Huntington´s disease (HD) is an incurable and fatal neurodegenerative disease, manifested by a progressive loss of neurons, causing neuronal motor dysfunction, cognitive decline and death. The disease is caused by a CAG repeat expansion in the Huntingtin (HTT) gene. The CAG expansion induces a neurotoxic effect, mainly thought to be caused by the expanded protein. However, also expanded RNA transcripts can cause toxicity by generation of small CAG-repeated RNAs. Generation of these small RNAs depends on Dicer ribonuclease activity and their toxic effects are modulated by Ago proteins. This links RNA toxicity with the RNA interference (RNAi) pathway, suggesting abnormal induction of gene silencing through small-repeated RNAs.The DNA damage response (DDR) plays a role in HD pathogenesis as well. Recently, it was shown that small RNAs derived from Dicer and Drosha processing can induce and control this response. The goal of this project is to address the possible link between small CAG-repeated RNA toxicity and the DDR pathway in HD.We hypothesize that small RNAs activate a DDR response and thereby induce neurotoxicity. We will address this by the following objectives: 1) Establish whether CAG-repeated RNAs activate the DDR pathway, 2) Establish whether DDR inhibition can modulate small RNA toxicity and whether DDR activation is RNAi pathway dependent, 3) Validation of our results using in vivo and ex vivo samples. We will use in vitro cell models combined with ex vivo brain samples from HD affected mice and humans. The effect of small CAG-repeated RNAs on activation of the DDR response will be analyzed by western blot, immuno-fluorescence and immuno-histochemistry. Small RNA pool analysis, knockdown studies and lentiviral transfection approaches will be used to reveal the specificity of the effect and the RNAi pathway dependence.This project will contribute to a better understanding of HD pathogenesis, RNA biology, small RNA toxicity and the DDR pathway.