Department of Molecular Genetics, Graduate School of Medicine, Kyoto University, Kyoto 606-8501, Japan.
Development. 2012 May;139(10):1734-43. doi: 10.1242/dev.076539. Epub 2012 Apr 4.
Fibroblast growth factor 2 (FGF2) and glial cell line-derived neurotrophic factor (GDNF) are required to recapitulate spermatogonial stem cell (SSC) self-renewal in vitro. Although studies have revealed the role of the GDNF signaling pathway in SSCs, little is known about how FGF2 is involved. In the present study, we assessed the role of the FGF2 signaling pathway using a mouse germline stem (GS) cell culture system that allows in vitro expansion of SSCs. Adding GDNF or FGF2 induced phosphorylation of MAPK1/3, and adding the MAP2K1 inhibitor PD0325091 reduced GS cell proliferation and MAPK1/3 phosphorylation. Moreover, GS cells transfected with an activated form of Map2k1 not only upregulated Etv5 and Bcl6b gene expression, but also proliferated in an FGF2-independent manner, suggesting that they act downstream of MAP2K1 signaling to drive SSC self-renewal. Although GS cells transfected with Map2k1, Etv5 or Bcl6b showed normal spermatogonial markers, transplanting GS cells expressing Bcl6b into infertile mouse testes resulted in the formation of a germ cell tumor, suggesting that excessive self-renewal signals causes tumorigenic conversion. These results show that FGF2 depends on MAP2K1 signaling to drive SSC self-renewal via upregulation of the Etv5 and Bcl6b genes.
成纤维细胞生长因子 2 (FGF2) 和胶质细胞源性神经营养因子 (GDNF) 是体外重现精原干细胞 (SSC) 自我更新所必需的。尽管研究已经揭示了 GDNF 信号通路在 SSCs 中的作用，但对于 FGF2 如何参与其中知之甚少。在本研究中，我们使用允许 SSCs 体外扩增的小鼠生殖干细胞 (GS) 细胞培养系统评估了 FGF2 信号通路的作用。添加 GDNF 或 FGF2 诱导 MAPK1/3 的磷酸化，添加 MAP2K1 抑制剂 PD0325091 可减少 GS 细胞增殖和 MAPK1/3 磷酸化。此外，转染激活型 Map2k1 的 GS 细胞不仅上调了 Etv5 和 Bcl6b 基因的表达，而且还可以在不依赖 FGF2 的情况下增殖，表明它们作为 MAP2K1 信号的下游作用来驱动 SSC 自我更新。尽管转染了 Map2k1、Etv5 或 Bcl6b 的 GS 细胞显示出正常的精原细胞标记物，但将表达 Bcl6b 的 GS 细胞移植到不育小鼠睾丸中会导致生殖细胞肿瘤的形成，表明过度的自我更新信号会导致肿瘤转化。这些结果表明，FGF2 通过上调 Etv5 和 Bcl6b 基因依赖 MAP2K1 信号来驱动 SSC 自我更新。
