Evolution of a Fundamental Morphogenetic Process in the Volvocine Algae

沃尔沃辛藻基本形态发生过程的进化

• 批准号: 0131565
• 负责人: David Kirk
• 金额: $ 40.5万
• 依托单位: Washington University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-01-01 至 2005-12-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0131565&HistoricalAwards=false
• 关键词: Evolution; Fundamental; Morphogenetic; Process; Volvocine

0131565KirkOur goal is to determine how an essential morphogenetic process requiring extensive intercellular coordination arose as multicellular Volvox evolved from a unicellular ancestor resembling its closest unicellular relative, Chlamydomonas. In order to be able to swim, at the end of embryogenesis Volvox must turn right-side-out in a gastrulation-like process called "inversion," which involves coordinated cytoskeletal rearrangements and changes in cell shape and motility. Transposon tagged mutants with a variety of inversion (inv) defects have been generated. The first of these to be analyzed was Inv61, in which inversion is arrested at the halfway point as a result of a transposon insertion in invA, a gene encoding a novel kinesin that is localized in the embryo near the cytoplasmic bridges that play a key role in inversion. We have cloned the C. reinhardtii orthologue of invA (iarA) and found that it encodes a kinesin (IarA) that is 90 % identical in aa sequence to InvA in both its motor and tail domains. Because invA encodes a microtubule motor, whereas invA mutants resemble wild-type embryos exposed to actomyosin inhibitors, we will explore the possibility that InvA acts as a link between the actin- and tubulin-based cytoskeletons in Volvox. Meanwhile, we will study expression and localization of its orthologue, IarA, throughout the Chlamydomonas cell cycle, and will study the effects of blocking IarA function, using a dominant negative construct, antibody inhibition, or RNA interference. We will use nuclear transformation to determine whether iarA can rescue the invA mutant. If it does not, we will test chimeric iarA-invA constructs to define regions of InvA that are required to confer an inversion function on IarA. The functional significance of such structural differences will then be tested with appropriate kinesin binding and/or motility assays. If iarA does rescue invA, we will compare the proteins with which IarA interacts in Chlamydomonas versus those with which it interacts in Volvox.

0131565KirkOur的目标是确定多细胞Volvox是如何从与其最接近的单细胞近亲衣藻相似的单细胞祖先进化而来的，这一必要的形态发生过程需要广泛的细胞间协调。为了能够游泳，在胚胎发育结束时，Volvox必须在一个类似原肠形成的过程中向右翻，这个过程被称为“倒置”，这个过程涉及细胞骨架的协调重排以及细胞形状和运动的变化。转座子标记的具有各种倒位(Inv)缺陷的突变体已经产生。其中第一个被分析的基因是INV61，它的倒位是由于在INVA中插入转座子而被阻止的，INVA是一种编码一种新的动蛋白的基因，位于胚胎中靠近细胞质桥的地方，在倒位中起关键作用。我们克隆了INVA(IARA)的C.reinhardtii同源基因，发现它编码一个激动素(IARA)，在其运动区和尾区与INVA在AA序列上有90%的同源性。由于invA编码微管马达，而invA突变体类似于暴露于肌动蛋白抑制剂的野生型胚胎，我们将探索invA在Volvox中作为基于肌动蛋白和微管蛋白的细胞骨架之间的联系的可能性。同时，我们将研究其同源基因IARA在衣藻细胞周期中的表达和定位，并将研究使用显性负结构、抗体抑制或RNA干扰来阻断IARA功能的效果。我们将使用核转化来确定Iara是否可以拯救inva突变体。如果没有，我们将测试嵌合的Iara-inva构造，以定义向Iara赋予反转函数所需的inva区域。这种结构差异的功能意义将通过适当的Kinesin结合和/或运动性分析来测试。如果IARA确实拯救了INVA，我们将比较IARA在衣藻中与之相互作用的蛋白质与它在Volvox中与之相互作用的蛋白质。