B6.129X1(FVB)-Nr1h4^tm1.2Gonz/J

Stock number: 028393
Product name: Fxr^fl (loxP::exon::loxP::Δneo)
Research areas: Cancer Research, Cardiovascular Research, Diabetes and Obesity Research, Metabolism Research, Neurobiology Research, Research Tools

Description

Fxr^fl (loxP::exon::loxP::Δneo) is a floxed allele - loxP sites flank the last exon of the Nr1h4 gene. Removal of the floxed sequence creates a null allele. These mice may be useful in studying metabolic disorders, as well as the regulation of bile acid, lipid and glucose metabolism. Δneo)

Detailed description

Nr1h4 encodes the farnesoid X receptor (FXR), a member of the nuclear receptor superfamily of transcription factors that functions to regulate bile acid, lipid and glucose metabolism.


The Fxr^fl (loxP::exon::loxP::Δneo) allele has loxP sites flanking the last exon (encoding a large portion of the ligand binding/dimerization domain and all of the 3' UTR) of the Nr1h4 gene. Mice homozygous for Fxr^fl are viable and fertile with no reported abnormalities.

When bred to mice that express Cre recombinase, the resulting offspring may be useful in generating a tissue-specific FXR knock-out (see comment on knock-out allele further below).
For example, breeding Fxr^fl to germline Cre-expressing mice (see Stock No. 003724) results in the global knock-out allele (FXR/BAR null ; available as Stock No. 007214). FXR/BAR null homozygotes exhibit elevated serum bile acid, cholesterol and triglycerides, increased hepatic cholesterol and triglycerides, and a proatherogenic serum lipoprotein profile.
Furthermore, when bred to Villin-Cre transgenic mice (see Stock No. 018963), the resulting Fxr^ΔIE homozygotes lack FXR function in intestinal intraepithelial cells. Such animals exhibit abnormal bile salt homeostasis and are useful for studying treatments of human metabolic disorders.

Comments on the knock-out allele: Because the floxed exon encodes a large portion of the ligand binding/dimerization domain and all of the 3' UTR (including the polyA signal), it was anticipated that no stable transcript would result from the knock-out allele. However, the knock-out allele expresses a null-specific mRNA predicting a 376 aa protein (compared to the wildtype 451 aa protein). Importantly, western blot detected no Nr1h4 protein product in liver tissue from mice homozygous for the knock-out allele.

Development

The Fxr^fl (loxP::exon::loxP::Δneo) allele was created in the laboratory of Dr. Frank J. Gonzalez (National Institutes of Health (NIH/NCI)) to have the last exon of the nuclear receptor subfamily 1 group H member 4 gene (Nr1h4; Fxr) flanked by loxP sites - this exon encodes a large portion of the ligand binding/dimerization domain (amino acids 406-484) and all of the 3'-untranslated region (including the polyA signal). First, a targeting vector was designed with a loxP site upstream of the last exon, and a loxP-flanked PGK-Neo cassette just downstream of the last exon of the Nr1h4 gene on chromosome 10. The construct was electroporated into undisclosed embryonic stem (ES) cells. Correctly targeted ES cells were injected into recipient blastocysts, and chimeras were bred with undisclosed mice for germline transmission - establishing the 3-loxP mutant mice (loxP::exon::loxP::PGK-Neo::loxP). The donating investigator reports they were bred to cre-expressing mice (on a FVB background) for germline deletion of the PGK-Neo cassette. The resulting Fxr^fl (loxP::exon::loxP::Δneo) mice were thereafter maintained on a C57BL/6N genetic background (with some FVB and/or 129 contributions possible from breeding to different cre-expressing lines) (see SNP results). In 2017, Fxr^fl males (devoid of any cre-expressing alleles) with black coat color were sent to The Jackson Laboratory Repository. Upon arrival, mice were bred to C57BL/6NJ females (Stock No. 005304) for one generation to establish our live colony.
Of note, it is not known if the Y chromosome was fixed to the C57BL/6N background.

In 2017, a 48 SNP (single nucleotide polymorphism) panel analysis, with 43 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 males sent to The Jackson Laboratory Repository. While the 43 markers throughout the genome suggested a C57BL/6 genetic background, 4 of 5 markers that determine C57BL/6J from C57BL/6N were found to be segregating in some mice (chromosomes 1 [~139 Mbp], 6 [~108 Mbp], 8 [~58 Mbp] and 15 [~57 Mbp]). These data suggest that the mice sent to The Jackson Laboratory Repository were on a mixed C57BL/6N;C57BL/6J genetic background (majority C57BL/6N).

Allele

Symbol: Nr1h4^tm1.2Gonz
Name: targeted mutation 1.2, Frank Gonzalez
MGI accession ID: MGI:6121372
Generation method: Targeted
Attributes: Conditional ready (e.g. floxed); No functional change
Marker symbol: Nr1h4
Marker name: nuclear receptor subfamily 1, group H, member 4
Chromosome: 10
Strain of origin: 129X1/SvJ
Molecular note: The targeting construct contains a loxP site inserted in the intron 5' to the last exon and a loxP-flanked neomycin cassette inserted 3' to the last exon. Germline Cre-mediated recombination removed the floxed neo cassette leaving the last exon floxed. The last exon encodes the ligand binding domain and all of the 3' untranslated region.
General note: The version of this allele that contains the neomycin selection cassette, Nr1h4^tm1.1Gonz, has been analyzed.