Pluripotency is the potential for a cell to give rise to any cell type in an organism. Multipotency is the potential for a cell to give rise to multiple different cell fates, but the scope of possible cell fates for a multipotent cell is narrower than that of a pluripotent cell. While the determinants of pluripotency are being heavily studied, not as much research is being done on the determinants of multipotency. In C. elegans, there are a pair of precursor cells that divide into two daughter cells with distinct fates. One daughter cell becomes a multipotent progenitor known as the somatic gonadal precursor (SGP) while the other becomes a terminally differentiated head mesodermal cell (hmc). The SGP gives rise to all of the tissues of the somatic gonad. This suggests that the factors specifying multipotency are segregated into the SGP daughters and away from the hmc daughters. Our goal is to identify these determinants of multipotency vs. terminal differentiation. We used fluorescence-activated cell sorting to isolate SGPs and hmcs from the same worms and performed RNA sequencing to identify ~6000 genes that are differentially expressed between the two cell types (Mathies et al., 2019). Of these 6,000 genes, 175 were transcription factors that were more highly expressed in SGPs than in hmcs. The goal of the project is to identify transcription factors that regulate multipotency in this lineage. We have taken the approach of inactivating the transcription factors by RNAi and observing if there is any change in the expression of SGP or hmc markers. The hmc is marked with
arg-1::GFP, while the SGPs are marked with
ehn-3::tdTomato. We are also examining L4 staged worms for changes in gonadal morphology indicative of a loss of multipotency. Each SGP generates one of the two gonadal arms; therefore, changes in the fate or potency of the SGPs could be reflected in gonadal shape. To date, we have screened 44 of the 175 transcription factor genes and found two genes in which expression of GFP is higher in SGPs compared to our negative control and three genes for which gonadal shape is altered. These genes are good candidate transcription factors that may play a role in determining cell fate and/or multipotency. Our plan going forward is to continue screening the list of 175 transcription factors to identify genes with differentiated expression for further analysis. Ultimately, we aim to identify the downstream targets of these transcription factors; among these are the direct mediators of multipotency. This project will contribute to our understanding of multipotency, and it opens the door for research into homologous genes in other organisms that may be conserved regulators of multipotency.