Drouin J, Sun Y L, Tremblay S, Lavender P, Schmidt T J, de Léan A, Nemer M
Institut de Recherches Cliniques de Montréal, Québec, Canada.
Mol Endocrinol. 1992 Aug;6(8):1299-309. doi: 10.1210/mend.6.8.1406707.
The glucocorticoid receptor (GR) is a hormone-inducible transcription factor which activates transcription of specific genes by binding to a DNA sequence present in the promoters of inducible genes. These glucocorticoid response elements (GREs) have a conserved palindromic sequence. Each half-GRE palindrome binds one subunit of GR. We have assessed the relative affinity of GR monomers and homodimers for GRE and determined whether homodimer formation is rate-limiting for high affinity GRE binding. The in vitro affinity of GRE binding by GR homodimers was approximately 2 x 10(-10) M, whereas it was approximately 1 nM for GR monomers. While homodimer:GRE complexes were very stable, monomer:GRE complexes appeared less stable in vitro. At low receptor concentration, GR preferentially bound GRE as a homodimer. Prior dilution of GR (equilibrium shifted to monomers) before addition to a GRE binding reaction resulted in slower kinetics of binding by comparison to parallel reactions in which concentrated (largely homodimeric) GR was added first. Taken together, these experiments suggest that homodimer formation is rate-limiting for high affinity GRE binding. A GRE mutant which contained only a half-binding site and which was unable to bind GR homodimers was also unable to confer glucocorticoid-inducible transcription. Taken together with previous work, these experiments support the model that GR homodimers are required for hormone-dependent activation of transcription and that receptor homodimer formation is rate-limiting for GRE binding.
糖皮质激素受体（GR）是一种激素诱导型转录因子，它通过与诱导基因启动子中存在的DNA序列结合来激活特定基因的转录。这些糖皮质激素反应元件（GREs）具有保守的回文序列。每个半GRE回文结合GR的一个亚基。我们评估了GR单体和同二聚体对GRE的相对亲和力，并确定同二聚体的形成是否是高亲和力GRE结合的限速步骤。GR同二聚体与GRE结合的体外亲和力约为2×10^(-10) M，而GR单体的亲和力约为1 nM。虽然同二聚体：GRE复合物非常稳定，但单体：GRE复合物在体外似乎不太稳定。在低受体浓度下，GR优先以同二聚体形式结合GRE。在加入GRE结合反应之前先对GR进行稀释（平衡转向单体），与首先加入浓缩的（主要是同二聚体的）GR的平行反应相比，结合动力学较慢。综上所述，这些实验表明同二聚体的形成是高亲和力GRE结合的限速步骤。一个只包含半个结合位点且无法结合GR同二聚体的GRE突变体也无法赋予糖皮质激素诱导的转录。与之前的工作一起，这些实验支持了这样一个模型，即GR同二聚体是激素依赖性转录激活所必需的，并且受体同二聚体的形成是GRE结合的限速步骤。
