DNA Damage Repair in Huntington’s Disease and Other Neurodegenerative Diseases

Abstract

Recent genome-wide association studies of Huntington’s disease (HD) primarily highlighted genes involved in DNA damage repair mechanisms as modifiers of age at onset and disease severity, consistent with evidence that more DNA repair genes are being implicated in late age–onset neurodegenerative diseases. This provides an exciting opportunity to advance therapeutic development in HD, as these pathways have already been under intense investigation in cancer research. Also emerging are the roles of other polyglutamine disease proteins in DNA damage repair mechanisms. A potential universal trigger of oxidative DNA damage shared in these late age–onset diseases is the increase of reactive oxygen species (ROS) in human aging, defining an age-related mechanism that has defied other hypotheses of neurodegeneration. We discuss the potential commonality of DNA damage repair pathways in HD and other neurodegenerative diseases. Potential targets for therapy that may prove beneficial across many of these diseases are also identified, defining nodes in the ataxia telangiectasia-mutated (ATM) complex, mismatch repair, and poly ADP-ribose polymerases (PARPs).

Huntington’s disease (HD) was precisely defined in 1993 when the huntingtin gene was cloned and the disease-causing CAG nucleotide expansion was defined, showing an inverse correlation between expansion length and age at onset [1]. The problem of HD was daunting, as the gene encoded one of the largest non-membrane proteins in the human proteome at 3144 amino acids or 350 kDa. Over the next 25 years, the normal function of huntingtin also eluded strict definition. Cell biology and biochemical studies revealed the protein to have no enzymatic activity and no conserved functional motifs, with the exception that proteins with polyglutamine tracts were often transcriptionally active and huntingtin contained HEAT repeats common to several protein scaffolds [2, 3]. Interactome studies showed huntingtin at the heart of several complexes [4] and it has been localized to the nucleus [5], endoplasmic reticulum [

Mismatch Repair and Somatic CAG Expansion

The mismatch repair (MMR) pathway is of particular interest because of its influence on somatic expansion of CAG repeats, which can undergo progressive length increases over time, particularly in the brain [46]. Somatic expansion of huntingtin is associated with earlier age of HD onset [47] and more severe symptoms [48]. In mouse models, expansion is prevented upon deletion of MMR genes, mutS homologs 2 and 3 (msh2 and msh3), mutL homolog 1 (mlh1), or PMS1 homolog 2 (pms2) [49,50,51,52]. These proteins normally work in concert to repair insertion-deletion loops in the DNA, but they also drive disease-associated repeat expansions [53].

The initial evidence from mouse models has since been bolstered by human data, clearly implicating the MMR pathway in disease progression. Although the chromosomal region bearing MLH1 was identified by GWAS, but did not initially reach genome-wide significance [17], a subsequent SNP genoty** study confirmed a modifier haplotype at the MLH1 locus [21]. The genomic region bearing MSH3 was identified by a second GWAS with more in-depth measures of disease progression [18], and PMS2 was among the SNP locations independently confirmed by genoty** [20]. Disease-associated SNPs near the MLH1, MSH3, PMS2, and PMS1 loci have recently been confirmed and the importance of uninterrupted CAG repeats within the huntingtin gene itself, which also impacts somatic instability, is now clear [19, 23]. PMS2 and MSH3 have also been implicated in SCA1 and myotonic dystrophy, respectively [20, 22]. Modifier genes are summarized in Table 1.

Although it is likely that MMR proteins influence pathology via somatic expansion, this may not be the sole mechanism, as tissue specificity of somatic expansion correlates with striatal neurodegeneration in HD, but not in SCAs [54], despite ubiquitous transcription of the CAG-containing proteins. It is possible that MMR proteins also contribute to disease modification through other DNA repair factors, including TP53 [55], breast cancer associated-1 (BRCA1) [56], and ATM [57]. The MMR machinery has also been implicated in the repair of several types of DNA damage [58,59,60], including ICLs [61].

In the patient population, pathology is likely to be caused by a combination of these mechanisms. The fact that MMR proteins, and scaffolds such as huntingtin and ATM are common components of many DNA repair complexes [91].

The relevance of PARylation to HD has yet to be determined, but from GWAS we may have a hint that ties hyper-PARylation to the ATM/huntingtin complex. Another major lead in HD GWAS is the ribonucleoside-diphosphate reductase subunit M2B, or RRM2B, also known as P53R2. RRM2B is a critical ribonucleotide salvager which nets a severe fatal pediatric disorder called mitochondrial DNA depletion syndrome (MDDS) that affects muscle, the brain, and the respiratory tract when null [92]. Like huntingtin, RRM2B is activated by the TP53 tumor suppressor [93], and is a known ATM interactor [94]. The ribonucleoside-diphosphate reductase activity could be critical downstream of PARG to salvage ADP back from hydrolyzed PAR chains during neuronal energy crisis. Thus, we can hypothesize a mechanism of RRM2B disease modification by catalyzing the conversion of poly ADP-ribose chains back to critical adenosine ribonucleosides.

HD GWAS in the Bigger Picture of Polyglutamine Diseases

Given the significance of HD genetic modifier SNPs to some spinocerebellar ataxias, we may find an intersection of molecular mechanisms of disease between HD and other CAG repeat disorders with respect to DNA repair. One node is the ATM complex, but huntingtin has also been defined at the TCR complex [45] along with ataxin-3, the affected protein in SCA3, and PKNP, the protein mutated in MCSZ [27]. Ataxin-3 has also been implicated by others in the double-strand break response [95]. CAG expansion in the androgen receptor, a transcription factor involved in DNA damage repair signaling, leads to spinal and bulbar muscle atrophy, SBMA, or Kennedy’s disease [96, 97]. CAG expansion in the TATA box-binding protein (TBP) causes SCA17, and TBP localizes to damaged DNA [98]. DNA repair has also been implicated in SCA1, as overexpression of DNA repair factors replication protein A1 and high mobility group box 1 in mouse and Drosophila models corrects motor phenotypes [99, 100]. Thus, we may anticipate increased relevance of DNA repair in many late age–onset neurodegenerative diseases, as the natural increased ROS stress during human aging and effects on DNA/RNA oxidation are tempting mechanisms to explain why these diseases typically occur later in life.

Increased DNA oxidation and hyper-PARylation appeared in neurodegenerative disease studies in the late 1990s [101, 102], but the early study of PAR chains and relevance of oxidation to disease mechanism were not further explored in favor of various amyloid hypotheses of late age–onset neurodegeneration. Although guanine oxidation products were the focus of these studies as a biomarker of DNA oxidation, it is not clear that all DNA bases are equally modified under oxidative stress nor processed in a similar manner. Adenosine bases are subject to nucleotide salvage in neurons and have unique utility after base excision repair to be salvaged back to adenosine nucleosides for energy production [85]. Neurons are highly metabolically active and thus generate high ROS loads, yet oxidative base damage to DNA cannot be repaired by DNA replication as in mitotic cell types. Brain subregions can also transiently flip to aerobic glycolysis or Warburg metabolism to generate ATP at times of energy stress [103], but this energy supply comes at a high cost of increased ROS stress. The burden of reactive oxygen levels on mitochondria, which have a diminished efficiency with human aging, might explain how mitochondrial dysfunction is a common aspect of all neurodegenerative diseases [104].

To fully understand the implications of defective DNA repair in neurodegeneration, it is important to understand how DNA repair imparts a severe energy stress on neurons. High rates of neuronal metabolism mean that even as energy is depleted, ROS by-products cause damage that further drains the energy supply. In this way, neurons must struggle to maintain energy levels and even a minor deficiency in a DNA repair protein would amount to undue neuronal death over time (Fig. 2).

Fig. 2

DNA repair and neuronal energy homeostasis. The combined energetic costs of high metabolic rate, and repair of metabolism by-product–mediated damage, make neurons vulnerable to minor deficiencies in DNA repair proteins with age

The involvement of DNA repair pathways in neurodegenerative diseases presents a number of opportunities for therapeutic intervention. Although drugs against key potential targets have been developed for cancer, preclinical data on their applicability for neurodegenerative diseases is needed as the goal of anticancer therapy is to kill affected cells, whereas the goal of antineurodegeneration therapy is to save affected cells. The immediate goals in researching DNA repair in neurodegeneration are to define the roles of DNA repair factors, in terms of their scaffolding versus enzymatic functions, to precisely define therapeutic mechanisms of either inhibiting enzymatic activity or modulating protein levels, or both. Testing in the most clinically relevant models and special attention to measures of genotoxicity will help pave the way to the clinic.