Methods for measuring ligand dissociation and nuclear receptor turnover in whole cells

Understanding the molecular mechanisms of steroid hormone action requires assays that measure rates of ligand dissociation and receptor degradation. Ligand dissociation is a pseudo-first order reaction of a high affinity [3H]-labeled ligand. Receptor turnover as described here is the rate of degradation of a radiolabeled receptor. The methods make use of transient expression of a nuclear receptor in cultured cells and are applicable to all nuclear receptors. Rates of ligand dissociation and receptor degradation provided the first insight into the interdomain interactions of the androgen receptor and the molecular basis for the phenotypic effects of naturally occurring androgen receptor loss-of-function germline mutations and gain-of-function somatic mutations, and for the potency differences between the biologically active androgens, testosterone, and dihydrotestosterone.