REGULATION OF PARATHYROID FUNCTION

描述（改编自申请人的摘要）：主要 甲状旁腺功能亢进是一种相对常见的内分泌功能障碍， 发病率为每10万人40-45例。 二次 甲状旁腺功能亢进是肾衰竭的一种表现。 有超过 在美国有10万名透析患者。 因此， 甲状旁腺功能给健康护理管理带来了很大负担 和劳动力的流失。 甲状旁腺的控制 激素分泌几乎是唯一的，因为钙最终抑制 而在大多数其他系统中，钙刺激分泌。 这 研究的重点是参与的分子机制， 控制钙介导的PTH分泌。 GTP结合信号的作用 正常甲状旁腺分泌中的转导蛋白和蛋白激酶C 将被研究。 合成寡核苷酸探针将用于选择 对于正常牛甲状旁腺表达文库中的DNA序列 在噬菌体λ gt 10中构建。 与信号相对应的DNA 转导蛋白将被亚克隆，并通过双脱氧核苷酸序列分析进行测序。 终止链法推断GTP结合蛋白的类型 在甲状旁腺细胞中表达。 这将确定是否 甲状旁腺细胞含有与其他细胞相同的信号转导蛋白， 分泌组织，或者如果有一个独特的转导蛋白， 甲状旁腺，这可以解释反向钙敏感性 激素分泌 基于衍生的氨基酸序列，肽 将合成特定序列。 这些肽将用于 产生特异于单个信号转导的多克隆抗血清 蛋白质，以便定量甲状旁腺细胞中的这些蛋白质。 的 作为百日咳毒素底物的转导蛋白， 霍乱毒素和一种与磷脂酶C连接的毒素不敏感蛋白 将被定性。 百日咳毒素和霍乱毒素对 钙、镁和锶诱导的PTH分泌抑制， 评价胞质钙的变化。 蛋白激酶的作用 甲状旁腺分泌中的C将通过监测酶活性来评估， 并通过使用对这三种蛋白特异的抗肽抗血清， 蛋白激酶C亚型。鞘氨醇碱代谢是 细胞外钙、镁和锶将被评估， 蛋白激酶C抑制剂的生产。 这些研究利用了 甲状旁腺表达文库、DNA测序和抗肽抗体 将为将来检查正常和异常的 甲状旁腺功能

DESCRIPTION (Adapted from the Applicant's Abstract): Primary hyperparathyroidism is a relatively common endocrine dysfunction with an incidence of 40-45 cases per 100,000 population. Secondary hyperparathyroidism is a manifestation of renal failure. There are over 100,000 patients on dialysis in the United States. Thus, disorders of parathyroid function impose a significant burden on health care management and loss from the labor force each year. The control of parathyroid hormone secretion is almost unique in that calcium ultimately inhibits secretion whereas in most other systems calcium stimulates secretion. This investigation focuses on molecular mechanisms that are involved in the control of calcium-mediated PTH secretion. The role of GTP-binding signal transducing proteins and protein kinase C in normal parathyroid secretion will be studied. Synthetic oligonucleotide probes will be used to select for DNA sequences in a normal bovine parathyroid expression library constructed in phage lambda gtl0. The DNA corresponding to the signal transduction proteins will be subcloned and sequenced by the dideoxy chain-termination method to deduce the types of GTP-binding proteins expressed in the parathyroid cell. This will establish whether the parathyroid cell contains signal transduction proteins common to other secretory tissues, or if there is a unique transducing protein to the parathyroid which can account for the reverse calcium sensitivity of hormone secretion. Based upon the derived amino acid sequences, peptides of specific sequence will be synthesized. These peptides will be used to generate polyclonal antisera specific to the individual signal transduction proteins in order to quantitate these proteins in parathyroid cells. The transducing proteins which serve as substrates for pertussis toxin and cholera toxin, and a toxin-insensitive protein linked to phospholipase C will be characterized. The effect of pertussis toxin and cholera toxin on calcium, magnesium and strontium-induced suppression of PTH secretion and changes in cytosolic calcium will be evaluated. The role of protein kinase C in parathyroid secretion will be evaluated by monitoring enzyme activity, and through the use of anti-peptide antisera specific for the three isoforms of protein kinase C. Sphingoid base metabolism as a function of extracellular calcium, magnesium and strontium will be evaluated for the production of protein kinase C inhibitors. These studies utilizing the parathyroid expression library, DNA sequencing and antipeptide antibodies will form a basis for the future examination of normal and abnormal parathyroid function.