B6N.129P2(Cg)-Itgal^tm1Hlz/J

Stock number: 028241
Product name: LFA-1^d
Research areas: Cancer Research, Cell Biology Research, Developmental Biology Research, Immunology, Inflammation and Autoimmunity Research, Research Tools

Description

The LFA-1^d mutant allele has the integrin αL cytoplasmic GFFKR sequence deleted; resulting in a constitutively active conformation. Homozygous mice exhibit defects in cell adhesion and costimulatory signaling, including reduced antigen-specific T cell activation, impaired leukocyte migration and recruitment during inflammatory responses (e.g. sepsis, delayed-type hypersensitivity or transplant rejection).

Detailed description

Integrin αL (Itgal; Cd11a or LFA-1) functions in cellular adhesion and costimulatory signaling. Deletion of the cytoplasmic GFFKR sequence in LFA-1 results in the disassembly of the αL- and the β2-subunit cytoplasmic tails; leading to a constitutively active conformation (LFA-1^d).

When maintained in a barrier unit with standard specific-pathogen free conditions, the donating investigator reports that homozygous (LFA-1^d/d) and heterozygous (LFA-1^d/+) mice are viable and fertile with no gross anatomic or behavioral abnormalities.

LFA-1^d/d mouse leukocytes show increased adhesion to the LFA-1 ligand ICAM-1 on endothelial cells or antigen presenting cells; resulting in reduced antigen-specific T cell activation, impaired leukocyte migration and recruitment during inflammatory responses (e.g. sepsis, delayed-type hypersensitivity or transplant rejection).

Defective LFA-1 deactivation significantly prolongs cardiac allograft survival. Specifically, LFA-1^d/d animals receiving cardiac transplants of fully MHC-mismatched wildtype hearts survive significantly longer than wildtype recipients.

Development

The LFA-1^d mutant allele was created by Dr. Bernhard Holzmann (Technische Universität München). A targeting vector was designed to delete sequences encoding the GFFKR conserved amino acid motif in the membrane-proximal segment cytoplasmic domain (exon 31) of the integrin αL gene (Itgal; Cd11a or LFA-1) on chromosome 7. The vector also inserted a loxP-flanked neo cassette just upstream of exon 31. The construct was electroporated into 129P2/OlaHsd-derived E14.1 embryonic stem (ES) cells. Correctly targeted ES cells were injected into recipient blastocysts, and chimeric males were bred with C57BL/6N females for germline transmission. This generated mice with the Lfa-1^d-neo allele, which were then bred to Cre deleter mice (Tg(CMV-cre)1Cgn on a C57BL/6J background; Stock No. 006054) for germline deletion of the neo cassette. The resulting LFA-1^d males were bred with inbred C57BL/6N females for 10 generations prior to sending males to The Jackson Laboratory Repository in 2015. Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6N oocytes (Stock No. 005304).

Of note, the donating investigator reports that the Y chromosome may not have been fixed to the C57BL/6N background during backcrossing.

Allele

Symbol: Itgal^tm1Hlz
Name: targeted mutation 1, Bernhard Holzmann
MGI accession ID: MGI:3582587
Generation method: Targeted
Attributes: Null/Knockout
Marker symbol: Itgal
Marker name: integrin alpha L
Chromosome: 7
Strain of origin: 129P2/OlaHsd
Molecular note: A constitutive germline deletion of the amino acid motif GFFKR in the membrane proximal segment of the cytoplasmic domain, was accomplished by insertion of a neomycin resistance cassette and a HSV-thymidine kinase cassette. The neo was later removed by crossing to a cre expressing strain.