FGF2 Sustains NANOG and Switches the Outcome of BMP4-Induced Human Embryonic Stem Cell Differentiation

Abstract

SUMMARY Here we show that as human embryonic stem (ES) cells exit the pluripotent state, NANOG can play a key role in determining lineage outcome. It has previously been reported that BMPs induce differentiation of human ES cells into extraembryonic lineages. Here we find that FGF2, acting through the MEK-ERK pathway, switches BMP4 induced human ES cell differentiation outcome to mesendoderm, characterized by the uniform expression of T (brachyury) and other primitive streak markers. We also find that MEK-ERK signaling prolongs NANOG expression during BMP-induced differentiation; that forced NANOG expression results in FGF independent BMP4 induction of mesendoderm; and that knockdown of NANOG greatly reduces T induction. Together, our results demonstrate that FGF2 signaling switches the outcome of BMP4 induced differentiation of human ES cells by maintaining NANOG levels through the MEKERK pathway.

MSX2 mediates entry of human pluripotent stem cells into mesendoderm by simultaneously suppressing SOX2 and activating NODAL signaling

“…Unexpectedly, the pluripotency marker NANOG was repressed upon MSX2 depletion in both H1 and H9 hESCs (Supplementary information, Figure S3A-S3D). We speculated that this might be related to NANOG function in suppressing neural differentiation and in patterning different subtypes of mesoderm cells after exit of hESCs from pluripotency, as described previously [41,42]. To further confirm the role of MSX2 in hESC early differentiation and exclude the potential ambiguity brought by shRNAs, we deleted MSX2 gene in hESCs using CRISPR-CAS9 technology.…”

Induction of pluripotency in human somatic cells via a transient state resembling primitive streak-like mesendoderm

“…To further explore this possibility, we compared the global gene expression profiles of TRA-1-60 ( þ ) cells purified on various days, original HDFs, established iPSCs and differentiated progenies (endoderm, EN; mesoderm, ME; neuroectoderm, NE; and primitive streak-like mesendoderm [11][12][13][14] , PSMN from ESCs/ iPSCs using DNA microarrays ( Supplementary Fig. 4).…”

Signals from the surface modulate differentiation of human pluripotent stem cells through glycosaminoglycans and integrins

“…To induce mesoderm differentiation, cells (H9) were cultured on GBP or Matrigel with activin A for 1 d and then with BMP4 and bFGF for 3 d (8,11). Compared with cells cultured on Matrigel, cells cultured on GBP underwent a more pronounced decrease in expression of the pluripotency markers (NANOG and POU5F1) and expressed higher levels of primitive streak (T and MIXL1) and mesoderm marker genes (PDGFRA and FOXF1; SI Appendix, Fig.…”

“…Matrigel is the most widely used substratum for hPS cell propagation (5) and differentiation (6)(7)(8)(9)(10)(11)(12); it is derived from mouse sarcoma cells, and although its principal components are laminin, collagen, and entactin, Matrigel consists of up to 1,800 different proteins-including encapsulated growth factors-whose levels vary significantly from batch to batch (13). Accordingly, the contributions of Matrigel-delivered signals to specific phenotypic outcomes-self-renewal or differentiation-are difficult to characterize or control.…”