In human neutrophils, we show C5L2 is predominantly intracellular, whereas C5aR is expressed on the plasma membrane. its role from that of a rapid recycling or scavenging receptor in this cell type. This is thus the first example of a naturally occurring seven-transmembrane segment receptor that is both obligately uncoupled from G proteins and a negative modulator of signal transduction through the -arrestin pathway. Physiologically, these properties provide the possibility for additional fine-tuning of host defense. Keywords:Chemotaxis, Complement, G Protein-coupled Receptors (GPCR), 7-Helix Receptor, Inflammation, Neutrophil, Receptors, Signal Transduction, Anaphylatoxin Receptors == Introduction == The complement anaphylatoxin C5a is one of the most potent inflammatory mediators of the innate immune system, with the ability to activate all classes of myeloid cells as well as cells of many other lineages. Multiple studies using experimental animals deficient in C5 or Ki8751 mice with targeted deletion of the C5a receptor (C5aR)4have demonstrated the critical role for this anaphylatoxin in host defense. Generation of C5a has also been associated with disease conditions, including asthma, contact sensitivity reactions, autoimmune arthritis, and sepsis Ki8751 (reviewed in Refs.13). C5a manifests its activities by interaction with two known receptors, C5aR and C5L2. Cellular stimulation of the C5aR through Gi2, Gi3, or G16results in intracellular calcium mobilization and activation of signaling pathways including phosphatidylinositol 3-kinase, diacylglycerol, MAPK, ERK, and others (4,5), (reviewed in Ref.6). Like the C5aR, C5L2 is a putative seven-transmembrane segment protein that was identified by Ohnoet al.(7) as a cDNA with homology to the C5aR. It has similar sequence homology with the formyl peptide receptor (FPR) and the chemokine receptor chemR23. C5L2 is expressed on neutrophils, macrophages, and immature dendritic cells in coordination with the C5aR, although its mRNA is present at significantly reduced levels (7,8). Expression has additionally been reported in adrenal gland, spinal cord, thyroid, liver, lung, spleen, brain, and heart (9,10). Studies of the distinct properties of C5L2-utilizing transfection systems demonstrate its ability to bind C5a with affinity similar to that of the C5aR. It binds the metabolite des-Arg C5a with higher avidity (11,12). In contrast to the C5aR, however, we and others (12) have demonstrated that C5L2 is devoid of the ability to couple to intracellular G Ki8751 proteins due to an amino acid replacement of arginine by leucine in the DRY motif located in the second intracellular domain. Following transfection in L1.2 or rat basophilic leukemia cells, which are permissive for C5a-mediated signal transduction by the C5aR, ligand binding to C5L2 failed to induce calcium mobilization and resulted in minimal receptor phosphorylation relative to the C5aR. As a result, questions have been raised whether C5L2 activates G protein-independent intracellular signaling pathways or serves an alternate, ligand scavenging or additional regulatory function similar to the chemokine receptors D6, CXCR7, and DARC (1315). To this end, a recent study by Scolaet al.(16), reported ligand-independent internalization of C5L2 in transfected rat basophilic leukemia cells, which resulted in intracellular accumulation and degradation of C5a and des-Arg C5a. Info gleaned from studies of mice with targeted deletion of C5L2 shows the anti-inflammatory part of the receptor. C5L2/animals exhibit significantly improved inflammation inside a model of pulmonary immune complex injury compared with wild type animals (16,17). Inside a rat model of sepsis induced by cecal ligation and puncture, antibody blockade of C5L2 was associated with a dramatic increase in circulatory interleukin-6 (10). An individually generated line of C5L2-deficient mice yielded contrasting results, that C5L2 isn’t just a positive regulator of the C5aR, but that it is also critical for signaling by both C5a and C3a (8). Among the signaling pathways affected were MAPK, ERK, and protein kinase B/Akt. Another study showed that genetic deficiency- or antibody-mediated blockade of C5L2 offered modest safety from cecal ligation and puncture-mediated sepsis (18). Here, protection from high grade sepsis, which resulted in 100% LATS1 lethality, was afforded only by the combined inhibition of C5L2 and the C5aR. This statement additionally showed that signaling through C5L2 but not the C5aR, led to release of the inflammatory protein, high mobility group package 1.