STOCK Proc^tm1Fjc Tg(F10-Proc)785Fjc/J

Stock number: 038809
Product name: low-PC transgenic, PC^-(PC^Tg785)
Research areas: Developmental Biology Research, Hematological Research, Reproductive Biology Research

Description

These low-PC transgenic mice employ a mouse protein C (Proc; PC) transgene that allows a low level of PROC expression in a PROC knock-out background. They are useful in studies of thrombosis, inflammation, and spontaneous abortion.

Detailed description

Although mice heterozygous for a knock-out of the protein C (Proc; PC) gene survive the early lethality seen in the homozygotes, live to adulthood, and can be successfully employed in some disease models, the approximately 50% level of PROC in these mice is often too high to observe spontaneous phenotypes and is not reflective of symptomatic patients (see Stock No. 038808). For this reason, the donating laboratory produced this low-PC transgenic compound mutant model combining the targeted knock-out of Proc with a minimally-expressed Proc transgene.

Mice homozygous for the targeted knock-out allele and hemizygous for the transgene (PC^-/-(PC^Tg785)) are born at predicted Mendelian ratios and survive beyond birth. Mice homozygous for the knock-out allele but lacking the transgene die soon after birth.

PC^-/-(PC^Tg785) mice express only ~3% of the plasma PROC levels found in wildtype mice. They are susceptible to thrombosis and hemorrhage, most frequently in the tails and feet after 3 months of age. Necrosis of the paws/legs is often accompanied by severe edematous swelling. Approximately 10% of the PC^-/-(PC^Tg785) mice die spontaneously or need to be euthanized due to advanced disease. At 12 weeks of age, these mice become more hemorrhagic, as bleeding times are significantly prolonged. Low-PC transgenic mice also develop proinflammatory phenotypes with elevated levels of IL6.

Although low levels of PROC have no impact on male or female fertility, a severe PROC insufficiency inhibits the ability of females to sustain pregnancy to full term. PC^-/-(PC^Tg785) females are prone to spontaneous abortion. Studies suggest that maternal PROC is vital for sustaining pregnancy beyond 7.5 days postcoitum, likely by regulating the balance of coagulation and inflammation during trophoblast invasion.

Development

Independently-generated targeted knock-out and transgenic mutant mice were intercrossed to produce this compound mutant strain.

For the targeted mutation (tm): Exons 2-9, containing the entire Proc coding region, were replaced by a neomycin cassette via homologous recombination in (129X1/SvJ x 129S1/Sv)F1- Kitl⁺-derived R1 embryonic stem (ES) cells. The resultant mice were backcrossed to C57BL/6J by the donating laboratory. The donating laboratory also notes CBA contributions to the genetic background. See Stock No. 038808.

The transgenic (Tg) allele incorporates: F7 (FVII; inactivated by a deletion in the promoter), the F7-F10 intergenic region, the F10 promoter, complete mouse Proc cDNA (mPC), SV40 poly A, and F10 exons 2-9 (not active). The transgenic vector was introduced to C57BL/6 embryos. The donating laboratory notes that this construct should allow appropriate temporal and spatial expression patterns, as well as proper γ-carboxylation, of the PROC cDNA, which has characteristics similar to those of F10. Since much less was then known about the characteristics of the promoter elements of the murine Proc gene, the use of the known proximal promoter of the F10 gene was their preferred approach. This is transgene founder line Tg785 (also called Ji or "Jimmy"), carrying at least 2 copies of the transgene, and expressing ~3% of the plasma PROC levels found in wildtype mice. The transgenic integration site has not been mapped.

The donating laboratory notes C57BL/6 and CBA contributions to the genetic background of this compound mutant strain. Upon arrival at The Jackson Laboratory, sperm was cryopreserved. Cryorecovery/live colony development will involve C57BL/6J (Stock No. 000664).

Allele

Symbol: Proc^tm1Fjc
Name: targeted mutation 1, Francis J Castellino
MGI accession ID: MGI:2183063
Generation method: Targeted
Attributes: Null/Knockout
Synonyms: PC^-
Marker symbol: Proc
Marker name: protein C
Marker synonyms: APC; inactivator of coagulation factors Va, VIII; PC; PROC1; THPH3; THPH4
Chromosome: 18
Strain of origin: (129X1/SvJ x 129S1/Sv)F1-Kitl⁺
Molecular note: Exons 2 through 9, containing the entire coding region, were deleted by the insertion of a neomycin selection cassette. Northern blot analysis showed that transcript encoded by the endogenous allele was absent in homozygous mutant embryos. Assays of plasma obtained from homozygous mutant mice showed a lack of amidolytic activity.

Allele

Symbol: Tg(F10-Proc)785Fjc
Name: transgene insertion 785, Francis J Castellino
MGI accession ID: MGI:7713362
Generation method: Transgenic
Attributes: Inserted expressed sequence
Marker symbol: Tg(F10-Proc)785Fjc
Marker name: transgene insertion 785, Francis J Castellino
Chromosome: UN
Strain of origin: C57BL/6
Molecular note: The transgenic (Tg) allele incorporates: F7 (FVII; inactivated by a deletion in the promoter), the F7-F10 intergenic region, the F10 promoter, complete mouse Proc cDNA (mPC), SV40 poly A, and F10 exons 2-9 (not active).