ACTIVATION FUNCTION-1 OF RETINOIC ACID RECEPTOR BETA-2 IS AN ACIDIC ACTIVATOR RESEMBLING VP16

The mechanisms underlying transcriptional activation are not very well understood, and knowledge is based on experiments with a small number of mostly viral activators. We have investigated the mechanism underl ying transactivation by the activation domain present in the N-termina l part of retinoic acid receptor (RAR) beta 2 (AF-1), We show that RAR beta 2 phosphorylation is not crucial for its activity although it ma y modulate AF-1 activity, Sequential mutation of the negatively charge d residues (Asp) resulted in a stepwise decrease in activity, while mu tation of all aspartic acid residues resulted in complete loss of acti vity, Comparison of the critical region for activation with other acti vators revealed moderate homology with the viral activator VP16., The hydrophobic amino acids surrounding the negatively charged residues re ported to be critical for activation by VP16 are all conserved in AF-1 , The hydrophobic residues are required for AF-1, since mutation of th ese residues resulted in a decrease in activity, Furthermore, the acti vity of this activator, VP16 and TA(1) of RelA, is squelched by overex pression of an AF-1-containing expression construct, indicating that A F-1 is an acidic activator, Squelching experiments further indicate th at AF-1 and AF-2 function by different mechanisms, Comparison of activ ation functions present in the AB region of other members of the stero id/thyroid hormone receptor family: RAR alpha 2, RAR gamma 2, and GR s uggested that also these receptors contain an acidic activation domain , The mechanism underlying activation by AF-1 is discussed.