Exploring evolutional origins of viper venom haemorrhagic factors

蛇毒主要分为两类：神经毒性毒液和出血性毒液。本研究解决了后一种毒液的问题，毒蛇出血性毒液蛋白的进化起源是什么，以及毒蛇在进化过程中如何获得它们。毒蛇出血因子是由两个有毒多肽结构域组成的前体可溶性因子，一个是出血活性结构域，另一个是抗凝血活性结构域。前体在结构上类似于跨膜蛋白，通常被称为ADAMs（一种分解素和金属蛋白酶）。我们假设毒蛇的毒液模仿了祖先脊椎动物的亚当斯，在血管生成/血管生成和/或血细胞行为中具有功能。我们之前通过对斑马鱼胚胎血管和红母细胞的实时成像发现，ADAM8参与了斑马鱼胚胎血液循环的开始（Iida等人，Curr Biol）。, 2010)。通过这种成像技术，我们研究了毒蛇毒液出血毒液是如何影响血管生成或血管完整性的，结果发现其中一些毒液抑制了斑马鱼胚胎的血管生成。然后我们研究了ADAM8是否在血管形成中发挥了一些作用。向卵内注射抗ADAM8的反义morpholino寡核苷酸，对卵段间血管的伸长有抑制作用。这些胚胎的实时成像和电子显微照片显示，这些胚胎的血细胞附着在血管上异常，血管中的基因表达明显改变。蛇毒蛋白的出血活性可能与ADAM8在血管发生中的调节作用有关。

Snake venoms are classified mainly into two categories : neurotoxic and hemorrhagic venoms. This study addressed questions on the latter venom, what are the evolutional origins of viper hemorrhagic venom proteins or how vipers acquired them during evolution. Viper haemorrhagic factors are produced as precursor soluble factors composed of two toxic polypeptide domains, a domain of haemorrhagic activity and that of anti-coagulation activity. The precursors are similar in structure to transmembrane proteins generically named as ADAMs(a disintegrin and metalloproteases). We hypothesized that viper venoms mimicked ADAMs of ancestral vertebrates that carried functions in vasculogenesis/angiogenesis and/or behaviors of blood cells. We previously showed that ADAM8 is involved in the onset of blood circulation in zebrafish embryos by live imaging of their blood vessels and erythroblasts(Iida et al., Curr Biol., 2010.). By using this imaging technique, we examined how viper venom haemorrhagic venoms affects vasculogenesis or integrity of blood vessels, As a result, we found that some of them inhibited vasculogenesis of zebrafish embryos. Then we examine whether ADAM8 ply some roles on blood vessel formation. Injection of antisense morpholino oligonucleotides against ADAM8 into eggs caused inhibitory effects on elongation of intersegmental vessels. Live imaging and electron micrographs of those embryos revealed that blood cells attached to blood vessels abnormally and gene expression in blood vessels was significantly altered in those embryos. Haemorrhagic activities of viper venom proteins could be related to regulatory roles of ADAM8 in vasculogenesis.