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Mismatch repair genes Mlh1 and Mlh3 modify CAG instability in Huntington's disease mice: genome-wide and candidate approaches.

Pinto RM, Dragileva E, Kirby A, Lloret A, Lopez E, St Claire J, Panigrahi GB, Hou C, Holloway K, Gillis T, Guide JR, Cohen PE, Li GM, Pearson CE, Daly MJ, Wheeler VC - PLoS Genet. (2013)

Bottom Line: 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 MutSβ (MSH2-MSH3).The Mlh1 locus is highly polymorphic between B6 and 129 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.

View Article: PubMed Central - PubMed

Affiliation: Center for Human Genetic Research, Massachusetts General Hospital, Boston, Massachusetts, United States of America.

ABSTRACT
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 strain-specific 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 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 MutLγ (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 MutSβ (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.

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Proposed model of MutS and MutL-dependent events leading to CAG•CTG somatic instability.CAG•CTG repeat structures are initially recognized by the MutSβ (MSH2-MSH3) complex [25], [98]. The loop in the CAG•CTG repeat tract represents a short slip-out, previously identified as the main substrate for MMR protein-dependent repair of CAG•CTG structures in cell free systems [50], [51]. However, the nature of the putative CAG•CTG structure(s) that leads to MutS and MutL-dependent somatic instability in vivo is unknown. Following ATP hydrolysis by DNA-bound MutSβ [27], a MutLγ (MLH1–MLH3) heterodimer is preferentially recruited to the complex (thick arrow) over the MutLα (MLH1-PMS2) heterodimer (thin arrow). The total absence of HTT CAG expansion in Mlh3−/− mice suggests that PMS2 plays no role at all in this process. However, PMS2 has been shown to play a role in the expansion of CTG repeats in a DM1 mouse model [24], suggesting that these events may be genetic locus and/or mouse strain dependent. Following MutLγ binding, various pathways, e.g. canonical mismatch repair (MMR), noncanonical mismatch repair (ncMMR) and/or other DNA repair processes may be engaged and process the repeats such that they ultimately undergo expansion. Other members of alternative DNA repair pathways, namely OGG1, XPA and NEIL1 have been directly implicated in CAG/CTG somatic instability in mice [76]–[78], however, how these proteins intersect with MMR protein-dependent pathways has yet to be demonstrated.
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pgen-1003930-g011: Proposed model of MutS and MutL-dependent events leading to CAG•CTG somatic instability.CAG•CTG repeat structures are initially recognized by the MutSβ (MSH2-MSH3) complex [25], [98]. The loop in the CAG•CTG repeat tract represents a short slip-out, previously identified as the main substrate for MMR protein-dependent repair of CAG•CTG structures in cell free systems [50], [51]. However, the nature of the putative CAG•CTG structure(s) that leads to MutS and MutL-dependent somatic instability in vivo is unknown. Following ATP hydrolysis by DNA-bound MutSβ [27], a MutLγ (MLH1–MLH3) heterodimer is preferentially recruited to the complex (thick arrow) over the MutLα (MLH1-PMS2) heterodimer (thin arrow). The total absence of HTT CAG expansion in Mlh3−/− mice suggests that PMS2 plays no role at all in this process. However, PMS2 has been shown to play a role in the expansion of CTG repeats in a DM1 mouse model [24], suggesting that these events may be genetic locus and/or mouse strain dependent. Following MutLγ binding, various pathways, e.g. canonical mismatch repair (MMR), noncanonical mismatch repair (ncMMR) and/or other DNA repair processes may be engaged and process the repeats such that they ultimately undergo expansion. Other members of alternative DNA repair pathways, namely OGG1, XPA and NEIL1 have been directly implicated in CAG/CTG somatic instability in mice [76]–[78], however, how these proteins intersect with MMR protein-dependent pathways has yet to be demonstrated.

Mentions: With regard to potential mechanisms of CAG expansion it is of interest that MSH3 and MLH3 appear to play relatively minor roles in classical MMR inasmuch as Msh3 and Mlh3 deficiencies result in weak mutator phenotypes and relatively low cancer predisposition phenotypes [42], [72]–[75]. In strong contrast, loss of either of these two proteins has a major impact on CAG/CTG expansion. Conversely, MSH6 and PMS2 play prominent roles in classical MMR [72]–[74]. However, MSH6 is either unnecessary for, or plays a very minimal role in mediating somatic CAG/CTG expansions [19], [22], [25], and knockout of Pms2 had a moderate effect of CTG expansion in DM1 mice [24], implicating a role for different MLH1 partners. In the present study the complete absence of HTT CAG expansion in HdhQ111/+Mlh3 mice argues against a role for PMS2 in generating expansions in these mice. Further genetic crosses in both DM1 and HdhQ111 mice would be needed to determine whether the relative contributions of Pms2 and Mlh3 genes in the two mouse models depends on the genomic locus of the repeat and/or strain background. While we do not expect PMS2 levels to be altered in Mlh3 knockout mice [74], additional experiments are needed in Mlh3 and Pms2 knockout mouse tissues to determine whether any compensatory changes in PMS2 or MLH3 proteins, respectively, occur. However, overall, the data thus far indicate that MLH3 is a more significant player than PMS2 in CAG/CTG expansion and suggest that CAG/CTG repeats may preferentially engage a pathway(s) involving MutSβ and MutLγ complexes, as illustrated in Figure 11.


Mismatch repair genes Mlh1 and Mlh3 modify CAG instability in Huntington's disease mice: genome-wide and candidate approaches.

Pinto RM, Dragileva E, Kirby A, Lloret A, Lopez E, St Claire J, Panigrahi GB, Hou C, Holloway K, Gillis T, Guide JR, Cohen PE, Li GM, Pearson CE, Daly MJ, Wheeler VC - PLoS Genet. (2013)

Proposed model of MutS and MutL-dependent events leading to CAG•CTG somatic instability.CAG•CTG repeat structures are initially recognized by the MutSβ (MSH2-MSH3) complex [25], [98]. The loop in the CAG•CTG repeat tract represents a short slip-out, previously identified as the main substrate for MMR protein-dependent repair of CAG•CTG structures in cell free systems [50], [51]. However, the nature of the putative CAG•CTG structure(s) that leads to MutS and MutL-dependent somatic instability in vivo is unknown. Following ATP hydrolysis by DNA-bound MutSβ [27], a MutLγ (MLH1–MLH3) heterodimer is preferentially recruited to the complex (thick arrow) over the MutLα (MLH1-PMS2) heterodimer (thin arrow). The total absence of HTT CAG expansion in Mlh3−/− mice suggests that PMS2 plays no role at all in this process. However, PMS2 has been shown to play a role in the expansion of CTG repeats in a DM1 mouse model [24], suggesting that these events may be genetic locus and/or mouse strain dependent. Following MutLγ binding, various pathways, e.g. canonical mismatch repair (MMR), noncanonical mismatch repair (ncMMR) and/or other DNA repair processes may be engaged and process the repeats such that they ultimately undergo expansion. Other members of alternative DNA repair pathways, namely OGG1, XPA and NEIL1 have been directly implicated in CAG/CTG somatic instability in mice [76]–[78], however, how these proteins intersect with MMR protein-dependent pathways has yet to be demonstrated.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3814320&req=5

pgen-1003930-g011: Proposed model of MutS and MutL-dependent events leading to CAG•CTG somatic instability.CAG•CTG repeat structures are initially recognized by the MutSβ (MSH2-MSH3) complex [25], [98]. The loop in the CAG•CTG repeat tract represents a short slip-out, previously identified as the main substrate for MMR protein-dependent repair of CAG•CTG structures in cell free systems [50], [51]. However, the nature of the putative CAG•CTG structure(s) that leads to MutS and MutL-dependent somatic instability in vivo is unknown. Following ATP hydrolysis by DNA-bound MutSβ [27], a MutLγ (MLH1–MLH3) heterodimer is preferentially recruited to the complex (thick arrow) over the MutLα (MLH1-PMS2) heterodimer (thin arrow). The total absence of HTT CAG expansion in Mlh3−/− mice suggests that PMS2 plays no role at all in this process. However, PMS2 has been shown to play a role in the expansion of CTG repeats in a DM1 mouse model [24], suggesting that these events may be genetic locus and/or mouse strain dependent. Following MutLγ binding, various pathways, e.g. canonical mismatch repair (MMR), noncanonical mismatch repair (ncMMR) and/or other DNA repair processes may be engaged and process the repeats such that they ultimately undergo expansion. Other members of alternative DNA repair pathways, namely OGG1, XPA and NEIL1 have been directly implicated in CAG/CTG somatic instability in mice [76]–[78], however, how these proteins intersect with MMR protein-dependent pathways has yet to be demonstrated.
Mentions: With regard to potential mechanisms of CAG expansion it is of interest that MSH3 and MLH3 appear to play relatively minor roles in classical MMR inasmuch as Msh3 and Mlh3 deficiencies result in weak mutator phenotypes and relatively low cancer predisposition phenotypes [42], [72]–[75]. In strong contrast, loss of either of these two proteins has a major impact on CAG/CTG expansion. Conversely, MSH6 and PMS2 play prominent roles in classical MMR [72]–[74]. However, MSH6 is either unnecessary for, or plays a very minimal role in mediating somatic CAG/CTG expansions [19], [22], [25], and knockout of Pms2 had a moderate effect of CTG expansion in DM1 mice [24], implicating a role for different MLH1 partners. In the present study the complete absence of HTT CAG expansion in HdhQ111/+Mlh3 mice argues against a role for PMS2 in generating expansions in these mice. Further genetic crosses in both DM1 and HdhQ111 mice would be needed to determine whether the relative contributions of Pms2 and Mlh3 genes in the two mouse models depends on the genomic locus of the repeat and/or strain background. While we do not expect PMS2 levels to be altered in Mlh3 knockout mice [74], additional experiments are needed in Mlh3 and Pms2 knockout mouse tissues to determine whether any compensatory changes in PMS2 or MLH3 proteins, respectively, occur. However, overall, the data thus far indicate that MLH3 is a more significant player than PMS2 in CAG/CTG expansion and suggest that CAG/CTG repeats may preferentially engage a pathway(s) involving MutSβ and MutLγ complexes, as illustrated in Figure 11.

Bottom Line: 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 MutSβ (MSH2-MSH3).The Mlh1 locus is highly polymorphic between B6 and 129 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.

View Article: PubMed Central - PubMed

Affiliation: Center for Human Genetic Research, Massachusetts General Hospital, Boston, Massachusetts, United States of America.

ABSTRACT
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 strain-specific 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 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 MutLγ (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 MutSβ (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.

Show MeSH
Related in: MedlinePlus