B6.129S4-Ccn1^tm4Lfl/J

Stock number: 037247
Product name: Ccn1^D125A
Research areas: Apoptosis Research, Cell Biology Research, Immunology, Inflammation and Autoimmunity Research, Internal/Organ Research

Description

Ccn1^D125A/D125A knock-in mice have a neo cassette replacing Ccn1 gene with Ccn1^D125A cDNA that encodes a single amino acid substitution (Aspartate -125 to Alanine). This point mutation disrupts the α_v β₃ /α_v β₅ binding sites for CCN1. These mice may be useful for studying cell proliferation, endothelial adhesion, angiogenesis, apoptosis, and extracellular matrix formation.

The donating investigator has made several Ccn1 mouse lines available: Ccn1^D125A (Stock No. 037247), Ccn1^DM (Stock No. 037248) and Ccn1^flox (Stock No. 037249).

Detailed description

Ccn1 gene encodes cellular communication network factor 1 (CCN1/CYR61) protein, which is a matricellular protein that interacts with several integrins and heparan sulfate proteoglycans (HSPGs). CCN1 is involved with cell proliferation, angiogenesis, cell adhesion, senescence, and apoptosis. CCN1 may also regulate aspects of inflammation and tissue regeneration. As such, mutations in this gene may play a role in a variety of inflammatory and fibrotic diseases as well as in certain types of cancers and wound healing.

Ccn1 knock-out mice are embryonic lethal due to vascular defects during early development. By contrast, these Ccn1^D125A/D125A knock-in mice are viable and fertile with no apparent abnormalities; however, if injury is induced in these homozygous mice then injury repair and tissue regeneration may be affected. The Ccn1^D125A/D125A knock-in creates a single amino acid substitution (Asp-125 to Ala) resulting in disruption of α_v β₃/α_v β₅ binding site. Ccn1^D125A/D125A knock-in mice demonstrate reduced capacity for cholangiocyte proliferation and repair after bile duct ligation surgery resulting in severe liver necrosis. CCN1 also induces phagocytic removal of apoptotic cells, Gram-positive and Gram-negative bacteria through binding integrin α_v β₃ in macrophages, and thus Ccn1^D125A/D125A knock-in mice are defective in bacterial clearance and removal of apoptotic neutrophils after infections. These mice may be useful for studying inflammation, cell proliferation and differentiation, and injury repair.

The donating investigator has made several Ccn1 mouse lines available: Ccn1^D125A (Stock No. 037247), Ccn1^DM (Stock No. 037248) and Ccn1^flox (Stock No. 037249).

Development

The Ccn1^D125A mutation was created by replacing endogenous Ccn1 transcription site, exon I and half of exon II with a DNA cassette containing the Ccn1 promoter, a cDNA encoding Ccn1^D125A and the selection marker PGKNeo. The construct was electroporated into 129S4/SvJae-derived J1 embryonic stem (ES) cells. Correctly targeted ES cells were injected into blastocysts and the resulting chimeric mice were identified by genotyping and were bred to C57BL/6J mice (Stock No. 000664). The resulting colony of Ccn1^D125A mice were backcrossed to C57BL/6J mice for at least 8 generations by the donating laboratory (see SNP note below). Upon arrival at The Jackson Laboratory, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J oocytes. Of note, the Y chromosome was fixed during back-crossing to C57BL/6J.

In 2022, a 48 SNP (single nucleotide polymorphism) panel analysis, with 43 markers covering all 19 chromosomes and the X chromosome, as well as 5 markers that distinguish between the C57BL/6J and C57BL/6N substrains, was performed on the males sent to The Jackson Laboratory Repository. One of the 43 markers throughout the genome was segregating with 129, on Chromosome 7. Also, 3 of the 5 markers that determine C57BL/6J from C57BL/6N were found to be segregating, including the rd8 recessive mutation associated with retinal degeneration on Chromosome 1 within the Crb1 locus. These data suggest the mice sent to The Jackson Laboratory Repository were on a mixed C57BL/6N;C57BL/6J genetic background.

Allele

Symbol: Ccn1^tm4Lfl
Name: targeted mutation 4, Lester F Lau
MGI accession ID: MGI:5763569
Generation method: Targeted
Synonyms: Ccn1^D125A
Marker symbol: Ccn1
Marker name: cellular communication network factor 1
Marker synonyms: CCN1; CYR61; GIG1; IBP-10; IGFBP10; IGFBP-10
Chromosome: 3
Strain of origin: 129S4/SvJae
Molecular note: Nucelotide substitution results in the amino acid substitution of alanine for aspartic acid at position 125 (D125A). This mutation disrupts integrin binding.