ABSTRACT
DNA damage repair genes are modifiers of disease onset in Huntington's disease (HD), but how this process intersects with associated disease pathways remains unclear. Here we evaluated the mechanistic contributions of protein inhibitor of activated STAT-1 (PIAS1) in HD mice and HD patient-derived induced pluripotent stem cells (iPSCs) and find a link between PIAS1 and DNA damage repair pathways. We show that PIAS1 is a component of the transcription-coupled repair complex, that includes the DNA damage end processing enzyme polynucleotide kinase-phosphatase (PNKP), and that PIAS1 is a SUMO E3 ligase for PNKP. Pias1 knockdown (KD) in HD mice had a normalizing effect on HD transcriptional dysregulation associated with synaptic function and disease-associated transcriptional coexpression modules enriched for DNA damage repair mechanisms as did reduction of PIAS1 in HD iPSC-derived neurons. KD also restored mutant HTT-perturbed enzymatic activity of PNKP and modulated genomic integrity of several transcriptionally normalized genes. The findings here now link SUMO modifying machinery to DNA damage repair responses and transcriptional modulation in neurodegenerative disease.
Subject(s)
DNA Repair Enzymes/genetics , DNA Repair , DNA/genetics , Huntingtin Protein/genetics , Huntington Disease/genetics , Phosphotransferases (Alcohol Group Acceptor)/genetics , Protein Inhibitors of Activated STAT/genetics , Protein Processing, Post-Translational , Small Ubiquitin-Related Modifier Proteins/genetics , Animals , Cell Differentiation , DNA/metabolism , DNA Damage , DNA Repair Enzymes/metabolism , Disease Models, Animal , Female , Humans , Huntingtin Protein/metabolism , Huntington Disease/metabolism , Huntington Disease/pathology , Male , Mice , Mice, Inbred C57BL , Mice, Transgenic , Neurons/metabolism , Neurons/pathology , Phosphotransferases (Alcohol Group Acceptor)/metabolism , Pluripotent Stem Cells/metabolism , Pluripotent Stem Cells/pathology , Primary Cell Culture , Protein Inhibitors of Activated STAT/antagonists & inhibitors , Protein Inhibitors of Activated STAT/metabolism , RNA, Small Interfering/genetics , RNA, Small Interfering/metabolism , Small Ubiquitin-Related Modifier Proteins/antagonists & inhibitors , Small Ubiquitin-Related Modifier Proteins/metabolism , Sumoylation , Transcription, GeneticABSTRACT
Polyglutamine (polyQ) repeat diseases are a class of neurodegenerative disorders caused by CAG-repeat expansion. There are diverse cellular mechanisms behind the pathogenesis of polyQ disorders, including transcriptional dysregulation. Interestingly, we find that levels of the long isoform of nuclear paraspeckle assembly transcript 1 (Neat1L) are elevated in the brains of mouse models of spinocerebellar ataxia types 1, 2, 7 and Huntington's disease (HD). Neat1L was also elevated in differentiated striatal neurons derived from HD knock-in mice and in HD patient brains. The elevation was mutant Huntingtin (mHTT) dependent, as knockdown of mHTT in vitro and in vivo restored Neat1L to normal levels. In additional studies, we found that Neat1L is repressed by methyl CpG binding protein 2 (MeCP2) by RNA-protein interaction but not by occupancy of MeCP2 at its promoter. We also found that NEAT1L overexpression protects from mHTT-induced cytotoxicity, while reducing it enhanced mHTT-dependent toxicity. Gene set enrichment analysis of previously published RNA sequencing data from mouse embryonic fibroblasts and cells derived from HD patients shows that loss of NEAT1L impairs multiple cellular functions, including pathways involved in cell proliferation and development. Intriguingly, the genes dysregulated in HD human brain samples overlap with pathways affected by a reduction in NEAT1, confirming the correlation of NEAT1L and HD-induced perturbations. Cumulatively, the role of NEAT1L in polyQ disease model systems and human tissues suggests that it may play a protective role in CAG-repeat expansion diseases.
Subject(s)
Huntington Disease/genetics , Methyl-CpG-Binding Protein 2/genetics , RNA, Long Noncoding/genetics , Spinocerebellar Ataxias/genetics , Alternative Splicing/genetics , Animals , Cell Differentiation/genetics , Corpus Striatum/metabolism , Corpus Striatum/pathology , Gene Knock-In Techniques , Humans , Huntingtin Protein/genetics , Huntington Disease/physiopathology , Mice , Neurons/metabolism , Neurons/pathology , Peptides/genetics , Promoter Regions, Genetic , Protein Isoforms/genetics , RNA-Binding Proteins/genetics , Spinocerebellar Ataxias/physiopathology , Trinucleotide Repeat Expansion/geneticsABSTRACT
OBJECTIVE: Spinocerebellar ataxia type 1 is an autosomal dominant fatal neurodegenerative disease caused by a polyglutamine expansion in the coding region of ATXN1. We showed previously that partial suppression of mutant ataxin-1 (ATXN1) expression, using virally expressed RNAi triggers, could prevent disease symptoms in a transgenic mouse model and a knockin mouse model of the disease, using a single dose of virus. Here, we set out to test whether RNAi triggers targeting ATXN1 could not only prevent, but also reverse disease readouts when delivered after symptom onset. METHODS: We administered recombinant adeno-associated virus (rAAV) expressing miS1, an artificial miRNA targeting human ATXN1 mRNA (rAAV.miS1), to a mouse model of spinocerebellar ataxia type 1 (SCA1; B05 mice). Viruses were delivered prior to or after symptom onset at multiple doses. Control B05 mice were treated with rAAVs expressing a control artificial miRNA, or with saline. Animal behavior, molecular phenotypes, neuropathology, and magnetic resonance spectroscopy were done on all groups, and data were compared to wild-type littermates. RESULTS: We found that SCA1 phenotypes could be reversed by partial suppression of human mutant ATXN1 mRNA by rAAV.miS1 when delivered after symptom onset. We also identified the therapeutic range of rAAV.miS1 that could prevent or reverse disease readouts. INTERPRETATION: SCA1 disease may be reversible by RNAi therapy, and the doses required for advancing this therapy to humans are delineated. Ann Neurol 2016;80:754-765.
