Cul3flox (loxP::frt-neo-frt::exons4-7::loxP) is a cullin 3 hypomorphic allele that is converted to a null allele after Cre recombinase exposure. These mice may be useful in studying the function of cullin-RING-based BTB-CUL3-RBX1 E3 ubiquitin-protein ligase complexes in multiple areas, including autism and cancer.
Jeffrey D Singer, Portland State University
Cul3 encodes the ubiquitin scaffold protein cullin 3; the core component of multiple cullin-RING-based BCR (BTB-CUL3-RBX1) E3 ubiquitin-protein ligase complexes that function to mediate the ubiquitination and subsequent proteasomal degradation of target proteins.
The Cul3flox allele has loxP sites flanking exons 4-7 of the Cul3 gene. The floxed region also contains a frt-flanked PGK-neo cassette upstream of exon 4. Cul3flox is a hypomorphic allele that is converted to a null allele (Cul3- or CR) after Cre recombinase exposure. Compared to wildtype (Cul3+/+) MEFs, the Cul3 expression levels are diminished to ~85% in Cul3flox/+ MEFs, ~70% in Cul3flox/flox MEFs. Removal of the frt-flanked PGK-neo via Flp recombinase generates the Cul3floxΔneo allele, which is also a hypomorph (Cul3 expression reduced to ~90% in Cul3floxΔN/floxΔN MEFs).
When bred to mice that express Cre recombinase, the resulting offspring may be useful in generating tissue-specific CUL3 knockout.
For example, when Cul3flox are bred to also harbor an Albumin-Cre transgene (see Stock No. 016832) and a p53flox allele (see Stock No. 008462), the resulting triple mutant mice with liver-specific simultaneous ablation of CUL3 and p53 are useful to study hepatic progenitor cell transformation into malignant tumor-initiating cells and the subsequent primary hepatocellular carcinoma.
Breeding Cul3flox mice to also have the Pax8-rtTA transgene (Stock No. 007176) and Tet-promoter-driven Cre recombinase transgene (see Stock No. 006234), the resulting triple mutant mice allow doxycycline-inducible renal tubule–specific CUL3 knockout. When temporally induced in adult animals, this triple mutant can be used to study familial hyperkalemic hypertension (FHHt) without the systemic/developmental effects of early CUL3-deficiency.
Mice homozygous for the floxed allele (Cul3flox/flox) are viable and fertile with no reported abnormalities (born at the expected rate and appear normal at birth and throughout development).
A targeting vector was designed by Dr. Jeffrey D. Singer (while at Brown University; now at Portland State University) to have a loxP site followed by a frt-flanked PGK-neo cassette (reverse orientation) upstream of exon 4, and a second loxP site in intron 7 of the cullin 3 gene (Cul3) on chromosome 1. The construct was electroporated into 129S4/SvJaeSor-derived AK7 embryonic stem (ES) cells. Correctly targeted ES cells were injected into recipient blastocysts, and chimeric mice were bred with undisclosed mice for germline transmission. The resulting Cul3flox mice were backcrossed with C57BL/6NCrl wildtype mice for three generations prior to sending to The Jackson Laboratory Repository in 2015. Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6NJ oocytes (Stock No. 005304).
Of note, it is not known if the Y chromosome has been fixed to the C57BL/6N background during backcrossing.
|Allele Name||targeted mutation 1, Jeffrey D Singer|
|Allele Type||Targeted (Conditional ready (e.g. floxed), No functional change)|
|Allele Synonym(s)||Cul3tm1Jdsr; targeted mutation 1, Jeffrey D Singer|
|Gene Symbol and Name||Cul3, cullin 3|
|Gene Synonym(s)||expressed sequence AW146203; AI467304; PHA2E; CUL-3; expressed sequence AI467304; AW146203|
|Strain of Origin||129S4/SvJaeSor|
|Molecular Note||Exons 4-7 were flanked with loxP sites. An FRT-flanked neo was included in intron 3.|
When maintaining a live colony, heterozygous mice may be bred together, to wildtype mice from the colony or to C57BL/6NJ inbred mice (Stock No. 005304). Alternatively, homozygous mice may be bred together.
When using the Cul3flox mouse strain in a publication, please cite the originating article(s) and include JAX stock #028349 in your Materials and Methods section.
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