Subcellular Mass Spectrometry Reveals Proteome Remodeling in an Asymmetrically Dividing (Frog) Embryonic Stem Cell

This project enhanced discovery subcellular proteomics sensitivity by integrating diaPASEF with capillary electrophoresis mass spectrometry (CE-MS). The method was developed, tested, and validated on ~200 pg of the HeLa proteome digest, ~80% of the total proteome of a single cell. Reproducible identification of at least 1,000 proteins marked robust performance to explore subcellular proteome heterogeneity as the dorsal-animal (D1) cell divides asymmetrically in the Xenopus laevis embryo to establish the D11 and D12 descendants in the 16-cell embryo. The label-free quantification (LFQ) values were normalized to compare equal proteome amounts. Relative quantification of the proteomes revealed molecular differences between the future D11 and future D12 poles of the D1 cell and the offspring cells. Functional experiments ventralizing the fertilized embryo using ultraviolet (UV) light revealed corroborated the proteome profiles and helped decipher the origin of the gradients in the context of dorsal-ventral patterning.