Glycobiology vol. 24 no. 9 pp. 807–817, 2014 doi:10.1093/glycob/cwu066 Advance Access publication on July 6, 2014 REVIEW CD22 and Siglec-G regulate inhibition of B-cell signaling by sialic acid ligand binding and control B-cell tolerance Downloaded from http://glycob.oxfordjournals.org/ Lars Nitschke1 co-receptors is regulated by extracellular-bound ligands, which Division of Genetics, Department of Biology, University of Erlangen, determine the association of the co-receptors to the BCR. B-cell Erlangen, Germany immunity depends on recognition of foreign antigens and a lack of responsiveness to self-antigens. Although central B-cell toler- Received on April 29, 2014; revised on June 26, 2014; accepted on June 27, 2014 ance is induced in the bone marrow by several mechanisms, many peripheral B cells in the periphery escape these tolerance CD22 and Siglec-G are two B-cell expressed members of the mechanisms, produce self-reactive antibodies and need to be con- Siglec (sialic acid-binding immunoglobulin (Ig)-like lectin) trolled by peripheral tolerance mechanisms in order to prevent family and are potent inhibitors of B-cell signaling. Genetic autoimmunity (Wardemann et al. 2003). Inhibitory co-receptors approaches have provided evidence that this inhibition of on B cells contribute to this control of B-cell tolerance. at Universitaet Erlangen-Nuernberg, Wirtschafts- und Sozialwissenschaftliche Z on August 3, 2016 B-cell antigen receptor (BCR) signaling by Siglecs is depend- CD22 (Siglec-2) and Siglec-G (or its human ortholog ent on ligand binding to sialic acids in specific linkages. The Siglec-10) are two B-cell expressed members of the Siglec (sialic cis-ligand-binding activity of CD22 leads to homo-oligomer acid-binding immunoglobulin (Ig)-like lectin) family and have formation, which are to a large extent found in membrane been shown to negatively regulate BCR signaling (Jellusova and domains that are distinct from those containing the BCR. In Nitschke 2012). Both are associated to some extent with the BCR contrast, Siglec-G is recruited via sialic acid binding to the (Peaker and Neuberger 1993; Zhang and Varki 2004; Müller et al. BCR. This interaction of Siglec-G with mIgM leads to an in- 2013) and are inhibitory co-receptors. Both Siglecs carry intracel- hibitory function that seems to be specific for B-1 cells. Both lular inhibitory ITIMs (immunoreceptor tyrosine-based inhibition CD22 and Siglec-G control B-cell tolerance and loss of these motifs) and recruit the tyrosine phosphatase SHP-1 that inhibits proteins, its ligands or its inhibitory pathways can increase signaling (Doody et al. 1995; Pfrengle et al. 2013). The inhibitory the susceptibility for autoimmune diseases. CD22 is a target functions of both Siglecs are regulated by ligand binding (Hutzler protein both in B-cell leukemias and lymphomas, as well as et al. 2014). Both Siglecs bind to sialic acids in specific linkages, in B-cell mediated autoimmune diseases. Both antibodies which are attached to glycoproteins and which are present on and synthetic chemically modified sialic acids are currently cellular surfaces, including the B-cell surface. The presence or tested to target Siglecs on B cells. absence of these ligands regulates the association of these two Siglecs to the BCR and thus influences the strength of inhibition. Keywords: autoimmunity / B lymphocytes / inhibitory Since sialic acids are abundantly expressed in vertebrates, but signaling / Siglecs often not found on microorganisms they are a part of “self- structures” and there is evidence that the sialic acid binding by B-cell Siglecs regulates B-cell tolerance and prevents autoimmun- Introduction to B-cell Siglecs ity (Crocker et al. 2007). B cells are an important part of the adaptive immune system and generate the humoral immune response by secretion of antigen- Regulation of B-cell signaling specific antibodies. The signaling by the B-cell antigen receptor (BCR), induced by antigen binding is crucial for triggering B-cell CD22 and Siglec-G are the only two Siglecs which are development, B-cell activation, as well as differentiation to expressed on murine B cells. Human B cells express CD22, antibody-secreting plasma cells. Accessory transmembrane mole- Siglec-10, as well as Siglec-6 (Crocker et al. 2007). CD22 has a cules or co-receptors on the B-cell surface modulate BCR signal- very B-cell restricted expression pattern (Torres et al. 1992). An ing. These B-cell co-receptors can be activatory or inhibitory and expression and function outside of the B-cell lineage has only fulfil these functions by binding additional signaling proteins been shown on intestinal eosinophils and on a DC subpopula- with their intracellular tails and thereby recruiting these effectors tion (Wen et al. 2012; Ma et al. 2013). Also Siglec-G shows the into the vicinity of the BCR. Very often the activity of highest expression pattern on B cells; however, it is also expressed on dendritic cells (DCs) and eosinophils as shown by use of newly generated monoclonal anti-Siglec-G antibodies 1To whom correspondence should be addressed: Tel: +49-9131-85-28453; (Pfrengle et al. 2013; Hutzler et al. 2014). Both Siglecs are able Fax: +49-9131-85-28526; e-mail: [email protected] to associate to the BCR on B cells and inhibit BCR-mediated © The Author 2014. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: [email protected] 807 L Nitschke signaling. While both Siglecs can inhibit BCR signaling on B 1996; Otipoby et al. 1996; Sato et al. 1996; Nitschke et al. cells, Siglec-G deficient mice show that Siglec-G is mainly a 1997). A second phosphatase, the inositol phosphatase SHIP, non-redundant inhibitory receptor on B-1 cells, a B-cell subpo- can also be bound to the tyrosine phosphorylated CD22 tail in a pulation with special functions (Ding et al. 2007; Hoffmann complex with the adaptor proteins Grb2 and Shc, but has a so et al. 2007). In contrast, CD22 seems to be the dominant inhibi- far unknown function (Poe et al. 2000). The CD22 ITIM motifs tory Siglec for conventional B cells (also called B-2 cells). are critical for the inhibitory function of CD22, as has been However, when transfected into a chicken B-cell line, which shown in B-cell lines, but also recently in mice in which the resembles immature conventional B cells, Siglec-G can also three ITIM tyrosines were mutated (Müller et al. 2013). These Downloaded from inhibit surface (s)IgM-triggered signaling there (Hoffmann CD22 ITIM-mutated knockin mice (called CD22-Y2,5,6F et al. 2007). Also when mouse B cells are treated with high- mice, for mutations in the 2nd, 5th and 6th tyrosine of the three affinity Siglec-G ligands, BCR signaling can be suppressed in ITIMs) had a similar phenotype as CD22-deficient mice. conventional B cells (Pfrengle et al. 2013). The inhibitory sig- Siglec-G is a negative regulator of Ca2+ signaling specifically naling by CD22 and Siglec-G has been the subject of several in B-1 cells, as Siglecg−/− mice have a B-1-cell restricted pheno- recent reviews and will therefore be covered here only briefly type and Siglecg−/− B-1 cells show increased anti-IgM-induced http://glycob.oxfordjournals.org/ (Nitschke 2009; Jellusova and Nitschke 2012). Ca2+ signaling, in contrast to B-2 cells of these mice (Hoffmann The signaling function of CD22 has been dissected in detail et al. 2007). Siglec-G and the human ortholog Siglec-10 both by several groups. The intracellular tail of CD22 carries six tyr- have one ITIM and one ITIM-like sequence and can bind SHP-1 osines, which are the only known signaling modules of CD22. (as shown for Siglec-G) (Pfrengle et al. 2013)orSHP-1andthe Three tyrosines are found within conventional ITIMs (Y783, related phosphatase SHP-2 (as shown for Siglec-10) (Whitney Y843 and Y863), one is part of an ITIM-like motif (Y817) and et al. 2001). How these SHP phosphatases regulate downstream one is a Grb-2 binding motif (Y828) (Doody et al. 1995; signaling in B-1 cells is not known in detail as crucial signaling Otipoby et al. 2001) (Figure 1). Upon BCR cross-linking CD22 molecules which are part of the BCR signalosome, such as is rapidly phosphorylated by the Src kinase Lyn (Smith et al. PLCγ, Btk or SLP65/BLNK showed unchanged tyrosine phos- at Universitaet Erlangen-Nuernberg, Wirtschafts- und Sozialwissenschaftliche Z on August 3, 2016 1998). The phosphorylated ITIM residues of the CD22 tail are phorylation in Siglecg−/− B-1 cells (Hoffmann et al. 2007). The bound by the tyrosine phosphatase SHP-1, a central inhibitory transcription factor NFATc1 has elevated expression levels in signaling molecule (Doody et al. 1995). SHP-1 is the main Siglecg−/− B-1 cells, which could be an autoregulatory process downstream effector of CD22 and CD22-deficient B cells show due to increased Ca2+ signaling (Jellusova and Nitschke 2012). increased tyrosine phosphorylation of several proteins which One study also reported increased NF-kB activation in Siglecg−/− are involved in triggering proximal BCR signaling responses, B-1 cells, a finding that was not confirmed by another group such as SLP65/BLNK, Vav1 and CD19 (Fujimoto et al. 1999; (Ding et al. 2007; Jellusova and Nitschke 2012). Gerlach et al. 2003). Correspondingly, BCR-induced Ca2+ sig- Both CD22 and Siglec-G also modulate TLR signaling in B naling is strongly enhanced in Cd22−/− B cells (O’Keefe et al. cells. TLRs are pattern recognition receptors for bacterial or viral Fig. 1. Regulation of BCR signaling by CD22 and Siglec-G binding to sialic acids in cis.(A) CD22 is an inhibitor of BCR signaling on conventional B cells (B-2 cells).