B6(SJL)-Pik3cd^tm1.1Draw/J

Stock number: 041804
Product name: PIK3CD-E1020K
Strain synonyms: aPIK3CD
Research areas: Cancer Research, Hematological Research, Immunology, Inflammation and Autoimmunity Research, Research Tools

Description

The PIK3CD-E1020K conditional knock-in mice have a Cre-inducible activated form of PIK3CD (E1020K) mutation introduced into the Pik3cd (phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit delta) gene. This mutation is synonymous with the human PIK3CD*E1021K activating mutation associated with Activated PI3K-Delta Syndrome (APDS). Homozygous PIK3CD-E1020K mice before Cre-induction are normal. The model may be useful in studying the role of Pik3cd mutations in the development of APDS.

Detailed description

The PIK3CD-E1020K Cre-inducible knock-in mice have the activating mutation E1020K in the Pik3cd (phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit delta) gene. The mutation is equivalent to the E1021K mutation in humans that causes Activated PI3K-Delta Syndrome (APDS), an immunodeficiency caused by gain-of-function mutations in PIK3CD. Homozygous PIK3CD-E1020K mice exhibit normal breeding, lifespan, and no abnormal phenotypes before Cre-recombination. The following phenotypes were observed in mice crossed with the B cell-specific promoter-regulated Cre recombinase-expressing strains: MB1-Cre (expressed early in B cell development) - bone marrow B lymphopenia, increased numbers of peritoneal B1a cells, splenic marginal zone B cells, and plasma B cells; increased serum IgM and IgG3. CD21-Cre (expressed in peripheral B cells early in maturation)- increased number of peritoneal B1a cells, splenic marginal zone cells; decrease in splenic follicular mature B cells; increased serum IgM; reduced class-switched antibodies following T cell-dependent immunization. AID-Cre (expressed in germinal center B cells)- reduced class-switched antibodies following T cell-dependent immunization. The model may be useful in studying the underlying cellular and molecular mechanisms of APDS and other areas of research, such as immunodeficiency, T cell senescence, lymphadenopathy, and B cell development, activation, and function.

Development

The targeting construct consisting of loxP::endogenous Pik3cd exon 24::frt-neo-frt::mloxP::inverted exon24*E1020K::inward-facing loxP::inward-facing mloxP was designed to insert mutant exon 24 bearing E1020K in 3'→5' orientation downstream of endogenous exon 24, flanked by two sets of loxP and mloxP sites. The mutation introduces Pik3cd c.3058G>A (p.E1020K) in endogenous mouse Pik3cd exon 24, which is equivalent to the human PIK3CD p.E1021K APDS mutation. The reference transcript (Ensembl ID: ENSMUST00000105690) was used for the exon numbering. The targeting vector also included a neo-cassette flanked by frt sites. The construct was electroporated into C57BL/6-derived embryonic stem (ES) cells. Correctly targeted ES cells were injected into recipient blastocysts. The resulting male chimeric mice were bred with C57BL/6 females to establish germline transmission, then heterozygous mice were mated to Flp-strain mice (Stock No. 005703) to excise the frt-flanked neo-selection cassette. The resulting mice with the PIK3CD-E1020K allele were backcrossed to C57BL/6 to establish the colony and were maintained by the donating laboratory. Upon arrival at The Jackson Laboratory, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J (Stock No. 000664) oocytes.

Allele

Symbol: Pik3cd^tm1.1Draw
Name: targeted mutation 1.1, David J Rawlings
MGI accession ID: MGI:8346996
Generation method: Targeted
Attributes: Conditional ready (e.g. floxed); Humanized sequence
Synonyms: PIK3CD-E1020K
Marker symbol: Pik3cd
Marker name: phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit delta
Marker synonyms: 2410099E07Rik; 2610208K16Rik; APDS; IMD14; IMD14A; IMD14B; p110D; P110DELTA; PI3K; PI3Kdelta; ROCHIS
Chromosome: 4
Strain of origin: C57BL/6
Molecular note: A loxP site was inserted into intron 23, and a cassette, consisting of an FRT site flanked neomycin selection cassette, a loxP511 site, a linker, an inverted exon 24 containing a p.E1021K mutation, an inverted loxP site, and an inverted loxP511 site, was inserted downstream of exon 24. The neo cassette was removed by subsequent Flp-mediated recombination. It is a conditional-ready knock-in allele and only after Cre-mediated recombination the endogenous exon 24 will be removed and the mutated exon 24 inverted and expressed. The mutation is equivalent to the same human mutation associated with activated PI3K-delta syndrome (APDS).