STOCK Atp4a^tm1Ges Atp12a^tm1Ges/GumzMmjax

Stock number: 039045
Product name: HKα₁ HKα₂ KO
Research areas: Endocrine Deficiency Research, Metabolism Research

Description

The HKα₁ HKα₂ null strain was created by combining Atp4a^tm1Ges (MMRRC Stock No. 036712) and Atp12a^tm1Ges (MMRRC Stock No. 036713). This strain may be useful for studies examining the role of HKα₁ and HKα₂ in mineralocrticoid-induced electrolyte and acid-base disturbances.

Detailed description

Atp4a (H+/K+ exchanging, gastric ATPase, alpha polypeptide or HKα1) encodes the alpha-subunit of a P-type ATPase (gastric proton pump) found in membranes of the gastric parietal cell and the kidney. Atp12a (H+/K+ transporting, nongastric ATPase, alpha polypeptide or HKα2) encodes the alpha-subunit of a P-type family of ion transport ATPases found in the colon, kidneys and prostate. ATP12A mediates K⁺ absorption in the colonic epithelium.

HKα₁ HKα₂ null mice have a neomycin selection cassette disrupting exon 8 of Atp4a (H+/K+ exchanging, gastric ATPase, alpha polypeptide) and a neomycin cassette disrupting exon 20 of Atp12a (H+/K+ transporting, nongastric ATPase, alpha polypeptide). Mice homozygous for the double knockout are viable and fertile. Administration of mineral corticoid desoxycorticosterone pivalate (DOCP) to HKα_1-2 null mice results in lower blood [HCO3-] and less Na+ and K+ retention than wild type. Excess mineralocorticoid is associated with hypertension through changes in Na+, K+ and acid-base homeostasis. In addition, mice exhibit an altered response to dietary challenges such a potassium deprivation, relative to wild type control mice. Dietary potassium deprivation results in a renal and colonic phenotype.

Of note, both alleles are distributed separately as B6.129X1(Cg)-Atp4a^tm1Ges/Mmjax (MMRRC Stock No. 036712) and C.129X1(Cg)-Atp12a^tm1Ges/Mmjax (MMRRC Stock No. 036713).

Development

The HKα₁ HKα₂ null strain was created by combining Atp4a^tm1Ges (MMRRC Stock No. 036712) and Atp12a^tm1Ges (MMRRC Stock No. 036713).

The HKα₁ targeting vector was designed to replace codons 360-390 in exon 8 of the Atp4a (H+/K+ exchanging, gastric ATPase, alpha polypeptide) on chromosome 7 with the neomycin resistance gene. The replacement abolishes the catalytic phosphorylation site required for enzyme activity. The construct was electroporated into 129X1/SvJ derived embryonic stem (ES) cells. Correctly targeted ES cells were injected into blastocysts. The resulting chimeric animals were crossed to Black Swiss mice and offspring were backcrossed to C57BL/6 more than 10 generations. Mice were transferred to the laboratory of Dr. Charles Wingo (University of Florida), and Dr. Michelle Gumz where they were maintained on the C57BL/6 background.

The HKα₂ targeting vector used the neomycin selection cassette to disrupt exon 20 of the Atp12a (H+/K+ transporting, nongastric ATPase, alpha polypeptide) gene on chromosome 14. Exon 20 contains the transmembrane domains critical for ion transport. The construct was electroporated into 129X1/SvJ derived embryonic stem (ES) cells. Correctly targeted ES cells were injected into C57BL/6 blastocysts. The resulting chimeric animals were crossed to Black Swiss mice. Mice from the colony were backcrossed to BALB/c for at least 10 generations. Mice were transferred to the laboratory of Dr. Charles Wingo (University of Florida), and then to the laboratory of Dr. Michelle Gumz (University of Florida) where they were backcrossed to the C57BL/6 background for at least 5 generations be the donating laboratory (see SNP note below). Upon arrival in February 2024, sperm was cryopreserved.

In 2024, 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. Four of the 43 markers throughout the genome were segregating with 129 and/or BALB/c, suggesting an incomplete backcross. Also, all 5 markers that determine C57BL/6J from C57BL/6N were found to be C57BL/6N allele-type.

Allele

Symbol: Atp4a^tm1Ges
Name: targeted mutation 1, Gary E Shull
MGI accession ID: MGI:2158246
Generation method: Targeted
Attributes: Null/Knockout
Synonyms: H/Kalpha^-
Marker symbol: Atp4a
Marker name: ATPase, H+/K+ exchanging, gastric, alpha polypeptide
Marker synonyms: ATP6A; H+/K+-ATPase alpha; H+K+-transporting alpha 1; Hka; Hkatpc
Chromosome: 7
Strain of origin: 129X1/SvJ
Molecular note: A part of exon 8 encoding amino acids 360-390 was replaced with a neomycin selection cassette. Northern blot analysis demonstrated that no detectable full length transcript was produced from this allele in stomach of homozygous mice, although trace levels of a shorter 1 kb transcript was detected. Immunocytochemistry experiments on stomach tissue of homozygous mice demonstrated that no detectable protein was present.

Allele

Symbol: Atp12a^tm1Ges
Name: targeted mutation 1, Gary E Shull
MGI accession ID: MGI:2177160
Generation method: Targeted
Attributes: Null/Knockout
Synonyms: cHKA -
Marker symbol: Atp12a
Marker name: ATPase, H+/K+ transporting, nongastric, alpha polypeptide
Marker synonyms: ATP1AL1; ATPase H+K+-transporting, alpha 2; cHKA; HK; HKalpha2; H-K-ATPase
Chromosome: 14
Strain of origin: 129X1/SvJ
Molecular note: A nenomycin selection cassette was inserted into exon 20. Northern blot and RT-PCR analysis on RNA derived from colon of homozygous mice demonstrated that aberrantly spliced transcripts were produced from this allele, but none of these produced a functional message.