Efficient Conversion of Human Fibroblasts to Functional Neurons by Sequential Inactivation of PTB/nPTB‐regulated RNA Programs

Cellular reprogramming can be accomplished by using a set of cell lineage‐specific transcription factors. Our recent work reveals a key RNA program regulated by the RNA binding protein PTB, which is not only necessary but also sufficient to trans‐differentiate diverse cell types into neuronal‐like cells or even functional neurons (Xue et al., Cell 152:82‐96, 2013). In this RNA program, PTB down‐regulation allows the neuronal‐specific microRNA miR124 to more efficiently target its substrates, including multiple components of the REST complex responsible for repressing a large array of neuronal‐specific transcription factors in non‐neuronal cells. Down‐regulated REST also further depresses miR124, thus creating a sustainable signal for reprogrammed cells to maintain the neuronal identity. One application of this new neuronal induction protocol is to convert patient fibroblasts into neurons for mechanistic studies of neurodegenerative diseases. Interesting, unlikely mouse cells, human fibroblasts can only be converted to neuronal‐like cells with complex neuronal morphology but without functional neuronal properties, suggesting the existence of another gatekeeper(s) that uniquely operates for neuronal maturation in human cells. We now demonstrate that this gatekeeper consists of the PTB paralog called nPTB and a specific transcription factor Brn2, which together forms another feedback loop critical for neuronal maturation. Either overexpression of Brn2 or down‐regulation of nPTB in PTB down‐regulated cells induces mature neuronal marks and a full spectrum of neuronal activities. These data demonstrate a sequential control mechanism for neuronal induction and maturation in human cells.