Polβ/XRCC1 heterodimerization dictates DNA damage recognition and basal Polβ protein levels without interfering with mouse viability or fertility

Koczor, Christopher A.; Thompson, Marlo K; Sharma, Nidhi; Prakash, Aishwarya; Sobol, Robert W.

doi:10.1016/j.dnarep.2023.103452

Cited by 5 publications

(15 citation statements)

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“…A follow‐up study using the interferon‐responsive promoter Mx for expression of Cre suggested that deletion of the Polb gene could be as high as 90%, depending on the tissue (Kuhn et al, 1995). Further, using a mosaic Cre model system that only partially deletes the gene (by crossing the Polb ‐Floxxed mouse with a nestin–Cre transgenic mouse line whereby Cre recombinase is expressed from the rat nestin promoter), the lethality due to the loss of Polβ expression was suppressed, yet the mice were smaller, similar to our functionally defective homozygous Polb L301R/V303R mouse model (Koczor et al, 2023).…”

Section: Gene Knock‐out Mouse Models For Dna Polymerase Betasupporting

confidence: 64%

“…(2) Tg(MMTV-cre)4Mam/J, whereby Cre is expressed in the virgin and lactating mammary gland (Regua et al 2021;Wagner et al 1997Wagner et al , 2001, and ( 3 In this first report, the Polb gene was deleted only in T-cells by virtue of the T-cell selective expression of Cre in the Cre lck transgenic mouse model (Gu et al, 1994;Orban et al, 1992) (see Table 1 gene could be as high as 90%, depending on the tissue (Kuhn et al, 1995). Further, using a mosaic Cre model system that only partially deletes the gene (by crossing the Polb-Floxxed mouse with a nestin-Cre transgenic mouse line whereby Cre recombinase is expressed from the rat nestin promoter), the lethality due to the loss of Polβ expression was suppressed, yet the mice were smaller, similar to our functionally defective homozygous Polb L301R/V303R mouse model (Koczor et al, 2023).…”

Section: Cre-mediated Polb Gene Deletionsupporting

confidence: 64%

“…The most recent of the Polb KI mice is our Polb L301R/V303R KI mouse (Koczor et al, 2023). Importantly, the recruitment of Polβ to sites of DNA damage in the cell is dependent on the interaction of Polβ with Xrcc1 (Figure 1) via a defined protein–protein interaction domain (Fang et al, 2014; Koczor et al, 2021).…”

Section: Gene Knock‐in Mouse Models For Polβmentioning

confidence: 99%

“…These studies identified the minimal amino acid residues required for the Polβ/Xrcc1 interaction in the mouse. The introduction of the double L301R/V303R mutation completely blocks the Polβ/Xrcc1 interaction (Koczor et al, 2023). We used CRISPR‐based gene editing for targeted mutation of these two codons in exon 13 that correspond to the essential amino acid residues for the interaction between Polβ and Xrcc1, modifying these as follows: L301R and V303R (Koczor et al, 2023).…”

Section: Gene Knock‐in Mouse Models For Polβmentioning

confidence: 99%

“…The introduction of the double L301R/V303R mutation completely blocks the Polβ/Xrcc1 interaction (Koczor et al, 2023). We used CRISPR‐based gene editing for targeted mutation of these two codons in exon 13 that correspond to the essential amino acid residues for the interaction between Polβ and Xrcc1, modifying these as follows: L301R and V303R (Koczor et al, 2023). The expressed mutant Polβ protein can no longer interact with Xrcc1, is proteolytically unstable and is defective in recruitment to sites of DNA damage (Koczor et al, 2023).…”

Section: Gene Knock‐in Mouse Models For Polβmentioning

confidence: 99%

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Mouse models to explore the biological and organismic role of DNA polymerase beta

Sobol

2024

Environ and Mol Mutagen

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Gene knock‐out (KO) mouse models for DNA polymerase beta (Polβ) revealed that loss of Polβ leads to neonatal lethality, highlighting the critical organismic role for this DNA polymerase. While biochemical analysis and gene KO cell lines have confirmed its biochemical role in base excision repair and in TET‐mediated demethylation, more long‐lived mouse models continue to be developed to further define its organismic role. The Polb‐KO mouse was the first of the Cre‐mediated tissue‐specific KO mouse models. This technology was exploited to investigate roles for Polβ in V(D)J recombination (variable‐diversity‐joining rearrangement), DNA demethylation, gene complementation, SPO11‐induced DNA double‐strand break repair, germ cell genome stability, as well as neuronal differentiation, susceptibility to genotoxin‐induced DNA damage, and cancer onset. The revolution in knock‐in (KI) mouse models was made possible by CRISPR/cas9‐mediated gene editing directly in C57BL/6 zygotes. This technology has helped identify phenotypes associated with germline or somatic mutants of Polβ. Such KI mouse models have helped uncover the importance of key Polβ active site residues or specific Polβ enzyme activities, such as the PolbY265C mouse that develops lupus symptoms. More recently, we have used this KI technology to mutate the Polb gene with two codon changes, yielding the PolbL301R/V303R mouse. In this KI mouse model, the expressed Polβ protein cannot bind to its obligate heterodimer partner, Xrcc1. Although the expressed mutant Polβ protein is proteolytically unstable and defective in recruitment to sites of DNA damage, the homozygous PolbL301R/V303R mouse is viable and fertile, yet small in stature. We expect that this and additional targeted mouse models under development are poised to reveal new biological and organismic roles for Polβ.

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Section: Gene Knock‐out Mouse Models For Dna Polymerase Betasupporting

confidence: 64%