This Car5a knockout strain is useful in studies of oxidative stress, ureagenesis (ammonia detoxification), gluconeogenesis and lipogenesis.
William S Sly, Saint Louis University Medical Center
The Car5a gene encodes for mitochondrial carbonic anhydrase 5a, a zinc metalloenzyme that catalyze the reversible hydration of carbon dioxide (to carbonic acid).
Carbonic anhydrase 5a is expressed primarily in the liver, is involved in ureagenesis (ammonia detoxification), gluconeogenesis, lipogenesis, and mitochondrial oxidative stress. These mice carry a knock out mutation for the Car5a gene, in which exon 3 and flanking sequence (150 bp of intron 2 and 3.05 kb of intron 3) have been replaced by a PGK-NEO cassette. Exon 3 encodes 2 of the 3 zinc binding histidine residues, and the mutation introduces a frame shift at exon 4. No gene product (mRNA or protein) is detected by RT-PCR or Western blot analysis on total RNA from liver and kidney. Although homozygotes are viable, they are smaller in size than wildtype controls. The Donating Investigator reports that homozygotes are fertile but do not breed well. Homozygotes exhibit elevated levels of circulating ammonia, alpha-ketoglutarate, fumarate, malate, acetoacetate, subaric acid, and sebacic acid. Urinary metabolites of homozygotes are also abnormal.
When bred to Car5b knockout mice (Stock No. 025570), the resulting double mutants die shortly after weaning, and show even greater impairment in ureagenesis and gluconeogenesis compared to the single mutant Car5a knockout mice.
A targeting vector containing a PGK-NEO cassette was used to disrupt exon 3 and flanking sequence (150 bp of intron 2 and 3.05 kb of intron 3). The construct was electroporated into 129X1/SvJ derived RW-4 embryonic stem (ES) cells. Correctly targeted ES cells were injected into C57BL/6J blastocysts. The resulting chimeric male animals were bred to wildtype C57BL/6J females to achieve germline transmission. Upon arrival at The Jackson Laboratory, the mice were crossed to C57BL/6J (Stock No. 000664) at least once to establish the colony.
|Allele Name||targeted mutation 1, William S Sly|
|Allele Type||Targeted (Null/Knockout)|
|Gene Symbol and Name||Car5a, carbonic anhydrase 5a, mitochondrial|
|Strain of Origin||129X1/SvJ|
|Molecular Note||All of exon 3 as well as 150 bp of intron 2 and 3.05 kb of intron 3 were replaced by a PGK-neomycin resistance gene. This removed two of the three zinc binding histidine residues and created a frameshift beginning in exon 4. No protein was detected by Western blot analysis.|
When maintaining a live colony, heterozygous mice may be bred together, to wildtype siblings, or to C57BL/6J inbred mice (Stock No. 000664). The Donating Investigator reports that homozygotes do not breed well.
When using the B6;129X1-Car5atm1Sly/J mouse strain in a publication, please cite the originating article(s) and include JAX stock #025331 in your Materials and Methods section.