The Wt1flox allele has loxP sites flanking exons 8-9 of the Wilms tumor 1 homolog gene. Removal of the floxed sequence creates a null allele. These mice may be useful for generating tissue-specific WT1-deletion for studying the role of this zinc finger nuclear transcription factor in leukemias and solid tumors.
Vicki Huff, University of Texas, M. D. Anderson Cancer Center
The Wilms' tumor (WT) suppressor gene (Wt1) encodes a zinc finger nuclear transcription factor. WT1 functions in cell growth and differentiation. Congenital WT1 mutations predispose to Wilms tumor (a tumor of the kidneys), and are observed in individuals with Denys-Drash Syndrome, Frasier Syndrome and WAGR (Wilms tumour, aniridia [absence of iris], genitourinary anomalies and retardation). A combination of germline and somatic mutations are observed in a subset of Wilms tumor where they are almost invariably homozygous - resulting in inactivation of WT1. Somatic WT1 mutations are present in acute and chronic myeloid leukemia, myelodysplastic syndrome and other blood neoplasms (including acute lymphoblastic leukemia). Wilms tumors display WT1 expression similar to fetal kidney, consistent with the fetal origin of Wilms tumors.
In mice, germline knockout of WT1 (Wt1-/-) results in apoptosis of intermediate mesoderm and subsequent failure of kidney and gonadal development. Wt1-/- embryos die during fetal development or at birth, depending upon strain background. Somatic mosaic inactivation of Wt1 results in the development of Wilms tumor in the background of biallelic expression of the normally imprinted Igf2 gene
The Wt1 floxed allele (Wt1flox) has loxP sites flanking Wt1 exons 8-9 (encoding the second and third of the four DNA-binding zinc-finger domains). Mice homozygous for this floxed allele (Wt1flox/flox) are viable and fertile with no reported gross physical or behavioral abnormalities. The donating investigator reports that homozygotes have normal expression levels of WT1 mRNA and protein.
When bred to mice that express Cre recombinase, the resulting offspring will have the floxed region deleted in cre-expressing tissues. The deletion allele (Wt1Δ) encodes an in-frame truncated protein that is expressed at levels similar to endogenous WT1, but the Wt1Δ protein is non-functional. Homozygous deletion (Wt1Δ/Δ) embryos are phenotypically like Wt1-/- embryos - consistent with Wt1Δ being a loss-of-function allele.
The Wt1flox allele was created in the laboratory of Dr. Vicki Huff (University of Texas, M. D. Anderson Cancer Center) to have loxP sites flanking exons 8-9. First, a targeting vector was designed to place a loxP site upstream of exon 8, and a loxP-flanked neo cassette just downstream of exon 9 of the Wilms tumor 1 homolog locus (Wt1) on chromosome 2. The construct was electroporated into 129S7/SvEvBrd-Hprt+-derived AB1 embryonic stem (ES) cells. Correctly targeted ES cells were injected into recipient blastocysts, and chimeric males were bred with C57BL/6 females to establish the colony. The resulting Wt1neo-loxP mice were bred with CMV-Cre transgenic mice (unspecified genetic background), and offspring with germline deletion of the neo cassette, but retaining the loxP-flanked exons 8-9, were selected (and the cre-expressing transgene was removed). The donating investigator reports Wt1flox mice (mixed C57BL/6;129 genetic background) were bred together for several years as homozygotes prior to sending males to The Jackson Laboratory Repository in 2017. Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J oocytes (Stock No. 000664).
|Allele Name||targeted mutation 2, Vicki Huff|
|Allele Type||Targeted (Conditional ready (e.g. floxed), No functional change)|
|Allele Synonym(s)||Wt1flox; Wt1fx|
|Gene Symbol and Name||Wt1, Wilms tumor 1 homolog|
|Strain of Origin||129S7/SvEvBrd-Hprt+|
|Molecular Note||Exons 8 and 9 were flanked by loxP sites and a neo resistance cassette with a loxP site was inserted downstream of the floxed exons via homologous recombination. The neo cassette was removed by mating with CMV-Cre transgenic mice.|
Mice homozygous for this floxed allele (Wt1flox/flox) are viable and fertile with no reported gross physical or behavioral abnormalities. 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 Wt1flox mouse strain in a publication, please cite the originating article(s) and include JAX stock #019554 in your Materials and Methods section.