The C-terminal Nonapeptide of Mature Chemerin Activates the Chemerin Receptor with Low Nanomolar Potency

Abstract

Chemerin is a novel protein identified as the natural ligand of ChemR23 (chemerinR), a previously orphan G protein-coupled receptor expressed in immature dendritic cells and macrophages. Chemerin is synthesized as a secreted precursor, prochemerin, which is poorly active, but converted into a full agonist of chemerinR by proteolytic removal of the last six amino acids. In the present work, we have synthesized a number of peptides derived from the C-terminal domain of human prochemerin and have investigated their functional properties as agonists or antagonists of human chemerinR. We found that the nonapeptide 149 YFPGQ-FAFS 157 (chemerin-9), corresponding to the C terminus of processed chemerin, retained most of the activity of the full-size protein, with regard to agonism toward the chemerinR. Extension of this peptide at its N terminus did not increase the activity, whereas further truncations rapidly resulted in inactive compounds. The C-terminal end of the peptide appeared crucial for its activity, as addition of a single amino acid or removal of two amino acids modified the potency by four orders of magnitude. Alanine-scanning mutagenesis identified residues Tyr 149 , Phe 150 , Gly 152 , Phe 154 , and Phe 156 as the key positions for chemerinR activation. A modified peptide (YHSFFFPGQFAFS) was synthesized and iodinated, and a radioligand binding assay was established. It was found that the ability of the various peptides to activate the chemerin receptor was strictly correlated with their affinity in the binding assay. These results confirm that a precise C-terminal processing is required for the generation of a chemerinR agonist. The possibility to restrict a medium sized protein to a nonapeptide, while keeping a low nanomolar affinity for its receptor is unusual among G protein-coupled receptors ligands. The identification of these short bioactive peptides will considerably accelerate the pharmacological analysis of chemerin-chemerinR interactions.

Recombinant adenovirus type 5 vectors that target DC-SIGN, ChemR23 and αvβ3 integrin efficiently transduce human dendritic cells and enhance presentation of vectored antigens

“…This peptide (designated here as Chemerin FAFS) was able to significantly improve the efficiency of rAd5 vector-mediated transduction of DC, while also increasing the phenotypic maturation of transduced DC and enhancing their ability to support antigen-specific activation of autologous human CD8 + T cells. Importantly, this enhancement was not observed when a mutated derivative of the Chemerin targeting peptide was used (this mutated peptide, designated here as Chemerin FAAS, contains a phenylalanine-to-alanine substitution at a reside known to be required for receptor binding; (35)). The use of the ChemR23/Chemerin-axis as a targeting strategy may also have effects on DC migration, which could represent an additional advantage of this approach -although additional studies will be needed to investigate the feasibility of this idea.…”

“…ChemR23 is a G protein coupled chemokine receptor present on human monocyte-derived DC and macrophages, as well as blood-derived plasmacytoid and myeloid DC (29,32). The natural ligand of ChemR23, Chemerin, has been shown to serve as a potent chemotactic factor for primary DC, and truncated forms of Chemerin as short as 9 amino acids have been demonstrated to bind to ChemR23 with nanomolar affinity (34,35). We therefore used a biotinylated 13-mer peptide derived from Chemerin to target our rAd5 vectors to primary human monocyte-derived DC.…”

“…FnFn10 contains the wild-type sequence of this peptide and binds with low affinity to α v β 3 , while 3JCLI4 contains a mutated derivative of this sequence, selected for its ability to bind with high (800 pM) affinity to α v β 3 (26). To target the ChemR23 receptor, we used a biotinylated 13-mer peptide (hereafter denoted as Chemerin FAFS) based on the recently described active C-terminal region of Chemerin, which has been shown to bind with nanomolar affinity to its cognate receptor, ChemR23 (32,35). We also included a biotinylated control peptide in which a critical phenylalanine residue with Chemerin FAFS was changed to alanine; this peptide (hereafter denoted as Chemerin FAAS) was used as a negative control.…”

The Role of ChemR23 in the Induction and Resolution of Cigarette Smoke-Induced Inflammation

“…Chemerin is widely expressed throughout the body in epithelium (9) and endothelium (8), but also in fibroblasts (12), adipocytes (13), and keratinocytes (8). Chemerin is synthesized as preprochemerin and secreted as poorly active prochemerin, gaining full bioactivity as a ligand for ChemR23 after C-terminal cleavage of 6 to 7 aa (9,14,15). Active chemerin is increased in several inflammatory secretions that are enriched in inflammatory cell-derived proteases such as neutrophil elastase and mast cell tryptase.…”

Endothelial Cell–Derived Chemerin Promotes Dendritic Cell Transmigration

“…Chemerin is secreted as a poorly active precursor that is converted into a bioactive agonist following the proteolytic removal of the last 6 or 7 aa from the C terminus (7). Various proteases mediate chemerin processing; these include neutrophil serine proteases and proteases from the coagulation and fibrinolytic cascades (8).…”