Differential Activation of CC Chemokine Receptors by AOP-RANTES

Jörn Elsner,Matthias Mack,Hilke Brühl,Yasmin Dulkys,Daniela Kimmig,Graham Simmons,Paul R Clapham,Detlef Schlöndorff,Alexander Kapp,Timothy N.C Wells,Amanda E.I Proudfoot

doi:10.1074/jbc.275.11.7787

Abstract

RANTES (regulated on activation normal T cell expressed) has been found at elevated levels in biological fluids from patients with a wide range of allergic and autoimmune diseases and is able to attract several subtypes of leukocytes including eosinophils and monocytes into inflamed tissue. Amino-terminal modifications of RANTES produce receptor antagonists which are candidates for blocking this cellular recruitment. Met-RANTES has been shown to modulate inflammation in vivo, while AOP-RANTES is a potent inhibitor of R5 human immunodeficiency virus type 1 (HIV-1) strains and has been shown to down-modulate CCR5 and prevent recycling of the receptor. We have studied the effect of AOP-RANTES in eosinophil activation and have found that it is able to efficiently elicit eosinophil effector functions through CCR3, as measured by the release of reactive oxygen species and calcium mobilization, whereas Met-RANTES is inactive in these assays. AOP-RANTES is found to inhibit CCR3-mediated HIV-1 infection with moderate potency, in contrast to its potent inhibition of CCR5-mediated HIV-1 infection. Furthermore, we have investigated the abilities of these modified proteins to down-modulate CCR1 and CCR3 from the surface of monocytes and eosinophils. We show here that AOP-RANTES is much less effective than RANTES in down-modulation of CCR1. Surprisingly, recycling of CCR1 was minimal after incubation with RANTES while there was complete recycling with AOP-RANTES. In the case of CCR3, no significant difference was found between RANTES and AOP-RANTES in down-modulation and recycling. It therefore appears that trafficking of RANTES receptors follows different patterns, which opens up potential new targets for therapeutic intervention.

Highlights

Chemokines are chemotactic proteins that play a central role in immune and inflammatory responses by the attraction and activation of leukocytes
We have studied the effect of AOP-RANTES in eosinophil activation and have found that it is able to efficiently elicit eosinophil effector functions through CCR3, as measured by the release of reactive oxygen species and calcium mobilization, whereas Met-RANTES is inactive in these assays
RANTES Receptor Expression on Normal Human Eosinophils—The eosinophils from healthy non-atopic donors used in these experiments were screened for their profile of the RANTES receptors CCR1, CCR3, and CCR5 by flow cytometry using the appropriate monoclonal antibodies

Summary

Introduction

Chemokines are chemotactic proteins that play a central role in immune and inflammatory responses by the attraction and activation of leukocytes. It was generally accepted that the chemokine subclasses differ in their biological activity to stimulate different kinds of leukocytes, so that CXC chemokines are mediators of acute inflammation through neutrophil activation while the CC chemokines mediate chronic inflammation by attracting leukocytes such as eosinophils, monocytes, lymphocytes, basophils, and dendritic cells. This paradigm has recently been shown to have exceptions; for example, CXCR3 is expressed on activated T cells [4] and neutrophils can be activated by CC chemokines following stimulation with interferon-␥ [5]. A virally produced chemokine, vMIP-II encoded by herpesvirus-8 has been shown to be effective in preventing nephritis [19]

Methods

Results

Conclusion