An ATF3-CreERT2 Knock-In Mouse for Axotomy-Induced Genetic Editing: Proof of Principle.

Authors:
Leanne M Ramer
Leanne M Ramer
The University of British Columbia
Canada
Stephen B McMahon
Stephen B McMahon
King's College London
United Kingdom
Franziska Denk
Franziska Denk
King's College London
United Kingdom
Matt S Ramer
Matt S Ramer
King's College London

eNeuro 2019 Mar-Apr;6(2). Epub 2019 Apr 9.

International Collaboration on Repair Discoveries, the University of British Columbia, Vancouver, British Columbia, Canada V5Z 1M9.

Genome editing techniques have facilitated significant advances in our understanding of fundamental biological processes, and the Cre-Lox system has been instrumental in these achievements. Driving Cre expression specifically in injured neurons has not been previously possible: we sought to address this limitation in mice using a Cre-ERT2 construct driven by a reliable indicator of axotomy, activating transcription factor 3 (ATF3). When crossed with reporter mice, a significant amount of recombination was achieved (without tamoxifen treatment) in peripherally-projecting sensory, sympathetic, and motoneurons after peripheral nerve crush in hemizygotes (65-80% by 16 d) and was absent in uninjured neurons. Importantly, injury-induced recombination did not occur in Schwann cells distal to the injury, and with a knock-out-validated antibody we verified an absence of ATF3 expression. Functional recovery following sciatic nerve crush in ATF3-deficient mice (both hemizygotes and homozygotes) was delayed, indicating previously unreported haploinsufficiency. In a proof-of-principle experiment, we crossed the ATF3-CreERT2 line with a floxed phosphatase and tensin homolog (PTEN) line and show significantly improved axonal regeneration, as well as more complete recovery of neuromuscular function. We also demonstrate the utility of the ATF3-CreERT2 hemizygous line by characterizing recombination after lateral spinal hemisection (C8/T1), which identified specific populations of ascending spinal cord neurons (including putative spinothalamic and spinocerebellar) and descending supraspinal neurons (rubrospinal, vestibulospinal, reticulospinal and hypothalamic). We anticipate these mice will be valuable in distinguishing axotomized from uninjured neurons of several different classes (e.g., via reporter expression), and in probing the function of any number of genes as they relate to neuronal injury and regeneration.

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Source
http://eneuro.org/lookup/doi/10.1523/ENEURO.0025-19.2019
Publisher Site
http://dx.doi.org/10.1523/ENEURO.0025-19.2019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6464513PMC
April 2019
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