| The Toll-like receptor/interleukin-1 receptor (TLR/IL-1R) superfamily and four TIR-containing adaptor proteins, MyD88, TIRAP/MAL, TRIF/TICAM-1 and TRAM/TIRP, play important roles in inflammation and the innate immune response to pathogens. We identified a fifth TIR-domain containing protein, MyD88-5 (also called SARM-1). MyD88-5 encodes a novel 724-aa cytosolic protein with at least seven predicted HEAT/Armadillo repeats in the N-terminus, two SAM domains, and a C-terminal TIR domain. The gene is highly conserved from Drosophila and nematode to human. Northern blots and quantitative RT-PCR showed that the tissue distribution of MyD88-5 differs from that of other MyD88-family proteins, being highest in brain, moderate in splenic lymphocytes and undetectable in macrophages, neutrophils and thymocytes. Antibody generated against recombinant MyD88-5 detected a protein of ∼80 kDa in mouse brain. Expression in mouse brain peaked at embryonic day 18. Unlike the other MyD88-family members, overexpression of MyD88-5 in macrophage-like cells did not activate NF-kappaB. When expressed in either myeloid or non-myeloid cells, MyD88-5 associated with (but was extrinsic to) mitochondria. A fraction of MyD88-5 from mouse brain was isolated with microtubules and mitochondria. JNK3a and MyD88-5 were co-immunoprecipitated. While transfected JNK3a was distributed in COS-1 cells mainly at the plasma membrane, the majority of JNK3a became localized with mitochondria when MyD88-5 was co-expressed. We used a bacterial artificial chromosome system to generate transgenic mice that expressed either green fluorescent protein (GFP) or a MyD88-5-GFP fusion protein under the endogenous MyD88-5 promoter/enhancer. Results revealed that MyD88-5 is expressed in the brain specifically in neurons, in a pattern consistent with a peri-mitochondrial localization. MyD88-5 null mice were generated by gene targeting and their responses are under study in model of ischemic and degenerative brain diseases. The significance of MyD88-5 expression in lymphocytes will be available to study after the null mice are backcrossed to a homogenous state. In summary, MyD88-5's structure, tissue distribution and intracellular associations support that it plays a distinctive biologic role compared to other known members of the TIR family. |
