Apaf1 (apoptotic peptidase activating factor 1) knock-out mice exhibit a high percentage of perinatal lethality, reduced apoptosis in the brain, craniofacial abnormalities and hyperproliferation of neuronal cells. These mice may be useful for studies of mitochondria-dependent apoptosis.
Dr. Tak Mak, University Health Network/Un of Toronto
The targeted Apaf1 (apoptotic peptidase activating factor 1) gene is involved in the initiation of apoptosis by binding and activating Casp9 in the presence of cytochrome c and dATP. Very few homozygous knock-out mice are produced from a heterozygote x heterozygote mating, and of the homozygotes that survive to birth almost all die within 12 hours of birth. Morphological abnormalities such as ectopic masses on the forehead, a cauliflower-like mass on the face, and a cone shaped head mass and exencephaly are observed in E12.5 homozygous embryos. TUNEL assays detect fewer apoptotic nuclei in the hindbrain, the roof of the midbrain, and the cortex of the forebrain at E14.5. Increased BrdU incorporation reflects hyperproliferation of cells in the hindbrain at E12.5 and the forebrain at E13/5. Apaf1-deficient cells are resistant to a variety of apoptotic stimuli, and the processing of Caspases 2, 3, and 8 is impaired. However, both thymocytes and activated T lymphocytes are sensitive to Fas-induced killing, indicating that Fas-mediated apoptosis in these cells is independent of Apaf1. These mice may be useful for studies of mitochondria-dependent apoptosis.
A targeting vector containing a neomycin cassette, in sense orientation, was used to replace a 4.0 kb fragment containing an exon encoding a portion of the nucleotide binding P loop.The construct was electroporated into 129P2/OlaHsd derived E14K embryonic stem (ES) cells. Correctly targeted ES cells were injected into C57BL/6J blastocysts. The resulting chimeric animals were bred to C57BL/6 to achieve germline transmission and then backcrossed to C57BL/6 for 10 generations. Northern blot analysis of RNA extracted from E9.5 embryos confirms the absence of Apaf1 expression. Upon arrival, sperm was cryopreserved. To establish a live colony, an aliquot of frozen sperm is used to fertilize C57BL/6J.
|Allele Name||targeted mutation 1, Tak W Mak|
|Allele Type||Targeted (Null/Knockout)|
|Allele Synonym(s)||apaf1-; Apaf1tm1Ari|
|Gene Symbol and Name||Apaf1, apoptotic peptidase activating factor 1|
|Strain of Origin||129P2/OlaHsd|
|General Note|| |
Null mutations are lethal, arresting development with phenotypes ascribed to the lack of proper apoptosis, such that too many cells are present, yielding exencephaly and webbed paws. Fas-induced apoptosis, however, appears unaffected in thymocytes, suggesting Apaf1 works by a different mechanism (J:49841). Apaf1 is essential for the activation of other caspase genes. Some phenotypes in Apaf1 mutants overlap with those in mice carrying mutations in Casp3 and Casp9 (J:49840, J:49841).
|Molecular Note||A 4 kb sequence containing an exon that encodes a portion of the nucleotide-binding P loop was replaced by a neomycin cassette.|
When maintaining a live colony, heterozygous mice may be bred to wildtype mice from the colony. Almost all mice homozygous for the mutation die prior to birth or within 12 hours after birth.
When using the Apaf1- mouse strain in a publication, please cite the originating article(s) and include MMRRC stock #44042 in your Materials and Methods section.
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