The role of mosquito microRNAs in blood digestion.

蚊子 microRNA 在血液消化中的作用。

• 批准号: 9272797
• 负责人: ALEXANDER SIMEON RAIKHEL
• 金额: $ 38.28万
• 依托单位: UNIVERSITY OF CALIFORNIA RIVERSIDE
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-06-15 至 2019-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9272797
• 关键词: 3' Untranslated Regions; Aedes; Anopheles gambiae; Antibodies; Binding Sites; Blood; Cell Line; Computational algorithm; Computer Analysis; Culicidae; Data; Digestion; Enzymes; Evaluation; Event; Female; Gene Expression; Gene Proteins; Gene Targeting; Genes; Human; Immunity; Knowledge; Link; Luciferases; MicroRNAs; Midgut; Molecular; Orthologous Gene; Peptide Hydrolases; Phenotype; Physiological; Play; Porifera; Process; Proteins; RNA; RNA Interference; Regulation; Reporter; Reproduction; Role; Specificity; Testing; Transfection; Transgenic Organisms; Untranslated RNA; Untranslated Regions; Vector-transmitted infectious disease; base; feeding; gene interaction; human disease; in vitro Assay; in vitro testing; in vivo; innovation; knock-down; loss of function; pathogen; programs; public health relevance; reproductive; spatiotemporal; vector control; vector mosquito

DESCRIPTION (provided by applicant): Female mosquitoes require a blood meal for reproduction, and during blood feeding spread many of the devastating vector-borne diseases in humans. A deeper understanding of the molecular mechanisms linked to mosquito blood meal processes, reproductive events and anti-pathogen immunity is of particular importance for devising innovative vector control strategies. Based on strong preliminary data, we suggest that microRNAs (miRNA) play important and conserved roles in blood digestive functions of vector mosquitoes. We have developed antagomirs for depletion of miRNAs and their functional characterization in Aedes aegypti and Anopheles gambiae. We have identified two miRNAs with strong effects on blood digestion. Evaluation of spatiotemporal specificity of these miRNAs will be achieved by utilizing GAL4-UAS miRNA Tough Decoy or sponge loss-of-function transgenic mosquitoes. An important step in understanding miRNA function is determining their targets. A multi-algorithm computational target-prediction approach will be used for determination of the miRNA targets. Luciferase reporter constructs carrying 3 2UTRs of the candidate miRNA targets will then be transfected into mosquito cell lines, and their expression levels will be tested after ectopic co-transfection of a miRNA that is being studied. A phenotype rescue will be achieved by RNA interference knockdown of a target gene in the transgenic loss-of-function miRNA mosquitoes. As an alternative approach, we will identify miRNAs complementary to 32UTRs of genes encoding key enzymes involved in blood digestion and study their respective roles. This application presents a comprehensive approach for functional characterization of miRNAs in two of the most important mosquito vectors for human diseases.

描述（由申请人提供）：雌性蚊子需要繁殖的血液餐，而在血液喂养期间，人类中的许多毁灭性载体疾病传播了许多灾难性的疾病。对与蚊子血粉过程，生殖事件和抗病原体免疫相关的分子机制的更深入了解对于制定创新的载体控制策略尤其重要。基于强大的初步数据，我们建议microRNA（miRNA）在载体蚊子的血液消化功能中起重要而保守的作用。我们已经开发了用于耗尽miRNA的Antagomirs及其在伊德斯埃及和冈比亚山脉的功能表征。我们已经确定了两个对血液消化有强大影响的miRNA。通过利用Gal4-UAS miRNA硬诱饵或海绵功能转基因蚊子来评估这些miRNA的时空特异性。理解miRNA功能的重要步骤是确定其目标。多算法计算目标预测方法将用于确定miRNA靶标。然后，携带3个候选miRNA靶标的荧光素酶报告基因构建体将转染到蚊子细胞系中，并在其表达水平后进行测试。 正在研究的miRNA的异位共转染。通过RNA干扰敲低靶基因在转基因功能丧失miRNA蚊子中，将实现表型救援。作为另一种方法，我们将确定与32个基因相互补充的miRNA，这些基因编码了参与血液消化的关键酶并研究其各自的作用。该应用为在人类疾病的两个最重要的蚊子媒介中提供了一种全面的miRNA功能表征的方法。