Endogenous TDP-43 mislocalization in a novel knock-in mouse model reveals DNA repair impairment, inflammation, and neuronal senescence

Abstract

Abstract TDP-43 mislocalization and aggregation are key pathological features of motor neuron diseases (MND) such as amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). However, existing overexpression animal models typically capture late-stage TDP-43 proteinopathies, leaving a gap in our understanding of early motor neuron-specific disease mechanisms during pre-symptomatic phases. We address this by generating a new endogenous knock-in (KI) mouse model using a combination of CRISPR/Cas9 and FLEX Cre-switch strategy for the conditional expression of a mislocalized Tdp-43∆NLS variant of mouse Tdp-43. This variant is either expressed conditionally in whole mice or specifically within the motor neurons. The mice exhibit loss of nuclear Tdp-43 with its concomitant cytosolic accumulation and aggregation in targeted cells, leading to increased DNA double-strand breaks (DSB), signs of inflammation and DNA damage-associated cellular senescence. Notably, unlike WT Tdp43 which functionally interacts with Xrcc4 and DNA Ligase 4, key DSB repair proteins in the non-homologous end-joining pathway, the Tdp-43∆NLS mutant sequesters them into cytosolic aggregates, exacerbating neuronal damage in mice brain. The mutant mice also exhibit myogenic degeneration in limb muscles and distinct motor deficits, consistent with the characteristics of MND. Our findings reveal progressive degenerative mechanisms in motor neurons expressing endogenous Tdp-43∆NLS mutant, independent of TDP-43 overexpression or other confounding etiological factors. Thus, this unique Tdp-43 KI mouse model, which displays key molecular and phenotypic features of Tdp-43 proteinopathy, offers a significant opportunity to further characterize the early-stage progression of MND and also opens avenues for developing DNA repair-targeted approaches for treating TDP-43 pathology-linked neurodegenerative diseases.