Dendritic cells (DC) are essential to the initiation of an immune response due to their unique ability to take-up and process Ag, translocate to lymph nodes, and present processed Ag to naive T cells. Many chemokines, chemokine receptors and other G protein-coupled receptors (GPCRs) are implicated in these various aspects of DC biology. Through microarray analysis, we compared expression levels of chemokines, their cognate receptors, and selected GPCRs in human monocytes and in vitro monocyte-derived immature and mature DC. Hierarchical clustering of gene expression clearly distinguishes the three cell types, most notably highlighting exceptional levels of expression of the GPCR GPR105 within the immature monocyte-derived DC (MDDC) gene cluster. Little or no expression was observed within the monocyte and mature MDDC cluster. Putative functionality of the GPR105 receptor was demonstrated by an observed calcium flux in immature MDDC treated with the potent GPR105 agonist, uridine 5′-diphosphoglucose (UDP-glucose), while no response to the nucleotide sugar was seen in monocytes and mature MDDC. This UDP-glucose-induced calcium response was, at least in part, pertussis toxin-sensitive. Moreover, immature MDDC from some donors treated with UDP-glucose exhibit an increase in expression of the costimulatory molecule CD86, which correlates with the intensity of the UDP-glucose-induced calcium flux. Together, these data demonstrate differential expression of GPR105 on immature and mature MDDC and suggest a role for the receptor and its agonist ligand in DC activation.

ADAMs (a disintegrin and metalloprotease domains) are metalloprotease and disintegrin domain-containing transmembrane glycoproteins with proteolytic, cell adhesion, cell fusion, and cell signaling properties. ADAM8 was originally cloned from monocytic cells, and its distinct expression pattern indicates possible roles in both immunology and neuropathology. Here we describe our analysis of its biochemical properties. In transfected COS-7 cells, ADAM8 is localized to the plasma membrane and processed into two forms derived either by prodomain removal or as remnant protein comprising the extracellular region with the disintegrin domain at the N terminus. Proteolytic removal of the ADAM8 propeptide was completely blocked in mutant ADAM8 with a Glu 330 to Gln exchange (EQ-A8) in the Zn 2؉ binding motif (HE 330 LGHNLGMSHD), arguing for autocatalytic prodomain removal. In co-transfection experiments, the ectodomain but not the entire MP domain of ADAM8 was able to remove the prodomain from EQ-ADAM8. With cells expressing ADAM8, cell adhesion to a substratebound recombinant ADAM8 disintegrin/Cys-rich domain was observed in the absence of serum, blocked by an antibody directed against the ADAM8 disintegrin domain. Soluble ADAM8 protease, consisting of either the metalloprotease domain or the complete ectodomain, cleaved myelin basic protein and a fluorogenic peptide substrate, and was inhibited by batimastat (BB-94, IC 50 ϳ50 nM) but not by recombinant tissue inhibitor of matrix metalloproteinases 1, 2, 3, and 4. Our findings demonstrate that ADAM8 processing by autocatalysis leads to a potential sheddase and to a form of ADAM8 with a function in cell adhesion. ADAM1 (a disintegrin and metalloprotease domain) proteins constitute a family of transmembrane glycoproteins and serve essential physiological roles in fertilization, myogenesis, and neurogenesis. These functions are due to distinct protein domains involved in cell-cell fusion, cell-cell interaction, or proteolysis of membrane proteins, a process termed ectodomain shedding (1). To date, the family of ADAM proteinases comprises more than 30 members in different species (2, 3), and 24 ADAM genes were found in the mouse genome. Fourteen of the murine ADAMs contain the catalytic consensus sequence HEXXHXXGXXHD in their metalloprotease domains and are therefore predicted to be proteolytically active (4). The cleavage of myelin basic protein (MBP) by ADAM10/MADM was the first demonstration of proteolysis by ADAMs (5). The tumor necrosis factor-␣ convertase (ADAM17) was purified on the basis of its ability to cleave tumor necrosis factor-␣ (6, 7) and a number of other peptide and protein substrates in vitro (1, 8). Proteolysis of membrane-bound surface molecules was also demonstrated for heparin-binding epidermal growth factor (9) and amyloid precursor protein (10,11), which are cleaved by ADAMs 9 and 10, respectively.Catalytically active ADAMs are usually activated by furincatalyzed removal of the prodomain or by other proprotein convertases. For cleavage by furin-like...

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