RNANEUROTOX

Small RNA toxicity and DNA damage response in Huntington´s disease

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 Obiettivo del progetto (Objective)

'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.'

Introduzione (Teaser)

A European consortium set out to dissect the molecular aetiology of Huntington's disease (HD). Their findings should lead to more specific and targeted therapies.

Descrizione progetto (Article)

HD is a genetic disorder associated with neuronal motor dysfunction. It is caused by a trinucleotide repeat expansion in the huntingtin gene, which leads to the production of a protein that is toxic to neuronal cells. The mutant product also induces protein aggregation and an abnormal DNA damage response (DDR).

Emerging evidence indicates that the trinucleotide expansion also produces small RNA molecules that are toxic to neuronal cells. However, the precise mechanism behind this RNA-mediated toxicity is not known.

Given the implication of the DDR in HD pathogenesis, the EU-funded 'Small RNA toxicity and DNA damage response in Huntington's disease' (RNANEUROTOX) project worked under the hypothesis that small RNAs somehow activate the DDR, causing severe neurotoxicity. The key objective was to decipher the link between the two.

Study activities were designed to provide evidence that mutant huntingtin RNA products induce a DDR in cultured cells. For this purpose, researchers obtained neuronal cells and treated them with the mutant huntingtin RNA. Microarray analysis of gene expression changes was focused on genes that are implicated in the DDR as well as on genes previously reported to be down-regulated in affected brain areas of HD patients. To further validate the association of RNA toxicity with induction of DNA damage, scientists inhibited DDR.

Collectively, the results of the RNANEUROTOX research project will contribute to a better understanding of the molecular mechanisms underlying HD. The generated knowledge could be translated to other disorders and lead to novel therapeutic interventions.