ION CHANNEL REGULATION BY THE CYTOPLASMIC TAIL OF PDK1

多囊肾病是继单基因突变后最常见的遗传性疾病。在受影响的队列中，PKD-1基因的杂合胚系突变以及随之而来的编码多肽多囊蛋白-1的氨基酸序列的改变与疾病共分离。第二个等位基因的体细胞突变可能是囊变的开始或加速所必需的。膀胱发生与生长控制失调和基质结构改变有关，并被认为需要这样做。这些变化伴随着从上皮净重吸收表型到净分泌表型的转变。然而，尽管人类疾病的某些方面在PKD-1-/-小鼠中的表型复制、候选多囊蛋白-1结合伙伴的发现、多囊蛋白-1亚域过表达引起的细胞信号变化的发现、多囊蛋白-1突变导致持续分泌表型伴随着囊肿大的机制仍不清楚。作为融合蛋白CD16.7-PKD-1(115-226)的多囊蛋白-1的部分C末端胞质结构域的过表达上调了非洲爪哇卵母细胞和HEK293细胞的阳离子通道活性。我们假设，在ADPKD-1杂合子的偶尔的肾小管上皮细胞中，这种或类似功能的扰动伴随着假定的正常生殖系多囊蛋白-1等位基因的体细胞突变，从而导致囊肿的产生。我们进一步假设，这种失调(或这一功能的丧失)促进或导致ADPKD从正常的净溶质重吸收表型向以囊上皮细胞为代表的净分泌表型的转变。我们建议将这些研究扩展到极化的正常和ADPKD上皮细胞，并研究这种活性的调节。我们将寻找新的相互作用蛋白质，并制备适合结构分析的蛋白质。

Polycystic kidney disease is the most common inherited disease secondary to mutation of a single gene. Heterozygous germline mutations in the PKD-1 gene and consequent alterations in the amino acid sequence of the encoded polypeptide, polycystin-1, cosegregate with disease in affected cohorts. Somatic mutations in the second allele may be required for onset of or acceleration of cystogenesis. Cystogenesis is associated with and is thought to require dysregulation of growth control and altered matrix structure. These changes are coupled with conversion from an epithelial phenotype of net reabsorption to one of net secretion. However, despite the phenotypic reproduction of aspects of the human disease in PKD1 -/- mice, the discovery of candidate polycystin-1 binding partners, the discovery of alterations in cell signalling produced by overexpression of a polycystin-1 subdomain, the mechanisms by which polycystin-1 mutations lead to a sustained secretory phenotype accompanied by cyst enlargement remain unknown. Overexpression of part of the C-terminal cytoplasmic domain of polycystin-1 as the fusion protein CD16.7-PKD-1(115-226) upregulates cation channel activity in Xenopus oocytes and in HEK293 cells. We hypothesize that perturbation of this or similar functions accompanies the postulated somatic mutation of the normal germline polycystin-1 allele in the occasional tubular epithelial cells of ADPKD-1 heterozygotes that give rise to cysts. We further hypothesize that this dysregulation (or loss of this function) promotes or causes transition from the normal phenotype of net solute reabsorption to that of net secretion typifying cyst epithelial cells in ADPKD. We propose to extend these studies to polarized normal and ADPKD epithelial cells, and to study regulation of this activity. We will search for novel interacting proteins, and prepare protein suitable for structural analysis.