STOCK Pdgfrb^{tm13(Pdgfrb)Sor}/J

Stock number: 018434
Research areas: Cancer Research, Cardiovascular Research, Cell Biology Research, Developmental Biology Research, Hematological Research, Immunology, Inflammation and Autoimmunity Research, Internal/Organ Research, Research Tools

Description

The PDGFRβ^(S)J conditional knockin allele has the endogenous PDGFRβ sequences replaced with a floxed-STOP cassette upstream of the constitutively active PDGFRβJ mutant isoform (V536A mutation in the juxtamembrane domain). Following Cre recombinase-mediated excision of the floxed-STOP cassette, expression of the constitutively active mutant PDGFRβ leads to high basal kinase activity without the addition of ligand. These mutant mice may be useful in studying PDGFR signaling in multipotent progenitor mural cells (pericytes and vascular smooth muscle cells), as well as in cardiovascular and obesity-related disease.

Detailed description

The PDGFRβ^(S)J conditional knockin allele has the endogenous PDGFRβ sequences replaced with a loxP-flanked PGKneo (lox-stop-lox) cassette upstream of the constitutively active PDGFRβJ mutant isoform (V536A mutation in the juxtamembrane domain disrupts a highly conserved juxtamembrane region in type III receptor tyrosine kinases that contacts the kinase domain important for full auto-inhibition, leading to constitutive PDGFRβ activity). Prior to Cre recombinase exposure, no expression of the constitutively active βJ mutant isoform is observed, and the mutant allele functions as a knockout. Mice homozygous for the PDGFRβ^(S)J allele are neonatal lethal. Heterozygous PDGFRβ^(S)J mice are viable and fertile with no reported abnormalities. Following Cre recombinase-mediated excision of the floxed-STOP cassette, high basal kinase activity without the addition of ligand is observed in the cre-expressing tissues.

Of note, pan-deletion of the floxed-STOP cassette can be achieved by breeding PDGFRβ^(S)J conditional knockin mice with mice expressing Cre recombinase in early embryonic development (Meox2-Cre [Stock No. 003755], Sox2-Cre [Stock No. 004783], or Prm-Cre [Stock No. 003328]). In the resulting double mutant offspring, widespread expression of the constitutively active PDGFRβ leads to normal embryonic development but eventual death by ~14 days of age. In these double mutant mice, constitutively active PDGFRβ induces a battery of immune response genes in pericytes and mesenchymal cells, and also inhibits in vitro differentiation of white adipocytes.

When PDGFRβ^(S)J conditional knockin mice are bred to Sm22-Cre mice [Stock No. 004746], constitutively active PDGFRβ drives cell proliferation and downregulates differentiation genes in aortic vascular smooth muscle tissues of the double mutant offspring.

Importantly, the donating investigator reports they have not observed any consistent phenotypic differences between the Cre recombinase-activated phenotypes of PDGFRβJ and PDGFRβK (see Stock No. 018435) mice, regardless of Cre recombinase tissue specificity.

Development

To create the PDGFRβ^(S)J targeting vector, Dr. Philippe Soriano (Mount Sinai School of Medicine) isolated a full-length mouse platelet derived growth factor β (Pdgfrb) cDNA sequence and introduced a valine to alanine substitution at codon 536 (V536A) of the juxtamembrane domain by site-directed mutagenesis. This βJ mutation results in a constitutively active mutant PDGFRβ. A loxP-flanked PGKneo cassette (lox-stop-lox) was placed between the splice acceptor and the initiating codon of the mutant cDNA sequence. This construct replaced the endogenous Pdgfrb exons 2-4 via electroporation into 129S4/SvJaeSor-derived AK7 embryonic stem (ES) cells. Correctly targeted ES cells were injected into recipient blastocysts, and the resulting chimeric males were mated with females to generate the PDGFRβ^(S)J knockin mouse line. Mutant mice were bred to B6;129 mice harboring other mutations/transgenes, and/or to C57BL/6J wildtype, and/or C57BL/6NTac wildtype mice for several generations prior to sending to The Jackson Laboratory Repository. The donating investigator reports selecting PDGFRβ^(S)J mice with no other mutations present for breeding. Upon arrival, mutant mice were bred to C57BL/6J inbred mice (Stock No. 000664) for at least one generation to establish The Jackson Laboratory Repository colony.

Allele

Symbol: Pdgfrb^{tm13(Pdgfrb)Sor}
Name: targeted mutation 13, Philippe Soriano
MGI accession ID: MGI:5140888
Generation method: Targeted
Attributes: Conditional ready (e.g. floxed); No functional change
Marker symbol: Pdgfrb
Marker name: platelet derived growth factor receptor, beta polypeptide
Chromosome: 18
Strain of origin: 129S4/SvJaeSor
Molecular note: Exons 2 and 3 were replaced with a splice acceptor, floxed neo/stop cassette, and the mouse cDNA with nucleotide substitution(s) that results in the amino acid substitution of alanine for valine at position 536 (V536A). This mutation is an activating mutation.