Abstract
HIV-1 envelope glycoprotein gp120 induces, independently of infection, the release of CCL2 from macrophages. In turn, this chemokine acts as an autocrine factor enhancing viral replication. In this study, we show for the first time that phosphoinositide-specific phospholipase C (PI-PLC) is required for the production of CCL2 triggered by gp120 in macrophages. Using a combination of confocal laser-scanner microscopy, pharmacologic inhibition, western blotting and fluorescence-activated cell sorter analysis, we demonstrate that gp120 interaction with CCR5 leads to nuclear localization of the PI-PLC β1 isozyme mediated by mitogen-activated protein kinase ERK-1/2. Notably, phosphatidylcholine-specific phospholipase C (PC-PLC), previously reported to be required for NF-kB-mediated CCL2 production induced by gp120 in macrophages, drives both ERK1/2 activation and PI-PLC β1 nuclear localization induced by gp120. PI-PLC β1 activation through CCR5 is also triggered by the natural chemokine ligand CCL4, but independently of ERK1/2. Finally, PI-PLC inhibition neither blocks gp120-mediated NF-kB activation nor overall accumulation of CCL2 mRNA, whereas it decreases CCL2 transcript level in the cytoplasm. These results identify nuclear PI-PLC β1 as a new intermediate in the gp120-triggered PC-PLC-driven signal transduction pathway leading to CCL2 secretion in macrophages. The finding that a concerted gp120-mediated signaling involving both PC- and PI-specific PLCs is required for the expression of CCL2 in macrophages suggests that this signal transduction pathway may also be relevant for the modulation of viral replication in these cells. Thus, this study may contribute to identify novel targets for therapeutic intervention in HIV-1 infection.
Highlights
CCL2, previously known as monocyte chemoattractant protein1 (MCP-1), is a member of the CC-chemokine family secreted by a variety of both hematopoietic and non-hematopoietic cells, with monocytes/macrophages representing the major source in the peripheral blood [1,2]
To evaluate whether phosphoinositide-specific phospholipase C (PI-PLC) is involved in the signal transduction pathway triggered by gp120 in monocyte-derived macrophages (MDM), we focused our attention on the b family of PI-PLCs, since the members of this family are activated by heterotrimeric G protein subunits after stimulation of G protein-coupled receptors (GPCRs) [21]
By performing confocal laser-scanner microscopy (CLSM) analysis, we found that PI-PLC b1 is highly expressed in control MDM, mainly localized throughout the cytoplasm in proximity to the plasma membrane and poorly in the nucleus (Figure 1A)
Summary
CCL2, previously known as monocyte chemoattractant protein (MCP-1), is a member of the CC-chemokine family secreted by a variety of both hematopoietic and non-hematopoietic cells, with monocytes/macrophages representing the major source in the peripheral blood [1,2]. From a clinical point of view, CCL2 is one of the most studied proinflammatory molecules among the chemokine family and represents a potential intervention point for the treatment of various inflammatory, autoimmune and infectious diseases. This chemokine is induced during a variety of human acute and chronic viral infections. In addition to infection itself, virus-derived proteins such as gp120, Nef, matrix protein p17 and transactivator protein Tat induce a significant increase in the expression and release of this chemokine [3]
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