Functional Analysis of Activating Receptor LMIR4 as a Counterpart of Inhibitory Receptor LMIR3

Abstract

The leukocyte mono-Ig-like receptor (LMIR) belongs to a new family of paired immunoreceptors. In this study, we analyzed activating receptor LMIR4/CLM-5 as a counterpart of inhibitory receptor LMIR3/CLM-1. LMIR4 is expressed in myeloid cells, including granulocytes, macrophages, and mast cells, whereas LMIR3 is more broadly expressed. The association of LMIR4 with Fc receptor-␥ among immunoreceptor tyrosinebased activation motif-bearing molecules was indispensable for LMIR4-mediated functions of bone marrow-derived mast cells, but dispensable for its surface expression. Cross-linking of LMIR4 led to Lyn-and Syk-dependent activation of bone marrow-derived mast cells, resulting in cytokine production and degranulation, whereas that of LMIR3 did not. The triggering of LMIR4 and TLR4 synergistically caused robust cytokine production in accordance with enhanced activation of ERK, whereas the co-ligation of LMIR4 and LMIR3 dramatically abrogated cytokine production. Notably, intraperitoneal administration of lipopolysaccharide strikingly up-regulated LMIR3 and down-regulated LMIR4, whereas that of granulocyte colony-stimulating factor upregulated both LMIR3 and LMIR4 in granulocytes. Cross-linking of LMIR4 in bone marrow granulocytes also resulted in their activation, which was enhanced by lipopolysaccharide. Collectively, these results suggest that the innate immune system is at least in part regulated by the qualitative and quantitative balance of the paired receptors LMIR3 and LMIR4.The Ig-like receptors provide positive and negative regulation of immune cells upon recognition of various ligands (1-5). We identified previously leukocyte mono-Ig-like receptors (LMIRs) 2 from a cDNA library of bone marrow-derived mast cells (BMMCs). We (6) and others (7-9) demonstrated that LMIR1/MAIRI (myeloid-associated Ig-like receptor I)/CLM-8 (CMRF-35-like molecules-8) and LMIR2/MAIRII/CLM-4/ DIgR1 (dendritic cell-derived Ig-like receptor 1) are expressed mainly in myeloid cells. The human homolog of LMIR1 is CMRF-35H/IRp60 (inhibitory receptor protein of 60 kDa)/ CD300a (10 -14). The inhibitory effects of LMIR1 on mast cells and eosinophils and the activatory roles of LMIR2 in macrophages have been described recently (6,7,11). In addition to LMIRs, a variety of Ig-like paired receptors are expressed by myeloid cells (2,(15)(16)(17), but the biological significance of a paired receptor remains incompletely understood. Despite the similarity in the extracellular Ig-like domains, a striking structural difference between activating and inhibitory receptors exists in the transmembrane and cytoplasmic regions. In general, the former associate with an immunoreceptor tyrosinebased activation motif (ITAM)-or the related activating motifbearing adaptor transmembrane protein, including DAP10, DAP12, or Fc receptor-␥ (FcR␥), via a positively charged residue in the transmembrane domain, whereas the latter include an immunoreceptor tyrosine-based inhibitory motif (ITIM) in the cytoplasmic domain (1,5,18,19). Cells of the myeloid lineage su...

Eμ/miR-125b transgenic mice develop lethal B-cell malignancies

“…The 3 0 UTR was cloned to downstream of the Renilla luciferase stop codon in pGL4.74 vector (Promega, Madison, WI, USA). To generate the mutant 3 0 UTR of the mouse Trp53inp1, two-step PCR mutagenesis was performed [24][25][26] using the WT-3 0 UTR as a template. The Rluc-2 Â 125b site vector contained two copies of a sequence designed to be perfectly complementary to the miR125b downstream of the Renilla luciferase stop codon.…”

An Activating and Inhibitory Signal from an Inhibitory Receptor LMIR3/CLM-1: LMIR3 Augments Lipopolysaccharide Response through Association with FcRγ in Mast Cells

“…Paired immune receptors are generally composed of several activating and inhibitory receptors with high homology in the extracellular domains (1,2). We have previously characterized leukocyte mono-Ig-like receptors 1-5 (LMIR1-5) 3 among a novel paired receptor family called LMIR/CMRF-35-like molecules (CLMs)/myeloid-associated Ig-like receptor (MAIR)/ dendritic cell-derived Ig-like receptor (DIgR)/CD300 (3)(4)(5)(6)(7)(8)(9). LMIR3/CLM-1/MAIR-V/DIgR2/CD300LF is an inhibitory receptor that possesses an ITIM in the cytoplasmic region (4,6,7,10,11).…”

“…We have previously characterized leukocyte mono-Ig-like receptors 1-5 (LMIR1-5) 3 among a novel paired receptor family called LMIR/CMRF-35-like molecules (CLMs)/myeloid-associated Ig-like receptor (MAIR)/ dendritic cell-derived Ig-like receptor (DIgR)/CD300 (3)(4)(5)(6)(7)(8)(9). LMIR3/CLM-1/MAIR-V/DIgR2/CD300LF is an inhibitory receptor that possesses an ITIM in the cytoplasmic region (4,6,7,10,11). On the other hand, LMIR4/CLM-5 or LMIR5/CLM-7, which is an activating receptor paired with LMIR3, associates with an ITAM-containing adaptor protein, FcR␥ or DNAX activating protein of 12 kDa (DAP12), respectively (4,11,12).…”

Expression of ADAMTS4 in Ewing's sarcoma

“…To date, various important molecules have been detected including SDF-1, a member of the tumor necrosis factor receptor superfamily TROY, Xenopus-Tsukushi, Vasorin and leukocyte mono-Ig-like receptor (LMIR) by the SST method (18)(19)(20)(21)(22)(23). In this study, we identified a secreted molecule ADAMTS4 (a disintegrin and metalloproteinase with thrombospondin motifs) from EWS cell lines by using the SST system based on retrovirus-mediated expression screening (SST-REX) (24,25).…”