Evolution of Placental Calcium Transport in Reptiles

陆生脊椎动物表现出两种基本的繁殖模式；胎生，即活的幼崽的出生，以及卵生，即在雌性体外进行胚胎发育的产卵。除了单孔目动物外，所有哺乳动物都是胎生的。海龟、鳄鱼和鸟类都是卵生动物。相比之下，有鳞爬行动物（蛇和蜥蜴）的生殖模式多样化，大约20%是胎生的，至少有三种是双峰繁殖的。鳞片的胎生已经独立进化了100多次，胎生的进化与胚胎营养模式的转变有关，从加强卵子的能量储存到胎盘的营养交换。所有脊椎动物胚胎的成功发育都需要钙。卵生物种和胎生物种胚胎钙供应的来源和模式不同。卵生物种的胚胎从蛋黄和蛋壳中获取钙，这些来源在卵黄形成和产卵前的脱壳过程中就已经存在了。胎生胚胎从卵黄中获得部分钙，但在发育后期也依赖于胎盘移植。钙转运到胚胎的时间、来源和机制是否影响了导致胎生的进化创新的潜力？该项目将探索在陆地脊椎动物中生殖模式多样性最高的一个谱系（鳞片爬行动物）中，胚胎钙供应模式与生殖模式（产卵或生育）之间的关系。本研究将比较卵生和胎生鳞片胚胎钙供应的模式和机制，为胎盘钙运输的进化建立模型。本项目将比较子宫分泌钙的时间和机制以及胚胎动员钙的时间和机制：1)两种新近进化为胎生的物种，2)一种胎盘钙运输较少的胎生物种，3)一种严重依赖胎盘钙的胎生物种。密切相关的产卵物种将被研究以进行比较。原子吸收光谱将用于确定胚胎钙摄取的模式。通过使用免疫印迹和免疫组织化学技术确定钙转运蛋白calbinin - d28k、Ca2+- atp酶的发育表达，研究子宫和胚胎膜钙转运的时间和机制。本研究通过研究一种高度保守的向胚胎输送营养物质的机制，将有助于更好地了解陆生脊椎动物的胚胎发育。该项目将通过在博士学位授予部门的研究环境中为主要来自本科部门的本科生提供机会，将研究和教育结合起来。通过与校园内罗纳德·e·麦克奈尔学者项目的管理人员合作，将积极招收来自社会弱势群体的本科生。这些努力将针对任何机构的任何符合条件的本科生。这些学生将接受培训，目的是增加他们未来在研究生课程中取得成功的潜力。

Terrestrial vertebrates exhibit two basic modes of reproduction; viviparity, or the birth of live young, and oviparity, or the laying of eggs in which embryonic development occurs outside the body of the female. All mammals, with the exception of monotremes, are viviparous. Turtles, crocodilians and birds are exclusively oviparous. In contrast, squamate reptiles (snakes and lizards) show a diversity of reproductive patterns with approximately 20% being viviparous and at least three species being reproductively bimodal. Viviparity in squamates has evolved independently more than 100 times and the evolution of viviparity is associated with a transition in the pattern of embryonic nutrition from fortification of energy stores in the egg to placental nutrient exchange. All vertebrate embryos require calcium for successful development. The source and pattern of calcium provision to embryos of oviparous species differs from that of viviparous species. Embryos of oviparous species derive calcium from yolk and eggshell, sources laid down during vitellogenesis and shelling prior to egglaying. Viviparous embryos derive some calcium from yolk but depend also on placental transfer late in development. Does the timing, source and mechanism of calcium transport to embryos influence the potential for evolutionary innovations leading to viviparity? This project will explore the relationship between the pattern of embryonic calcium provision and reproductive mode (egg-laying or live-bearing) in a lineage (squamate reptiles) that exhibits the highest diversity of reproductive patterns among terrestrial vertebrates. The pattern and mechanism of calcium provision to embryos of selected oviparous and viviparous squamates will be compared to develop a model for the evolution of placental calcium transport. The project will compare the timing and mechanism of calcium secretion by the uterus and the timing and mechanism of calcium mobilization by embryos of: 1) two species that have recently evolved viviparity, 2) a viviparous species with little placental calcium transport, and 3) a viviparous species that is heavily dependent on placental calcium. Egg-laying species that are closely related will be studied for comparison. Atomic absorption spectroscopy will be used to define the pattern of calcium uptake by embryos. The timing and mechanism of calcium transport in the uterus and embryonic membranes will be studied by defining the developmental expression of the calcium transporting proteins, calbindin-D28K, Ca2+-ATPase, using immunoblotting and immunohistochemistry. This research will contribute a greater understanding of embryonic development of terrestrial vertebrates through study of a highly conserved mechanism for delivery of nutrients to embryos. The project will integrate research and education by providing opportunities for undergraduate students from predominantly undergraduate departments in the research environment of a Ph.D. granting department. Undergraduate students from underrepresented segments of society will be actively recruited through collaboration with administrators of the Ronald E. McNair Scholars Program on campus. These efforts will target any eligible undergraduate from any institution. These students will receive training with an aim to increase their potential for future success in graduate programs.