UC Santa Barbara

Proper specification of cell fates and organ identity is critical to the creation of a fully functional organism during animal development. Uncovering the dynamic events underlying deployment of gene regulatory networks (GRNs) that dictate cell fate and organ identity is critical to understanding the mechanisms that ensure developmental robustness and the relationship between the genesis and function of organs. In C. elegans, the entire intestine arises from a single cell, the E (endoderm) blastomere, which is specified by the seven-cell stage of embryogenesis. As with nearly all other cells throughout development of the animal, the E cell undergoes a stereotyped pattern of cell divisions, giving rise to 20 intestinal cells in the fully developed embryo before hatching into a first-stage larva. While many of the key, phylogenetically conserved regulatory factors comprising the endoderm GRN, including a cascade of GATA-type transcription factors, have been identified, the function and identity of modulators that regulate faithful execution of the GRN during endoderm development have not been comprehensively revealed. To access the broad set of genetic modulators underlying the endoderm GRN, I have developed a high-throughput pipeline based on bulk sorting of C. elegans embryos and large-scale genomic sequencing and applied it to identifying genomic regions responsible for natural variation in its execution. This method revealed quantitative trait loci (QTLs) that overlap with those identified by classical strategies, thus validating the approach, and also identified additional QTLs underlying variation in execution of endoderm development. I found that one candidate gene, encoding BRAP-2 (BRCA1-associated protein-2), regulates endoderm specification by modulating expression of a core regulator, the END-1 GATA factor, likely by tuning the activity of key maternal regulatory inputs into the GRN. I further applied this method to the bulk analysis of embryos following mutagenesis to assess genetic regions that, when mutated, alter gut development. This latter strategy revealed that components in actin networks play a previously unknown role in ensuring the proper number of endoderm cells. This study contributes to our understanding of how regulatory inputs fine-tune an embryonic GRN, resulting in the robust production of an entire germ layer, the endoderm, and a crucial organ, the gut.