The Apc15lox floxed allele has loxP sites flanking the adenomatosis polyposis coli gene last coding exon (encoding the functional domains involved in β-catenin regulation, the C-terminal domains needed for microtubule-binding and the polyA sequences). Removal of the floxed sequence results in Apc inactivation and concomitant, constitutive activation of the Wnt/β-catenin signal transduction pathway. These mice may be useful in studying Cre recombinase-inducible intestinal tumorigenesis, as well as Apc deletion in other tissues.
Ron M.J.M Smits, Erasmus University Medical Center
Riccardo Fodde, Erasmus Medical Center
Apc encodes the adenomatosis polyposis coli protein with diverse cellular functions including cellular proliferation, differentiation, cytoskeleton regulation, migration and apoptosis. APC mutations are commonly associated with intestinal tumorigenesis, and increasingly implicated with phenotypes outside of the intestine.
The Apc15lox floxed allele has loxP sites flanking the Apc last exon that encodes ~2200 amino acids with all the functional domains involved in β-catenin regulation, the C-terminal domains needed for microtubule-binding and the polyA sequences [Robanus-Maandag et al. 2010 Carcinogenesis 31:946].
Prior to Cre recombinase exposure, mice homozygous for the floxed allele (Apc15lox/15lox) are viable and fertile with no reported abnormalities.
When bred to mice that express Cre recombinase, the resulting offspring may be useful in generating tissue-specific Apc inactivation and concomitant, constitutive activation of the Wnt/β-catenin signal transduction pathway. Specifically, deletion of the Apc last exon results in a truncated protein (ApcΔ15) that is devoid of its main function (β-catenin regulation) and, because the polyA-deficient mRNA is unstable, is expressed at very low levels (~5% of wildtype).
For example, when bred to germline Cre-expressing mice (EIIa-Cre; see Stock No. 003724), the resulting C57BL/6 ApcΔ15/+ mice develop multiple tumors/adenomas in the small intestine at an early age - similar to those reported in small intestine of the C57BL/6J ApcMin/+ mice (Stock No. 002020).
In addition, to study tumorigenesis in the large intestine, one may breed Apc15lox mice to animals with cre expression directed more specifically to large intestinal epithelium - for example, CDX2P-NLS Cre transgenic mice (Stock No. 009350). Those resulting mice may be useful in studying familial adenomatous polyposis (FAP)-associated colorectal cancer and sporadic colorectal cancer.
A targeting vector was designed by Drs. Ron M.J.M. Smits and Riccardo Fodde (while at Leiden University Medical Center, The Netherlands; later at Erasmus Medical Center) to have a loxP-flanked PGK-neo cassette just upstream of, and a loxP site just downstream of the last coding exon of the adenomatosis polyposis coli locus (Apc) on chromosome 18. See exon numbering note below. The construct was electroporated into 129P2/OlaHsd-derived IB10 embryonic stem (ES) cells. ES cells with the Apcneo-15lox genotype were transiently transfected with a Cre recombinase-expressing plasmid. The resulting ES cells with the Apc15lox genotype (loxP-flanked last coding exon with the PGK-neo cassette deleted) were injected into recipient blastocysts. Chimeric males were bred with C57BL/6J females for germline transmission and to establish the colony. The Apc15lox colony was backcrossed to C57BL/6J inbred mice for at least 30 generations prior to sending males to The Jackson Laboratory Repository in 2015. Upon arrival, sperm was cryopreserved. To establish a live colony, an aliquot of frozen sperm may be used to fertilize C57BL/6J oocytes (Stock No. 000664).
Of note, the donating investigator reports that, at least once during backcrossing, a heterozygous female was bred to a C57BL/6J inbred male (thus the Y chromosome of the congenic strain is of C57BL/6J origin).
The original publication describing the Apc15lox mice identified the Apc locus to have 15 coding exons, the last of which encodes ~2200 amino acids with all the functional domains involved in β-catenin regulation, the C-terminal domains needed for microtubule-binding and the polyA sequences [Robanus-Maandag et al. 2010 Carcinogenesis 31:946]. In 2018, genomic structure analysis of the Apc locus revealed it to have an additional upstream exon (with the ATG start site in exon 2). As such, this last exon number is changed from 15 to 16. Because the Apc15lox allele has the last coding exon floxed, an alternate synonym may be Apc16lox.
|Allele Name||targeted mutation 1, Ron Smits|
|Allele Type||Targeted (Conditional ready (e.g. floxed), No functional change)|
|Allele Synonym(s)||Apc15lox; Apc16lox; Apcex15lox; Apctm1Ecrm|
|Gene Symbol and Name||Apc, adenomatosis polyposis coli|
|Strain of Origin||129P2/OlaHsd|
|Molecular Note||A floxed neo cassette was inserted upstream of exon 16 (formerly 15) and an additional loxP site was inserted downstream of exon 16. Cre mediated recombination removed the neo cassette leaving exon 16 floxed.|
When maintaining a live colony, heterozygous mice may be bred together, to wildtype mice from the colony or to C57BL/6J inbred mice (Stock No. 000664). Alternatively, homozygous mice may be bred together.
When using the Apc15lox mouse strain in a publication, please cite the originating article(s) and include JAX stock #029275 in your Materials and Methods section.