Format

Send to

Choose Destination
Cell Rep. 2019 Apr 16;27(3):708-718.e10. doi: 10.1016/j.celrep.2019.03.076.

Single-Cell RNA-Sequencing-Based CRISPRi Screening Resolves Molecular Drivers of Early Human Endoderm Development.

Author information

1
Program in Molecular Medicine, Diabetes Center of Excellence, University of Massachusetts Medical School, Worcester, MA 01605, USA.
2
Department of Translational Psychiatry, Max Planck Institute of Psychiatry, 80804 Munich, Germany.
3
Program in Molecular Medicine, Diabetes Center of Excellence, University of Massachusetts Medical School, Worcester, MA 01605, USA. Electronic address: rene.maehr@umassmed.edu.

Abstract

Studies in vertebrates have outlined conserved molecular control of definitive endoderm (END) development. However, recent work also shows that key molecular aspects of human END regulation differ even from rodents. Differentiation of human embryonic stem cells (ESCs) to END offers a tractable system to study the molecular basis of normal and defective human-specific END development. Here, we interrogated dynamics in chromatin accessibility during differentiation of ESCs to END, predicting DNA-binding proteins that may drive this cell fate transition. We then combined single-cell RNA-seq with parallel CRISPR perturbations to comprehensively define the loss-of-function phenotype of those factors in END development. Following a few candidates, we revealed distinct impairments in the differentiation trajectories for mediators of TGFβ signaling and expose a role for the FOXA2 transcription factor in priming human END competence for human foregut and hepatic END specification. Together, this single-cell functional genomics study provides high-resolution insight on human END development.

KEYWORDS:

CRISPRi; chromatin accessibility; dCas9-KRAB; endoderm; hepatic endoderm; human development; perturbation screen; pluripotent stem cells; single-cell RNA-seq; stem cell differentiation

Supplemental Content

Full text links

Icon for Elsevier Science Icon for PubMed Central
Loading ...
Support Center