Delineation of two distinct type 1 activation functions in the androgen receptor amino-terminal domain.
Based on the finding that some transcription factors contain multiple transcriptional regulatory activities, we constructed a panel of rat androgen receptor (AR) mutants containing small internal deletions and point mutations within the amino-terminal region of the receptor. Trans-activation assays in CV-1 cells using AR-responsive reporter genes were performed and led to the identification of two noncontiguous trans-activation regions in the AR amino terminus. One of these regions, termed activator function 1a (AF-1a) is a highly-conserved 14-amino acid segment that is predicted to form a beta-turn followed by an acidic amphipathic alpha-helix. Point mutagenesis within AF-1a revealed that two adjacent hydrophobic residues were required for full AR trans-activation function, as arginine substitutions resulted in a 60% reduction in transcriptional activity. A second amino-terminal region was also identified and has been designated AF-1b. Deletion of the 65-amino acid AF-1b segment, which contains numerous glutamate and aspartate residues, caused a 55% decrease in trans-activation function. An AF-1a/AF-1b double mutant retains less than 10% trans-activation function compared with wild-type AR, suggesting that AF-1a and AF-1b may each contribute separately to maximal AR activity. To determine whether AF-1a and AF-1b play a role in AR-mediated trans-repression of AP-1 function, we tested single and double AF-1a/AF-1b mutants in a transient trans-repression assay. Our results showed that neither AF-1a nor AF-1b was required for AP-1 trans-repression, demonstrating that AR-mediated trans-repression and trans-activation are discrete functions.
Department of Biochemistry, University of Arizona, Tucson, Arizona 85721, USA.
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NL
Chamberlain
D C
DC
Whitacre
R L
RL
Miesfeld
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002460:Cell Line; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010449:Peptide Mapping; D010641:Phenotype; D011487:Protein Conformation; D051381:Rats; D011944:Receptors, Androgen / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion; D014158:Transcription, Genetic
D011944:Receptors, Androgen