B6;129S1-Aptx^tm1Pmc/PmthJ

Stock number: 037331
Product name: Aptx^-
Research areas: Cancer Research, Cell Biology Research, Mouse/Human Gene Homologs

Description

Aptx^-/- mice lack the two exons encoding the HIT domain of the aprataxin gene. This strain may be useful for studying the role of oxidative stress and DNA break repair in the onset of neurological conditions such as Ataxia Telangiectasia (A-T) and Ataxia oculomotor apraxia 1 (AOA1).

Detailed description

Aptx^-/- mice lack the two exons encoding the histidine triad (HIT) domain of the aprataxin (Aptx) gene. Aptx, a member of the HIT superfamily, encodes a DNA-binding protein involved in single-strand DNA break repair, double-strand DNA break repair, and base excision repair which protects cells from DNA lesions caused by damage. The HIT domain contains the catalytic residue responsible for the DNA hydrolase activity of APTX. Removal of the HIT domain results in a frame-shift that causes the loss of the C-terminal 181 amino acids including the HIT domain as well as the zinc finger domains, and also results in a stop codon at amino acid 161. Mice that are homozygous for this allele are viable and fertile.

Oxidative stress induced damage to DNA has been suggested to be a significant factor in the onset of some neurological conditions including Alzheimer's disease, Parkinson's disease, and Huntington's disease, as well as Ataxia Telangiectasia (A-T) and Ataxia oculomotor apraxia 1 (AOA1), the most common recessive spinocerebellar ataxia. AOA1 is characterized by ataxia and oculomotor apraxia. A lack of functional aprataxin disrupts DNA repair and can lead to an accumulation of damage in cells, particularly affecting cells of the cerebellum, which can lead to cell death in the cerebellum, causing the characteristic movement problems of AOA1.

Astrocytes from these Aptx^-/- mice fail to support short-patch repair of adenylated oxidative single strand breaks unlike wild-type cells. Following treatment with hydrogen peroxide and aphidicolin, single strand break repair in astrocytes is slowed compared to in similarly treated wild-type cells.

When bred to Atm^R35X/R35X mice (Stock No. 037332), resulting double mutant mice develop a progressively severe ataxic phenotype associated with atrophy of the cerebellar molecular layer. They also exhibit a high predisposition to developing cancer (thymomas) and immune abnormalities such as impaired early thymocyte development and T-cell maturation, symptoms characteristic of A-T.

Development

The Aptx^- targeting vector was designed to flank two exons encompassing the HIT domain of the aprataxin (Aptx) gene. The targeting vector also included a downstream loxP-flanked neomycin resistance (neo) cassette. The construct was electroporated into 129S1/Sv-Oca2⁺ Tyr⁺ Kitl⁺-derived W9.5 embryonic stem (ES) cells. Correctly targeted ES cells were transiently transfected with a Cre expression plasmid to delete the neo cassette and the two floxed exons. Correctly recombined ES cells, with the neo cassette and floxed exons removed, were injected into blastocysts and resulting chimeric males were bred with C57BL/6J females (Stock No. 000664). Resulting offspring were bred to establish a colony of Aptx^-/- mice, and these mice were further backcrossed to C57BL/6J mice for a total of 10 generations by the donating laboratory (see SNP note below). Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J oocytes.

In 2022, a 48 SNP (single nucleotide polymorphism) panel analysis, with 43 markers covering all 19 chromosomes and the X chromosome, as well as 5 markers that distinguish between the C57BL/6J and C57BL/6N substrains, was performed on the males sent to The Jackson Laboratory Repository. Many of the 43 markers throughout the genome were segregating with 129, suggesting an incomplete backcross.

Allele

Symbol: Aptx^tm1Pmc
Name: targeted mutation 1, Peter J McKinnon
MGI accession ID: MGI:3686875
Generation method: Targeted
Attributes: Null/Knockout
Marker symbol: Aptx
Marker name: aprataxin
Chromosome: 4
Strain of origin: 129S1/Sv-Oca2⁺ Tyr⁺ Kitl⁺
Molecular note: The two exons encompassing the HIT domain were deleted using cre mediated recombination. The deletion resulted in a frame-shift that caused the loss of the C-terminal 181 amino acids including the histidine triad and zinc finger domains. The mutant transcript was found to be expressed in various tissues, including brain, by Northern blot analysis.