Introduction: Huntington’s disease (HD) is a dominantly inherited disorder. HD is a
neurodegenerative disease, with cognitive decline, behavioral symptoms and chorea. The
Huntingtin gene (HTT) normally contains 6-35 CAG repeats encoding a glutamine tract in the huntingtin protein. Expansion of this repeat leads to HD, with longer CAG tracts associated with earlier disease onset and greater severity. The expanded HTT CAG repeat further expands in individuals’ somatic cells over time. Specific modifier genes influence the somatic CAG expansion and the disease process in HD knock-in mouse models. Loss of mismatch repair (MMR) gene Mlh1 abrogates instability of the CAG repeat expansion and delays onset of disease-associated changes in mouse striatum. Notably, Mlh1 was identified as a candidate modifier of disease onset in patients. For HD therapeutic development, it is necessary to determine how Mlh1 may be involved in disease modulation. In the current study, two mice lines are used to explore specific actions of Mlh1: knock-in mice in which the huntingtin polyglutamine tract is encoded by an interrupted CAG tract (HttCAG105IR), or by a pure CAG repeat (HttCAG105Pure) of approximately the same length. The interrupted repeat does not exhibit somatic instability, while the pure repeat does. The goal is to compare the rate of disease-associated changes in striatal neurons and to compare the effect of Mlh1 knockout on each strain. If the interrupted mice do not show a delay of onset-associated changes, this would suggest that Mlh1 alters the disease process via a mechanism that is dependent on somatic instability. If, on the other hand, the interrupted mice show a similar delay of onset as the pure mice, this would indicate that Mlh1 modifies disease via another mechanism that is instability-independent.
Methods: This translational study included mice with either a pure or interrupted repeat,
encoding 105 glutamines, which were crossed with Mlh1 knockout mice. Progeny of different Mlh1 genotypes were investigated for somatic instability using PCR amplification assay, and for the rate of disease progression using immunohistochemical staining of nuclear huntingtin in medium- spiny neurons in the striatum.
Results: We showed that HttCAG105IR mice have stable repeats, while HttCAG105Pure mice have unstable repeats, providing appropriate models to distinguish effects of Mlh1 on stable and unstable alleles. Both Mlh1 knockout and CAG interruption stabilize the repeat. Analyses of nuclear huntingtin immunoreactivity in striatum show that both Mlh1 and repeat interruption reduce nuclear huntingtin, but indicate that Mlh1 knockout might, in-part, act via a mechanism independent of repeat expansion.
Conclusion: The present findings provide an indication that Mlh1-/- and HttCAG105IR/+ are
separately correlated with stabilizing the CAG repeat and reducing nuclear accumulation of
huntingtin. These outcomes suggest that combining these effects may further reduce nuclear huntingtin accumulation. Additional mice will be needed to test this prediction. Overall, the pure and interrupted CAG mouse models provide great tools for testing other modifier genes and dissecting disease mechanisms
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