iRhom2 is required for the secretion of mouse TNFα

Abstract

TNFα is a powerful inflammatory stimulus, central both to the control of infection, and as an agent of inflammatory disease. The most potent inducers of TNFα secretion signal through the Toll-like receptors, and we describe here a chemically-induced mutation that impairs this response in macrophages. A missense mutation was revealed in the gene encoding the inactive rhomboid protease iRhom2, which was not complemented by a null allele of the same gene. Neither the missense nor the null allele affected TLR-induced secretion of IL-6. Moreover, unlike a mutation in TNFα, the iRhom2 missense mutation did not cause enhanced susceptibility to colitis induced by dextran sodium sulfate. These results establish a specific role for iRhom2 in the secretion of TNFα, and present a new target for the modulation of inflammation.

Figure 1

A recessive mutation in Rhbdf2 is associated with reduced TLR-induced TNFα secretion. (A) Phenotype of the index sinecure mouse. Peritoneal macrophages from 62 G3 descendants of ENU-mutagenized sires were stimulated with a panel of TLR ligands, and TNFα production measured by L-929 bioassay. Genome-wide (B) and fine (C) mapping of the sinecure mutation to distal chromosome 11. (D) A homozygous transversion mutation in Rhbdf2, corresponding to an isoleucine to phenylalanine missense mutation (E) in the N-terminal transmembrane domain (TM) of iRhom2 protein. Predicted phosphorylation (P) and glycosylation sites are also indicated. (F) Topology of the iRhom2 protein, indicating the position of the sinecure missense mutation.

Figure 2

Mutant alleles of Rhbdf2 are noncomplementary and cause a specific block in TNFα secretion. Thioglycollate-elicited peritoneal cells were isolated from mice of the indicated genotypes, and cultured in the presence of 200 pg/mL MALP-2, 1 ng/mL LPS, or media alone (-) for 4 hours. TNFα (A) and IL-6 (B) was then measured in the culture supernatant.