PUBLICATION
Transient, flexible gene editing in zebrafish neutrophils and macrophages for determination of cell-autonomous functions
- Authors
- Isiaku, A.I., Zhang, Z., Pazhakh, V., Manley, H.R., Thompson, E.R., Fox, L.C., Yerneni, S., Blombery, P., Lieschke, G.J.
- ID
- ZDB-PUB-210729-4
- Date
- 2021
- Source
- Disease models & mechanisms 14(7): (Journal)
- Registered Authors
- Manley, Harriet, Pazhakh, Vahid
- Keywords
- CRISPR-Cas9, Cell autonomy, Gene editing, Macrophages, Neutrophils, Zebrafish
- MeSH Terms
-
- Animals
- Animals, Genetically Modified
- CRISPR-Cas Systems/genetics
- Gene Editing*
- Humans
- Macrophages/metabolism
- Neutrophils/metabolism
- Transcription Factors/metabolism
- Zebrafish*/genetics
- Zebrafish*/metabolism
- PubMed
- 34296745 Full text @ Dis. Model. Mech.
Citation
Isiaku, A.I., Zhang, Z., Pazhakh, V., Manley, H.R., Thompson, E.R., Fox, L.C., Yerneni, S., Blombery, P., Lieschke, G.J. (2021) Transient, flexible gene editing in zebrafish neutrophils and macrophages for determination of cell-autonomous functions. Disease models & mechanisms. 14(7):.
Abstract
Zebrafish are an important model for studying phagocyte function, but rigorous experimental systems to distinguish whether phagocyte-dependent effects are neutrophil or macrophage specific have been lacking. We have developed and validated transgenic lines that enable superior demonstration of cell-autonomous neutrophil and macrophage genetic requirements. We coupled well-characterized neutrophil- and macrophage-specific Gal4 driver lines with UAS:Cas9 transgenes for selective expression of Cas9 in either neutrophils or macrophages. Efficient gene editing, confirmed by both Sanger and next-generation sequencing, occurred in both lineages following microinjection of efficacious synthetic guide RNAs into zebrafish embryos. In proof-of-principle experiments, we demonstrated molecular and/or functional evidence of on-target gene editing for several genes (mCherry, lamin B receptor, trim33) in either neutrophils or macrophages as intended. These new UAS:Cas9 tools provide an improved resource for assessing individual contributions of neutrophil- and macrophage-expressed genes to the many physiological processes and diseases modelled in zebrafish. Furthermore, this gene-editing functionality can be exploited in any cell lineage for which a lineage-specific Gal4 driver is available. This article has an associated First Person interview with the first author of the paper.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping