Molecular evolution of latrotoxins: neurotoxins from black widow spider venom

描述(由申请人提供)：黑寡妇蜘蛛(Latrodectus属)的毒液含有一种强有力的神经毒素--α-乳杆菌毒素，它能引起脊椎动物神经分泌细胞大量释放递质。尽管α-乳杆菌毒素能够引起人类的严重中毒，但它在脊椎动物神经递质释放的研究中也发挥着重要作用。对一种欧亚物种(L.tredecimgutatus)蛇毒的生化特征研究表明，它除了含有α-乳杆菌毒素外，还含有其他与α-乳杆菌毒素相似的神经毒素，但对无脊椎动物有选择性的作用。总而言之，α-乳杆菌毒素和这些无脊椎动物毒素被认为是共同基因家族中功能不同的成员，统称为乳杆菌毒素。虽然有证据表明，乳杆菌毒素可能存在于其他乳杆菌(31种)和相关物种中，但乳杆菌毒素的多样性几乎不为人所知。这项拟议的研究试图通过描述不同蜘蛛物种之间的多样性，来研究拉特罗毒素基因家族的分子和功能进化。比较分子分析还将与a-乳杆菌毒素的功能研究相结合，以确定导致其脊椎动物毒性的进化步骤。拟议的研究有三个具体目标：(1)利用基因组和蛋白质组方法表征三个重点物种的乳杆菌毒素的多样性；(2)通过对脊椎动物细胞进行重组α-乳杆菌毒素的功能分析，比较这三个物种的α-乳杆菌毒素的兴奋特性；以及(3)利用生物信息学分析来研究乳杆菌毒素的全球系统发育分布和分子进化，以确定α-乳杆菌毒素起源的时间和环境。实现这些目标将提供关于这个具有医学意义的基因家族的基本信息，并将建立合成危险莱特罗毒素的物种范围。相关性：α-乳杆菌毒素是研究脊椎动物神经递质释放的重要工具。相关毒素的表征将提供更广泛的分子探针，以更准确地确定神经元和神经胶质信号的机制基础，有助于神经发育和疾病的研究。这项拟议的研究还将允许生物医学研究人员合成抗蛇毒毒素，以中和不同蜘蛛物种的α-乳杆菌毒素；由于α-乳杆菌毒素中毒而住院的情况并不少见，目前的治疗方法降低了治疗某些物种叮咬的有效性。

DESCRIPTION (provided by applicant): The venom of black widow spiders (genus Latrodectus) contains a potent neurotoxin, a-latrotoxin, that evokes mass release of transmitters from vertebrate neurosecretory cells. Though capable of causing severe envenomation in humans, a-latrotoxin also plays an instrumental role in the study of vertebrate neurotransmitter release. Biochemical characterization of one Eurasian species' (L. tredecimguttatus) venom shows that in addition to a-latrotoxin, it contains other neurotoxins that are similar to a-latrotoxin in protein sequence, but which selectively affect invertebrates. Together, a-latrotoxin and these invertebrate toxins are considered functionally distinct members of a common gene family collectively known as latrotoxins. While evidence suggests that latrotoxins may occur in other Latrodectus (31 species) and related species, latrotoxin diversity is virtually unknown outside of L. tredecimguttatus. The proposed study seeks to investigate the molecular and functional evolution of the latrotoxin gene family by characterizing its diversity across spider species. Comparative molecular analyses will also be coupled to functional studies of a-latrotoxin in order to identify the evolutionary steps leading to its vertebrate toxicity. The proposed study has three specific aims: (1) to characterize the diversity of latrotoxins in three focal species using genomic and proteomic approaches; (2) to compare the excitatory properties of a-latrotoxin from these three species by conducting functional assays of recombinant a-latrotoxins on vertebrate cells; and (3) to investigate the global phylogenetic distribution and molecular evolution of latrotoxins with bioinformatic analyses to pinpoint the timing and circumstances surrounding the origin of a-latrotoxin. Achieving these aims will provide fundamental information about this medically significant gene family, and will also establish the range of species that synthesize hazardous latrotoxins. Relevance: a-latrotoxin is an essential tool for the study of vertebrate neurotransmitter release. Characterization of related toxins will provide a wider breadth of molecular probes to more precisely determine the mechanistic basis of neuronal and glial signaling, aiding studies of neurological development and disorders. The proposed research will also allow biomedical researchers to synthesize antivenoms tailored to neutralize a-latrotoxin from different spider species; hospitalization due to a-latrotoxin envenomation is not uncommon and current therapies have reduced effectiveness at treating bites from certain species.