The role of the TGF-B family in the cell differentiation from pluripotent bone marrow stem cells to immunocytes

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The present study was examined to clarify the role of the TGF-β family in the cell differentiation from pluripotent bone marrow stem cells to immunocytes, essentially by using mouse P19 embryonal carcinoma cells which resembled the inner cell mass of the early embryo and had pluripotency, and A-6 which was a hematopoietic stem cell line established from mouse ES cells. When P19 cells were cultured in a tissue culture dish in the presence of serum, Brachyury (Bra) and Goosecoid (Gsc) were unexpectedly expressed. Expression of Bra and Gsc, mesodermal genes, was induced by the stimulation of the Nodal and Cripto pathway, indicating the importance of control of Nodal and Cripto expression for mesodermal formation. This Nodal signaling was also mediated via Smad2/3 through Alk4/5/7, mouse TGF-β type I receptors. Furthermore, it was suggested that unidentified molecules other than Lefty were involved in inhibition of the Nodal pathway during monolayer P19 cell culture, leading to down-regulation of Nodal and subsequently Bra and Gsc. In A-6 cells, there was significant gene expression of not only various TGF-β receptors, various Smads and undifferentiated markers but also tissue-specific transcription factors such as GATA-1, Mitf and so on. The examination of the expression and function of alternative splice variants of Mitf isoforms revealed a diverse pattern of gene expression and different transcriptional activities of Mitf isoforms, suggesting discrete regulation of gene transcription in specific tissues by Mitf. The expression and function of alternative splice variants of Alk5 were examined. Ubiquitous expression of Alk5 isoforms in mouse tissues and cells including A-6 cells and insignificant effects of alternative splicing on transcriptional activation and signaling induced by TGF-β were shown.