REGULATION OF A DEVELOPMENTALLY CRITICAL INSECT PROTEIN

建议拨款的目的是利用荷尔蒙调节 变态相关蛋白转录调控机制的研究 昆虫发育过程中保幼激素对基因的调控。在……里面 粉纹夜蛾和其他许多昆虫的JH酯酶活性都很强 受自身激素JH的正向调节。这套系统对 一项关于这种或其他倍半萜激素如何作用的研究 调节蛋白质的基因表达以及荷尔蒙如何作用于 模式昆虫发育过程中基因表达的调控及其医学意义 重要的无脊椎动物。 拟议研究的具体目标和程序如下： 1)JH酯酶基因克隆(S)的分离和测序 木霉属(Trichoplesia ni.) 2)通过体内转化果蝇或体外转化果蝇 对T.ni细胞系的特定序列进行的转染分析 是激素激活JH酯酶表达所必需的。 3)通过凝胶延迟、DNAase I足迹和甲基 与JH酯酶相关的干扰特定序列或区域 荷尔蒙激活后结合蛋白质的基因。 4.启动蛋白质的纯化和鉴定 激素激活后，与JH酯酶两侧的序列结合 吉恩。 这些实验结果将是新的和原始的数据 JH刺激基因表达的分子机制。它的用途 在变态过程中高度表达的蛋白质，如 研究模型确保研究结果将加深我们的理解 属于并有能力颠覆一个基本的和对发展至关重要的 昆虫发育的调节点。的发生和重要性 JH在广泛的无脊椎动物群体中确保了结果将 对医学无脊椎动物具有广泛的意义和适用性 重要性。

The objective of the proposed grant is to use the hormonal regulation of a metamorphosis-associated protein to study the mechanisms of transcriptional regulation of genes by juvenile hormone (JH) during insect development. In Trichoplusia ni and many other insects JH esterase is strongly and positively regulated by its own hormone, JH. This system is attractive for a study on how this, and perhaps other, sesquiterpenoid hormones act to regulate gene expression of a protein and how the hormone can act to regulate gene expression during development of model insects and medically important invertebrates. The specific aims and procedures of the proposed study are: 1)isolate and sequence cDNA and genomic clone(s) for JH esterases(s) in Trichoplusia ni. 2)define through in vivo transformation of Drosophila or in vitro transfection assays on T. ni cell lines specific sequences which are necessary for hormonal activation of expression of JH esterase. 3)define through gel retardation, DNAase I footprinting and methyl interference specific sequences or regions associated with the JH esterase gene which bind proteins following hormonal activation. 4.initiate purification and characterization of proteins identified as binding, after hormonal activation, to sequences flanking the JH esterase gene. The results of these experiments will be new and original data on the molecular mechanisms of action of JH to stimulate gene expression. The use of a protein which becomes highly expressed during metamorphosis as the model for study assures that the findings will further our understanding of, and ability to disrupt, a fundamental and developmentally critical point of regulation of insect development. The occurrence and importance of JH in a wide range of invertebrate groups ensures that the results will have wide significance and applicability towards invertebrates of medical importance.