Reprogramming cell fate with a genome-scale library of artificial transcription factors.

Eguchi A, Wleklinski MJ, Spurgat MC, Heiderscheit EA, Kropornicka AS, Vu CK, Bhimsaria D, Swanson SA, Stewart R, Ramanathan P, Kamp TJ, Slukvin I, Thomson JA, Dutton JR, Ansari AZ
Proc Natl Acad Sci U S A. 2016 113 (51): E8257-E8266

PMID: 27930301 · PMCID: PMC5187731 · DOI:10.1073/pnas.1611142114

Artificial transcription factors (ATFs) are precision-tailored molecules designed to bind DNA and regulate transcription in a preprogrammed manner. Libraries of ATFs enable the high-throughput screening of gene networks that trigger cell fate decisions or phenotypic changes. We developed a genome-scale library of ATFs that display an engineered interaction domain (ID) to enable cooperative assembly and synergistic gene expression at targeted sites. We used this ATF library to screen for key regulators of the pluripotency network and discovered three combinations of ATFs capable of inducing pluripotency without exogenous expression of Oct4 (POU domain, class 5, TF 1). Cognate site identification, global transcriptional profiling, and identification of ATF binding sites reveal that the ATFs do not directly target Oct4; instead, they target distinct nodes that converge to stimulate the endogenous pluripotency network. This forward genetic approach enables cell type conversions without a priori knowledge of potential key regulators and reveals unanticipated gene network dynamics that drive cell fate choices.

MeSH Terms (19)

Animals Binding Sites Cell Lineage Cellular Reprogramming Chaperonin Containing TCP-1 Epigenesis, Genetic Fibroblasts Gene Expression Regulation, Neoplastic Gene Regulatory Networks Genomic Library HEK293 Cells Humans Mice Protein Domains Protein Engineering Sequence Analysis, RNA Transcription, Genetic Transcription Factors Zinc Fingers

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