Cytoplasmic Redistribution of Mutant TDP-43 in a BAC Based Mouse Model of Amyotrophic Lateral Sclerosis (P3.012)

OBJECTIVE:To create a novel model of ALS pathogenesis based on physiological expression of mutant TDP-43 BACKGROUND:Mutations in TAR DNA-binding protein 43 (TDP-43) are associated with ALS and frontotemporal dementia (FTD). Conventional TDP-43 cDNA overexpression models driven by ubiquitous or neuronally expressed promoters display highly variable disease phenotypes. To more accurately model ALS, we have generated transgenic mice using BAC constructs expressing human wild-type (WT) or mutant (M337V) TDP-43 under the control of the endogenous human promoter. DESIGN/METHODS:Two BAC vectors containing the full length human genomic locus of the WT and M337V mutation were generated. Embryonic stem cells (ESCs) were transfected with purified BAC constructs using PhiC31 integrase-mediated cassette exchange to selectively target insertion to the ROSA26 locus. Candidate ESCs were identified by PCR screening and immunoblotting to confirm transgene insertion and expression. TDP-43-Ypet-positive ESC clonal lines were used to generate chimeric mice by pronuclear injection. Retention of the M337V mutation was confirmed by sequencing DNA from chimeric mice. Founder (F1) mice were produced by crossing chimeric males and female C57Bl/6J. Mice were subjected to behavioural phenotyping, RNA expression analysis in the spinal cord, and systematic neuropathology at 3,6,9 and 12 months. Embryonic motor neurons were cultured from E13 spinal cord and analysed for TDP-43 mislocalisation. RESULTS:We have confirmed that a single BAC copy of the TDP-43-WT-Ypet and TDP-43-M337V-Ypet can successfully drive physiologically-relevant levels of expression in the transgenic mice by immunoblotting. TDP-43 harboring the M337V mutation is mislocalised to the cytoplasm in resting motor neurons. Cellular stress experiments indicate that mutant TDP-43 may inhibit the formation of stress granules. CONCLUSIONS:In this physiologically relevant mouse model of ALS pathogenesis cellular redistribution of TDP-43 occurs in a manner in keeping with the pathological signature seen in human autropy cases of ALS. Embryonic motor neuron culture from this model is a potential tool both to explore the pathogenesis and to screen neuroprotective drugs. Study Supported by:Motor Neurone Disease Association (UK) Disclosure: Dr. Talbot has received personal compensation for activities with Biogen Idec, Avanir Pharmaceuticals, and UCB Pharma. Dr. Gordon has nothing to disclose. Dr. Mutihac has nothing to disclose. Dr. Alegre-Abarrategui has nothing to disclose. Dr. Farrimond has nothing to disclose. Dr. Davies has nothing to disclose. Dr. Wade-Martins has nothing to disclose.