These MECL-1 KO mice may be useful for studying the role of the immunoproteasomes in T cell proliferation.
Alice Sijts, University Utrecht
In this strain, a neo cassette replaces exons 5-7 of the proteasome (prosome, macropain) subunit, beta type 10 (Psmb10) gene, abolishing gene expression.
Psmb10 encodes multicatalytic endopeptidase complex-like-1 (MECL-1), one of three catalytic immunosubunits that make up immunoproteasomes. In response to interferon-gamma (IFNγ) or tumor necrosis factor-alpha (TNF-α), cytokines that are released in the early stages of viral infections, catalytic immunosubunits are rapidly assembled to form immunoproteasomes. These specialized proteasomes are formed in most cells, including lymphoid cells, and have a relatively short life span of a few days. They process MHC class I Ag and play a role in CD8+ T cell proliferation. Homozygous mice are viable and fertile. These MECL-1-/- mice exhibit a 20% reduction in CD8+ T cell numbers in the spleen, with normal numbers of CD8+ and CD4+ T cells in the thymus.
A targeting vector was designed to replace exons 5-7 of the proteasome (prosome, macropain) subunit, beta type 10 (Psmb10) gene with a neomycin resistance (neo) cassette. The construct was electroporated into 129S2/SvPas-derived D3 embryonic stem (ES) cells. Correctly targeted ES cells were injected into blastocysts and the resulting chimeric males were bred to C57BL/6 females.
Another targeting vector was designed to replace exons 1-5 of the proteasome (prosome, macropain) subunit, beta type 8 (Psmb8) gene with a neomycin resistance (neo) cassette. The construct was electroporated into 129P2/OlaHsd-derived E14.1 embryonic stem (ES) cells. Correctly targeted ES cells were injected into blastocysts and the resulting chimeric males were bred to C57BL/6 females to generate a colony of Psmb8tm1Hjf knockout mice.
Mice carrying the mutations were crossed, and the resulting double mutants were backcrossed for at least 10 generations to C57BL/6 mice by the donating lab (see SNP note below). Upon arrival at The Jackson Laboratory, mice were bred to C57BL/6J (Stock No. 000664) for at least one generation to establish the colony. The Psmb8tm1Hjf allele was selected against, and was maintained as a separate strain (Stock No. 021201).
A 32 SNP (single nucleotide polymorphism) panel analysis, with 27 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 rederived living colony at The Jackson Laboratory Repository. Five of the 27 markers throughout the genome were segregating, suggested an incomplete backcross. One marker on Chromosome 11 was segregating from an unknown source.
|Allele Name||targeted mutation 1, Marcus Groettrup|
|Allele Type||Targeted (Null/Knockout)|
|Allele Synonym(s)||beta2i-; MECL-1-|
|Gene Symbol and Name||Psmb10, proteasome (prosome, macropain) subunit, beta type 10|
|Strain of Origin||129S2/SvPas|
|Molecular Note||A neomycin selection cassette replaced exons 5 through 7. Homozygous mice had no protein expression of the gene, as determined by 2D gel electrophoresis of 20S proteasomes purified from spleen and virally infected liver.|
When maintaining the live Psmb10tm1Mgro mutant colony, homozygous mice may be bred together.
When using the B6;129-Psmb10tm1Mgro/J mouse strain in a publication, please cite the originating article(s) and include JAX stock #021607 in your Materials and Methods section.