Differential Activation of Formyl Peptide Receptor-Like 1 by Peptide Ligands

Abstract

Formyl peptide receptor-like 1 (FPRL1) plays a key role in the regulation of immune responses. The activation of FPRL1 induces a complicated pattern of cellular signaling, which results in the regulation of several immune responses, such as chemotactic migration and the production of reactive oxygen species (ROS). Because some of these cellular responses are not beneficial to the host, ligands that selectively modulate these cellular responses are useful. His-Phe-Tyr-Leu-Pro-Met (HFYLPM) is a synthetic peptide that binds to FPRL1. In this study, we generated various HFYLPM analogues and examined their effects on cellular responses via FPRL1 in FPRL1-expressing rat basophilic leukemia-2H3 cells or in primary human neutrophils. Among the HXYLPM analogues, His-Arg-Tyr-Leu-Pro-Met (HRYLPM) activated a broad spectrum of cellular signaling events, including an intracellular Ca2+ concentration increase, phosphoinositide 3-kinase, extracellular signal-regulated kinase, and Akt activation, however, His-Glu-Tyr-Leu-Pro-Met (HEYLPM) activated only intracellular Ca2+ concentration and Akt but did not increase Ca2+. In addition, HRYLPM was found to stimulate chemotaxis and ROS generation via phosphoinositide 3-kinase and an intracellular Ca2+ concentration increase, respectively, whereas HEYLPM stimulated chemotaxis but not ROS generation. With respect to the molecular mechanisms involved in the differential action of HRYLPM and HEYLPM, we found that HRYLPM but not HEYLPM competitively inhibited the binding of 125I-labeled Trp-Lys-Tyr-Met-Val-d-Met-NH2 (WKYMVm, a FPRL1 ligand) to FPRL1. This study demonstrates that the important chemoattractant receptor, FPRL1, may be differentially modulated by distinct peptide ligands. We also suggest that HRYLPM and HEYLPM may be used to selectively modulate FPRL1.

Lysophosphatidylglycerol inhibits formyl peptide receptor like-1-stimulated chemotactic migration and IL-1β production from human phagocytes

“…Intracellular calcium concentration ([Ca 2+ ]i) was determined by Grynkiewicz's method using fura-2/AM (Bae et al, 2003). Briefly, the prepared cells were incubated with a 3 μM fura-2/AM at 37 o C for 50 min in a fresh serum-free RPMI 1640 medium with continuous stirring.…”

“…Superoxide anion generation was determined by measuring cytochrome c reduction using a microtiter 96-well plate ELISA reader (Bio-Tek instruments, EL312e, Winooski, VT) as previously described (Bae et al, 2003). Human neutrophils or monocytes (2 × 10 6 cells in RPMI 1640 medium) were preincubated with 50 μM of cytochrome c at 37 o C for 1 min and then incubated with each peptide.…”

The Synthetic Peptide Trp-Lys-Tyr-Met-Val-D-Met Inhibits Human Monocyte-Derived Dendritic Cell Maturation via Formyl Peptide Receptor and Formyl Peptide Receptor-Like 2

“…Protein samples were prepared for electrophoresis and then separated using a 10% SDS-polyacrylamide gel and the buffer system described previously (24). Following electrophoresis, they were blotted onto nitrocellulose membrane, and were then blocked by incubation in TBST (TBS, 0.05% Tween 20) containing 5% nonfat dried milk.…”

LL-37 inhibits serum amyloid A-induced IL-8 production in human neutrophils

“…We also demonstrated that FPRL1 could be differentially activated by distinct peptide ligands via superoxide anion generation, matrix-metalloproteinase-9 production, and lipid mediator production (Bae et al, 2003;Lee et al, 2005Lee et al, , 2006. Although FPRL1 activation by each ligand has been studied for several years, the effect of FPRL1 ligands on a strong FPRL1 agonist (SAA)-induced cellular signal has not been examined.…”

“…Previously, it has been demonstrated that FPRL1 can be differentially stimulated to express ligand-selective properties (Chiang et al, 2000;Bae et al, 2003;Lee et al, 2005Lee et al, , 2006. Furthermore, Chiang et al (2000) reported that the stimulation of FPRL1 by its peptide ligands resulted Figure 1.…”