BMP4 promotes mouse iPS cell differentiation to male germ cells via Smad1/5, Gata4, Id1 and Id2

Abstract

Generation of male germ cells from pluripotent cells could provide male gametes for treating male infertility and offer an ideal model for unveiling molecular mechanisms of spermatogenesis. However, the influence and exact molecular mechanisms, especially downstream effectors of BMP4 signaling pathways, in male germ cell differentiation of the induce pluripotent stem (iPS) cells, remain unknown. This study was designed to explore the role and mechanism of BMP4 signaling in the differentiation of mouse iPS cells to male germ cells. Embryoid body (EB) formation and recombinant BMP4 or Noggin were utilized to evaluate the effect of BMP4 on male germ cell generation from mouse iPS cells. Germ cell-specific genes and proteins as well as the downstream effectors of BMP4 signaling pathway were assessed using real-time PCR and Western blots. We found that BMP4 ligand and its multiple receptors, including BMPR1a, BMPR1b and BMPR2, were expressed in mouse iPS cells. Real-time PCR and Western blots revealed that BMP4 could upregulate the levels of genes and proteins for germ cell markers in iPS cells-derived EBs, whereas Noggin decreased their expression in these cells. Moreover, Smad1/5 phosphorylation, Gata4 transcription and the transcripts of Id1 and Id2 were enhanced by BMP4 but decreased when exposed to Noggin. Collectively, these results suggest that BMP4 promotes the generation of male germ cells from iPS cells via Smad1/5 pathway and the activation of Gata4, Id1 and Id2. This study thus offers novel insights into molecular mechanisms underlying male germ cell development.