Somatic cell nuclear transfer, cell fusion, or expression of lineage-specific factors have been shown to induce cell-fate changes in diverse somatic cell types. We recently observed that forced expression of a combination of three transcription factors, Brn2 (also known as Pou3f2), Ascl1 and Myt1l, can efficiently convert mouse fibroblasts into functional induced neuronal (iN) cells. Here we show that the same three factors can generate functional neurons from human pluripotent stem cells as early as 6 days after transgene activation. When combined with the basic helix-loop-helix transcription factor NeuroD1, these factors could also convert fetal and postnatal human fibroblasts into iN cells showing typical neuronal morphologies and expressing multiple neuronal markers, even after downregulation of the exogenous transcription factors. Importantly, the vast majority of human iN cells were able to generate action potentials and many matured to receive synaptic contacts when co-cultured with primary mouse cortical neurons. Our data demonstrate that non-neural human somatic cells, as well as pluripotent stem cells, can be converted directly into neurons by lineage-determining transcription factors. These methods may facilitate robust generation of patient-specific human neurons for in vitro disease modelling or future applications in regenerative medicine.
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Acknowledgements We would like to thank Y. Kokubu for technical assistance in molecular cloning and Y. Zhang for assistance in iPS cell induced neuron culture. We also thank Y. Sun for providing the microRNAs expression lentiviral vectors and S. Majumder for the REST-VP16 construct. This work was enabled by start-up funds from the Institute for Stem Cell Biology and Regenerative Medicine at Stanford (M.W.), the Ellison Medical Foundation (M.W.), the Stinehard-Reed Foundation (M.W.), the Donald E. and Delia B. Baxter Foundation (M.W.), the NIH grants 1R01MH092931 (M.W. and T.C.S.) andRC4 NS073015 (M.W.), and a Robertson Investigator Award from the New York Stem Cell Foundation. Z.P.P. is supported by 2008 and 2010 NARSAD Young Investigator Awards. T.V. is supported by the Ruth and Robert Halperin Stanford Graduate Fellowship. A.C. is supported by the AXA research fund and D.R.F. is supported by BioX Undergraduate Fellowship.