Sex-Specific Gene Activity in the Mosquito Larva

描述：(申请人提供)：全球范围内的毒力死灰复燃 蚊媒疾病的抗药性增加了对 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 幼蚊体内的活动，以开发新的灭蚊方法 通过转基因操作的策略。大多数性别特异基因的研究 活动主要集中在成虫体内的表达。在果蝇身上， 成虫基因激活的分子基础--黑腹果蝇 雌性和雄性的压抑是众所周知的转录 调节器，双性(DSX)。 我们对六聚体基因的研究使我们能够开发一种模型系统来 研究蚊子幼虫的性别特异性基因活性。在全变态中 昆虫、六聚体储存蛋白(六聚体储存蛋白)是由 晚期幼虫的脂肪体。最近我们展示了其中一个 六聚体，AatHex-1.2，由蚊子四龄幼虫产生， 黑斑伊蚊，是真正的雌性特有的。在该基因5‘侧翼区域 AatHex-1.2基因，我们确定了可能的调控元件(包括dsx 结合位点)，我们认为它控制着它的组织、阶段和性别特异性 活动。这一地区的700个基点足以将海平面上升10倍 报告基因在中华绒螯蟹雌幼虫脂肪体中的独特活性 异源双翅目，黑腹蝇D. 为了阐明性二态AatHex-1.2活性的分子基础，我们 建议鉴定必需的调控DNA序列和转录 因子结合位点。调控序列将通过基因转移在 蚊子，埃及伊蚊和埃及伊蚊。以及在果蝇身上。 转录因子将用凝胶迁移率来表征 移位分析和DNA酶脚印。实现这些目标不会 只会更好地理解性行为和脂肪的分子基础 昆虫幼虫体内的身体特异性基因活性，但也将定义最低限度的 推动改进昆虫不育技术(SIT)以控制昆虫和 开发将转基因引入自然种群的新方法。

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.