Molecular defect in activation of a mutant Stat5b

最近的一份报告发现，一名患者表现出生长激素不敏感的临床特征，他在Stat5b基因上存在纯合子错义突变，导致SH2结构域附近630位氨基酸的丙氨酸替换为脯氨酸。突变的Stat5b蛋白造成的确切分子缺陷尚不清楚。初步实验表明，突变的Stat5b蛋白在培养的哺乳动物细胞中瞬时转染后被检测到，但始终保持在野生型Stat5b蛋白水平的50%。在重组实验中，突变体Stat5b在生长激素刺激下未能增强报告基因的活性。这项研究提案试图确定突变的Stat5b在其失活周期中的特定分子缺陷，强调两个假设的功能障碍领域。第一个特定目标是研究突变的Stat5b的蛋白质稳定性和亚细胞定位。第二个特定目的是研究附近SH2结构域的功能是否被破坏，使得突变的Stat5b在二聚化过程中不能与激活的生长激素受体/JAK2复合体或其他Stat5b分子相互作用。

A recent report identified a patient presenting with clinical features of growth hormone insensitivity, who harbored a homozygous mis-sense mutation in the Stat5b gene that resulted in an alanine to proline substitution at amino acid 630, adjacent to the SH2 domain. The precise molecular defect caused by the mutant Stat5b protein has not been elucidated. Preliminary experiments have demonstrated that the mutant Stat5b protein is detected upon transient transfection in cultured mammalian cells, however at consistently <50% the level of wild-type Stat5b protein. In reconstitution experiments, mutant Stat5b failed to enhance reporter gene activity upon growth hormone stimulation. This research proposal attempts to identify the specific molecular defect of the mutant Stat5b in its inactivation cycle, highlighting two hypothesized areas of dysfunction. The first specific aim addresses protein stability and subcellular location of mutant Stat5b. The second specific aim focuses on whether the function of the nearby SH2 domain is disrupted, such that the mutant Stat5b can not interact with the activated growth hormone receptor/Jak2 complex or with other Stat5b molecules in the dimerization process.