NOVEL HOMEOBOX GENES IN DEVELOPING GENITOURINARY TRACT

许多影响肾脏和肾脏的发育障碍的原因 泌尿生殖道(例如，肾发育不全、发育不良和囊性 )仍然未知，很大程度上是因为人们对 正常肾脏发育的分子机制。 含有同源异型盒的基因负责器官的分化 许多物种，其中一个基因(Hox-1.5)的破坏产生 小鼠的先天畸形。最近，我们克隆了一本小说 一类小鼠的同源异型盒基因(命名为Cux)。Cux基因 都与果蝇的切割基因有关，这是必需的 为了昆虫排泄器官--马氏管的正常发育 这是一个原始的肾脏。Cux的推测结构 基因产品表明，它们也可能编码核 参与基因调控的转录因子。此外，我们发现， 至少有一个基因(Cux-1)在肾脏、睾丸、 和肾上皮细胞提示它可能在 哺乳动物肾脏和泌尿生殖道的正常发育。 因此，Cux-1表达的异常可能参与了 泌尿生殖系统异常或肿瘤的发病机制。进一步测试 这些假说，编码Cux-1、Cux-2和相关基因的全长cDNA 将通过文库筛选和聚合酶链式反应克隆小鼠切割的同源物。这个 卵巢癌组织中Cux-1和Cux-2表达的时空分布 小鼠生殖道发育有待进一步研究 采用原位杂交和半定量聚合酶链式反应。特定的 Cux融合蛋白的抗血清将用于鉴定和 Cux基因产物在成熟和发育肾脏中的免疫定位 还有睾丸。Cux-1和Cux-2的启动子/调控区 基因将被克隆、测序和表达。Cux-1基因是否 绑定到自己的启动子/监管区域的产品将被评估为 鉴定其DNA结合特异性的第一步。这个 Cux-1基因的功能作用将通过特定的 反义寡核苷酸对肾上皮细胞的抑制作用。 从细胞增殖、形态、极性、上皮细胞 组织和基因表达的功能作用可以归因于 Cux-1基因。Cux-1基因是否是卵巢癌发生所必需的 小鼠的肾脏和泌尿生殖道将通过破坏进行测试 在转基因小鼠中进行同源重组和表达。我们的 长期目标将是解决Cux基因在一系列 肾脏发生的基因调控机制。在这些 未来的研究，步入更靠近CUX的通路 (Cux基因和基因产物的调控)和远端的步骤 (靶基因)将被更详细地研究。这样的研究应该 有助于我们对泌尿生殖道的基本了解 发展。此外，对这一类小说的进一步刻画 含有同源异型盒的基因将会与整体研究有关 真核基因调控。

The causes of many developmental disorders affecting the kidney and genitourinary tract (e.g., renal agenesis, hypoplasia, and cystic dysplasia) remain unknown, largely because little is known about the molecular mechanisms underlying development of the normal kidney. Homeobox-containing genes are responsible for organ differentiation in many species, and disruption of one such gene (Hox-1.5) produces congenital abnormalities in the mouse. Recently, we cloned a novel class of homeobox-containing genes (named cux) from mouse. Cux genes are related to the cut gene of Drosophila melanogaster which is required for normal development of Malpighian tubules, the insect excretory organ that serves as a primitive kidney. The inferred structures of the cux gene products indicate that they also are likely to encode nuclear transcription factors involved in gene regulation. Moreover, we found that at least one of the genes (cux-1) is expressed in kidney, testis, and renal epithelial cells suggesting that it may have a role in the normal development of the mammalian kidney and genitourinary tract. Abnormalities of cux-1 expression, then, might be involved in the pathogenesis of urogenital abnormalities or neoplasms. To further test these hypotheses, full-length cDNAs encoding cux-1, cux-2, and related murine cut homologs will be cloned by library screening and PCR. The temporal and spatial patterns of expression of cux-1 and cux-2 in the developing genitourinary tract of mouse will be further investigated using in situ hybridization and semi-quantitative PCR. Specific antisera raised to cux fusion proteins will be used to identify and immunolocalize the cux gene products in the mature and developing kidney and testis. The promoter/regulatory regions of the cux-1 and cux-2 genes will be cloned, sequenced, and expressed. Whether the cux-1 gene product binds to its own promoter/regulatory region will be evaluated as a first step towards identifying its DNA-binding specificity. The functional roles of the cux-1 gene will be assessed by specific inhibition in renal epithelial cells using antisense oligonucleotides. From the effects on cell proliferation, morphology, polarity, epithelial organization, and gene expression, functional roles may be ascribed to the cux-1 gene. Whether the cux-1 gene is required for development of the kidney and genitourinary tract in mouse will be tested by disruption using homologous recombination and expression in transgenic mice. Our long-term aims will be to address the role of cux genes in a cascade of genetic regulatory mechanisms responsible for nephrogenesis. In these future studies, steps in pathways that are more proximal to cux (regulation of cux genes and gene products) and steps that are distal (target genes) will be examined in greater detail. Such studies should contribute to our fundamental understanding of genitourinary tract development. Moreover, further characterization of this novel class of homeobox-containing genes would be relevant to the overall study of eukaryotic gene regulation.