Evolution of Dimorphic Morphogenesis in Rhabditid Nematodes

性别二态性是性别之间在特征上的差异，如颜色，身体部位的形状和大小（形态学），甚至行为。本研究项目的主要目的是了解二形结构的形态多样性是如何演变的。这一目标解决了发育和进化生物学中两个基本上重要但基本上没有答案的问题：（1）基因调控网络（GRNs）的变化如何导致形态多样性？以及（2）GRNs如何影响这种进化变化？一个主要的假设是，一些基因，称为“热点”基因，反复参与形态进化，可能是由于他们在GRNs的中心位置。该研究项目将使用一种模型结构--线虫尾尖来检验这一“热点”假说，这种结构已经反复进化出二型性。这项研究不仅将增加对基本进化机制的理解，而且将有助于理解形态发生，这是发育，癌症转移和伤口愈合的基本过程。关于控制性别差异和控制发育时间的基因，也有望获得新的知识。更广泛的影响包括在发育遗传学、基因组学和进化方面培训新的研究生和本科生。由于这些学生将从不同的人才库中招募，妇女和代表性不足的少数民族的参与可能会在这一科学学科中得到丰富。在这项研究的过程中，新的资源（基因构建体，菌株和数据库）和新的方法（基因表达的组织特异性分析）将被产生和共享，可能对其他研究形态发生和细胞生物学有用。最后，一个独特的生活，策划收集不同的线虫物种将保持作为一个宝贵的资源，比较生物学，基因组学和education.The尾部尖端的男性和女性的许多小杆线虫物种（以及小杆线虫祖先）有一个尖锐的形态，但男性特有的尾尖形态发生（TTM）产生一个圆形的形状已反复获得和失去在进化过程中。两种不同的方法将被用来测试的“热点”基因假说的TTM二态性的演变：候选基因的方法和公正的基因组水平的方法。首先，DMD-3，在秀丽隐杆线虫TTM的“主调节器”，将作为候选的“热点”基因进行测试。smFISH将用于确定DMD-3是否在来自独立进化TTM的系统发育谱系的雄性中表达。贝叶斯方法将用于检测DMD-3表达变化与TTM二型性之间的任何系统发育相关性。如果存在这种相关性，DMD-3的功能需求将通过CRISPR/Cas9敲除或RNAi进行测试。第二种方法将使用尾尖特异性转录组谱的系统发育分析来鉴定与TTM进化一致地改变表达的任何其他基因。将获得在L3至L4的几个阶段从雄性和雌性激光解剖的尾尖的RNA-seq图谱。对于每个物种，这些谱将鉴定在L3和L4阶段之间动态表达、在不同性别之间差异表达并且与胚胎发育中TTM的获得或丧失相关的尾尖基因。

Gender dimorphism is the difference between genders in features such as coloration, shapes and sizes of body parts (morphology), or even behavior. The primary objective of this research project is to understand how morphological diversity in dimorphic structures has evolved. This objective addresses two fundamentally important but largely unanswered questions in developmental and evolutionary biology: (1) How do changes in gene regulatory networks (GRNs) result in morphological diversity? and (2) How might GRNs bias such evolutionary change? A major hypothesis is that some genes, called "hot-spot" genes, are involved repeatedly in morphological evolution, perhaps due to their central positions in GRNs. The research project will test this "hot-spot" hypothesis using a model structure, the nematode tail tip, which has repeatedly evolved dimorphism. This research will not only add to the understanding of fundamental evolutionary mechanisms, but will aid in the understanding of morphogenesis, a fundamental process in development, cancer metastasis and wound healing. New knowledge is also expected regarding genes that govern gender differences and control the timing of developmental events. Broader impacts include the training of new graduate and undergraduate students in developmental genetics, genomics and evolution. Because such students will be recruited from diverse pools, the participation of women and underrepresented minorities is likely to be enriched in this scientific discipline. In the course of this research, new resources (gene constructs, strains and databases) and novel methods (tissue-specific analysis of gene expression) will be generated and shared, likely to be useful to others researching morphogenesis and cell biology. Finally, a unique living, curated collection of diverse nematode species will be maintained as a valuable resource for comparative biology, genomics and education.The tail tips of both males and females of many rhabditid nematode species (as well as the rhabditid ancestor) have a pointed morphology, but male-specific tail tip morphogenesis (TTM) producing a rounded shape has been repeatedly gained and lost during evolution. Two different approaches will be used to test the "hot-spot" gene hypothesis for the evolution of TTM dimorphism: a candidate-gene approach and an unbiased genomics-level approach. First, DMD-3, the "master-regulator" of TTM in Caenorhabditis elegans, will be tested as a candidate "hot-spot" gene. smFISH will be used to determine if DMD-3 is expressed in males from phylogenetic lineages that independently evolved TTM. Bayesian methods will be used to test for any phylogenetic correlation between changes in DMD-3 expression and TTM dimorphism. If such correlation exists, the functional requirement for DMD-3 will be tested by CRISPR/Cas9 knockout or RNAi. The second approach will use phylogenetic analysis of tail-tip-specific transcriptome profiles to identify any other genes that changed expression in concert with TTM evolution. RNA-seq profiles will be obtained for tail tips that have been laser-dissected from males and females at several stages from L3 through L4. For each species, these profiles will identify tail tip genes that are dynamically expressed between L3 and L4 stages, differentially expressed between the different genders, and correlate with the gain or loss of TTM in the phylogeny.