Phosphorylation and intramolecular stabilization of the ligand binding domain in the nuclear receptor steroidogenic factor 1.
Steroidogenic factor 1 (SF-1) is an orphan nuclear receptor with no known ligand. We showed previously that phosphorylation at serine 203 located N'-terminal to the ligand binding domain (LBD) enhanced cofactor recruitment, analogous to the ligand-mediated recruitment in ligand-dependent receptors. In this study, results of biochemical analyses and an LBD helix assembly assay suggest that the SF-1 LBD adopts an active conformation, with helices 1 and 12 packed against the predicted alpha-helical bundle, in the apparent absence of ligand. Fine mapping of the previously defined proximal activation function in SF-1 showed that the activation function mapped fully to helix 1 of the LBD. Limited proteolyses demonstrate that phosphorylation of S203 in the hinge region mimics the stabilizing effects of ligand on the LBD. Moreover, similar effects were observed in an SF-1/thyroid hormone LBD chimera receptor, illustrating that the S203 phosphorylation effects are transferable to a heterologous ligand-dependent receptor. Our collective data suggest that the hinge together with helix 1 is an individualized specific motif, which is tightly associated with its cognate LBD. For SF-1, we find that this intramolecular association and hence receptor activity are further enhanced by mitogen-activated protein kinase phosphorylation, thus mimicking many of the ligand-induced changes observed for ligand-dependent receptors.
Desclozeaux
Marion
M
Departments of Physiology. Cellular and Molecular Pharmacology. Biochemistry and Biophysics University of California San Francisco, San Francisco, California 94143-0444, USA.
Krylova
Irina N
IN
Horn
Florence
F
Fletterick
Robert J
RJ
Ingraham
Holly A
HA
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051717:Fushi Tarazu Transcription Factors; D018257:Helix-Loop-Helix Motifs; D018398:Homeodomain Proteins; D006801:Humans; D008024:Ligands; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010766:Phosphorylation; D017434:Protein Structure, Tertiary; D018160:Receptors, Cytoplasmic and Nuclear; D011988:Receptors, Thyroid Hormone / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D054339:Steroidogenic Factor 1; D037042:Thyroid Hormone Receptors beta; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D004268:DNA-Binding Proteins; D051717:Fushi Tarazu Transcription Factors; D018398:Homeodomain Proteins; D008024:Ligands; C515057:NR5A1 protein, human; D018160:Receptors, Cytoplasmic and Nuclear; D011988:Receptors, Thyroid Hormone; D011993:Recombinant Fusion Proteins; D054339:Steroidogenic Factor 1; D037042:Thyroid Hormone Receptors beta; D014157:Transcription Factors; C515058:steroidogenic factor 1, mouse; D020928:Mitogen-Activated Protein Kinases