kinases / Q000235:genetics / Q000378:metabolism
D006728:Hormones; D008024:Ligands; C488666:MED13 protein, human; D056892:Mediator Complex; D046912:Multiprotein Complexes; D018167:Receptors, Calcitriol; D047490:Retinoid X Receptor alpha; D014157:Transcription Factors; D014815:Vitamins; D012694:Serine; D048490:raf Kinases; D048049:Extracellular Signal-Regulated MAP Kinases; D015689:Oncogene Protein p21(ras); D002117:Calcitriol