Temperature-dependent inactivation of nucleic acid binding and aggregation of the 1,25-dihydroxyvitamin D3 receptor.

ABSTRACT

The interaction of the 1 alpha,25-dihydroxyvitamin D3 receptor with immobilized calf thymus DNA has been compared with its sedimentation properties on hypotonic sucrose gradients. Forty to sixty percent of total hormonereceptor complexes formed at 4 degrees C were retained by DNA-cellulose and could be eluted by 0.18 to 0.2 M KCl. In contrast, heating preparations to 25 degrees C rapidly and irreversibly converted receptor to a form which bound hormone and DEAE-cellulose normally, but was unable to associate with DNA. Similarly, the ability of receptor to aggregate to a 6 S species was labile at 25 degrees C. Stabilization of receptor in the DNA binding aggregating form was accomplished using Ca2+, Mg2+, Mn2+, or Na2MoO4 while several protease and phosphatase inhibitors were ineffective. An examination of DNA binding properties of aggregating and nonaggregating receptor forms revealed that only receptor competent to enter into aggregates could bind DNA suggesting that a functional nucleic acid binding site, and, hence, a nucleic acid interaction is necessary for aggregate formation. Consistent with this view, an RNAreceptor interaction appears to be involved in formation of the 6 S complex since removal of RNA by ribonuclease treatment or purification of receptor reduced aggregation, an effect that could be reversed by addition of purified RNA.