TDP-43 subcellular mislocalisation is correlated with loss of optineurin binding for frontotemporal dementia and amyotrophic lateral sclerosis associated TBK1 missense variants

BackgroundFrontotemporal dementia (FTD) is one of the most common forms of younger-onset dementia. FTD is genetically, pathologically and clinically related to amyotrophic lateral sclerosis (ALS), a rapidly progressive neurodegenerative disorder. Mutations in TANK-binding kinase 1 (TBK1) have been identified as a rare cause of FTD and ALS. TBK1 has known roles in inflammation and autophagy and interacts with other FTD and ALS proteins such as optineurin (OPTN): however, which of its roles are important to FTD/ALS pathogenesis remains undetermined. To date, >90 TBK1 rare variants have been identified in FTD/ALS patients: >50% of these are missense variants of unknown significance (VUS). MethodsIn this study, we have used a functional assay pipeline to investigate the effect of 16 TBK1 VUS with in-silico evidence of pathogenicity, together with two known pathogenic mutations and a common benign TBK1 polymorphism. Our assay pipeline evaluated the effect of TBK1 VUS on steady-state levels of TBK1, kinase activity and binding to OPTN. We also assessed the impact of TBK1 VUS on a key neuropathological feature of FTD and ALS cases: mislocalisation of neuronal TDP-43 from the nucleus to the cytoplasm. ResultsWe observed some TBK1 VUS that had similar effects to TBK1 loss-of-function mutations, demonstrating decreased kinase activity and loss of OPTN binding. Both known pathogenic mutations and several TBK1 VUS also increased the cytoplasmic/nuclear ratio of TDP-43 and this inversely correlated with their degree of OPTN binding but not with kinase activity. ConclusionsThese results suggest that loss of the direct interaction between TBK1 and OPTN is more critical to FTD and ALS pathogenesis than TBK1s kinase activity. However, further studies are needed to elucidate exactly how loss of TBK1 binding to OPTN leads to TDP-43 pathology and ultimately neurodegeneration.