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.

项目摘要/摘要细菌蛋白毒素及其靶标的共同进化导致发现了许多新型蛋白修改和分子生物学应用的发展。节肢动物，最多样化的门在地球上，是许多细菌共生体的宿主感染。孕妇传播的细胞内寄生虫螺旋体Poulsonii编码一种毒素（螺旋藻雄性固醇或SPAID）导致雄性在其天然宿主果蝇中杀死男性。这拟议的项目旨在了解细菌毒素如何认识到宿主细胞环境特别杀死男性。为此，我们使用了螺旋体 - 嗜醇模型系统和遗传学和分子生物学，以鉴定SPAID靶向的特定宿主蛋白以及毒素如何结合这些蛋白质。先前的工作表明，螺旋体通过SPAID检测到雄性特异性致命的存在复合体。该综合大楼是雄性苍蝇的剂量补偿，缺少第二份 X的X。它是在男性的早期发展过程中形成的，是果蝇性确定的一部分路径。鉴于此结果，我们假设果蝇中存在自然抗Spaid 种群，SPAID的目标是剂量补偿综合体（DCC）的一部分，并直接spaid 结合DCC组件，改变其活性。我们强大的初步数据表明我们是正确的，因为我们查找果蝇的自然变体（在DGRP内），在存在的情况下会产生雄性 Spaid和/或螺旋体。我们还发现，DCC组件的组织特异性敲低部分抑制在雄性组织中的毒性。提出这一假设，并在强大的初步数据的指导下，我们建议追求两个具体的目标识别直接与SPAID毒素相互作用的宿主蛋白，并表征宿主内部的变化蛋白质和SPAID会影响男性在分子和细胞水平上的杀戮。我们将（1）使用酵母2-杂交筛选和共免疫沉淀，（2）使用天然识别spaid毒性的抑制剂果蝇和遗传筛查的变化。这些实验的结果将说明细菌毒素如何操纵宿主细胞环境。