Genomic Approaches to Unravel Virulence and Resistance Determinants of Vector-transmitted Viruses in Tomato

主要研究者：S. P. Dinesh-Kumar（加州大学戴维斯分校）CoPI：Robert Gilbertson，Diane Ullman和Ilias Tagkopoulos（加州大学戴维斯分校）番茄是世界上第二大蔬菜作物。美国和世界范围内的番茄生产受到两种毁灭性昆虫传播病毒的威胁-番茄斑萎病毒（TSWV）和番茄黄化曲叶病毒（TYLCV）。这些病毒的管理很困难，往往严重依赖使用杀虫剂来控制病媒。室内控制往往无效，对人类健康和环境产生不利影响，并可能导致选择对杀虫剂有抗药性的病媒。在番茄中，TYLCV的Ty 1基因座和TSWV的Sw 5基因座是主要的遗传抗性来源，并且两者都已经渗入到商业番茄品种中。然而，很少有人知道Ty 1和Sw 5介导的抗性的机制。该项目将提供新的见解的性质和相互作用的机制，植物抗性和易感性感染的昆虫传播的病毒。这些研究将提供有关RNA（TSWV）与DNA（TYLCV）病毒在感染和/或宿主防御过程中是否靶向相似或不同途径的见解。然后，该项目将使用这些研究的结果来优化和验证一种称为miRNA诱导的基因沉默（MIGS）的新方法，该方法可用于评估番茄中的基因功能。 如果成功，该项目的成果可能会导致针对这些病毒的新一代抗病毒策略。该项目还将每年为来自萨克拉门托城市学院的本科生和戴维斯高中的高中生提供为期10周的培训项目。由野生番茄近缘种渗入栽培番茄的Sw 5和Ty 1基因座介导的遗传抗性分别为TSWV和TYLCV提供了最佳的保护来源。该项目的研究人员假设Sw 5和Ty 1介导的TSWV和TYLCV识别触发了涉及常见和特异性免疫信号网络的早期和可辨别的转录变化。此外，这些病毒对易感番茄基因型的感染诱导转录变化，促进病毒毒力。为了深入了解转录变化，研究人员将使用RNAseq方法来鉴定番茄TSWV和TYLCV抗性和易感反应期间差异表达的转录本。计算方法将用于分析RNASeq数据，以鉴定在TSWV和TYLCV抗性和易感相互作用期间差异调节的关键共同和特异性基因。该项目将优化和验证MIGS方法，通过靶向已知的候选基因，在转基因番茄中进行基因功能研究。优化后的MIGS方法将用于评价所选差异表达基因在番茄TYLCV和TSWV抗感互作中的功能。该项目的结果可能会为设计新一代策略提供见解，不仅控制TYLCV和TSWV，还控制其他重要的植物病原体。所有数据和生物资源将向公众开放。所有RNAseq数据将通过GEO保存和提供。所有MIGS载体和候选靶基因构建体将根据要求并通过Addgene（https：//www.addgene.org/）和ABRC（http：//www.biosci.ohio-state.edu/plantbio/Facilities/abrc/abrchome.htm）获得。 转基因番茄系的种子可通过加州大学戴维斯分校的番茄遗传资源中心（http：//tgrc.ucdavis.edu/）获得。

PI: S.P. Dinesh-Kumar (University of California-Davis)CoPIs: Robert Gilbertson, Diane Ullman, and Ilias Tagkopoulos (University of California-Davis)Tomato is the second most important vegetable crop in the world. Tomato production in the US and worldwide is threatened by two devastating insect-transmitted viruses - tomato spotted wilt virus (TSWV) and tomato yellow leaf curl virus (TYLCV). Management of these viruses is difficult and often relies heavily on the use of insecticides to control the vectors. Insecticidal control is often ineffective, has negative human health and environmental effects, and can lead to selection of insecticide-resistant forms of the vectors. In tomato, the Ty1 locus for TYLCV and the Sw5 locus for TSWV are major genetic resistance sources, and both have been introgressed into commercial tomato varieties. However, very little is known about the mechanisms involved in Ty1- and Sw5-mediated resistance. This project will provide new insights into the nature of and interplay between mechanisms of plant resistance and susceptibility to infection by insect-transmitted viruses. These studies will provide insights into whether similar or different pathways are targeted by RNA (TSWV) versus DNA (TYLCV) viruses during infection and/or host defense. The project will then use the results from these studies to optimize and validate a new approach called miRNA-induced gene silencing (MIGS) that can be used to assess gene function in tomato. If successful, the outcomes of this project will likely lead to a new generation of antiviral strategies against these viruses. The project will also provide a 10-week training program every year for undergraduate students from Sacramento City College and high school students from Davis Senior High School.The genetic resistance mediated by Sw5 and Ty1 loci introgressed from wild tomato relatives into cultivated tomato provides the best source of protection against TSWV and TYLCV, respectively. The investigators of this project hypothesize that Sw5- and Ty1-mediated recognition of TSWV and TYLCV trigger early and discernable transcriptional changes that involve common and specific immune signaling networks. In addition, infection of these viruses on susceptible tomato genotypes induces transcriptional changes that promote virus virulence. To gain insights into the transcriptional changes, the investigators will use RNAseq approach to identify differentially expressed transcripts during TSWV and TYLCV resistance and susceptible responses in tomato. Computational methods will be used to analyze RNASeq data to identify key common and specific genes that are differentially regulated during TSWV and TYLCV resistance and susceptible interaction. The project will optimize and validate the MIGS approach for gene function studies in transgenic tomato by targeting known candidate genes. The optimized MIGS approach will be used to evaluate function of selected differentially expressed genes during TYLCV and TSWV resistance and susceptible interactions in tomato. Results from the project will likely provide insights into designing new generation of strategies to control not only TYLCV and TSWV but also other important plant pathogens. All data and biological resources will be accessible to the public. All RNAseq data will be deposited and available through GEO. All MIGS vector and candidate target gene constructs will be available upon request and through Addgene (https://www.addgene.org/) and ABRC (http://www.biosci.ohio-state.edu/~plantbio/Facilities/abrc/abrchome.htm). Seeds from transgenic tomato lines will be available through the Tomato Genetics Resource Center at UC Davis (http://tgrc.ucdavis.edu/).