A mutation in the endonuclease domain of mouse MLH3 reveals novel roles for MutLγ during crossover formation in meiotic prophase I

Toledo, Manuel; Sun, Xianfei; Brieño‐Enríquez, Miguel A.; Raghavan, Vandana; Gray, Stephen; Pea, Jeffrey; Venkatesh, Anita; Patel, Lekha; Borst, P.; Alani, Eric; Cohen, Paula E.

doi:10.1101/517748

Cited by 9 publications

(30 citation statements)

References 69 publications

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“…The original repeat numbers were 280 and 282 respectively. The lack of suitable antibodies for detecting mouse MLH3 has been reported previously [ 38 ] and our own testing of multiple MLH3 antibodies also did not identify any suitable for verifying the Mlh3 -/- lines by western blot. Loss of MLH3 had no effect on the levels of MLH1, PMS1 or PMS2 ( Fig 2C ), consistent with previous reports and with the idea that loss of MLH3 does not affect the levels of MutLα and MutLβ [ 39 ].…”

Section: Resultsmentioning

confidence: 49%

All three mammalian MutL complexes are required for repeat expansion in a mouse cell model of the Fragile X-related disorders

et al. 2020

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Expansion of a CGG-repeat tract in the 5' untranslated region of the FMR1 gene causes the fragile X-related disorders (FXDs; aka the FMR1 disorders). The expansion mechanism is likely shared by the 35+ other diseases resulting from expansion of a disease-specific microsatellite, but many steps in this process are unknown. We have shown previously that expansion is dependent upon functional mismatch repair proteins, including an absolute requirement for MutLγ, one of the three MutL heterodimeric complexes found in mammalian cells. We demonstrate here that both MutLα and MutLβ, the two other MutL complexes present in mammalian cells, are also required for most, if not all, expansions in a mouse embryonic stem cell model of the FXDs. A role for MutLα and MutLβ is consistent with human GWA studies implicating these complexes as modifiers of expansion risk in other Repeat Expansion Diseases. The requirement for all three complexes suggests a novel model in which these complexes cooperate to generate expansions. It also suggests that the PMS1 subunit of MutLβ may be a reasonable therapeutic target in those diseases in which somatic expansion is an important disease modifier.

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Section: Resultsmentioning

confidence: 49%

All three mammalian MutL complexes are required for repeat expansion in a mouse cell model of the Fragile X-related disorders

et al. 2020

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“…Alternatively, excessive stabilisation impairs the resolution of recombination intermediates as crossovers or non-crossovers, which manifests in a persistence of unrepaired DSBs and abnormally high numbers of RNF212/MSH4-marked recombination foci beyond mid pachytene, as seen in HEI10-and CNTD1-deficient meiocytes 15,16,18 . In contrast, if only crossover maturation is defective, paring down of RNF212/MSH4 to a few recombination complexes per chromosome occurs 14,31 , but mature crossover-specific recombination foci (marked by CDK2, HEI10 and MutLγ) and/or crossovers do not form as observed in Mlh3 −/− mice 6,15,31 . Hence, to test PRR19 involvement in crossover differentiation, we examined DSB repair kinetics in Prr19 −/− spermatocytes by detecting markers of unrepaired DSBs.…”

Section: Resultsmentioning

confidence: 96%

“…Whereas maturation of crossover precursors into crossovers requires MutLγ, differentiation of crossover precursors from noncrossovers does not, as judged by the paring down of RNF212/ MSH4 foci to a few per chromosome in mid pachytene Mlh3 −/− spermatocytes 14,31 . Whereas MLH1 foci depend on MLH3, MLH3 focus formation is only enhanced by MLH1 6,21 .…”

Section: Resultsmentioning

confidence: 99%

Proline-rich protein PRR19 functions with cyclin-like CNTD1 to promote meiotic crossing over in mouse

et al. 2020

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Orderly chromosome segregation is enabled by crossovers between homologous chromosomes in the first meiotic division. Crossovers arise from recombination-mediated repair of programmed DNA double-strand breaks (DSBs). Multiple DSBs initiate recombination, and most are repaired without crossover formation, although one or more generate crossovers on each chromosome. Although the underlying mechanisms are ill-defined, the differentiation and maturation of crossover-specific recombination intermediates requires the cyclin-like CNTD1. Here, we identify PRR19 as a partner of CNTD1. We find that, like CNTD1, PRR19 is required for timely DSB repair and the formation of crossover-specific recombination complexes. PRR19 and CNTD1 co-localise at crossover sites, physically interact, and are interdependent for accumulation, indicating a PRR19-CNTD1 partnership in crossing over. Further, we show that CNTD1 interacts with a cyclin-dependent kinase, CDK2, which also accumulates in crossover-specific recombination complexes. Thus, the PRR19-CNTD1 complex may enable crossover differentiation by regulating CDK2.

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“…Htt Q111/+ and Mlh3 WT/DN mice were crossed together to generate Htt Q111/+ Mlh3 WT/DN and Htt +/+ Mlh3 WT/DN mice, which were subsequently intercrossed to generate heterozygous Htt Q111/+ mice (hereafter referred to as Htt Q111 , for simplicity) that were either wild type (Mlh3 WT/WT ), heterozygous (Mlh3 WT/DN ), or homozygous (Mlh3 DN/DN ) for the Mlh3 DN point mutation. Genotyping of the Htt Q111 and Mlh3 DN alleles in genomic DNA extracted from tail or ear biopsies at weaning, or in adult tissues for genotype confirmation, was carried out as previously described (16,42). All animal procedures were carried out to minimize pain and discomfort, under approved IACUC protocols of the Massachusetts General Hospital.…”

Section: Micementioning

confidence: 99%

Somatic CAG expansion in Huntington’s disease is dependent on the MLH3 endonuclease domain, which can be excluded viaMLH3splice redirection to suppress expansion

Roy

Vitalo

Andrew

et al. 2020

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Somatic expansion of the CAG repeat tract that causes Huntington's disease (HD) is thought to contribute to the rate of disease pathogenesis. Therefore, factors influencing repeat expansion are potential therapeutic targets. Genes in the DNA mismatch repair pathway are critical drivers of somatic expansion in HD mouse models. Here, we have tested, using genetic and pharmacological approaches, the role of the endonuclease domain of the mismatch repair protein MLH3 in somatic CAG expansion in HD mice and patient cells. A point mutation in the MLH3 endonuclease domain completely eliminated CAG expansion in the brain and peripheral tissues of a HD knock-in mouse model (HttQ111). To test whether the MLH3 endonuclease could be manipulated pharmacologically, we delivered splice switching oligonucleotides in mice to redirect Mlh3 splicing to exclude the endonuclease domain. Splice redirection to an isoform lacking the endonuclease domain was associated with reduced CAG expansion. Finally, CAG expansion in HD patient-derived primary fibroblasts was also significantly reduced by redirecting MLH3 splicing to the endogenous endonuclease domain-lacking isoform. These data indicate the potential of targeting the MLH3 endonuclease domain to slow somatic CAG repeat expansion in HD, a therapeutic strategy that may be applicable across multiple repeat expansion disorders.

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A mutation in the endonuclease domain of mouse MLH3 reveals novel roles for MutLγ during crossover formation in meiotic prophase I

Cited by 9 publications

References 69 publications

All three mammalian MutL complexes are required for repeat expansion in a mouse cell model of the Fragile X-related disorders

All three mammalian MutL complexes are required for repeat expansion in a mouse cell model of the Fragile X-related disorders

Proline-rich protein PRR19 functions with cyclin-like CNTD1 to promote meiotic crossing over in mouse

Somatic CAG expansion in Huntington’s disease is dependent on the MLH3 endonuclease domain, which can be excluded viaMLH3splice redirection to suppress expansion

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