Stoichiometry of Base Excision Repair Proteins Correlates with Increased Somatic CAG Instability in Striatum over Cerebellum in Huntington's Disease Transgenic Mice (original) (raw)
Related papers
Neurobiology of Disease, 2009
Modifying the length of the Huntington's disease (HD) CAG repeat, the major determinant of age of disease onset, is an attractive therapeutic approach. To explore this we are investigating mechanisms of intergenerational and somatic HD CAG repeat instability. Here, we have crossed HD CAG knockin mice onto backgrounds deficient in mismatch repair genes, Msh3 and Msh6, to discern the effects on CAG repeat size and disease pathogenesis. We find that different mechanisms predominate in inherited and somatic instability, with Msh6 protecting against intergenerational contractions and Msh3 required both for increasing CAG length and for enhancing an early disease phenotype in striatum. Therefore, attempts to decrease inherited repeat size may entail a full understanding of Msh6 complexes, while attempts to block the age-dependent increases in CAG size in striatal neurons and to slow the disease process will require a full elucidation of Msh3 complexes and their function in CAG repeat instability.
Genetic Modifiers of CAG.CTG Repeat Instability in Huntington's Disease Mouse Models
2017
Huntington's disease (HD) is a dominantly inherited neurodegenerative disorder whose characterstics were first described by George Huntington in 1872. Several decades later, in 1993, the mutation behind this disease was found to be an unstable expanded CAG repeat within exon 1 of the HTT gene localized on the short arm of chromosome 4. The majority of HD patients carry more than 40 CAG repeats, which become unstable and usually increase in size in successive generations and in tissues. In order to dissect the molecular mechanisms underlying CAG repeat instability, several HD mouse models have been created in the 1990s. Significant data have revealed that the absence of proteins from the mismatch repair (MMR) or the base and nucleotide excision repair decreased the pathogenic expansion‐biased somatic mosaicism and/or intergenerational expansions. Some polymorphic variants of MMR genes have also been associated with reduced somatic expansions. Since expansion‐ biased somatic mosai...
Mitochondrion, 2013
Oxidative DNA damage results from the attack by reactive oxygen and nitrogen species (ROS/RNS) on human genome. This includes base modifications such as oxidized bases, abasic (AP) sites, and single-strand breaks (SSBs), all of which are repaired by the base excision repair (BER) pathway, one among the six known repair pathways. BER-pathway in mammalian cells involves several evolutionarily conserved proteins and is also linked to genome replication and transcription. The BER-pathway enzymes, namely, DNA glycosylases (DGs) and the end-processing proteins such as abasic endonuclease (APE1), form complexes with downstream repair enzymes via protein-protein and DNA-protein interactions. An emerging concept for BER proteins is their involvement in non-canonical functions associated to RNA metabolism, which is opening new interesting perspectives. Various mechanisms that are underlined in maintaining neuronal cell genome integrity are identified, but are inconclusive in providing protection against oxidative damage in neurodegenerative disorders, main emphasis is given towards the role played by the proteins of BER-pathway that is discussed. In addition, mechanisms of action of BER-pathway in nuclear vs. mitochondria as well as the non-canonical functions are discussed in connection to human neurodegenerative diseases.
Human Molecular Genetics, 2014
Expansion of CAG/CTG trinucleotide repeats causes numerous inherited neurological disorders, including Huntington's disease (HD), several spinocerebellar ataxias and myotonic dystrophy type 1. Expanded repeats are genetically unstable with a propensity to further expand when transmitted from parents to offspring. For many alleles with expanded repeats, extensive somatic mosaicism has been documented. For CAG repeat diseases, dramatic instability has been documented in the striatum, with larger expansions noted with advancing age. In contrast, only modest instability occurs in the cerebellum. Using microarray expression analysis, we sought to identify the genetic basis of these regional instability differences by comparing gene expression in the striatum and cerebellum of aged wild-type C57BL/6J mice. We identified eight candidate genes enriched in cerebellum, and validated four-Pcna, Rpa1, Msh6 and Fen1-along with a highly associated interactor, Lig1. We also explored whether expression levels of mismatch repair (MMR) proteins are altered in a line of HD transgenic mice, R6/2, that is known to show pronounced regional repeat instability. Compared with wildtype littermates, MMR expression levels were not significantly altered in R6/2 mice regardless of age. Interestingly, expression levels of these candidates were significantly increased in the cerebellum of control and HD human samples in comparison to striatum. Together, our data suggest that elevated expression levels of DNA replication and repair proteins in cerebellum may act as a safeguard against repeat instability, and may account for the dramatically reduced somatic instability present in this brain region, compared with the marked instability observed in the striatum.
DNA instability in postmitotic neurons
Proceedings of the …, 2008
Huntington's disease (HD) is caused by a CAG repeat expansion that is unstable upon germ-line transmission and exhibits mosaicism in somatic tissues. We show that region-specific CAG repeat mosaicism profiles are conserved between several mouse models of HD and therefore develop in a predetermined manner. Furthermore, we demonstrate that these synchronous, radical changes in CAG repeat size occur in terminally differentiated neurons. In HD this ongoing mutation of the repeat continuously generates genetically distinct neuronal populations in the adult brain of mouse models and HD patients. The neuronal population of the striatum is particularly distinguished by a high rate of CAG repeat allele instability and expression driving the repeat upwards and would be expected to enhance its toxicity. In both mice and humans, neurons are distinguished from nonneuronal cells by expression of MSH3, which provides a permissive environment for genetic instability independent of pathology. The neuronal mutations described here accumulate to generate genetically discrete populations of cells in the absence of selection. This is in contrast to the traditional view in which genetically discrete cellular populations are generated by the sequence of random variation, selection, and clonal proliferation. We are unaware of any previous demonstration that mutations can occur in terminally differentiated neurons and provide a proof of principle that, dependent on a specific set of conditions, functional DNA polymorphisms can be produced in adult neurons.
PLoS Genetics, 2013
The Huntington's disease gene (HTT) CAG repeat mutation undergoes somatic expansion that correlates with pathogenesis. Modifiers of somatic expansion may therefore provide routes for therapies targeting the underlying mutation, an approach that is likely applicable to other trinucleotide repeat diseases. Huntington's disease Hdh Q111 mice exhibit higher levels of somatic HTT CAG expansion on a C57BL/6 genetic background (B6.Hdh Q111 ) than on a 129 background (129.Hdh Q111 ). Linkage mapping in (B6x129).Hdh Q111 F2 intercross animals identified a single quantitative trait locus underlying the strainspecific difference in expansion in the striatum, implicating mismatch repair (MMR) gene Mlh1 as the most likely candidate modifier. Crossing B6.Hdh Q111 mice onto an Mlh1 null background demonstrated that Mlh1 is essential for somatic CAG expansions and that it is an enhancer of nuclear huntingtin accumulation in striatal neurons. Hdh Q111 somatic expansion was also abolished in mice deficient in the Mlh3 gene, implicating MutLc (MLH1-MLH3) complex as a key driver of somatic expansion. Strikingly, Mlh1 and Mlh3 genes encoding MMR effector proteins were as critical to somatic expansion as Msh2 and Msh3 genes encoding DNA mismatch recognition complex MutSb (MSH2-MSH3). The Mlh1 locus is highly polymorphic between B6 and 129 strains. While we were unable to detect any difference in base-base mismatch or short slipped-repeat repair activity between B6 and 129 MLH1 variants, repair efficiency was MLH1 dose-dependent. MLH1 mRNA and protein levels were significantly decreased in 129 mice compared to B6 mice, consistent with a dose-sensitive MLH1-dependent DNA repair mechanism underlying the somatic expansion difference between these strains. Together, these data identify Mlh1 and Mlh3 as novel critical genetic modifiers of HTT CAG instability, point to Mlh1 genetic variation as the likely source of the instability difference in B6 and 129 strains and suggest that MLH1 protein levels play an important role in driving of the efficiency of somatic expansions.
Putting the Brakes on Huntington Disease in a Mouse Experimental Model
PLoS genetics, 2015
Huntington disease (HD) is a hereditary neurodegenerative disorder that causes a progressively debilitating impact on movement, cognition, speech, and mood. It most commonly develops during adulthood and worsens over a 10-15-year period. The genetic basis of HD is an expansion of the (CAG) n trinucleotide repeat in the first exon of the HTT gene [1,2]. Although the function of the normal HTT protein is not well established, in-frame repeat expansion results in the accumulation of an abnormally long polyglutamine tract, which is believed to contribute to mutant protein toxicity and neural degeneration [3]. Consequently, CAG repeat length is inversely correlated with age of onset and severity of disease. Disease-size CAG repeats are prone to further lengthening, which leads to two distinct aspects of their instability: expansions during intergenerational transmissions and somatic expansions occurring throughout the lifetime of an individual (Fig 1A). An outstanding question in the field is whether somatic expansions contribute to disease. The problem is that somatic expansions occur in the context of expressing an already toxic protein, making it difficult to address this question. Two commonly discussed ideas are that (i) disease arises with time because of expression of the toxic protein or RNA or that (ii) the onset of disease depends on gradual accumulation of somatic expansions in patients' tissues throughout life. To distinguish between these scenarios, it is critical to assess to what extent, if any, somatic expansions accelerate disease progression. Since 1993 when the cause of HD was first reported, substantial effort has been made to understand the molecular basis of repeat expansions and the mechanisms of disease pathophysiology. While these studies were conducted in various model systems [4,5], mouse models appeared to be particularly well suited for unraveling disease pathophysiology. Despite a few caveats, such as the much longer CAG repeat lengths (i.e., >100 repeats) needed to recapitulate disease symptoms and the relatively small-scale of expansions, mouse models of HD have led to principal contributions in the field. First, they exhibit both intergenerational and somatic repeat expansions. Second, candidate gene analysis uncovered a surprising role of the mismatch repair (MMR) complex MutSβ in promoting rather than protecting against CAG repeat expansions (Fig 1) [6,7]. A breakthrough in distinguishing between the molecular mechanisms of intergenerational and somatic expansions came with the discovery that loss of 8-oxoguanine glycosylase (OGG1) specifically decreased CAG expansions in somatic cells, but it had no effect on intergenerational transmissions [8]. The main function of OGG1 is to remove 8-oxoguanine (8-oxoG), a mutagenic base by-product accumulating in DNA after its exposure to reactive oxygen species (ROS). It was hypothesized, therefore, that aberrant repair of DNA oxidative damage specifically elevates repeat instability in long-lived differentiated cells, such as neurons. OGG1 excises 8-oxoG, generating a nick in the damaged DNA strand, which is further processed to permit
Human Molecular Genetics, 2011
Huntington's disease (HD) involves marked early neurodegeneration in the striatum, whereas the cerebellum is relatively spared despite the ubiquitous expression of full-length mutant huntingtin, implying that inherent tissue-specific differences determine susceptibility to the HD CAG mutation. To understand this tissue specificity, we compared early mutant huntingtin-induced gene expression changes in striatum to those in cerebellum in young Hdh CAG knock-in mice, prior to onset of evident pathological alterations. Endogenous levels of full-length mutant huntingtin caused qualitatively similar, but quantitatively different gene expression changes in the two brain regions. Importantly, the quantitatively different responses in the striatum and cerebellum in mutant mice were well accounted for by the intrinsic molecular differences in gene expression between the striatum and cerebellum in wild-type animals. Tissue-specific gene expression changes in response to the HD mutation, therefore, appear to reflect the different inherent capacities of these tissues to buffer qualitatively similar effects of mutant huntingtin. These findings highlight a role for intrinsic quantitative tissue differences in contributing to HD pathogenesis, and likely to other neurodegenerative disorders exhibiting tissue-specificity, thereby guiding the search for effective therapeutic interventions.
Age-dependent decline of DNA base excision repair activity in rat cortical neurons
Synthetic oligonucleotide duplexes containing a single uracil (U) or 8-oxoguanine (8-oxoG) were used as a model substrates to assess the base excision repair (BER) ability of neuronal extracts prepared from the cerebral cortex of young (7 days), adult (180 days) and old (720 days) rats. Our results demonstrate that BER activity in neurons markedly declines with age. The decline in BER could be attributed to decrease in the expression levels and activities of BER enzymes. Supplementing neuronal extracts with uracil DNAglycosylase (UDG), 8-oxoguanine DNA glycosylase (OGG1), apurinic endonuclease 1, pol b and T4 DNA ligase independently could not restore the loss of BER activity in adult and old neuronal extracts. However, supplementation of pol b in combination of T4 DNA ligase to neuronal extract, improved the BER in adult and old neuronal extracts. Additional supplementation of the extracts with UDG or OGG1 apart from pol b and T4 DNA ligase, or with all pure enzymes restored very markedly the loss of BER in aging neurons. These results suggest the age-dependent decline in BER is due to an overall deficiency of the various factors needed for BER but pol b and DNA ligase seem to be the most limiting factors. 2012 Elsevier Ireland Ltd. All rights reserved.
The molecular biology of Huntington's disease
Psychological Medicine, 2001
Background. Huntington's disease (HD) is a fatal neurodegenerative disorder with an autosomal dominant mode of inheritance. It leads to progressive dementia, psychiatric symptoms and an incapacitating choreiform movement disorder, culminating in premature death. HD is caused by an increased CAG repeat number in a gene coding for a protein with unknown function, called huntingtin. The trinucleotide CAG codes for the amino acid glutamine and the expanded CAG repeats are translated into a series of uninterrupted glutamine residues (a polyglutamine tract).Methods. This review describes the epidemiology, clinical symptomatology, neuropathological features and genetics of HD. The main aim is to examine important findings from animal and cellular models and evaluate how they have enriched our understanding of the pathogenesis of HD and other diseases caused by expanded polyglutamine tracts.Results. Selective death of striatal and cortical neurons occurs. It is likely that the HD mutati...