HORMONE RESISTANCE IN PATIENTS WITH PSEUDOHYPOPARATHYROIDISM

许多激素的作用需要它们与特定的激素相互作用。 通过鸟嘌呤核苷酸偶联的细胞表面受体- 结合蛋白Gs对腺苷酸环化酶的刺激作用。的表达 生物活性改变的荷尔蒙、受体或Gs分子 会导致荷尔蒙作用的增加或减少。该协议检查 激素在信号转导中作用改变的分子基础 利用Gs蛋白作为信号转导的通路。这些 疾病包括假性甲状旁腺功能减退症1a型和McCune Albright 甲状旁腺功能减退症 由于钙敏感受体或甲状旁腺素基因的缺陷，以及短 由于生长激素释放激素受体缺陷导致的身高。 患者表型的生化和临床特征 促进了以实验室为基础的方法来阐明分子 疾病的机制。在过去的一年里，我们主要专注于 甲状旁腺激素抵抗的分子基础阐明 假性甲状旁腺功能减退症(PHP)Lb型，一种常染色体综合征，其中 激素抵抗仅限于甲状旁腺激素靶组织，并与 G蛋白活性正常。我们已经研究并描述了 甲状旁腺激素抵抗患者的生化表型及其临床意义 来自几个多层家庭的未受影响的亲属。外周血 用Epstein-Barr病毒转化淋巴母细胞，基因组DNA 和总RNA从转化的B-杆菌培养物中分离得到。 淋巴细胞。我们之前的工作排除了甲状旁腺素受体基因 这些家系中甲状旁腺激素抵抗的基础。为了鉴定基因 这解释了PHP1b型对PTH的抗性，我们已经执行了 利用半自动荧光聚合酶链式反应技术进行全基因组扫描 分析微卫星标记基因座的连锁关系。我们已经完成了 所有常染色体间隔20 cM的连锁图谱，并使用 一项仅受影响的分析，我们发现PHP类型lb与上一个基因座存在连锁 10q染色体，LOD分数为2.94。我们最近开始 分析另一个大型的，多元化的家庭，我们预计 增加来自该家庭的受影响对象的信息量将会增加 我们的LOD得分大于3，因此确认链接在5厘米以内 正确的轨迹。未来的研究将集中在识别 这一区域的候选基因。

The action of many hormones requires their interaction with specific cell surface receptors that are coupled by the guanine nucleotide- binding protein Gs to stimulation of adenylyl cyclase. Expression of hormones, receptors, or Gs molecules with altered biological activity can result in gain or loss of hormone action. This protocol examines the molecular basis for altered hormone action in signal transduction pathways that utilize Gs protein as a signal transducer. These disorders include pseudohypoparathyroidism type la and McCune Albright syndrome due to altered expression or function of Gs, hypoparathyroidism due to defects in the calcium-sensing receptor or PTH gene, and short stature due to defects in the growth hormone-releasing hormone receptor. Biochemical and clinical characterization of the patient's phenotype facilitates a laboratory-based approach to elucidating the molecular mechanism of disease. Over the past year we have focused primarily on elucidation of the molecular basis for PTH resistance in subjects with pseudohypoparathyroidism (PHP) type lb, an autosomal syndrome in which hormone resistance is limited to PTH target tissues and associated with normal G protein activity. We have examined and characterized the biochemical phenotype of subjects with PTH resistance and their unaffected relatives from several multiplex kindreds. Peripheral blood lymphoblasts were transformed with Epstein-Barr virus, and genomic DNA and total RNA were isolated from cultures of the transformed B- lymphocytes. Our previous work had excluded the PTH receptor gene as the basis for PTH resistance in these families. To identify the gene that accounts for PTH resistance in PHP type lb, we have performed a genome-wide scan using semi-automated fluorescence-based PCR technology to analyze linkage of microsatellite marker loci. We have completed a linkage map at 20 cM intervals of all autosomal chromosomes, and using an affected-only analysis we found linkage of PHP type lb to a locus on chromosome 10q that has a LOD score of 2.94. We have recently begun to analyze another large, multiplex family, and we anticipate that the addition of informative affected subjects from this family will increase our LOD score to greater than 3, thus confirming linkage to within 5 cm of the correct locus. Future studies will focus on identification of the candidate gene in this region.