A Transcriptomic View of the Proteome Variability of Newborn and Adult Bothrops jararaca Snake Venoms

BackgroundSnake bite is a neglected public health problem in communities in rural areas of several countries. Bothrops jararaca causes many snake bites in Brazil and previous studies have demonstrated that the pharmacological activities displayed by its venom undergo a significant ontogenetic shift. Similarly, the venom proteome of B. jararaca exhibits a considerable variation upon neonate to adult transition, which is associated with changes in diet from ectothermic prey in early life to endothermic prey in adulthood. Moreover, it has been shown that the Brazilian commercial antibothropic antivenom, which is produced by immunization with adult venom, is less effective in neutralizing newborn venom effects. On the other hand, venom gland transcripts of newborn snakes are poorly known since all transcriptomic studies have been carried out using mRNA from adult specimens. Methods/Principal FindingsHere we analyzed venom gland cDNA libraries of newborn and adult B. jararaca in order to evaluate whether the variability demonstrated for its venom proteome and pharmacological activities was correlated with differences in the structure of toxin transcripts. The analysis revealed that the variability in B. jararaca venom gland transcriptomes is quantitative, as illustrated by the very high content of metalloproteinases in the newborn venom glands. Moreover, the variability is also characterized by the structural diversity of SVMP precursors found in newborn and adult transcriptomes. In the adult transcriptome, however, the content of metalloproteinase precursors considerably diminishes and the number of transcripts of serine proteinases, C-type lectins and bradykinin-potentiating peptides increase. Moreover, the comparison of the content of ESTs encoding toxins in adult male and female venom glands showed some gender-related differences. Conclusions/SignificanceWe demonstrate a substantial shift in toxin transcripts upon snake development and a marked decrease in the metalloproteinase P-III/P-I class ratio which are correlated with changes in the venom proteome complexity and pharmacological activities.研究背景：蛇咬伤是多个国家农村社区中被忽视的公共卫生问题。三色矛头蝮（Bothrops jararaca）在巴西造成了大量蛇咬伤病例，既往研究表明，其蛇毒所展现出的药理活性会发生显著的个体发育转变。类似地，该物种的蛇毒蛋白质组（venom proteome）在从幼体到成体的过渡过程中也存在明显差异，这与其食性变化相关：幼体阶段以变温猎物为食，成体阶段则转为捕食恒温猎物。此外，已有研究显示，采用成体蛇毒免疫制备的巴西商用抗矛头蝮蛇毒血清（antibothropic antivenom），其中和幼体蛇毒毒性效应的效果较差。另一方面，由于此前所有转录组学研究均使用成体标本的mRNA，幼体蛇毒的毒腺转录本相关研究仍极为匮乏。 方法与主要研究结果：本研究对幼体与成体三色矛头蝮的毒腺cDNA文库进行了分析，旨在探究其蛇毒蛋白质组与药理活性所呈现的差异性，是否与毒素转录本的结构差异存在关联。分析结果表明，三色矛头蝮毒腺转录组的差异性表现为数量层面的差异：幼体毒腺中金属蛋白酶（metalloproteinase）的含量极高。此外，该差异性还体现在幼体与成体转录组中蛇毒金属蛋白酶（Snake Venom Metalloproteinase, SVMP）前体的结构多样性上。然而在成体转录组中，金属蛋白酶前体的含量显著降低，而丝氨酸蛋白酶（serine proteinase）、C型凝集素（C-type lectin）以及缓激肽增强肽（bradykinin-potentiating peptide）的转录本数量则有所上升。此外，对成体雌雄个体毒腺中编码毒素的表达序列标签（Expressed Sequence Tag, EST）含量进行比较后，发现了部分与性别相关的差异。 结论与意义：本研究证实，随着蛇类发育，其毒素转录本会发生显著变化，且金属蛋白酶P-III/P-I类的比例显著降低，这一变化与蛇毒蛋白质组的复杂性及药理活性的改变密切相关。