Embryonic stem cells (ES) are able to grow indefinitely while maintaining pluripotency and keeping the ability to differentiate. Several transcription factors are related to the maintenance of pluripotency, and some genes that are commonly found upregulated in tumors are associated with the long- term maintenance of the ES phenotype and culture proliferation ability. In this study, investigators Takahashi and Yamanaka from Kyoto University, analyzed which factors are crucial for inducing pluripotency. To this end, they first selected 24 candidate genes, and transfected mouse embryonic fibroblasts (MEF) with these into the Fbx15 gene locus using a beta-geo cassette and a helper retrovirus vector. They found that only transfection of all 24 genes together produced G418 resistance, in contrast of using single genes alone. Then, they evaluated which of these genes were critical individually, and found that Oct3/4, Klf4, Sox2 and c-Myc were relevant to generate the induced pluripotent stem (iPS) cells. RT- PCR showed that iPS cells expressed ES markers, and global expression profiles -using microarrays- revealed that iPS cells clusters with ES cells but not with fibroblasts, although some genes seemed to be more efficiently regulated in ES cells -thus, iPS were similar but not identical to ES cells. Evaluation of pluripotency of the obtained clones was performed by subcutaneous injection to form teratomas. Histology, immuno- staining and RT- PCR confirmed that some clones differentiated into all three germ layers without expressing trophoblast markers. Pluripotency was confirmed by growing the clones in culture dishes: the embryoids attached to the surface and initiated differentiation. The experiments were repeated using adult mouse fibroblasts, and again the same four factors succeeded in producing pluripotent cells. This research results interesting for the advance that it represents for stem cell in vitro differentation, although more research is required to establish the role of each of the four factors towards the generation and control of pluripotency.
Takahashi, K., and S.Yamanaka. 2006. Cell 126:663-676
epigenetics
More comments about this paper can be found in the same issue of Cell, at the Faculty of 1000 website, and here.
Takahashi, K., and S.Yamanaka. 2006. Cell 126:663-676
epigeneticsMore comments about this paper can be found in the same issue of Cell, at the Faculty of 1000 website, and here.
