B6.129-Kras^tm1Bbd/J

Stock number: 026924
Product name: Kras^LSLG12Vgeo
Research areas: Cancer Research, Developmental Biology Research, Research Tools

Description

Mice bearing the Cre recombinase-inducible Kras^LSLG12Vgeo allele may be useful for studying K-RAS driven lung adenocarcinomas.

Detailed description

Kras^LSLG12Vgeo mice contain a 5’ floxed-STOP followed by a G12V mutation in the Kras (v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog) gene, and a 3’ IRES-β-geo cassette. Ras genes are small GTPases that act downstream of receptors and nonreceptor tyrosine protein kinases in multiple pathways to transfer information to the nucleus. K-ras has been found to be activated in human tumors in pancreatic, colon, breast, and lung cancer. The unrecombined Kras^LSLG12Vgeo allele is null. Hence, homozygous embryos die at midgestation as the Kras^–/– mice. Cre-dependent removal of the floxed-STOP cassette allows concomitant expression of the Kras^G12V oncoprotein along with expression of the chimeric Geo protein which has lacZ activity. Expression of the Kras^G12V oncoprotein in germ line, by breeding to B6.C-Tg(CMV-cre)1Cgn/J mice (Stock No. 006054), is embryonic lethal with embryos dying at around E9.5. In some cases partial recombination leading to mosaic embryos was observed. Embryonic lethality in these cases depended on the extent of KrasG12V oncoprotein expression. No such mosaic embryos were observed when crossing to the EIIa-Cre transgenic strain, which resulted in more efficient cleavage of the STOP cassette in the germ line.

Ubiquitous expression of the Kras^G12V oncoprotein in adult mice, by crossing to strains that express the inducible CreERT2 recombinase under the control of various ubiquitous promoters followed by exposure to a tamoxifen diet, selectively results in the induction of lung adenomas and adenocarcinomas. In some occasions, benign lesions have been observed in other tissues such as stomach and skin papillomas. Lung adenocarcinomas are also observed upon tracheal infection with Adeno-Cre particles. Treated mice display multifocal lesions in lungs, ranging from small patches of bronchiolo-alveolar hyperplasias to large bronchiolo-alveolar adenomas and adenocarcinomas that compress adjacent lung structures. Alveolar type II (ATII) cells yield malignant adenocarcinomas while Clara cells required an inflammatory response to yield hyperplasias and adenomas.

When Kras^LSLG12Vgeo mice were bred to bitransgenic Elas-tTA/tetO-Cre mice that express the Cre recombinase under the control of the Elastase promoter in a tet-off system, they developed pancreatic intraepithelial neoplasias (PanIN) with 100% incidence. They also developed full blown pancreatic ductal adenocarcinomas albeit with lesser frequency (20% by one year of age). Exposure of these mice to the antibiotic doxycyclin in order to prevent expression of the Cre recombinase until the mice reach adulthood, completely prevents the development of PanIN lesions in spite of wide spread expression of the Kras^G12V oncoprotein in acinar cells. Induction of PanIN lesions and occasional PDAC tumors in these adult mice can be achieved by inducing pancreatic damage by treating them with low doses of caerulein.

Finally, embryonic fibroblasts (MEFs) from these mice do not undergo proliferative senescence and proliferate continuously as immortal cells upon Cre-mediated recombination of the Kras^LSLG12Vgeo allele.

Development

A targeting vector was designed to insert an internal ribosome entry site (IRES)-β-geo cassette into the 3’ untranslated region of the Kras (v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog) gene. A second targeting event inserted a floxed STOP and hygromycin (hygro) cassette upstream of the Kras initiation codon. This event also introduced two point mutations in codon 12 of exon 1 (GGT to GTA) replacing a glycine residue with a valine (G12V), a mutation commonly found in human tumors. The construct was electroporated 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 C57BL/6 females to generate a colony of Kras^LSLG12Vgeo mice. These mice were bred to C57BL/6.OlaHsd mice for at least 15 generations. Upon arrival at The Jackson Laboratory, mice were bred to C57BL/6J (Stock No. 000664) for at least one generation to establish the colony.

Allele

Symbol: Kras^tm1Bbd
Name: targeted mutation 1, Mariano Barbacid
MGI accession ID: MGI:3582830
Generation method: Targeted
Marker symbol: Kras
Marker name: Kirsten rat sarcoma viral oncogene homolog
Chromosome: 6
Strain of origin: (129X1/SvJ x 129S1/Sv)F1-Kitl⁺
Expressed genes: Bgeo,fusion of beta-galactosidase and neomycin phosphotransferase genes,E. coli; Kras,KRAS proto-oncogene, GTPase,rat
Site of expression: Bi-cistronic expression of Kras and lacZ will occur in cells where promoters driving Cre recombinase are expressed.
Molecular note: In one homologous recombination event an IRES Beta-geo cassette was inserted into the 3' untranslated region and in a second homologous recombination event a mutation changing glycine codon 12 in exon 1 into a valine (p.G12V) was inserted along with a loxP site flanked transcriptional stop and hygromycin resistance gene cassette. This resulted in bicistronic Cre recombination-dependent expression of the oncogenic Kras allele and Beta-geo.