Heterozygosity for a Hypomorphic Polβ Mutation Reduces the Expansion Frequency in a Mouse Model of the Fragile X-Related Disorders

Abstract

The Fragile X-related disorders (FXDs) are members of the Repeat Expansion Diseases, a group of human genetic conditions resulting from expansion of a specific tandem repeat. The FXDs result from expansion of a CGG/CCG repeat tract in the 5’ UTR of the FMR1 gene. While expansion in a FXD mouse model is known to require some mismatch repair (MMR) proteins, our previous work and work in mouse models of another Repeat Expansion Disease show that early events in the base excision repair (BER) pathway play a role in the expansion process. One model for repeat expansion proposes that a non-canonical MMR process makes use of the nicks generated early in BER to load the MMR machinery that then generates expansions. However, we show here that heterozygosity for a Y265C mutation in Polβ, a key polymerase in the BER pathway, is enough to significantly reduce both the number of expansions seen in paternal gametes and the extent of somatic expansion in some tissues of the FXD mouse. These data suggest that events in the BER pathway downstream of the generation of nicks are also important for repeat expansion. Somewhat surprisingly, while the number of expansions is smaller, the average size of the residual expansions is larger than that seen in WT animals. This may have interesting implications for the mechanism by which BER generates expansions.

Multidrug Resistance-associated Protein-1 (MRP-1)-dependent Glutathione Disulfide (GSSG) Efflux as a Critical Survival Factor for Oxidant-enriched Tumorigenic Endothelial Cells

“…ab12148, lot GR162790-3, Abcam) (35), rabbit polyclonal anti-Apex-1 antibody (1:500 dilution, catalog no. ab 137708, lot GR109895-7, Abcam) (8,36), anti-mouse ␤-actin (1:10,000 dilution, catalog no. A5441, lot 055K4854, Sigma) (8, 37), anti-mouse monoclonal p84 (1:5000 dilution, catalog no.…”

Human MutLγ, the MLH1–MLH3 heterodimer, is an endonuclease that promotes DNA expansion

“…The incision of the loop-containing DNA by the activated MutLα endonuclease occurs in the loop-containing or loop-lacking strand, whereas the cleavage of the loop-containing DNA by the activated MutLγ endonuclease only takes place in the loop-lacking strand. Pol β (66,67) or another DNA polymerase utilizes the generated strand break to perform a DNA synthesis reaction that leads to triplet repeat expansion or contraction in the event that involves MutLα endonuclease and to triplet repeat expansion in the event that entails MutLγ endonuclease. After the DNA synthesis step, the nick is sealed by DNA ligase I or III (68).…”

Close encounters: Moving along bumps, breaks, and bubbles on expanded trinucleotide tracts

“…For longer gaps, proliferating cell nuclear antigen (PCNA) and the flap endonuclease 1 (FEN-1) are required, and may be catalyzed by pol β or by pol δ or pol ε [84,90,93]. Indeed, TNRs expansion is reduced in transgenic mice lacking 8-oxo-guanine glycosylase (OGG1) [94], NEIL1 [95] pol β with normal activity [96] and XPA [97], strongly supporting the notion that excision repair machinery mediates the instability during removal of oxidative damage in vivo (Fig. 3A,B).…”

“…Indeed, both pol β and MSH2-MSH3 are critical to the expansion mechanisms. In mice, a heterozygous Y265C mutation in pol β is sufficient to significantly reduce both the number of expansions in paternal gametes, and the extent of somatic expansion in some tissues of the FXS mouse, which harbor CGG repeats [96]. In mice and/or cells, loss of either MSH2 or MSH3 (and some polymorphisms) [100] also abolishes expansion within exon 1 of the human HD allele [100–104], in the DM1 allele [105–109], in the CGG Fragile X syndrome (FXS) [110,111].…”