The aim of this project is to investigate early steps in the pathogenesis of SPG4 and SPG31 in a neuronal cell culture model. Using CRISPR/Cas9 technology, we will generate both SPAST and REEP1 knockout lines from isogenic controls. In addition, iPSCs from SPG4- and SPG31-patients will be used, which will then be differentiated into human neurons. By comparative transcriptome analysis of knockout cells with their isogenic controls aberrant metabolic pathways will be identified and their pathophysiological relevance will further be validated by comparison with patient lines. Suitable candidates identified by transcriptome analysis will then examined by mass spectrometry in biomaterials of patients for corresponding changes in the proteome, which can serve as biomarkers for future therapy studies. The most promising metabolic pathway will be selected for pharmacological or genetic interventions and the therapeutic effect in human neurons will be functionally validated.