Discovering Biomedically Significant Spider Venom Toxins with Genomics

描述（由申请人提供）：动物毒液是复杂的生化鸡尾酒，能够产生多种极其有效的生理效应。毒液也被高度珍视为药物的新来源，并且对于救生抗蛇毒血清的开发至关重要。尽管其科学和临床意义，许多生物医学上重要的毒液的分子组成和功能活动仍然知之甚少。这项提案的长期目标是了解黑寡妇蜘蛛毒液的分子复杂性，功能和进化。黑寡妇蜘蛛拥有一种高度神经毒性的毒液，会给人类带来剧烈的疼痛。虽然黑寡妇毒液在脊椎动物神经传递的研究中发挥了关键作用，但这种毒液的分子组成及其功能和生物医学应用尚未使用现代分子技术进行研究。拟议的工作将通过三个具体目标解决这一重大知识差距。首先，将使用下一代测序方法组装一个全面的黑寡妇毒腺转录组，以表征毒腺组织中表达的基因。第二，从转录组中鉴定参与毒液功能的转录物组。这将涉及肽指纹图谱的蛋白质组技术，使用质谱法和转录组数据库的生物信息学筛选来确定毒液样本中的蛋白质。标签分析的下一代测序应用也将用于鉴定表现出毒液特异性表达模式的基因。第三个具体目标将通过毒液基因变异的跨物种比较来研究黑寡妇毒液的功能进化，量化选择模式，以及基因表达的比较分析。对功能重要的毒素蛋白的系统发育分析也将揭示导致黑寡妇蜘蛛毒液极端毒性的分子机制。完成后，这项工作将为黑寡妇毒液的分子多样性、进化和生物医学用途提供无与伦比的视角。该项目的结果还将产生重要的公共基因组资源，并为旨在了解毒素功能进化的分子机制的未来研究提供基础。 公共卫生相关性：该计划的主要目标是确定黑寡妇蜘蛛毒液的分子组成，功能活性和进化。该项目与公共卫生有着重要的相关性，因为黑寡妇毒液对人类极其危险，但它也是研究脊椎动物神经生理学的重要工具。这项拟议中的工作将为黑寡妇蜘蛛极端毒性的分子基础提供前所未有的理解，并将有助于开发新的药物和改进的抗蛇毒血清。

DESCRIPTION (provided by applicant): Animal venoms are complex biochemical cocktails capable of diverse and extremely potent physiological effects. Venoms are also highly prized as novel sources of pharmaceuticals and are essential for the development of lifesaving antivenoms. Despite their scientific and clinical significance, the molecular composition and functional activities of many biomedically important venoms remains poorly known. The long-term objective of this proposal is to understand the molecular complexity, function and evolution of black widow spider venom. Black widow spiders possess a highly neurotoxic venom that causes severe pain in humans. While black widow venom has played a critical role in the study of vertebrate neurotransmission, the molecular composition of this venom and its functional and biomedical applications have not been investigated using modern molecular techniques. The proposed work will address this significant knowledge gap through three specific aims. First, a comprehensive black widow venom gland transcriptome will be assembled using next-generation sequencing methods to characterize the genes expressed in venom gland tissue. Second, the set of transcripts involved in venom function will be identified from the transcriptome. This will involve the proteomic technique of peptide fingerprinting to determine the proteins in venom samples using mass spectrometry and bioinformatic screens of a transcriptome database. The next-generation sequencing application of Tag-Profiling will also be used to identify genes that exhibit venom-specific patterns of expression. The third specific aim will examine the functional evolution of black widow venom through cross-species comparisons of venom gene variation that quantify patterns of selection, as well as comparative analyses of gene expression. Phylogenetic analyses of functionally important toxin proteins will also reveal the molecular mechanisms that have contributed to the extreme toxicity of black widow spider venom. When completed, the proposed work will provide an unparalleled view of the molecular diversity, evolution and biomedical utility of black widow venom. The results of this project will also generate significant public genomic resources and provide a foundation for future studies aimed at understanding the molecular mechanisms of toxin functional evolution. PUBLIC HEALTH RELEVANCE: The major goal of this proposal is to determine the molecular composition, functional activities and evolution of black widow spider venom. The project has significant relevance for public health because black widow venom is extremely hazardous to humans, but it is also an essential tool in the study of vertebrate neurophysiology. The proposed work will provide an unprecedented understanding of the molecular basis for the extreme toxicity of black widow spiders and will aid in the development of novel pharmaceuticals and improved antivenoms.