3). is usually constitutively recycling via clathrin-coated pits and able to internalize CCL19 as well as anti-CRAM antibodies. As this absence of classical chemokine receptor responses and the recycling and internalization features are characteristic for non-classical chemokine receptors, we suggest that CRAM is the newest member of this group. As CCL19 is known to be critically involved in lymphocyte and dendritic cell trafficking, CCL19-binding competition by CRAM might be involved in modulating these processes. Keywords:B cells, chemokine receptor, chemotaxis, transmission transduction == Introduction == Chemokines and their receptors play a major part in homing of leucocytes and recruiting them to sites of inflammation.1There are approximately 50 different human chemokines and 21 identified receptors expressed on specific subpopulations of leucocytes. Most chemokine receptors bind different chemokines with variable affinities.2Classically, chemokine receptors react to ligand binding by triggering an intracellular signalling cascade mediated by G-protein activation. This prospects to receptor phosphorylation by a G-protein-coupled receptor kinase, which results in uncoupling of the receptor from G-proteins and finally desensitization through -arrestin binding to the phosphorylated receptor.3,4This in turn promotes receptor internalization by linking it to the clathrin coat together with adaptor proteins.5,6 The complex system of chemokine receptors is supplemented and modulated by the group of so-called atypical or non-classical chemokine receptors [including D6, duffy antigen receptor for chemokines (DARC), chemocentryx chemokine receptor (CCX-CKR)], which are believed to be non-signalling or chemokine-regulating receptors, thereby modulating local chemokine concentrations (examined by Haraldsen and Rot7). Despite some common features like the lack of signalling and the altered DRYLAIV motif, the users of this group are heterogeneous in their behaviour. For example, D6 is usually a receptor expressed on endothelium 3-Methylcytidine and some leucocytes and is able to bind, scavenge and degrade most pro-inflammatory CC-chemokines. This activity is usually mediated by a constitutive internalization process.8CCX-CKR is restricted to the homeostatic chemokines CCL19, CCL21 and CXCL13 and specializes in their degradation.9On the other hand, DARC is expressed on erythrocytes and vascular endothelium and does not scavenge chemokines per se but acts as a buffer for both CXC and CC pro-inflammatory chemokines. It also effectively mediates transcytosis of its ligands through endothelial layers.10,11 The human geneCCRL2encodes a putative chemokine receptor first described in 1998 and produces two splice variants known as CRAM-A (chemokine receptor on activated macrophages) and the 36 base pair shorter, more common variant, CRAM-B (also known as human chemokine receptor; HCR12). CRAM shares over 40% sequence homology with known chemokine receptors 3-Methylcytidine such as CCR1, -2, -3 and -5, but bears a Mouse monoclonal to CD3/CD19/CD45 (FITC/PE/PE-Cy5) mutation in the highly conserved DRYLAIV motif (Table 1), with a glutamine (Q) at position 127 instead of aspartic acid (D), replacing an acidic residue with a neutral one and rendering coupling to the Gi protein highly unlikely. Until recently, no competitive ligand of CRAM had been found, although considerable data on its expression in tissues and haematopoietic cell types are available.13 == Table 1. == Comparison of amino acid sequences of the conserved DRYLAIV motif between chemokine receptor on activated macrophages (CRAM) and other atypical chemokine receptors Overall identity is usually indicated in the second column and sequence of the highly conserved second intracellular loop is in the third column, with conserved amino acids in strong in reference to the classical sequence. Considerable work has also been carried out around the murine homologue to CRAM, L-CCR (lipopolysaccharide inducible C-C chemokine receptor related gene), which shares only 51% sequence identity with the human gene, a very low value of identity compared with other chemokine receptor orthologues.1416It has been reported that L-CCR binds chemokines such as CCL2, -5, -7 and -8, but this remains controversial and has never been confirmed (discussed by Mantovaniet al.17). Only recently, murine and human CRAM have been found to bind to human chemerin, an adipokine previously described as an agonist for the G-protein-coupled receptor CMKLR1 (also known as ChemR23).15,18 Binding of 3-Methylcytidine chemerin was not followed by classical receptor activation and it was postulated that CRAM concentrates and presents its ligands without degrading them. We have previously explained CRAM as being widely expressed on B cells in a maturation-stage-dependent manner, with the highest expression on.