Mitochondrial dysfunction is associated with neuronal loss in Huntington’s disease (HD), a neurodegenerative disease caused by an abnormal polyglutamine expansion in huntingtin (Htt). However, the mechanisms linking mutant Htt and mitochondrial dysfunction in HD remain unknown. We identify an interaction between mutant Htt and the TIM23 mitochondrial protein import complex. Remarkably, recombinant mutant Htt directly inhibited mitochondrial protein import in vitro. Furthermore, mitochondria from brain synaptosomes of presymptomatic HD model mice and from mutant Htt-expressing primary neurons exhibited a protein import defect, suggesting that deficient protein import is an early event in HD. The mutant Htt-induced mitochondrial import defect and subsequent neuronal death were attenuated by overexpression of TIM23 complex subunits, demonstrating that deficient mitochondrial protein import causes mutant Htt-induced neuronal death. Collectively, these findings provide evidence for a direct link between mutant Htt, mitochondrial dysfunction and neuronal pathology, with implications for mitochondrial protein import-based therapies in HD.

SUMMARY Glioblastoma harbors a dynamic subpopulation of glioblastoma stem-like cells (GSCs) that can propagate tumors in vivo and is resistant to standard chemoradiation. Identification of the cell-intrinsic mechanisms governing this clinically important cell state may lead to the discovery of novel therapeutic strategies for this challenging malignancy. Here, we demonstrate that the mitotic E3 ubiquitin ligase CDC20-Anaphase-Promoting Complex (CDC20-APC) drives invasiveness and self-renewal in patient tumor-derived GSCs. Moreover, CDC20 knockdown inhibited and CDC20 overexpression increased the ability of human GSCs to generate brain tumors in an orthotopic xenograft model in vivo. CDC20-APC control of GSC invasion and self-renewal operates through pluripotency-related transcription factor SOX2. Our results identify a CDC20-APC/SOX2 signaling axis that controls key biological properties of GSCs, with implications for CDC20-APC-targeted strategies in the treatment of glioblastoma.

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Although epigenetic abnormalities have been described in Huntington’s disease (HD), the causal epigenetic mechanisms driving neurodegeneration in HD cortex and striatum remain undefined. Using an epigenetic pathway-targeted drug screen, we report that inhibitors of DNA methyltransferases (DNMTs), decitabine and FdCyd, block mutant huntingtin (Htt)-induced toxicity in primary cortical and striatal neurons. In addition, knockdown of DNMT3A or DNMT1 protected neurons against mutant Htt-induced toxicity, together demonstrating a requirement for DNMTs in mutant Htt-triggered neuronal death and suggesting a neurodegenerative mechanism based on DNA methylation-mediated transcriptional repression. Inhibition of DNMTs in HD model primary cortical or striatal neurons restored the expression of several key genes, including Bdnf, an important neurotrophic factor implicated in HD. Accordingly, the Bdnf promoter exhibited aberrant cytosine methylation in mutant Htt-expressing cortical neurons. In vivo, pharmacological inhibition of DNMTs in HD mouse brains restored the mRNA levels of key striatal genes known to be downregulated in HD. Thus, disturbances in DNA methylation play a critical role in mutant Htt-induced neuronal dysfunction and death, raising the possibility that epigenetic strategies targeting abnormal DNA methylation may have therapeutic utility in HD.

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Huntington's disease (HD) is a fatal neurodegenerative disorder caused by an abnormal expansion of polyglutamine repeats in the huntingtin protein (Htt). Transcriptional dysregulation is an early event in the course of HD progression and is thought to contribute to disease pathogenesis, but how mutant Htt causes transcriptional alterations and subsequent cell death in neurons is not well understood. RNA-Seq analysis revealed that expression of a mutant Htt fragment in primary cortical neurons leads to robust gene expression changes before neuronal death. Basic helix-loop-helix transcription factor Twist1, which is essential for embryogenesis and is normally expressed at low levels in mature neurons, was substantially up-regulated in mutant Htt-expressing neurons in culture and in the brains of HD mouse models. Knockdown of Twist1 by RNAi in mutant Htt-expressing primary cortical neurons reversed the altered expression of a subset of genes involved in neuronal function and, importantly, abrogated neurotoxicity. Using brain-derived neurotrophic factor (), which is known to be involved in HD pathogenesis, as a model gene, we found that Twist1 knockdown could reverse mutant Htt-induced DNA hypermethylation at the regulatory region and reactivate expression. Together, these results suggest that Twist1 is an important upstream mediator of mutant Htt-induced neuronal death and may in part operate through epigenetic mechanisms.

Recent studies have shown that the major mitotic regulator CDC20-Anaphase Promoting Complex (APC) is required for the maintenance of human glioblastoma stem-like cells (GSCs), a clinically important subpopulation of glioblastoma cells that are believed to underlie tumor recurrence. We have previously shown that CDC20-APC operates through pluripotency transcription factor SOX2 to promote GSC self-renewal and invasion in vitro (Mao, Gujar et al. Cell Reports. 2015). Using patient-derived GSCs, we herein show that APC co-activator CDC20 is required for GSC tumorigenicity in an orthotopic xenograft model. We further demonstrate through in vivo epistasis experiments that CDC20 acts through SOX2 to control GSC tumorigenicity. To test if CDC20-APC activity might dictate GSC responsiveness to standard-of-care chemotherapy agent temozolomide (TMZ), we utilized pharmacological and genetic approaches to inhibit CDC20-APC activity. Intriguingly, both APC inhibitor ProTAME and lentiviral CDC20 RNA interference augmented TMZ cytotoxicity in human GSCs. Collectively, these data suggest that CDC20-APC controls the tumor-initiating potential of GSCs in vivo through SOX2 and that CDC20-APC inhibitory strategies may not only disrupt the GSC state but also enhance the efficacy of chemotherapy.