SHIP-2, a recently identified inositol 5'-phosphatase, shares high level homology with SHIP-1. Although the role of SHIP-1 has been extensively studied, the role of SHIP-2 in myeloid cell functions is not known. Here, we have analyzed the expression patterns, molecular mechanism of activation, and function of SHIP-2 in human myeloid cell Fcgamma receptor (FcgammaR) signaling. We report that SHIP-2 is expressed in transformed myeloid cells and in primary macrophages, but not in peripheral blood monocytes. Treatment of peripheral blood monocytes with bacterial lipopolysaccharide induced expression of SHIP-2 in a dose-dependent manner. FcgammaRIIa clustering in THP-1 cells induced SHIP-2 tyrosine phosphorylation, suggesting a role for SHIP-2 in modulating FcgammaR-mediated function. Consistent with this notion, overexpression of wild-type SHIP-2 (but not catalytically deficient SHIP-2) in THP-1 cells almost completely abrogated NFkappaB-mediated gene transcription in response to FcgammaRIIa clustering. Furthermore, FcgammaRIIa-induced Akt activation was blocked by wild-type SHIP-2, but not by a catalytically deficient mutant of SHIP-2. Additional experiments analyzing the molecular mechanism of SHIP-2 induction by FcgammaRIIa revealed that SHIP-2 associated with the phosphorylated FcgammaRIIa immunoreceptor tyrosine-based activation motif via the SHIP-2 SH2 domain. Thus, an SH2 domain mutant of SHIP-2 failed to associate with FcgammaRIIa or to become tyrosine-phosphorylated upon FcgammaRIIa clustering. Finally, we also demonstrate that SHIP-2 phosphorylation was induced by FcgammaRI clustering in THP-1 cells. These findings unravel a novel level of regulation of FcgammaR-mediated activation of human myeloid cells by the expression and function of the inositol phosphatase SHIP-2.
"Instead, conformational changes in the LPC-bound CRP pentamer may distort binding sites for FcγRs, resulting in less efficient binding of the CRP-LPC complex to FcγRs. This may lead to less potent activation of FcγRs, thus reducing NF-kB activation  and the generation of ROS , as observed in this study. "
[Show abstract][Hide abstract] ABSTRACT: Rationale
C-reactive protein (CRP) and lysophosphatidylcholine (LPC) are phosphorylcholine-(PC)-containing oxidized phospholipids (oxPLs) found in oxidized LDL (oxLDL), which trigger pro-atherogenic activities of macrophages during the process of atherosclerosis. It has been previously reported that CRP binds to the PC head group of oxLDL in a calcium-dependent manner. The aim of this study was to investigate the importance of binding between CRP and LPC to the pro-atherogenic activities of macrophages.
Objectives and findings
A chemiluminescent immunoassay and HPLC showed that human recombinant CRP formed a stable complex with LPC in the presence of calcium. The Kd value of the binding of the CRP-LPC complex to the receptors FcγRIA or FcγRIIA was 3–5 fold lower than that of CRP alone. The CRP-LPC complex triggered less potent generation of reactive oxygen species and less activation of the transcription factors AP-1 and NF-kB by human monocyte-derived macrophages in comparison to CRP or LPC alone. However, CRP did not affect activities driven by components of oxLDL lacking PC, such as upregulation of PPRE, ABCA1, CD36 and PPARγ and the enhancement of cholesterol efflux by human macrophages. The presence of CRP inhibited the association of Dil-labelled oxLDL to human macrophages.
The formation of complexes between CRP and PC-containing oxPLs, such as LPC, suppresses the pro-atherogenic effects of CRP and LPC on macrophages. This effect may in part retard the progression of atherosclerosis.
Journal of Inflammation 10/2012; 9(1):42. DOI:10.1186/1476-9255-9-42 · 2.02 Impact Factor
"This process is accompanied by the release of pro-inflammatory cytokines and reactive oxygen species that, although required for optimal clearance of immune complex (IC), will lead to tissue damage if not tightly regulated. FcγR activity and phagocytosis are governed by phosphatases such as the SH2 (Src Homology 2) domain-containing inositol phosphatases SHIP and SHIP-2 , . These phosphatases share a high degree of homology within the catalytic domains. "
[Show abstract][Hide abstract] ABSTRACT: SHIP and SHIP-2 are inositol phosphatases that regulate FcγR-mediated phagocytosis through catalytic as well as non-catalytic mechanisms. In this study we have used two-dimensional fluorescence difference gel electrophoresis (DIGE) analysis to identify downstream signaling proteins that uniquely associate with SHIP or SHIP-2 upon FcγR clustering in human monocytes. We identified LyGDI as a binding partner of SHIP, associating inducibly with the SHIP/Grb2/Shc complex. Immunodepletion and competition experiments with recombinant SHIP domains revealed that Grb2 and the proline-rich domain of SHIP were necessary for SHIP-LyGDI association. Functional studies in primary human monocytes showed that LyGDI sequesters Rac in the cytosol, preventing it from localizing to the membrane. Consistent with this, suppression of LyGDI expression resulted in significantly enhanced FcγR-mediated phagocytosis.
PLoS ONE 06/2011; 6(6):e21175. DOI:10.1371/journal.pone.0021175 · 3.23 Impact Factor
"High expression of MMR is found on DC and lower expression on LC (Turville et al., 2002). CD16 is expressed by some DC, but not by LC, and ligation of CD16 may be associated with signalling of NFnB (Pengal et al., 2003). DC-associated, HS proteoglycandependent integrin ligand (DC-HIL) (Shikano et al., 2001b) is expressed at low levels intracellularly by DC, but not by LC (Fig. 6B). "
[Show abstract][Hide abstract] ABSTRACT: As human papillomavirus-like particles (HPV-VLP) represent a promising vaccine delivery vehicle, delineation of the interaction of VLP with professional APC should improve vaccine development. Differences in the capacity of VLP to signal dendritic cells (DC) and Langerhans cells (LC) have been demonstrated, and evidence has been presented for both clathrin-coated pits and proteoglycans (PG) in the uptake pathway of VLP into epithelial cells. Therefore, we compared HPV-VLP uptake mechanisms in human monocyte-derived DC and LC, and their ability to cross-present HPV VLP-associated antigen in the MHC class I pathway. DC and LC each took up virus-like particles (VLP). DC uptake of and signalling by VLP was inhibited by amiloride or cytochalasin D (CCD), but not by filipin treatment, and was blocked by several sulfated and non-sulfated polysaccharides and anti-CD16. In contrast, LC uptake was inhibited only by filipin, and VLP in LC were associated with caveolin, langerin, and CD1a. These data suggest fundamentally different routes of VLP uptake by DC and LC. Despite these differences, VLP taken up by DC and LC were each able to prime naive CD8(+) T cells and induce cytolytic effector T cells in vitro.
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