Diversity Supplement: Cytokinesis and RNA segregation in zebrafish development

描述（由申请人提供）：生殖质核糖核粒（GP RNPs）的母体遗传导致原始生殖细胞规范的保守基因表达程序的激活，我们使用斑马鱼作为脊椎动物模型系统来研究这一过程。斑马鱼母系遗传的GP RNPs通过细胞骨架机制达到渐进的多聚化水平：在沟槽形成之前聚集（预聚集），在沟槽形成期间向沟槽招募，以及在成熟过程中沿着沟槽的远端压实。这些连续的过程导致形成四个大团聚集的GP RNPs，这将最终赋予生殖细胞规格。该建议的总体假设是，在沟形成之前和期间，GP RNP多聚性的增加是基于肌动球蛋白依赖的细胞骨架重排，由与GP RNP相关的肌球蛋白II马达介导。我们假设，导致这一过程不同阶段的重排，如预聚集、沟槽补充和远端压实，具有共同的潜在机制基础。我们将测试肌动球蛋白相互作用的模型，这些相互作用可能导致GP RNP在犁沟开始之前和期间（目标1）以及在犁沟成熟期间（目标2）聚合。我们进一步假设，这些潜在的机制被不同地修改以产生不同的细胞输出。在Aim 1中，我们将验证这样的假设，即在沟启动之前和期间，GP RNP相关的f-肌动蛋白网络被向外生长的星状微管修饰，这一过程与GP RNP相关的肌球蛋白II的全局激活相关联。在Aim 2中，我们将验证这样的假设，即在沟成熟过程中，与细胞分裂相关的慢钙波赋予了肌球蛋白II活性和/或细胞骨架动力学的中向远端偏倚，从而导致沟远端GP RNP压实。我们的发现将为细胞命运决定因素分离的新基本机制提供见解。最近的研究表明，生殖系基因与多能性和致瘤性之间存在联系，了解种质分离的机制将为生殖、再生和癌症生物学提供适用的知识。

DESCRIPTION (provided by applicant): Maternal inheritance of germ plasm ribonucleoparticles (GP RNPs) results in the activation of a conserved gene expression program for primordial germ cell specification, and we use the zebrafish as a vertebrate model system to study this process. Zebrafish maternally inherited GP RNPs have co-opted the cytoskeletal machinery to reach progressive levels of multimerization: aggregation prior to furrow initiation (pre- aggregation), recruitment to the furrow during its initiation, and distal compaction along th furrow undergoing maturation. These sequential processes result in the formation of four large masses of aggregated GP RNPs, which will eventually confer germ cell specification. The overarching hypothesis of this proposal is that increases in GP RNP multimerization prior to and during furrow formation are based on actomyosin-dependent rearrangements of the cytoskeleton, mediated by myosin II motors associated with GP RNPs. We hypothesize that rearrangements leading to various stages of this process, pre-aggregation, furrow recruitment and distal compaction, have a common underlying mechanistic basis. We will test models of actomyosin interactions that may result in GP RNP multimerization prior to and during furrow initiation (Aim 1) and during furrow maturation (Aim 2). We further hypothesize that these underlying mechanisms are modified differently to produce different cellular outputs. In Aim 1, we will test the hypothesis that, prior to and during furrow initiation, a GP RNP- associated f-actin network is modified by outwardly growing astral microtubules, a process that is coupled to the global activation of GP RNP-associated myosin II. In Aim 2, we will test the hypothesis that, during furrow maturation, slow calcium waves associated with cytokinesis confer a medial-to-distal bias of myosin II activity and/or cytoskeletal dynamics that result in GP RNP compaction at distal ends of the furrow. Our findings will provide insights into novel fundamental mechanisms for the segregation of cell fate determinants. Recent studies have shown a link between germ line genes and pluripotency and tumorigenicity, and understanding mechanisms of germ plasm segregation will provide knowledge applicable to reproductive, regenerative and cancer biology.