An Inducible Lentiviral Guide RNA Platform Enables the Identification of Tumor-Essential Genes and Tumor-Promoting Mutations InVivo

BJ Aubrey; GL Kelly; AJ Kueh; MS Brennan; L O'Connor; L Milla; S Wilcox; L Tai; A Strasser; MJ Herold

Journal article

An Inducible Lentiviral Guide RNA Platform Enables the Identification of Tumor-Essential Genes and Tumor-Promoting Mutations InVivo

BJ Aubrey, GL Kelly, AJ Kueh, MS Brennan, L O'Connor, L Milla, S Wilcox, L Tai, A Strasser, MJ Herold

Cell Reports | Published : 2015

DOI: 10.1016/j.celrep.2015.02.002

Abstract

The CRISPR/Cas9 technology enables the introduction of genomic alterations into almost any organism; however, systems for efficient and inducible gene modification have been lacking, especially for deletion of essential genes. Here, we describe a drug-inducible small guide RNA (sgRNA) vector system allowing for ubiquitous and efficient gene deletion in murine and human cells. This system mediates the efficient, temporally controlled deletion of MCL-1, both invitro and invivo, in human Burkitt lymphoma cell lines that require this anti-apoptotic BCL-2 protein for sustained survival and growth. Unexpectedly, repeated induction of the same sgRNA generated similar inactivating mutations in the h..

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University of Melbourne Researchers

Gemma Kelly Author

Andreas Strasser Author

Marco Herold Author

Grants

Awarded by Leukemia and Lymphoma Society

Funding Acknowledgements

We thank all members of the M.J.H. laboratory for their support and advice; D. Gray for critical reading of the manuscript; G. Siciliano, C. Gatt, and their team for animal husbandry; S. Monard and his team for help with the flow cytometry unit; A.B. Rickinson and M. Rowe (University of Birmingham) for the kind gift of BL cell lines; and K. Rogers and the WEHI imaging facility. This work was supported by a Leukemia Foundation National Research Program Clinical PhD Scholarship (to B.J.A.), a Kay Kendall Leukemia Fund Intermediate Fellowship (KKL331 to G.L.K.); the National Health and Medical Research Council, Australia program grant 1016701 and fellowship 1020363, Leukemia and Lymphoma Society SCOR grant 7001-13 (to A.S.), and project grant GNT1049720 (to M.J.H.). This work was made possible through Victorian State Government Operational Infrastructure Support and Australian Government National Health and Medical Research Council Independent Research Institutes Infrastructure Support Scheme.