Current Projects
Our goal is to use C. elegans to develop a comprehensive understanding of how cellular metabolism diversifies and changes during embryonic development and the impact of disrupting different cellular metabolic processes on key developmental events including cell fate specification, cell migrations, and cell-cell communication. We are utilizing the advantages of our model system to answer the following questions:
What are the metabolic states across embryogenesis?
The limited information available about cellular metabolism in embryos indicates that cells do not have uniform metabolism. Furthermore, extensive studies from the field of cancer biology implicate a switch from oxidative phosphorylation (OxPhos) to aerobic glycolysis in enabling cell migration, but it remains to be determined whether a similar switch occurs for migrating cells in developing embryos. We hypothesize that different cells transiently adopt distinct metabolic profiles (the collection of metabolic reactions taking place at a given time in a cell) to accomplish specific developmental processes. For example, gastrulating cells may switch to glycolysis during their migration and switch back to OxPhos once they have established cell adhesions, or cells secreting Wnt ligand may upregulate cholesterol synthesis, but in their daughter cells which downregulate Wnt ligand expression, cholesterol synthesis decreases. To test this hypothesis, we are taking advantage of the transparency of C. elegans embryos and the ease of transgenesis to collect time lapse confocal images of genetically encoded fluorescent reporters for cellular metabolic processes like glycolysis across wild-type embryonic development. Due to the invariant lineage of the worm, we can determine the identity of these cells, their future fates, and their gene expression from existing single cell sequencing datasets. With our time lapse data, we can determine how metabolic states change over the course of a cell’s lifetime and in its subsequent daughter cells. We are working to image reporters that measure glycolysis, OxPhos, glutamine catabolism, and total energy usage as well as synthesis of key molecules like cholesterol, fatty acid, and glycans.
What developmental processes are disrupted when a metabolic pathway is disrupted?
Our understanding of the role of cellular metabolism in developmental processes has been limited by a disconnect between the two fields: those studying metabolism do not describe phenotypes in any detail beyond “embryonic lethal” while those studying development tend to ignore metabolic genes identified with unbiased approaches because they are “too fundamental” to basic cellular processes. We aim to close this gap by using our time-lapse confocal imaging approach to collect quantitative, single cell resolution data on the defects observed in embryos with disrupted metabolism. Using existing data, we have identified all previously reported metabolic genes that cause embryonic lethality when knocked down with maternal RNAi but develop beyond the first two divisions of embryogenesis; this identified genes in 26 of 62 pathways, plus 12 additional genes not associated with a particular pathway. We are using maternal RNAi to knock these genes down in a fate reporter strain that enables us to quantify TFs associated with intestine, pharynx and hypodermis (skin) fates during time lapse imaging in addition to the position and division timing in all cells. Thus we can quantify the role of 38 required metabolic genes in cell fate specification and cell migration. The embryonic phenotypes of major signaling pathways are known, so we can evaluate each metabolic phenotype, and if it shares sufficient overlap with known signaling phenotype, we can utilize signaling reporters, such as fluorescently labeled Notch receptor or β-catenin, to determine if knockdown of the metabolism gene is affecting these signaling pathways. Thus we can also determine the role of metabolism in embryonic cell-cell signaling.
How do transcription and other metabolic pathways change when a metabolic pathway is disrupted?
While research is evolving to understand how metabolism changes when key factors are absent in other life stages, our previous work emphasizes the fact that embryogenesis is a unique developmental stage. Oocytes are packed with nutrients and energy for the developing embryo and with the exception of mammals, more cannot be obtained until the organism hatches and eats. Because of this we hypothesize that distinct alternative pathways may have evolved to cope with low levels of key micronutrients. To test this hypothesis, we are performing single cell sequencing and gas chromatography–mass spectrometry metabolite analysis on embryos from mothers on diets deficient in essential micronutirents vitamin B12, vitamin A, folate and riboflavin. We will use the combination of single cell expression data for metabolic genes and global metabolite levels to determine the metabolic state of individual cells and identify changes in metabolism in these mutants to determine if any changes are distinct from those observed in adult worms. We are also working with collaborators to use this data to determine the metabolic flux for the embryo at different stages, the first such analysis for a metazoan embryo.
