Figure 3.

Mouse Embryonic Stem Cells Lose Expression of the Pluripotency Markers Oct4, Sox2, and Nanog and Express Trophoblast Markers on Depletion of Geminin and Emi1

(A) Giant cell formation and induction of trophectoderm markers Troma-1 and P-cadherin (both in green) is evident in B6/Blu-1 mouse embryonic stem (ES) cells depleted of Emi1, geminin, and Oct4 at 2 days. Total DNA is shown in red. Scale bars represent 20 μm.

(B) Emi1 and geminin depletion in mouse B6/Blu-1 ES cells result in downregulation of the pluripotency transcription factors Oct4, Sox2, and Nanog (in white). Total DNA is shown in red. Scale bars represent 20 μm.

(C) In B6/Blu-1 ES cells, geminin is coexpressed with Oct4 (geminin and Oct4 both in white). Total DNA is shown in red. Scale bars represent 20 μm. The bar chart shows that geminin is present in a higher proportion of asynchronous ES and EC cells than in 3T3 fibroblasts, and notably in a far great proportion of cells that are not actively replicating DNA. Error bars represent 5% standard error.

(D) Table summarizing cell-cycle analyses by immunofluorescence and flow cytometry. In asynchronous populations, the majority of 3T3 fibroblasts are in G1, in contrast to ES and EC cells in which G1 comprises a small fraction of the cell cycle and the majority of cells are in S phase.

(E) Geminin is present during G1 in ES cells harvested at different time points following mitotic shake-off. Approximately 50% of ES cells enter S phase at 2 hr, suggesting that the duration of G1 phase is likely to be between 2 and 4 hr.

For additional related data, see Figure S3.