RNA-DNA hybrids formed in the absence of Senataxin drive non-coding RNA expression and protein aggregation in the nucleolus

Neurodegeneration in the human population is often associated with loss of protein homeostasis that is driven by accumulation of specific aggregated proteins. Here we investigate whether deficiency in Senataxin, an RNA-DNA helicase associated with cerebellar ataxia and ALS, induces destabilization of intrinsically disordered proteins as we previously found with Ataxia-telangiectasia (A-T) cells and tissue. We find that Senataxin loss does generate protein aggregates but, unlike in A-T, these are associated with the nucleolus and are independent of oxidative stress and PARP activity. Non-coding RNA expression from the intergenic spacer region of ribosomal DNA is induced in the absence of Senataxin and drives the association and aggregation of many proteins known to be prone to aggregation in neurodegenerative disease. Both non-coding RNAs and protein aggregates are eliminated by overexpression of RNaseH1, implicating RNA-DNA hybrids as the key driver of these outcomes. We find that hybrids are high in the absence of Senataxin at intergenic sites, not at annotated protein-coding genes; these include sites of Senataxin binding in the genome, ribosomal DNA, and peri-centromeric regions. These findings suggest that destabilization of the proteome is driven by Senataxin loss and that RNA-DNA hybrids and non-coding RNAs play a critical role in this process.