Qualitative and Quantitative Comparison of the Proteome of Erythroid Cells Differentiated from Human iPSCs and Adult Erythroid Cells by Multiplex TMT Labelling and NanoLC-MS/MS

Induced pluripotent stem cells (iPSC) are an attractive progenitor source for the generation of in vitro blood products. However, before iPSC-derived erythroid cells can be considered for therapeutic use their similarity to adult erythroid cells must be confirmed. We have analysed the proteome of erythroid cells differentiated from the iPSC fibroblast derived line (C19) and showed they express hallmark RBC proteins, including all those of the ankyrin and 4.1R complex. We next compared the proteome of erythroid cells differentiated from three iPSC lines (C19, OCE1, OPM2) with that of adult and cord blood progenitors. Of the 1989 proteins quantified,3 % differed in level by 2-fold or more between the different iPSC-derived erythroid cells. When compared to adult cells, 11 % of proteins differed in level by 2-fold or more, falling to 1.9 % if a 5-fold threshold was imposed to accommodate slight inter-cell line erythropoietic developmental variation. Notably, the level of.30 hallmark erythroid proteins was consistent between the iPSC lines and adult cells. In addition, a sub-population (10–15%) of iPSC erythroid cells in each of the iPSC lines completed enucleation. Aberrant expression of some cytoskeleton proteins may contribute to the failure of the majority of the cells to enucleate since we detected some alterations in cytoskeletal protein abundance. In conclusion, the proteome of erythroid cells differentiated from iPSC lines is very similar to that of normal adult erythroid cells, but further work to improve the induction of erythroid cells in existing iPSC lines or t