Mitochondrial Dysfunction In Huntington's Disease Research Articles

Mitochondrial and Redox-Based Therapeutic Strategies in Huntington's Disease.

Significance: The molecular processes that determine Huntington's disease (HD) pathogenesis are not yet fully understood, and until now no effective neuroprotective therapeutic strategies have been developed. Mitochondria are one of most important organelles required for neuronal homeostasis, by providing metabolic pathways relevant for energy production, regulating calcium homeostasis, or controlling free radical generation and cell death. Because augmented reactive oxygen species (ROS) accompanied by mitochondrial dysfunction are relevant early HD mechanisms, targeting these cellular mechanisms may constitute relevant therapeutic approaches. Recent Advances: Previous findings point toward a close relationship between mitochondrial dysfunction and redox changes in HD. Mutant huntingtin (mHTT) can directly interact with mitochondrial proteins, as translocase of the inner membrane 23 (TIM23), disrupting mitochondrial proteostasis and favoring ROS production and HD progression. Furthermore, abnormal brain and muscle redox signaling contributes to altered proteostasis and motor impairment in HD, which can be improved with the mitochondria-targeted antioxidant mitoquinone or resveratrol, an SIRT1 activator that ameliorates mitochondrial biogenesis and function. Critical Issues: Various antioxidants and metabolic enhancers have been studied in HD; however, the real outcome of these molecules is still debatable. New compounds have proven to ameliorate mitochondrial and redox-based signaling pathways in early stages of HD, potentially precluding selective neurodegeneration. Future Directions: Unraveling the molecular etiology of deregulated mitochondrial function and dynamics, and oxidative stress opens new prospects for HD therapeutics. In this review, we explore the role of redox unbalance and mitochondrial dysfunction in HD progression, and further describe advances on clinical trials in HD based on mitochondrial and redox-based therapeutic strategies.

VCP cooperates with UBXD1 to degrade mitochondrial outer membrane protein MCL1 in model of Huntington's disease

Proteasome-dependent turnover of mitochondrial outer membrane (OMM)-associated proteins is one of the mechanisms for maintaining proper mitochondrial quality and function. However, the underlying pathways and their implications in human disease are poorly understood. Huntington's disease (HD) is a fatal, inherited neurodegenerative disorder caused by expanded CAG repeats in the N terminal of the huntingtin gene (mutant Huntingtin, mtHtt). In this study, we show an extensive degradation of the OMM protein MCL1 (Myeloid cell leukemia sequence 1) in both HD mouse striatal cells and HD patient fibroblasts. The decrease in MCL1 level is associated with mitochondrial and cellular damage. Valosin-containing-protein (VCP) is an AAA-ATPase central to protein turnover via the ubiquitin proteasome system (UPS). We found that VCP translocates to mitochondria and promotes MCL1 degradation in HD cell cultures. Either down-regulation of VCP by RNA interference or inhibition of VCP by a dominant negative mutant abolishes MCL1 degradation in HD cell cultures. We further show that UBX-domain containing protein 1 (UBXD1), a known co-factor of VCP assisting in the recognition of substrates for protein degradation, selectively binds to MCL1 and interacts with VCP to mediate MCL1 extraction from the mitochondria. These results indicate that the OMM protein MCL1 is degraded by the VCP-UBXD1 complex and that the process is promoted by the presence of mtHtt. Therefore, our finding provides a new insight into the mechanism of mitochondrial dysfunction in HD.

Inhibition of mitochondrial protein import by mutant huntingtin.

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.

Mitochondrial Membrane Fluidity is Consistently Increased in Different Models of Huntington Disease: Restorative Effects of Olesoxime

Huntington disease (HD) is a fatal neurodegenerative disorder caused by a CAG repeat expansion in exon 1 of the huntingtin gene (HTT). One prominent target of the mutant huntingtin protein (mhtt) is the mitochondrion, affecting its morphology, distribution, and function. Thus, mitochondria have been suggested as potential therapeutic targets for the treatment of HD. Olesoxime, a cholesterol-like compound, promotes motor neuron survival and neurite outgrowth in vitro, and its effects are presumed to occur via a direct interaction with mitochondrial membranes (MMs). We examined the properties of MMs isolated from cell and animal models of HD as well as the effects of olesoxime on MM fluidity and cholesterol levels. MMs isolated from brains of aged Hdh Q111/Q111 knock-in mice showed a significant decrease in 1,6-diphenyl-hexatriene (DPH) anisotropy, which is inversely correlated with membrane fluidity. Similar increases in MM fluidity were observed in striatal STHdh Q111/Q111 cells as well as in MMs isolated from brains of BACHD transgenic rats. Treatment of STHdh cells with olesoxime decreased the fluidity of isolated MMs. Decreased membrane fluidity was also measured in olesoxime-treated MMs isolated from brains of HD knock-in mice. In both models, treatment with olesoxime restored HD-specific changes in MMs. Accordingly, olesoxime significantly counteracted the mhtt-induced increase in MM fluidity of MMs isolated from brains of BACHD rats after 12 months of treatment in vivo, possibly by enhancing MM cholesterol levels. Thus, olesoxime may represent a novel pharmacological tool to treat mitochondrial dysfunction in HD.

E01 Analysis of mitochondrial genes as potential genetic modifiers of the age-at-onset in Huntington disease

Background The major determinant of age-at-onset (AAO) in Huntington disease (HD) is the length of the causative polyglutamine expansion in the huntingtin protein, which is inversely correlated with the AAO and the course of the disease. However, both environmental and genetic factors are thought to modify the disease. Here, genetic factors are proposed to have an essential influence and especially polymorphism in genes associated with HD pathogenesis are thought to act as genetic modifiers. One characteristic feature in HD pathogenesis is mitochondrial dysfunction, which is marked by an impairment of mitochondrial integrity and trafficking leading to a disruption of mitochondrial function and an inhibition of energy supply in the cell. Amongst others, mutant huntingtin was shown to lead to oxidative stress, increased fragmentation of mitochondria and reduced ATP levels. A regulated expression of proteins, involved in mitochondrial fission and fusion (eg, Mfn1, Mfn2) antagonises the pathogenic effect, thus demonstrating to be interesting potential candidates for a modifying role in HD. Aim/method In order to identify genetic modifiers participating in mitochondrial function, we investigated several single nucleotide polymorphisms in mitochondrial fusion (Mfn1, Mfn2, Opa1) and fission (Fis1) genes for their frequency of ≥1%. Subsequently, we aim to analyse these polymorphic markers for a disease-modifying effect in more than 1400 European HD patients. Additionally, to establish a connection between mutant huntingtin and mitochondrial genes, we intend to examine a possible interaction between huntingtin and the fission protein Fis1 by co-immunoprecipitation experiments in cell culture. Conclusion The present study provides the opportunity to identify new genetic modifiers influencing the AAO and the pathogenic course of HD. These would further support the important pathophysiological role of mitochondrial dysfunction in HD and may offer new basis for further clarification of HD pathology.

Huntingtin protein interactions altered by polyglutamine expansion as determined by quantitative proteomic analysis

Huntington disease (HD) is a neurodegenerative disorder caused by an expansion of a polyglutamine repeat within the HD gene product, huntingtin. Huntingtin, a large (347 kDa) protein containing multiple HEAT repeats, acts as a scaffold for protein-protein interactions. Huntingtin-induced toxicity is believed to be mediated by a conformational change in expanded huntingtin, leading to protein misfolding and aggregation, aberrant protein interactions and neuronal cell death. While many non-systematic studies of huntingtin interactions have been reported, they were not designed to identify and quantify the changes in the huntingtin interactome induced by polyglutamine expansion. We used tandem affinity purification and quantitative proteomics to compare and quantify interactions of normal or expanded huntingtin isolated from a striatal cell line. We found that proteins preferentially interacting with expanded huntingtin are enriched for intrinsically disordered proteins, consistent with previously suggested roles of such proteins in neurodegenerative disorders. Our functional analysis indicates that proteins related to energy production, protein trafficking, RNA post-transcriptional modifications and cell death were significantly enriched among preferential interactors of expanded huntingtin. Expanded huntingtin interacted with many mitochondrial proteins, including AIFM1, consistent with a role for mitochondrial dysfunction in HD. Furthermore, expanded huntingtin interacted with the stress granule-associated proteins Caprin-1 and G3BP and redistributed to RNA stress granules under ER-stress conditions. These data demonstrate that a number of key cellular functions and networks may be disrupted by abnormal interactions of expanded huntingtin and highlight proteins and pathways that may be involved in HD cellular pathogenesis and that may serve as therapeutic targets.

Transducer of regulated CREB-binding proteins (TORCs) transcription and function is impaired in Huntington's disease

Huntington's disease (HD) is an incurable neurological disorder caused by an abnormal glutamine repeat expansion in the huntingtin (Htt) protein. In the present studies, we investigated the role of Transducers of Regulated cAMP response element-binding (CREB) protein activity (TORCs) in HD, since TORCs play an important role in the expression of the transcriptional co-regulator peroxisome proliferator-activated receptor gamma coactivator 1α (PGC-1α), whose expression is impaired in HD. We found significantly decreased TORC1 expression levels in STHdhQ111 cells expressing mutant Htt, in the striatum of NLS-N171-82Q, R6/2 and HdhQ111 HD transgenic mice and in postmortem striatal tissue from HD patients. TORC1 overexpression in wild-type (WT) and Htt striatal cells increased CREB mRNA and protein levels, PGC-1α promoter activity, mRNA expression of the PGC-1α, NRF-1, Tfam and CytC genes, mitochondrial DNA content, mitochondrial activity and mitochondrial membrane potential. TORC1 overexpression also increased the resistance of striatal cells to 3-nitropropionic (3-NP) acid-mediated toxicity. In cultured WT and mutant Htt striatal cells, small hairpin RNA-mediated TORC1 knockdown resulted in decreased PGC-1α expression and increased susceptibility to 3-NP-induced toxicity. Overexpression of PGC-1α partially prevented TORC1 knockdown-mediated increased susceptibility of Htt striatal cells to 3-NP. Specific knockdown of TORC1 in the striatum of NLS-N171-82Q HD transgenic mice induced neurodegeneration. Lastly, knockdown of Htt prevents transcriptional repression of TORC1 and CREB in Htt striatal cells. These findings show that impaired expression and function of TORC1, which results in a reduction in PGC-1α, plays an important role in mitochondrial dysfunction in HD.

A13 Mutant huntingtin induces activation of Bcl-2/adenovirus E1B 19-kDa interacting protein (BNIP3)

Huntington's disease (HD) is a neurodegenerative disorder characterised by progressive neuronal death in the basal ganglia and cortex. Although increasing evidence supports a pivotal role for mitochondrial dysfunction in the...

Mutant Huntingtin induces activation of the Bcl-2/adenovirus E1B 19-kDa interacting protein (BNip3)

Huntington's disease (HD) is a neurodegenerative disorder characterized by progressive neuronal death in the basal ganglia and cortex. Although increasing evidence supports a pivotal role of mitochondrial dysfunction in the death of patients' neurons, the molecular bases for mitochondrial impairment have not been elucidated. We provide the first evidence of an abnormal activation of the Bcl-2/adenovirus E1B 19-kDa interacting protein 3 (BNip3) in cells expressing mutant Huntingtin. In this study, we show an abnormal accumulation and dimerization of BNip3 in the mitochondria extracted from human HD muscle cells, HD model cell cultures and brain tissues from HD model mice. Importantly, we have shown that blocking BNip3 expression and dimerization restores normal mitochondrial potential in human HD muscle cells. Our data shed light on the molecular mechanisms underlying mitochondrial dysfunction in HD and point to BNip3 as a new potential target for neuroprotective therapy in HD.

PGC-1alphaas modifier of onset age in Huntington disease

Although there is a strong correlation between CAG repeat length and age at onset (AO) of motor symptoms, individual Huntington disease (HD) patients may differ dramatically in onset age and disease manifestations despite similar CAG repeat lengths. This has led to a search for genetic factors that influence AO. In order to identify such a genetic modifier, we analysed polymorphisms in the PGC-1alpha gene. Recent data indicate inhibition of PGC-1alpha function by mutant Htt supporting a link between transcriptional deregulation and mitochondrial dysfunction in HD. In > 400 HD patients, a polymorphism located within intron 2, a potential recombination hot spot, explains a small, but statistically significant, amount of the variability in AO. Our data suggest that PGC-1alpha has modifying effects on the pathogenic process in HD.

Mitochondrial DNA damage is a hallmark of chemically induced and the R6/2 transgenic model of Huntington's disease

Many forms of neurodegeneration are associated with oxidative stress and mitochondrial dysfunction. Mitochondria are prominent targets of oxidative damage, however, it is not clear whether mitochondrial DNA (mtDNA) damage and/or its lack of repair are primary events in the delayed onset observed in Huntington's disease (HD). We hypothesize that an age-dependent increase in mtDNA damage contributes to mitochondrial dysfunction in HD. Two HD mouse models were studied, the 3-nitropropionic acid (3-NPA) chemically induced model and the HD transgenic mice of the R6/2 strain containing 115–150 CAG repeats in the huntingtin gene. The mitochondrial toxin 3-NPA inhibits complex II of the electron transport system and causes neurodegeneration that resembles HD in the striatum of human and experimental animals. We measured nuclear and mtDNA damage by quantitative PCR (QPCR) in striatum of 5- and 24-month-old untreated and 3-NPA treated C57BL/6 mice. Aging caused an increase in damage in both nuclear and mitochondrial genomes. 3-NPA induced 4–6 more damage in mtDNA than nuclear DNA in 5-month-old mice, and this damage was repaired by 48 h in the mtDNA. In 24-month-old mice 3NPA caused equal amounts of nuclear and mitochondrial damage and this damage persistent in both genomes for 48 h. QPCR analysis showed a progressive increase in the levels of mtDNA damage in the striatum and cerebral cortex of 7–12-week-old R6/2 mice. Striatum exhibited eight-fold more damage to the mtDNA compared with a nuclear gene. These data suggest that mtDNA damage is an early biomarker for HD-associated neurodegeneration and supports the hypothesis that mtDNA lesions may contribute to the pathogenesis observed in HD.

Mutant Huntingtin Expression Induces Mitochondrial Calcium Handling Defects in Clonal Striatal Cells

Huntington disease (HD) is caused by a pathological elongation of CAG repeats in the huntingtin protein gene and is characterized by atrophy and neuronal loss primarily in the striatum. Mitochondrial dysfunction and impaired Ca2+ homeostasis in HD have been suggested previously. Here, we elucidate the effects of Ca2+ on mitochondria from the wild type (STHdhQ7/Q7) and mutant (STHdhQ111/Q111) huntingtin-expressing cells of striatal origin. When treated with increasing Ca2+ concentrations, mitochondria from mutant huntingtin-expressing cells showed enhanced sensitivity to Ca2+, as they were more sensitive to Ca2+-induced decreases in state 3 respiration and DeltaPsim, than mitochondria from wild type cells. Further, mutant huntingtin-expressing cells had a reduced mitochondrial Ca2+ uptake capacity in comparison with wild type cells. Decreases in state 3 respiration were associated with increased mitochondrial membrane permeability. The DeltaPsim defect was attenuated in the presence of ADP and the decreases in Ca2+ uptake capacity were abolished in the presence of Permeability Transition Pore (PTP) inhibitors. These findings clearly indicate that mutant huntingtin-expressing cells have mitochondrial Ca2+ handling defects that result in respiratory deficits and that the increased sensitivity to Ca2+ induced mitochondrial permeabilization maybe a contributing mechanism to the mitochondrial dysfunction in HD.

Thermoregulatory and metabolic defects in Huntington's disease transgenic mice implicate PGC-1α in Huntington's disease neurodegeneration

Huntington's disease (HD) is a fatal, dominantly inherited disorder caused by polyglutamine repeat expansion in the huntingtin (htt) gene. Here, we observe that HD mice develop hypothermia associated with impaired activation of brown adipose tissue (BAT). Although sympathetic stimulation of PPARgamma coactivator 1alpha (PGC-1alpha) was intact in BAT of HD mice, uncoupling protein 1 (UCP-1) induction was blunted. In cultured cells, expression of mutant htt suppressed UCP-1 promoter activity; this was reversed by PGC-1alpha expression. HD mice showed reduced food intake and increased energy expenditure, with dysfunctional BAT mitochondria. PGC-1alpha is a known regulator of mitochondrial function; here, we document reduced expression of PGC-1alpha target genes in HD patient and mouse striatum. Mitochondria of HD mouse brain show reduced oxygen consumption rates. Finally, HD striatal neurons expressing exogenous PGC-1alpha were resistant to 3-nitropropionic acid treatment. Altered PGC-1alpha function may thus link transcription dysregulation and mitochondrial dysfunction in HD.