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.
