STOCK Gpr108^tm1.1Brs/J

Stock number: 032823
Product name: Gpr108-null , Gpr108^-
Research areas: Cell Biology Research, Immunology, Inflammation and Autoimmunity Research, Neurobiology Research

Description

Gpr108-null mice have the G protein-coupled receptor 108 knock-out allele. These mice may be useful for studying the interrelationships between GPR108 and innate immune signaling components, including activation of NF-κB, suppression by TIRAP and negative regulation of Toll-like receptor (TLR)-triggered immune responses.

Detailed description

GPR108 is a seven-transmembrane family protein that functions as an NF-κB activator and negative regulator of Toll-like receptor (TLR)-triggered immune responses. GPR108 can be suppressed by TIRAP (an adaptor protein for TLR and MyD88). The interrelationships between GPR108 and innate immune signaling components are multifactorial and point to a membrane-associated signaling structure of significant complexity.
The Gpr108-null allele (Gpr108^-) abolishes endogenous gene expression. Both heterozygous mice (Gpr108^+/-) and homozygous mice (Gpr108^-/- ; Gpr108-null) are viable and fertile with normal breeding, and no gross phenotypic abnormalities. Homozygous MEFs contain no Gpr108 mRNA, while heterozygous MEFs show approximately half the transcript levels compared to wildtype. No Gpr108 mRNA detected in the adult lung and spleen tissue of homozygous mice.
Cells from Gpr108-null mice exhibit enhanced cytokine secretion and NF-κB and IRF3 signaling. Specifically, the transcriptional profile of Gpr108-null MEFs reveal elevated transcript abundance of TLR genes and genes associated with responses to TLR agonists in BMDMs. Enhanced TLR signaling is observed in Gpr108-null MEFs and BMDMs. Although Gpr108^-/- mice were more susceptible to LPS lethal challenge, there was only slight difference observed in serum IL-6 level, and other tested cytokines in the panel were similar to wildtype.

Development

The Gpr108-null allele (Gpr108^-) was created in the laboratory of Dr. Brian Seed (Massachusetts General Hospital, Harvard Medical School). First, a targeting vector was designed to delete a 2 kb upstream region and the entire coding region (17 exons) of the G protein-coupled receptor 108 gene (Gpr108) on chromosome 17 and replace it with an attB/attP-flanked tk-2A-neo cassette. The construct was electroporated into 129S6/SvEvTac-derived W4 embryonic stem (ES) cells, which were then transiently transfected with an R4 integrase-expressing vector to remove the attB/attP-flanked selection cassette (which leaves an attL sequence in its place). Correctly targeted ES cells were injected into recipient blastocysts, and chimeric males were bred with C57BL/6J females for germline transmission. The donating investigator reported that the resulting mice with the Gpr108-null allele were bred with C57BL/6J inbred mice for at least 10 generations (see SNP results) prior to sending homozygous males (black coat color) to The Jackson Laboratory Repository in 2019. Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J oocytes (Stock No. 000664).
Of note, it is not known if the Y chromosome has been fixed to the C57BL/6J background during backcrossing.

In 2019, 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. This revealed a total of 2 markers that were not fixed for C57BL/6 allele-type - one marker on chromosome 3 [~37 Mbp] was 129 allele-type for both males and one on chromosome 7 [~36 Mbp] was 129 allele-type for both males. In addition, 5 of 5 markers that determine C57BL/6 substrains were found to be informative: 4/5 makers were homozygous for C57BL/6N;129 allele-type for both males, while one male was segregating for the other marker. Taken together, these data suggest that the mice sent to The Jackson Laboratory Repository were on a subcongenic C57BL/6J;C57BL/6N genetic background.

Allele

Symbol: Gpr108^tm1.1Brs
Name: targeted mutation 1.1, Brian Seed
MGI accession ID: MGI:6267426
Generation method: Targeted
Attributes: Null/Knockout
Marker symbol: Gpr108
Marker name: G protein-coupled receptor 108
Marker synonyms: 1810015L19Rik; LUSTR2
Chromosome: 17
Strain of origin: 129S6/SvEvTac
Molecular note: An attB and attP-flanked neomycin resistance cassette replaced 17 exons and 2 kb upstream. R4 integrase expression removed the selection cassette. RT-PCR confirmed the absence of transcript expression in spleen and lungs.