This PKD2(S707A/S711A) knock-in strain expressing a functionally diminished enzyme may be useful in studies of T-cell-dependent antibody responses and TCR signaling.
Doreen Cantrell, University of Dundee
These mutant mice carry two point mutations, S707A and S711A (alanine substitution for serine), in the catalytic domain activation loop region, which are phosphorylated by protein kinase C and activate PKD catalytic activity, of the endogenous gene. Mice that are homozygous for the targeted mutation are viable, fertile, normal in size and do not display any gross physical or behavioral abnormalities. Although protein expression levels are not changed, total PRKD1 and PRKD2 activity is impaired (residual level of activity detected) in homozygotes, as measured by immunoprecipitation assay of thymocytes. Phorbol-ester-activated mutant protein produced in splenocytes or thymocytes from homozygotes does not effectively autophosphorylate on Ser873. Homozygotes exhibit normal T cell development and maturation and baseline Ig serum levels, but have a diminished antigen-specific IgM response. Mutant homozygous mice immunized with with NP-OVA have reduced antigen-specific IgM and IgG1 response compared to wildtype controls. Lymphocytes isolated from homozygotes and stimulated with TCR produce reduced levels of IL2 and IFN-gamma.
A targeting vector containing a floxed PGK-NEO cassette, the Ser707 and Ser711 point mutations, was used to insert the point mutations into exon 16 of the gene. The construct was electroporated into C57BL/6 derived Bruce4 embryonic stem (ES) cells. Correctly targeted ES cells were injected into BALB/c blastocysts. Germline transmission was confirmed and the mice were crossed to Oz Cre Deleter mice on the C57BL/6 background to remove the floxed selection cassette. The cre allele was then removed by selective breeding.
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 1.1, Doreen A Cantrell|
|Allele Type||Targeted (Not Applicable)|
|Allele Synonym(s)||PKD2(S707A/S711A); PKD2SSAA|
|Gene Symbol and Name||Prkd2, protein kinase D2|
|Promoter||Prkd2, protein kinase D2, mouse, laboratory|
|Strain of Origin||B6.Cg-Thy1a|
|Molecular Note||Exon 16 was replaced with one containing nucleotide substitutions that result in the amino acid substitution of alanine for serine at positions 707 and 711 (S707A, S711A). A floxed neo cassette inserted downstream of the modified exon 16 was removed by cre-mediated recombination.|
|Mutations Made By|| |
Elizabeth Emslie, University of Dundee
When maintaining a live colony, these mice can be bred as homozygotes.
When using the B6.Cg-Prkd2tm1.1Daca/J mouse strain in a publication, please cite the originating article(s) and include JAX stock #017285 in your Materials and Methods section.