CRISPR-mediated direct mutation of cancer genes in the mouse liver
Joshi, Nikhil S.
Crowley, Denise G.
Sharp, Phillip A.
Jacks, TylerNote: Order does not necessarily reflect citation order of authors.
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CitationXue, W., S. Chen, H. Yin, T. Tammela, T. Papagiannakopoulos, N. S. Joshi, W. Cai, et al. 2014. “CRISPR-mediated direct mutation of cancer genes in the mouse liver.” Nature 514 (7522): 380-384. doi:10.1038/nature13589. http://dx.doi.org/10.1038/nature13589.
AbstractThe study of cancer genes in mouse models has traditionally relied on genetically-engineered strains made via transgenesis or gene targeting in embryonic stem (ES) cells1. Here we describe a new method of cancer model generation using the CRISPR/Cas system in vivo in wild-type mice. We have used hydrodynamic injection to deliver a CRISPR plasmid DNA expressing Cas9 and single guide RNAs (sgRNAs)2–4 to the liver and directly target the tumor suppressor genes Pten5 and p536, alone and in combination. CRISPR-mediated Pten mutation led to elevated Akt phosphorylation and lipid accumulation in hepatocytes, phenocopying the effects of deletion of the gene using Cre-LoxP technology7, 8. Simultaneous targeting of Pten and p53 induced liver tumors that mimicked those caused by Cre-loxP-mediated deletion of Pten and p53. DNA sequencing of liver and tumor tissue revealed insertion or deletion (indel) mutations of the tumor suppressor genes, including bi-allelic mutations of both Pten and p53 in tumors. Furthermore, co-injection of Cas9 plasmids harboring sgRNAs targeting the β-Catenin gene (Ctnnb1) and a single-stranded DNA (ssDNA) oligonucleotide donor carrying activating point mutations led to the generation of hepatocytes with nuclear localization of β-Catenin. This study demonstrates the feasibility of direct mutation of tumor suppressor genes and oncogenes in the liver using the CRISPR/Cas system, which presents a new avenue for rapid development of liver cancer models and functional genomics.
Citable link to this pagehttp://nrs.harvard.edu/urn-3:HUL.InstRepos:15034908
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