Analysis of second messenger pathways stimulated by different chemokines acting at the chemokine receptor CCR5

The chemokine receptor, CCR5, responds to several chemokines leading to changes in activity in several signalling pathways. Here, we investigated the ability of different chemokines to provide differential activation of pathways. The effects of five CC chemokines acting at CCR5 were investigated for their ability to inhibit forskolin-stimulated 3′–5′-cyclic adenosine monophosphate (cAMP) accumulation and to stimulate Ca2+ mobilisation in Chinese hamster ovary (CHO) cells expressing CCR5. Macrophage inflammatory protein 1α (D26A) (MIP-1α (D26A), CCL3 (D26A)), regulated on activation, normal T-cell expressed and secreted (RANTES, CCL5), MIP-1β (CCL4) and monocyte chemoattractant protein 2 (MCP-2, CCL8) were able to inhibit forskolin-stimulated cAMP accumulation, whilst MCP-4 (CCL13) could not elicit a response. CCL3 (D26A), CCL4, CCL5, CCL8 and CCL13 were able to stimulate Ca2+ mobilisation through CCR5, although CCL3 (D26A) and CCL5 exhibited biphasic concentration–response curves. The Ca2+ responses induced by CCL4, CCL5, CCL8 and CCL13 were abolished by pertussis toxin, whereas the response to CCL3 (D26A) was only partially inhibited by pertussis toxin, indicating Gi/o-independent signalling induced by this chemokine. Although the rank order of potency of chemokines was similar between the two assays, certain chemokines displayed different pharmacological profiles in cAMP inhibition and Ca2+ mobilisation assays. For instance, whilst CCL13 could not inhibit forskolin-stimulated cAMP accumulation, this chemokine was able to induce Ca2+ mobilisation via CCR5. It is concluded that different chemokines acting at CCR5 can induce different pharmacological responses, which may account for the broad spectrum of chemokines that can act at CCR5.