Small molecule can replace Sox2, c-Myc in reprogramming
The standard technique for making differentiated cells that behave like embryonic stem cells uses viruses to insert the genes c-Myc, Klf4, Oct4 and Sox2. However, adding these genes into cells makes the cells less predictable and more likely to form tumours. Researchers have been able to reprogram neural stem cells using only Oct4, but these cells are not readily available from patient biopsies; thus, researchers are searching for alternative techniques. New work published in Cell Stem Cell shows that a small drug-like molecule can effectively replace two of the four genes typically used to generate induced pluripotent stem cells1.
To begin their hunt for compounds that could help reprogram cells, researchers led by Kevin Eggan and Lee Rubin of the Harvard Stem Cell Institute in Cambridge, Massachusetts used cultures of mouse skin cells engineered to express green fluorescent protein as a marker of pluripotency. They first screened for small molecules that allowed mouse cells to be reprogrammed without adding Sox2. When three such molecules were identified, the researchers tried again and found that one of the molecules could reprogram cells even in the absence of c-Myc, a tumour-promoting gene that, although not required for reprogramming, greatly boosts success rates.
To make sure the cells were really reprogrammed, the researchers performed a series of tests, including mixing the cells with mouse embryos and demonstrating that the cells could contribute to every type of tissue in chimeric mice. They named the identified molecule RepSox for its ability to replace Sox2 and also after the Boston Red Sox, the local major league baseball team. Previous studies had shown that this molecule inhibits a kinase in the TGFβ signaling pathway. Careful work showed that RepSox did not work by activating Sox2 in fibroblasts, as might be expected. Instead, the molecule functions in partially reprogrammed cells that accumulate in the absence of Sox2, apparently by inducing and stabilizing Nanog expression. Thus, the researchers wrote in the paper, the discovery of RepSox is important not only for replacing one of the reprogramming factors, but also for illuminating a new strategy for identifying such molecules. “There need not always be a discrete, one-to-one mapping between the functions of the reprogramming factors and their chemical replacements.”
Robert Blelloch, who studies reprogramming at the University of California, San Francisco, praised the team's strategy of only screening compounds whose mechanisms are at least partly understood. “They find a small molecule that replaces a factor, but they take it further and use it to understand the biology.”
Ichida, J. K. et al. A small-molecule inhibitor of Tgf-β signaling replaces Sox2 in reprogramming by inducing Nanog. Cell Stem Cell 10.1016/j.stem.2009.09.012 (published online 8 October 2009).
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Baker, M. RepSox hits a home run. Nat Rep Stem Cells (2009). https://doi.org/10.1038/stemcells.2009.130