iTRAQ proteome analysis reflects a progressed differentiation state of epiblast derived versus inner cell mass derived murine embryonic stem cells.

UNLABELLED

Mouse embryonic stem cells (mESC) and mouse epiblast stem cells (mEpiSC) share similar pluripotency factors like NANOG or POU5F1, however, their state of pluripotency differs significantly. mESC and mEpiSC can be derived from embryos generated by fertilization (FT) or by somatic cell nuclear transfer (NT). In this study we performed a 4-plex iTRAQ LC-MS/MS based approach, facilitating the multiplexed comparison of the four indicated types of stem cells. From four replicates of each cell type, 1650 proteins were quantified. 234 non redundant proteins with significant abundance alterations between FT/NT-mESC and FT/NT-mEpiSC, and 44 between FT and NT derived cells were detected. Bioinformatic analysis revealed that several pluripotency associated proteins, among them POU5F1, DNMT3L, TIF1B, and proteins involved in DNA repair like MSH2 and MSH6, are more abundant in mESC compared to mEpiSC. The abundance level of these proteins is not affected by the mode of embryo generation, whereas several cytoskeleton proteins show a higher abundance in NT-mESC compared to FT-mESC. In addition, a number of cytoskeletal proteins are enriched in mEpiSC, e.g., myosins, filamins and intermediate filament proteins, reflecting the progressed differentiation state of epiblast derived versus inner cell mass derived murine pluripotent stem cells.

BIOLOGICAL SIGNIFICANCE

This study aims to get new insights in the pluripotency state of stem cells and to deepen the knowledge of early cell differentiation. In an iTRAQ MS approach, we quantitatively compared proteomes of inner cell mass derived stem cells (mESC) with epiblast derived stem cells (mEpiSC). These stem cell types are derived from embryos of different developmental stages, and therefore vary considerably in their state of pluripotency and reflect different stages of early differentiation. The proteins which show significant abundance differences between the two stem cell lines represent (i) promising targets to further decipher molecular processes during early embryo development and (ii) useful molecular markers to monitor early differentiation events of stem cells by targeted approaches.