Karl M, Lamberts S W, Detera-Wadleigh S D, Encio I J, Stratakis C A, Hurley D M, Accili D, Chrousos G P
Developmental Endocrinology Branch, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892.
J Clin Endocrinol Metab. 1993 Mar;76(3):683-9. doi: 10.1210/jcem.76.3.8445027.
The clinical syndrome of generalized, compensated glucocorticoid resistance is characterized by increased cortisol secretion without clinical evidence of hyper- or hypocortisolism, and manifestations of androgen and/or mineralocorticoid excess. This condition results from partial failure of the glucocorticoid receptor (GR) to modulate transcription of its target genes. We studied the molecular mechanisms of this syndrome in a Dutch kindred, whose affected members had hypercortisolism and approximately half of normal GRs, and whose proband was a young woman with manifestations of hyperandrogenism. Using the polymerase chain reaction to amplify and sequence each of the nine exons of the GR gene alpha, along with their 5'- and 3'-flanking regions, we identified a 4-base deletion at the 3'-boundary of exon 6 in one GR allele (delta 4), which removed a donor splice site in all three affected members studied. In contrast, the sequence of exon 6 in the two unaffected siblings was normal. A single nucleotide substitution causing an amino acid substitution in the amino terminal domain of the GR (asparagine to serine, codon 363) was also discovered in exon 2 of the other allele (G1220) in the proband, in one of her affected brothers and in her unaffected sister. The functional importance of this mutation was tested in a cotransfection study using the recombinant expression vector pRShGR-Ser363 and the glucocorticoid responsive vector mouse mammary tumor virus-chloramphenicol transferase. This amino acid substitution did not alter the function of the glucocorticoid receptor. Using reverse transcription-polymerase chain reaction we could only identify messenger RNA transcripts of the G1220-allele but not of the delta 4-allele in the affected members of this family who were heterozygous for the G1220 mutation. This deletion in the glucocorticoid receptor gene was, thus, associated with the expression of only one allele and a decrease of GR protein by 50% in affected members of this glucocorticoid resistant family. The mutation identified in exon 2 did not segregate with the disease and appears to be of no functional significance. The presence of the null allele was apparently compensated for by increased cortisol production at the expense of concurrent hyperandrogenism.
全身性、代偿性糖皮质激素抵抗的临床综合征的特征是皮质醇分泌增加，但无皮质醇增多或减少的临床证据，以及雄激素和/或盐皮质激素过多的表现。这种情况是由于糖皮质激素受体（GR）调节其靶基因转录的部分功能失效所致。我们在一个荷兰家族中研究了这种综合征的分子机制，该家族中受影响的成员存在皮质醇增多症且GR数量约为正常水平的一半，先证者是一名有高雄激素血症表现的年轻女性。我们使用聚合酶链反应扩增并测序GR基因α的九个外显子及其5'和3'侧翼区域，在一个GR等位基因的外显子6的3'边界处发现了一个4碱基缺失（δ4），在所有三名受研究的受影响成员中，该缺失去除了一个供体剪接位点。相比之下，两名未受影响的兄弟姐妹的外显子6序列正常。在先证者、她的一名受影响的兄弟和她未受影响的姐妹的另一个等位基因（G1220）的外显子2中，还发现了一个单核苷酸替换，导致GR氨基末端结构域中的一个氨基酸替换（天冬酰胺替换为丝氨酸，密码子363）。在一项共转染研究中，使用重组表达载体pRShGR-Ser363和糖皮质激素反应性载体小鼠乳腺肿瘤病毒-氯霉素转移酶，测试了该突变的功能重要性。这种氨基酸替换并未改变糖皮质激素受体的功能。通过逆转录-聚合酶链反应，我们在该家族中对G1220突变呈杂合状态的受影响成员中，只能鉴定出G1220等位基因的信使RNA转录本，而未鉴定出δ4等位基因的转录本。因此，在这个糖皮质激素抵抗家族的受影响成员中，糖皮质激素受体基因中的这种缺失与仅一个等位基因的表达以及GR蛋白减少50%相关。在外显子2中鉴定出的突变与疾病不相关，似乎没有功能意义。无效等位基因的存在显然通过增加皮质醇产生得到了代偿，代价是同时出现高雄激素血症。
