Molecular regulation of diapause in Culex pipiens

淡色库蚊滞育的分子调控

• 批准号: 7089099
• 负责人: DAVID L DENLINGER
• 金额: $ 21.9万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-07-01 至 2010-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7089099
• 关键词: Culex; RNA interference; West Nile virus; complementary DNA; diapause; genetic regulation; in situ hybridization; northern blottings; nucleic acid sequence; photostimulus; polymerase chain reaction; subtraction hybridization

DESCRIPTION (provided by applicant): This proposal focuses on the molecular basis for diapause (dormancy) in the northern house mosquito, Culex pipiens. In the early autumn adult females of this species seek protected sites and remain there throughout the winter months in a reproductively inactive state know as diapause. Although many aspects of the physiology and environmental control of diapause are well known, there is little information concerning the molecular basis for this diapause or the diapause of any other vector species. Obtaining such information is critical not only for understanding regulation of seasonal cycles of mosquitoes, but also for understanding the seasonality of pathogens the mosquito transmit. In the case of Cx. pipiens the overwintering females appear to harbor West Nile virus, and thus serve as the mechanism for reintroducing the virus into bird populations the following spring. Our initial studies using suppressive subtractive hybridization (SSH) have identified four categories of genes that are upregulated in early diapause. These include genes involved in the diapause-associated switch from blood feeding to sugar feeding, the arrest of development, cold-hardening, and responding to juvenile hormone. Similar SSH experiments will examine mid and late stages of diapause. Clones isolated with this technique will be further characterized by full-length sequencing, by evaluating expression patterns throughout diapause using northern blotting and real-time PCR, by examining tissue-specific expression patterns, and by evaluating gene function using RNA interference. We also plan to use SSH to search for genes associated with the programming phase of diapause (early pupal stage). Such genes are likely to be involved in the decision to either enter or avert diapause. At the conclusion of this study we expect to have identified major gene pathways involved in the regulation of diapause in Cx. pipiens, thus allowing us to begin the construction of a regulatory hierarchy for diapause in this species. We anticipate that these results will suggest vulnerable targets for disrupting diapause and may provide clues for how viral replication is halted during the winter months.

描述（由申请人提供）：本提案重点研究北方家蚊（淡色库蚊）滞育（休眠）的分子基础。在初秋，该物种的成年雌性会寻找受保护的地点，并在整个冬季保持在那里，处于一种繁殖不活跃的状态，即滞育。尽管滞育的生理学和环境控制的许多方面是众所周知的，但关于这种滞育或任何其他媒介物种滞育的分子基础的信息很少。获取此类信息不仅对于了解蚊子季节性周期的调节至关重要，而且对于了解蚊子传播的病原体的季节性也至关重要。以 Cx 为例。 pipiens 越冬雌性似乎携带西尼罗河病毒，因此成为来年春天将病毒重新引入鸟类种群的机制。 我们使用抑制消减杂交（SSH）进行的初步研究已经确定了四类在滞育早期上调的基因。这些基因包括与滞育相关的从血液喂养到糖喂养的转变、发育停滞、冷硬化和对保幼激素的反应有关的基因。类似的 SSH 实验将检查滞育的中期和晚期。使用该技术分离的克隆将通过全长测序、使用 Northern 印迹和实时 PCR 评估整个滞育期间的表达模式、检查组织特异性表达模式以及使用 RNA 干扰评估基因功能来进一步表征。我们还计划使用 SSH 来搜索与滞育编程阶段（蛹早期）相关的基因。这些基因可能参与进入或避免滞育的决定。 在这项研究的结论中，我们希望能够确定参与 Cx 滞育调节的主要基因途径。 pipiens，从而使我们能够开始构建该物种滞育的监管等级。我们预计这些结果将提出破坏滞育的脆弱目标，并可能为如何在冬季停止病毒复制提供线索。