Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are two clinically different neurodegenerative diseases that lie on a pathogenic spectrum. The most common pathology present in ALS and FTD cases is cytoplasmic mislocalisation and aggregation of the RNA-binding protein TDP-43.
This PhD aims to dissect the contributions of RNA-binding and liquid-liquid phase separation to TDP-43 toxicity using novel iPSC models of ALS/FTD. This includes rational design of mutations in the TDP-43 gene that disrupt RNA binding while conserving the domain structures, as well as the generation of phase separation deficient mutants. The generated mutants will then be knocked into human induced pluripotent stem cell (iPSC) lines using CRISPR/Cas9. Resultant iPSCs will be differentiated into neurons and the effect on neuronal health (longitudinal dendritic branching and survival) and TDP-43 functionality (nucleocytoplasmic transport, phase separation, stress granule association and RNA metabolism) assessed. This will give insights into the molecular basis of TDP-43 toxicity, inform on the initiating factors in disease pathogenesis, and provide potential targets for therapeutic intervention.
This project combines the TDP-43 and microscopy (including super-resolution) expertise of the Mizielinska group with the RNA, genome editing and iPSC expertise of the Ruepp group. This will provide the student with a wealth of skills in stem cell culture, genome editing, molecular biology, biochemistry, and microscopy.
Year 1: Design and generation of TDP-43 mutants and characterisation in vitro
Year 2: Genome editing of iPSCs, neuronal differentiation and characterisation of generated lines
Year 3+: Study of toxicity and TDP-43 biology-based phenotypes in mutant neurons