B6.129(Cg)-Foxp3^{tm4(YFP/icre)Ayr}/J

Stock number: 016959
Product name: Foxp3^YFP-cre
Research areas: Immunology, Inflammation and Autoimmunity Research, Research Tools

Description

Foxp3^YFP/cre mutant mice express a knocked-in yellow fluorescent protein/iCre-recombinase fusion protein from the Foxp3 locus without disrupting expression of the endogenous Foxp3 gene. These mice may be useful for studying regulatory T cells suppression of autoimmunity and immune dysfunction.

Detailed description

Foxp3^YFP/cre mutant mice contain an internal ribosome entry site (IRES) and a yellow fluorescent protein (YFP) fused to a codon optimized Cre Recombinase (iCre) sequence downstream of the internal stop codon of the forkhead box P3 (Foxp3) gene. FOXP3 is a transcription factor required for the development and function of regulatory T (T _reg) cells, which regulate suppression of self-reactive T cells. In these mice, YFP expression is evident in FOXP3⁺ T_reg cells. Homozygous females and hemizygous males are viable, fertile and normal in size. When bred with mice containing a loxP-flanked sequence, Cre-mediated recombination will result in deletion of the floxed sequences in T _reg cells of the offspring.

For example, when these mice are bred with mice containing a floxed-interleukin 10 allele, deletion of IL10 in T _reg cells results in spontaneous colitis, and immune-mediated inflammation in the lungs and skin.

When bred to IR^Lox mice (Stock No. 006955), 50% of the F1 generation mice show evidence of ectopic recombination and 10% of the F2 generation mice have germline Cre recombination activity. This spontaneous ectopic recombination is seen in both somatic and germ tissues, leading to germline deletion of the insulin receptor (Insr) (Wu et al. Immunology 2020).

Development

The Foxp3^YFP/cre targeting vector was designed with an internal ribosome entry site (IRES), and a yellow fluorescent protein (YFP) fused to a codon optimized Cre Recombinase (iCre) sequence, followed by a frt-flanked neomycin (neo) resistance cassette inserted downstream of the internal stop codon of the X-linked forkhead box P3 (Foxp3) gene. This construct was injected into (129X1/SvJ x 129S1/Sv)F1-Kitl⁺-derived R1 embryonic stem (ES) cells. Correctly targeted ES cells were injected into C57BL/6 blastocysts, and the resulting chimeric males were bred to FLP-deleter mice (on a B6 background) to remove the neo cassette. The donating investigator reported that these mice were then backcrossed to C57BL/6J mice for at least 8 generations (see SNP note below). Upon arrival at The Jackson Laboratory, mice were bred to C57BL/6J inbred mice (Stock No. 000664) for at least one generation.

A 32 SNP (single nucleotide polymorphism) panel analysis, with 27 markers covering all 19 chromosomes and the X chromosome, as well as 5 markers that distinguish between the C57BL/6J and C57BL/6N substrains, was performed on the rederived living colony at The Jackson Laboratory Repository. While the 27 markers throughout the genome suggested a C57BL/6 genetic background, 4 markers that determine C57BL/6J from C57BL/6N were found to be segregating. These data suggest the mice sent to The Jackson Laboratory Repository were on a C57BL/6 ; C57BL/6N mixed genetic background.

 

Allele

Symbol: Foxp3^{tm4(YFP/icre)Ayr}
Name: targeted mutation 4, Alexander Y Rudensky
MGI accession ID: MGI:3790499
Generation method: Targeted
Attributes: Recombinase-expressing
Marker symbol: Foxp3
Marker name: forkhead box P3
Chromosome: X
Strain of origin: (129X1/SvJ x 129S1/Sv)F1-Kitl⁺
Expressed genes: cre,cre recombinase,bacteriophage P1; YFP,Yellow Fluorescent Protein,jellyfish
Site of expression: YFP florescence is detected in regulatory T cells.
Molecular note: A YFP/icre fusion construct with an upstream IRES element and a downstream frt-flanked neomycin resistance cassette was placed into the downstream untranslated region of the gene. Chimeric mice were crossed with FLPe expressing mice to remove the neomycin selection cassette. YFP-cre expression based on YFP flouresecene was only detected in regulatory T cells. Crossing these mice with mice that contain a cre-mediated reporter gene also demonstrated low level cre expression in about 2-10% of hematopoietic lineage cells including immature and mature B cells, T cells, myeloid cells and bone marrow precursor cells.