Mechanism of a Male Killing Toxin in a Drosophila endosymbiont

果蝇内共生体中雄性致死毒素的机制

基本信息

批准号：
10609906
负责人：
Irene Newton
金额：
$ 19.81万
依托单位：
TRUSTEES OF INDIANA UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-04-15 至 2024-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10609906
关键词：
Affect Alleles Amino Acids Animals Ankyrin Repeat Ankyrins Apoptosis Arthropods Bacterial Proteins Bacterial Toxins Binding Binding Proteins Biological Biological Assay Biological Models Biology Candidate Disease Gene Cells Cellular biology Chromatin Structure Co-Immunoprecipitations Complex Coupled Data Development Dosage Compensation (Genetics)Drosophila genus Drosophila melanogaster Educational process of instructing Embryo Embryonic Development Environment Eukaryotic Cell Event Exhibits Female Genes Genetic Genetic Crosses Genetic Polymorphism Genetic Screening Human Hybrids Immunity Infection Length Libraries Link Mass Spectrum Analysis Mediating Molecular Molecular Biology Mutation N-terminal Natural Resistance Open Reading Frames Ovary Parasites Pathway interactions Phenotype Planet Earth Population Post-Translational Protein Processing Proteins Resistance Scaffolding Protein Shapes Single Nucleotide Polymorphism Spiroplasma Stretching Structure Testing Tissues Toxic effect Toxin Variant Virulence Work Yeasts arms race endosymbiont experimental study fly genetic variant genome wide association study in vivo knock-down male mimicry novel offspring ovarian neoplasm pathogen protein function protein protein interaction sex determination symbiont tool transmission process

项目摘要

PROJECT SUMMARY/ABSTRACT The co-evolution of bacterial protein toxins and their targets has led to the discovery of many novel protein modifications and the development of applications in molecular biology. Arthropods, the most diverse phylum on Earth, are hosts to many bacterial symbionts that secrete diverse toxins of unknown function during infection. The maternally transmitted, intracellular parasite Spiroplasma poulsonii encodes one such toxin (Spiroplasma androcidin or Spaid) that causes male killing in its natural host Drosophila melanogaster. This proposed project aims to understand how the bacterial toxin Spaid recognizes the host cellular environment to specifically kill males. To that end, we use the Spiroplasma-Drosophila model system and a combination of genetics and molecular biology to identify specific host proteins targeted by Spaid and how the toxin binds these proteins. Previous work has suggested that Spiroplasma, via Spaid, detects the presence of the male specific lethal complex in flies. This complex is responsible for dosage compensation in male flies, which lack a second copy of the X. It is formed during early development in males and is part of the Drosophila sex determination pathway. Given this result, we hypothesized that natural resistance to Spaid would exist in Drosophila populations, that the target of Spaid is part of the dosage compensation complex (dcc), and that Spaid directly binds to dcc components, altering their activity. Our strong preliminary data suggest that we are correct, as we find natural variants of Drosophila melanogaster (within the DGRP), which produce males in the presence of Spaid and/or Spiroplasma. We also find that tissue specific knockdown of dcc components partially suppresses Spaid toxicity in male tissue. Towards this hypothesis, and guided by strong preliminary data, we propose to pursue two Specific Aims to identify host proteins that interact directly with the Spaid toxin and characterize how variation within host proteins and Spaid affect male killing at the molecular and cellular level. We will (1) Identify Spaid targets using a yeast 2-hybrid screen and co-immunoprecipitation, (2) identify suppressors of Spaid toxicity using natural variation in Drosophila and a genetic screen. Results from these experiments will illustrate how a bacterial toxin manipulates the host cellular environment.

项目概要/摘要细菌蛋白毒素及其靶标的共同进化导致许多新蛋白的发现分子生物学的修改和应用的开发。节肢动物门，最多样化的门在地球上，是许多细菌共生体的宿主，这些细菌共生体在感染。母体传播的细胞内寄生虫 Spiroplasma poulsonii 编码一种这样的毒素（螺原体雄激素或 Spaid）会导致其自然宿主黑腹果蝇的雄性死亡。这拟议的项目旨在了解细菌毒素 Spaid 如何识别宿主细胞环境专门杀死雄性。为此，我们使用螺原体-果蝇模型系统并结合遗传学和分子生物学，以确定 Spaid 靶向的特定宿主蛋白以及毒素如何结合这些蛋白质。先前的研究表明螺原体通过 Spaid 检测男性特异性致死细胞的存在苍蝇的复杂性。该复合物负责雄性果蝇的剂量补偿，而雄性果蝇缺乏第二个拷贝 X 的。它是在雄性早期发育过程中形成的，是果蝇性别决定的一部分途径。鉴于这一结果，我们假设果蝇中存在对 Spaid 的天然抗性人群，Spaid 的目标是剂量补偿复合体 (dcc) 的一部分，并且 Spaid 直接与 dcc 成分结合，改变其活性。我们强有力的初步数据表明我们是正确的，因为我们发现果蝇（Drosophila melanogaster）的自然变种（在 DGRP 内），这种变种在存在以下物质的情况下产生雄性斯佩德和/或螺原体。我们还发现 dcc 成分的组织特异性敲低部分抑制对雄性组织产生毒性。针对这一假设，并以强有力的初步数据为指导，我们建议追求两个具体目标识别与 Spaid 毒素直接相互作用的宿主蛋白并表征宿主内的变异蛋白质和 Spaid 在分子和细胞水平上影响雄性杀戮。我们将 (1) 使用以下方式识别 Spaid 目标酵母 2 杂交筛选和免疫共沉淀，(2) 使用天然产物鉴定 Spaid 毒性的抑制剂果蝇变异和遗传筛选。这些实验的结果将说明细菌毒素如何操纵宿主细胞环境。