Dissecting direct reprogramming from fibroblast to neuron using single-cell RNA-seq

Barbara Treutlein, Qian Yi Lee, J. Gray Camp, Moritz Mall, Winston Koh, Seyed Ali Mohammad Shariati, Sopheak Sim, Norma F. Neff, Jan M. Skotheim, Marius Wernig, Stephen R. Quake

Research output: Contribution to journalArticlepeer-review

319 Scopus citations

Abstract

Direct lineage reprogramming represents a remarkable conversion of cellular and transcriptome states. However, the intermediate stages through which individual cells progress during reprogramming are largely undefined. Here we use single-cell RNA sequencing at multiple time points to dissect direct reprogramming from mouse embryonic fibroblasts to induced neuronal cells. By deconstructing heterogeneity at each time point and ordering cells by transcriptome similarity, we find that the molecular reprogramming path is remarkably continuous. Overexpression of the proneural pioneer factor Ascl1 results in a well-defined initialization, causing cells to exit the cell cycle and re-focus gene expression through distinct neural transcription factors. The initial transcriptional response is relatively homogeneous among fibroblasts, suggesting that the early steps are not limiting for productive reprogramming. Instead, the later emergence of a competing myogenic program and variable transgene dynamics over time appear to be the major efficiency limits of direct reprogramming. Moreover, a transcriptional state, distinct from donor and target cell programs, is transiently induced in cells undergoing productive reprogramming. Our data provide a high-resolution approach for understanding transcriptome states during lineage differentiation.

Original languageEnglish
Pages (from-to)391-395
Number of pages5
JournalNature
Volume534
Issue number7607
DOIs
StatePublished - 8 Jun 2016
Externally publishedYes

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