The Membrane-Proximal Immunoreceptor Tyrosine-Based Inhibitory Motif Is Critical for the Inhibitory Signaling Mediated by Siglecs-7 and -9, CD33-Related Siglecs Expressed on Human Monocytes and NK Cells

Division of Cell Biology and Immunology, The Wellcome Trust Biocentre, University of Dundee, Dundee, United Kingdom.
The Journal of Immunology (Impact Factor: 4.92). 01/2005; 173(11):6841-9. DOI: 10.4049/jimmunol.173.11.6841
Source: PubMed


Siglec-7 and Siglec-9 are two members of the recently characterized CD33-related Siglec family of sialic acid binding proteins and are both expressed on human monocytes and NK cells. In addition to their ability to recognize sialic acid residues, these Siglecs display two conserved tyrosine-based motifs in their cytoplasmic region similar to those found in inhibitory receptors of the immune system. In the present study, we use the rat basophilic leukemia (RBL) model to examine the potential of Siglecs-7 and -9 to function as inhibitory receptors and investigate the molecular basis for this. We first demonstrate that Siglecs-7 and -9 are able to inhibit the FcepsilonRI-mediated serotonin release from RBL cells following co-crosslinking. In addition, we show that under these conditions or after pervanadate treatment, Siglecs-7 and -9 associate with the Src homology region 2 domain-containing phosphatases (SHP), SHP-1 and SHP-2, both in immunoprecipitation and in fluorescence microscopy experiments using GFP fusion proteins. We then show by site-directed mutagenesis that the membrane-proximal tyrosine motif is essential for the inhibitory function of both Siglec-7 and -9, and is also required for tyrosine phosphorylation and recruitment of SHP-1 and SHP-2 phosphatases. Finally, mutation of the membrane-proximal motif increased the sialic acid binding activity of Siglecs-7 and -9, raising the possibility that "inside-out" signaling may occur to regulate ligand binding.

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    • "The postulated inhibitory activity of Siglec 9 on the immune response through host and bacterial sialoglycans recognition [16] [17] prompted us to verify its expression on monocyte subsets at baseline and after IVIg. "
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    • "Siglec-9 is also expressed on monocytes and MUC16 promotes binding of monocytes to cancer cells. Upon binding to its ligand, Siglec-9 is phosphorylated on its Immunoreceptor Tyrosine-based Inhibition Motif (ITIM) tail, triggering an inhibitory signaling cascade that results in inhibition of the NK cell response [121]. Ovarian cancer cells are therefore likely to be protected from NK cell and monocyte attack due to the negative signaling induced via MUC16-Siglec-9 interaction (Figure 5). "
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    • "Moreover, we also demonstrate that constitutive expression of Siglec-7 on MDMs markedly contributes to their HIV-1 infection, since specific Siglec-7 blockade induced a reduction of HIV-RNA in MDMs in a 4-hour entry assay. The degree of surface levels of Siglec-7 on monocytes and macrophages is still being debated, as there is one study showing that expression of this molecule on monocytes and MDMs is very low [29], while several other reports demonstrated that monocytes express high levels of Siglec-7 [9,11,12,46,52]. In our experiments, the constitutive expression of Siglec-7 on monocytes paralleled the ones observed on NK cells [13], and remained very high on MDMs even after 7 days of culture with GM-CSF although at lower levels if compared to freshly isolated monocytes. "
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