B6.Cg-Padi4^tm1.2Kmow/J

Stock number: 026708
Product name: Pad4 flox
Research areas: Immunology, Inflammation and Autoimmunity Research, Internal/Organ Research, Research Tools

Description

Pad4^fl/fl floxed mice may be useful for studying the formation of neutrophil extracellular traps during acute infection.

Detailed description

Pad4^fl/fl floxed mice possess loxP sites flanking exons 9-10 of the peptidyl arginine deiminase, type IV (Padi4) gene. These exons contain part of the PAD4 active site, as well as four additional residues that are essential for Ca²⁺ binding. PAD4 is an enzyme responsible for the citrullination and deimination conversion of arginine to citrulline residues on histones. PAD4-mediated deimination of histone H3 and H4 is required for the formation of neutrophil extracellular traps (NETs), specialized anti-microbial structures comprised of decondensed chromatin decorated with microbicidal agents. NETs are responsible for killing gram-positive and gram-negative bacteria as well as fungi. Increased amounts of NETs are detrimental in chronic neutrophil-mediated inflammation such as the lung diseases chronic obstructive pulmonary disease (COPD) and cystic fibrosis (CF) due to increased severity of lung obstruction. Homozygous mice are viable and fertile. When bred to mice that express tissue-specific Cre recombinase, resulting offspring will have exons 9-10 deleted in the cre-expressing tissues.

When bred to B6.C-Tg(CMV-cre)1Cgn/J mice (Stock No. 006054) with widespread Cre recombinase expression, histone deimination is absent in neutrophils of resulting offspring, and they are defective in NET formation. PAD4 KO mice lose significantly more weight during influenza A infection, but retain normal survival. The number of lung infiltrating leukocytes was similar to that seen in wildtype mice during infection.

 

Development

A targeting vector was designed by Dr. Kerri Mowen (The Scripps Research Institute) to insert a loxP site followed by a frt-flanked neomycin resistance (neo) cassette upstream of exon 9, and a second loxP site downstream of exon 10 of the peptidyl arginine deiminase, type IV (Padi4) gene. The construct was electroporated into B6.Cg-Thy1^a-derived Bruce 4 embryonic stem (ES) cells. Correctly targeted ES cells were injected into B6(Cg)-Tyr^c-2J/J albino blastocysts and resulting chimeric males were bred with B6(Cg)-Tyr^c-2J/J albino females (Stock No. 000058). Offspring were bred with B6.Cg-Tg(ACTFLPe)9205Dym/J transgenic mice (Stock No. 005703) to delete the neo cassette, and progeny were crossed to remove the Flp-expressing transgene. Pad4^fl/fl mice were bred to C57BL/6J mice for at least 3 generations. Upon arrival, sperm was cryopreserved that still contained the Tyr^c-2J. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J oocytes (Stock No. 000664). The Tyr^c-2J allele was not maintained in the live colony.

Allele

Symbol: Padi4^tm1.2Kmow
Name: targeted mutation 1.2, Kerri A Mowen
MGI accession ID: MGI:5695766
Generation method: Targeted
Attributes: Conditional ready (e.g. floxed)
Marker symbol: Padi4
Marker name: peptidyl arginine deiminase, type IV
Chromosome: 4
Strain of origin: B6.Cg-Thy1^a
Molecular note: A targeting vector was designed to insert a loxP site followed by an FRT-flanked neomycin resistance (neo) cassette upstream of exon 9, and a second loxP site downstream of exon 10. Flp-mediated recombination removed the FRT-flanked neo cassette leaving exons 9 through 10 floxed.