Despite decades of research, molecular mechanisms that guide tissue differentiation during early embryonic development are not fully understood. Using custom-built microanalytical mass spectrometry instruments, our laboratory has recently found metabolic differences between identified cells in early developing embryos that give rise to different types of tissues in the frog (<i>Xenopus laevis)</i>. We further demonstrated that altering endogenous metabolite concentration impacted the cell’s tissue fate during development. For example, the D11 cell, which is known to primarily give rise to the lens, spinal cord, and the brain formed epidermal tissue when injected with metabolites that were more abundant in epidermal precursor cells (V11). However, the effect of the metabolite induced fate-changes on the cellular proteome and its role in changing the tissue fate of the D11 cells remains to be explored. Here we have developed a workflow to study the proteomic changes that occur following the alteration of metabolite concentration in early embryo. It combines cell lineage tracing, fluorescent activated cell sorting (FACS), tandem mass tag (TMT) labeling of proteomic samples and liquid chromatography-mass spectrometric (LC-MS) analysis. We injected D11 cells with a mixture of metabolite and fluorescent dye to trace D11 cell clones in later stages. We cultured the embryos to neurula stage, dissociated the embryos to live cells and sorted the fluorescent live cell clones using FACS. We labeled metabolically altered and control samples with TMT followed by LC-MS analysis, to quantify changes in protein abundances. The results from this study will advance our understanding of how proteome changes dynamically in the developing embryo and will help identify protein markers responsible for tissue differentiation.<br/>

Proteome remodeling in response to metabolite injection of embryonic cells

Macromolecular Chemistry: The Second Century

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ACS Spring 2020 National Meeting & Expo

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.