Combinatorial regulation by developmentally regulated transcription factors play a central role in defining which genes are regulated in each cell during development and disease, and allows the same factors to play different roles in different cells. The C. elegans embryo is an ideal system for a comprehensive study of the role of lineage history in the context-dependent regulation of cell fate because of its invariant lineage and powerful experimental tools. We recently developed automated lineage tracing and expression mapping methods for C. elegans embryogenesis and have built the ?Expression Patterns in Caenorhabditis? (EPIC) database that contains the expression of over 250 fluorescent reporters for transcription factor (TF) expression in every cell of developing embryos. In our past work, we have shown the potential for this resource as a starting point for defining mechanisms controlling development. While we will continue to map the expression of novel regulators, the main focus of this proposal is to use this database and our methods to address poorly understood questions in developmental biology. 1) How do cells know their lineage history and translate this information into correct terminal cell fates? We have identified a set of ~15 lineage-specific TFs whose expression distinguishes a group of progenitor cells for diverse tissues descended from the ?ABpxp? blastomeres. We plan to test whether these TFs are necessary and sufficient individually and in combination to specify the lineage identity of these progenitor cells, and to identify their targets, by imaging and genomics analysis of loss and gain-of-function mutants. We will complement this by a detailed analysis of the cis- regulatory sequences controlling terminal differentiation genes to identify their upstream regulators. These ?top-down? and ?bottom-up? approaches should eventually converge to a common regulatory network. 2) What mechanisms allow reuse of the same regulator(s) for different purposes in different developmental contexts, such as different lineages? We have identified two sets of genes, expressed in either posterior or anterior daughter cells after cell division, both of which are regulated by the Wnt pathway. We will combine enhancer fine-mapping, expression mapping of synthetic enhancers, and genome-wide binding assays for the Wnt-regulated TF POP-1/TCF to determine how each gene's response (activation or repression) to this pathway is regulated differently in different cells. 3) How does redundancy of genes and enhancers influence developmental robustness? Redundancy is extremely common in the early embryo, and we will test the hypothesis that this redundancy exists to ensure robust development in the face of environmental variability by detailed phenotyping of mutants in different conditions. In summary, the work we propose will begin to complete the early embryonic regulatory network and answer important questions about principles of development that are likely to be conserved across animals.
In animal development the single-celled zygote divides, and its progeny differentiate to produce an incredible diversity of cell types, but we know little about the molecular changes that occur in cells as they divide and differentiate. This project will use cutting- edge microscopy and genomics tools to study this process in the C. elegans embryo, whose stereotyped development allow rapid discovery of new mechanisms and whose conservation with humans suggests many of these mechanisms will be shared. We will focus on how the same gene regulates different genes in different cells, how redundant genes act to minimize developmental mistakes, and how multiple genes combine to specify each cell type.
