This strain carries a mutant allele with alanine substitutions for prolines 187 and 190 in exon 4; homozygotes fail to develop germinal centers in the spleen, have reduced OVA-specific antibody levels, display reduced airway hyperresponsiveness and exhibit less severe experimental autoimmune encephalomyelitis than wildtype controls. This mutant mouse strain may be useful in studies of humoral immune response and T cell proliferation.
Jonathan M Green, Washington University School of Medicine
Mice that are homozygous for the targeted mutation are viable, fertile, normal in size and do not display any gross physical or behavioral abnormalities. These mice carry a mutant allele with alanine substitutions for prolines 187 and 190 in exon 4 (CD28 AYAA, or P187,190A), which encodes the SH3 domain-containing protein interaction motif. Mutant gene product (mRNA) is detected by RT-PCR analysis of thymocytes and peripheral T cells. Mutant protein expression levels are similar to wildtype protein levels in control mice, as assayed by flow cytometry. T cells from mice homozygous for the CD28 AYAA KI mutation have impaired IL-2 production and secretion. Homozygotes fail to develop germinal centers in the spleen, reduced OVA-specific antibody level and display reduced airway hyperresponsiveness after sensitization and challenge with OVA. Homozygotes exhibit less severe experimental autoimmune encephalomyelitis than wildtype controls, although CNS inflammatory infiltration is similar to wildtype. This mutant mouse strain may be useful in studies of humoral immune response and T cell proliferation.
A targeting vector containing a floxed neo cassette was used to insert alanine substitutions for prolines 187 and 190 in exon 4, which encodes the SH3 domain?containing protein interaction motif. The construct was electroporated into 129X1/SvJ derived RW-4 embryonic stem (ES) cells. Correctly targeted ES cells were injected into C57BL/6 blastocysts. The resulting chimeric animals were crossed to a transgenic EllA-cre strain on the C57BL/6 background to remove the floxed neo selection cassette. The mice were then backcrossed to C57BL/6 for 10 generations. Upon arrival at The Jackson Laboratory, the mice were crossed to C57BL/6J once to establish the colony.
|Allele Name||targeted mutation 1, Jonathan M Green|
|Allele Type||Targeted (Not Specified)|
|Gene Symbol and Name||Cd28, CD28 antigen|
|Promoter||Cd28, CD28 antigen, mouse, laboratory|
|Strain of Origin||129X1/SvJ|
|Molecular Note||A targeting vector was designed to insert alanine substitutions for prolines 187 and 190 in exon 4 (P187A, P190A), thereby disrupting the motif responsible for interaction with SH3 domain-containing proteins. A floxed neo was included with the vector and subsequently removed via cre mediated recombination.|
|Mutations Made By|| |
Jonathan Green, Washington University School of Medicine
When maintaining a live colony, these mice can be bred as homozygotes.
When using the CD28-AYAA mouse strain in a publication, please cite the originating article(s) and include MMRRC stock #32039 in your Materials and Methods section.