Salivary Gland Genetic Markers

果蝇唾液腺的胚胎发育为分析参与决定和分化的基因提供了几个有利条件。在胚带延伸胚胎的外胚层中，腺体在一个特定的位置开始发育，这表明有几个已知的模式调节基因参与其中。然后在几个小时内发生分化，这意味着从承诺到分化几乎没有步骤。这项建议提出了一些方法，可以在早期阶段识别作用于唾液腺的基因，并将它们排序为唾液腺发育的途径。唾液腺标记基因将使用P-LacZ转座子来识别，该转座子在包括唾液腺在内的组织特定模式中表达β-半乳糖苷酶。致命转座子插入的纯合子胚胎将被检查是否有改变唾液腺发育的表型。对于非致命性插入，将通过不精确的转座子切除来诱导突变，以测试唾液腺的发育是否需要相邻的基因。对于一些被证明位于唾液腺发育重要基因内或附近的插入片段，插入片段两侧的区域将通过质粒拯救进行克隆，并在这些区域搜索胚胎唾液腺转录本。为了获得基因功能的线索，外显子区域将被测序，并检查与已知基因的同源性。最后，将检查所需基因的突变对几个增强子陷阱系中的β-半乳糖苷酶表达的影响。这些测试的结果将有助于确定胚胎唾液腺发育的途径。对果蝇早期发育中形成模式的基因的研究已经惊人地成功地解开了一个长期存在的谜团，即单个受精卵细胞是如何发展出复杂而有规律的模式的，这是成熟胚胎的特征。这项提议试图将这些研究向下游扩展到一个分化的器官--唾液腺的实际形成。

The embryonic development of the Drosophila salivary glands provides several advantages for analysis of the genes involved in determination and differentiation. In the ectoderm of the germ band extended embryo, the glands begin development at a defined position which suggests the involvement of several known pattern regulating genes. Differentiation then occurs within a few hours, implying that there are few steps leading from commitment to differentiation. This proposal suggests methods to identify genes acting in the salivary glands at this early stage and to order them into a pathway for salivary gland development. Salivary gland marker genes will be identified using P-lacZ transposons that express beta-galactosidase in tissue specific patterns that include the salivary glands. Embryos homozygous for lethal transposon inserts will be examined for phenotypes that alter salivary gland development. For non-lethal inserts, mutations will be induced by imprecise transposon excision to test whether an adjacent gene is required for the development of salivary glands. For some inserts that are shown to be within or near to a gene that is important for salivary gland development, regions flanking the insert will be cloned by plasmid rescue and these regions searched for embryonic salivary gland transcripts. To get clues to gene function, exonic regions will be sequenced and examined for homology with known genes. Finally, mutations in required genes will be examined for effects on beta-galactosidase expression in several enhancer trap lines. Results form these tests will help define a pathway for embryonic salivary gland development. %%% Investigations of the genes that form patterns in early drosophila development have been spectacularly successful in solving the longstanding riddle of how a single fertilized egg cell develops the intricate and regular pattern that characterizes the mature embryo. This proposal is an attempt to extend those studies downstream to the actual formation of a differentiated organ, the salivary gland.