The Pld2Flox allele has loxP sites flanking exons 13-15 (including the catalytic activity-dependent first HKD motif) of the phospholipase D2 gene. Removal of the floxed sequences creates a null allele (see Stock No. 028668). These mice may be useful in studying membrane trafficking, membrane fusion, cancer, neurodegeneration and Alzheimer's disease.
Gilbert Di Paolo, Columbia University Medical Center
Phospholipase D2 (Pld2) and its related isoform phospholipase D1 (Pld1) hydrolyze phosphatidylcholine to generate bioactive lipid phosphatidic acid (PA), are implicated in membrane trafficking/membrane fusion, and are elevated/up-regulated in various human cancers.
The Pld2Flox allele has loxP sites flanking Pld2 exons 13-15 of the Pld2 gene. Mice homozygous for Pld2Flox are viable and fertile with no reported abnormalities. Following exposure to Cre recombinase, the floxed sequences (including the catalytic activity-dependent first HKD motif) are deleted in the cre-expressing tissues; creating a null allele.
For example, breeding Pld2Flox mice to germline Cre-expressing mice results in the PLD2 global knockout strain: Pld2- mice are available as Stock No. 028668. Those homozygotes (Pld2-/-) are viable and fertile with no reported abnormalities. No protein expression from the knockout allele is detected by western blot analysis of homozygous brain tissue.
Furthermore, Pld2- animals may be used to study the protective effect of PLD2-deficiency on Alzheimer's disease. Specifically, when Pld2- animals are bred to have a transgene expressing the human APP with Familial Alzheimer's Disease Swedish mutations K670N/M671L, the resulting PLD2-deficient APPSw transgenic mice exhibit reduced Alzheimer's disease characteristics (amyloid-beta-induced synaptic dysfunction and cognitive deficits).
In addition, on a C57BL/6J genetic background, PLD2 ablation does not significantly reduce intestinal tumorigenesis in the ApcMin model (Stock No. 002020). This is contrary to the protective effect observed for PLD1-deficiency in ApcMin/+ mice (see Stock No. 028665 for description).
The Pld2Flox allele was created by Dr. Gilbert Di Paolo (Columbia University Medical Center). A targeting vector was designed to have a loxP site upstream of exon 13, and a frt-flanked neo cassette with 3' loxP site just downstream of exon 15 of the phospholipase D2 gene (Pld2) on chromosome 11. The construct was electroporated into C57BL/6 x 129/SvJ embryonic stem (ES) cells. Correctly targeted ES cells were injected into recipient blastocysts, and chimeric males were bred with C57BL/6J females to establish the Pld2Flox Neo colony.
The donating investigator reports that mice were then bred to the C57BL/6J-congenic FLP deleter strain (Stock No. 005703) for germline removal of the frt-flanked neo. The resulting Pld2Flox colony was backcrossed to C57BL/6J wildtype mice for at least six generations (and the Flp-expressing transgene was removed) prior to sending males with black coat color to The Jackson Laboratory Repository in 2016. Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J oocytes (Stock No. 000664).
Of note, the donating investigator reports that, at least once during backcrossing, a heterozygous female was bred to a C57BL/6J inbred male (thus the Y chromosome of the congenic strain is of C57BL/6J origin).
|Allele Name||targeted mutation 1.2, Gilbert Di Paolo|
|Allele Type||Targeted (Conditional ready (e.g. floxed), No functional change)|
|Gene Symbol and Name||Pld2, phospholipase D2|
|Strain of Origin||C57BL/6 x 129/SvJ|
|Molecular Note||A targeting vector was designed to have a loxP site upstream of exon 13, and an FRT-flanked neo cassette with 3' loxP site just downstream of exon 15 of the gene. Flp-mediated recombination removed the FRT-flanked neo cassette leaving exons 13-15 floxed.|
Mice homozygous for Pld2Flox are viable and fertile with no reported abnormalities. When maintaining a live colony, heterozygous mice may be bred together, to wildtype mice from the colony or to C57BL/6J inbred mice (Stock No. 000664). Alternatively, homozygous mice may be bred together.
When using the Pld2Flox mouse strain in a publication, please cite the originating article(s) and include JAX stock #028666 in your Materials and Methods section.