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

Asuka Eguchi; Matthew J. Wleklinski; Mackenzie C. Spurgat; Evan A. Heiderscheit; Anna S. Kropornicka; Catherine K. Vu; Devesh Bhimsaria; Scott A. Swanson; Ron Stewart; Parameswaran Ramanathan; Timothy J. Kamp; Igor Slukvin; James A. Thomson; James R. Dutton; Aseem Z. Ansari

doi:10.1073/pnas.1611142114

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

Asuka Eguchi, Matthew J. Wleklinski, Mackenzie C. Spurgat, Evan A. Heiderscheit, Anna S. Kropornicka, Catherine K. Vu, Devesh Bhimsaria, Scott A. Swanson, Ron Stewart, Parameswaran Ramanathan, Timothy J. Kamp, Igor Slukvin, James A. Thomson, James R. Dutton, Aseem Z. Ansari

Genetics, Cell Biology and Development (TMED)

Research output: Contribution to journal › Article › peer-review

24 Scopus citations

Abstract

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.

Original language	English (US)
Pages (from-to)	E8257-E8266
Journal	Proceedings of the National Academy of Sciences of the United States of America
Volume	113
Issue number	51
DOIs	https://doi.org/10.1073/pnas.1611142114
State	Published - Dec 20 2016

Bibliographical note

Funding Information:
We thank Sandra Tseng and Graham Erwin for help with ChIP-seq analysis. We thank José Rodríguez-Martínez for CSI analysis and Christina Shafer with RNA-seq analysis. We also acknowledge Mitchell Probasco with help with flow cytometry, Jennifer Bolin for help with Illumina sequencing, and Bret Duffin for the teratoma assay. We thank Laura Vanderploeg for help with figure graphics. We are also grateful to Judith Kimble, Sushmita Roy, Garrett Lee, and Fang Wan for helpful discussions. This work was supported by the NIH Grant HL099773, W. M. Keck Medical Research Award, and Progenitor Cell Biology Consortium Jump-Start Award 5U01HL099997-05 (Subaward 101330A). A.E. was supported by the Morgridge Biotechnology Wisconsin Distinguished Fellowship Award and the Stem Cell and Regenerative Medicine Training Award. A.S.K. was supported by Genomic Sciences Training Program Grant 5T32HG002760. D.B. was supported by a National Science Foundation-Nanoscale Science and Engineering Center grant.

Publisher Copyright:
© 2016, National Academy of Sciences. All rights reserved.

Keywords

Artificial transcription factor
Cell fate
Gene regulatory networks
Genome-scale library
Reprogramming

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Eguchi, A., Wleklinski, M. J., Spurgat, M. C., Heiderscheit, E. A., Kropornicka, A. S., Vu, C. K., Bhimsaria, D., Swanson, S. A., Stewart, R., Ramanathan, P., Kamp, T. J., Slukvin, I., Thomson, J. A., Dutton, J. R., & Ansari, A. Z. (2016). Reprogramming cell fate with a genome-scale library of artificial transcription factors. Proceedings of the National Academy of Sciences of the United States of America, 113(51), E8257-E8266. https://doi.org/10.1073/pnas.1611142114

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 2016, 'Reprogramming cell fate with a genome-scale library of artificial transcription factors', Proceedings of the National Academy of Sciences of the United States of America, vol. 113, no. 51, pp. E8257-E8266. https://doi.org/10.1073/pnas.1611142114

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

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Keywords

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