Sex-Specific Gene Activity in the Mosquito Larva

描述：（由申请人提供）：全球范围内毒力 蚊子传播疾病的抗药性增加了 新的蚊子控制策略。拟议研究的长期目标 是为了阐明女性特有基因的分子机制， 活性，以开发新的蚊子控制 通过转基因操作的策略。大多数性别特异性基因的研究 活动集中在成年昆虫中的表达。在果蝇中， 果蝇成年期基因激活的分子基础 女性和男性的抑制是很好理解的转录 调节器，Doubleex（DSX）。 我们对六聚体基因的研究使我们能够开发一个模型系统， 研究蚊子幼虫的性别特异性基因活性。在全变态的 在昆虫中，六聚体（六聚体储存蛋白）大量合成， 后期幼虫的脂肪体。最近我们发现， 六聚体，AatHex-1.2，由蚊子的四龄幼虫产生， 黑须伊蚊是雌性特有的在5 '侧翼区， AatHex-1.2基因，我们确定了推定的调控元件（包括DSX 结合位点），我们认为这决定了其组织特异性，阶段特异性和性别特异性 活动该区域的700 bp足以指导10倍高的 报告基因活性独特的雌性幼虫的脂肪体的 异源双翅目D.黑腹菌 为了阐明性二态AatHex-1.2活性的分子基础，我们 建议确定必要的调控DNA序列和转录 因子结合位点。调控序列将通过基因转移绘制， 蚊子，埃及伊蚊和Ae. atropalpus和果蝇。 转录因子将使用电泳迁移率来表征 移位测定和DNA酶足迹法。这些目标的实现不会 只会让我们更好地理解性和脂肪的分子基础 身体特异性基因活性，但也将定义一个最小的 启动子以改进用于昆虫控制的昆虫不育技术（SIT），以及 开发新的方法将转基因导入自然种群。

DESCRIPTION: (provided by the applicant): The worldwide resurgence of virulent and resistant forms of mosquito-borne diseases has increased the urgency for novel mosquito control strategies. The long-term goal of the proposed research is to elucidate the molecular mechanisms underlying female-specific gene activity in the immature mosquito in order to develop novel mosquito control strategies by transgenic manipulation. Most studies of sex-specific gene activity have focused on expression in adult insects. In the fruit fly, Drosophila melanogaster, the molecular basis for gene activation in adult females and repression in males is well understood for the transcriptional regulator, Doublesex (DSX). Our studies of hexamerin genes have allowed us to develop a model system to study sex-specific gene activity in the larval mosquito. In holometabolous insects, hexamerins (hexameric storage proteins) are abundantly synthesized by the fat body of late-stage larvae. Recently we showed that one of the hexamerins, AatHex-1 .2, produced by fourth-instar larvae of the mosquito, Aedes atropalpus, is truly female-specific. In the 5'-flanking region of the AatHex-1.2 gene, we identified putative regulatory elements (including DSX binding sites), which we believe govern its tissue-, stage- and sex-specific activity. 700 bp of this region are sufficient to direct 10-fold higher reporter gene activity uniquely in the fat body of female larvae of the heterologous dipteran, D. melanogaster. To elucidate the molecular basis for sexually dimorphic AatHex-1.2 activity, we propose to identify the requisite regulatory DNA sequences and transcription factor binding sites. Regulatory sequences will be mapped by gene transfer in the mosquitoes, Aedes aegypti and Ae. atropalpus, and in Drosophila. Transcription factors will be characterized using electrophoretic mobility shift assays and DNase foot printing. Accomplishment of these aims will not only provide a better understanding of the molecular basis for sex- and fat body-specific gene activity in an insect larva but will also define a minimal promoter to improve the sterile insect technique (SIT) for insect control and to develop novel means for driving transgenes into natural populations.