CRISPR/Cas9-engineered human ES cells harboring heterozygous and homozygous c-KIT knockout

Marcelo A S de Toledo; Xuhuang Fu; Frederick Kluge; Katrin Götz; Susanne Schmitz; Paul Wanek; Herdit M Schüler; Kristina Pannen; Nicolas Chatain; Steffen Koschmieder; Tim H Brümmendorf; Martin Zenke

doi:10.1016/j.scr.2022.102732

CRISPR/Cas9-engineered human ES cells harboring heterozygous and homozygous c-KIT knockout

Stem Cell Res. 2022 Apr:60:102732. doi: 10.1016/j.scr.2022.102732. Epub 2022 Mar 1.

Authors

Affiliations

¹ Institute for Biomedical Engineering, Department of Cell Biology, RWTH Aachen University Medical School, Aachen, Germany; Helmholtz Institute for Biomedical Engineering, RWTH Aachen University, Aachen, Germany; Department of Hematology, Oncology, Hemostaseology and Stem Cell Transplantation, RWTH Aachen University Hospital, Aachen, Germany; Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf (CIO ABCD), Aachen, Germany.
² Institute for Biomedical Engineering, Department of Cell Biology, RWTH Aachen University Medical School, Aachen, Germany; Helmholtz Institute for Biomedical Engineering, RWTH Aachen University, Aachen, Germany.
³ Institute for Human Genetics, RWTH Aachen University Hospital, Aachen, Germany.
⁴ Department of Hematology, Oncology, Hemostaseology and Stem Cell Transplantation, RWTH Aachen University Hospital, Aachen, Germany; Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf (CIO ABCD), Aachen, Germany.
⁵ Institute for Biomedical Engineering, Department of Cell Biology, RWTH Aachen University Medical School, Aachen, Germany; Helmholtz Institute for Biomedical Engineering, RWTH Aachen University, Aachen, Germany. Electronic address: martin.zenke@rwth-aachen.de.

PMID: 35279545
DOI: 10.1016/j.scr.2022.102732

Abstract

The receptor tyrosine kinase c-KIT (CD117) has a key role in hematopoiesis and is a marker for endothelial and cardiac progenitor cells. In vivo, deficiency of c-KIT is lethal and therefore using CRISPR/Cas9 editing we generated heterozygous and homozygous c-KIT knockout human embryonic stem cell (ES cell) lines. The c-KIT knockout left ES cell pluripotency unaffected as shown by immunofluorescence and trilineage differentiation potential. Heterozygous and homozygous c-KIT knockouts showed complete loss of exon 17, resulting in ablation of c-KIT protein from the cell surface. c-KIT knockout ES cells provide a valuable tool for further investigating c-KIT biology.

Keywords: CD117; CRISPR; ES cells; Embryonic stem cells; Stem cell factor receptor; c-KIT.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

CRISPR-Cas Systems / genetics
Cell Line
Heterozygote
Homozygote
Human Embryonic Stem Cells* / metabolism
Humans