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Sialic acid binding Ig-like lectin family
Identifiers
SymbolSIGLEC
Membranome210

Siglecs (Sialic acid-binding immunoglobulin-type lectins) are cell surface proteins that bind sialic acid. They are found primarily on the surface of immune cells and are a subset of the I-type lectins. There are 14 different mammalian Siglecs, providing an array of different functions based on cell surface receptor-ligand interactions.[1]

History

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The first described candidate Siglec was Sialoadhesin (Siglec-1/CD169) a lectin-like adhesion protein on macrophages.[2] Parallel studies by Ajit Varki and colleagues on the previously cloned CD22 (a B cell surface protein involved in adhesion and activation) showed direct evidence for sialic acid recognition. The subsequent cloning of Sialoadhesin by Crocker revealed homology to CD22 (Siglec-2), CD33 (Siglec-3) and myelin-associated glycoprotein (MAG/Siglec-4), leading to the proposal for a family of "Sialoadhesins". Varki then suggested the term Siglec as a better alternative and as a subset of I-type (Ig-type) lectins. This nomenclature was agreed upon and has been adopted by almost all investigators working on these molecules (by convention, Siglecs are always capitalised.) Several additional Siglecs (Siglecs 5–12) have been identified in humans that are highly similar in structure to CD33 and so are collectively referred to as "CD33-related Siglecs".[3] Further Siglecs have been identified including Siglec-14 and Siglec-15. Siglecs have been characterized into two distinct groups: the first and highly conserved-across-mammals group composed of Sialoadhesins, CD22, MAG, and Siglec-15, and a second group comprising Siglecs closely related to CD33.[4][5] Others such as Siglec-8 and Siglec-9 have homologues in mice and rats (Siglec-F and Siglec-E respectively in both). Humans have a higher number of Siglecs than mice and so the numbering system was based on the human proteins.[6]

Structure

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Sialoadhesin's variable immunoglobulin domain in complex with a sialylated glycan, focusing on the conserved salt bridge found in all siglecs
Sialoadhesin's variable immunoglobulin domain in complex with a sialylated glycan. Glycan carbons are in purple, protein carbons in green, oxygens in red, nitrogens in blue and hydrogens in white.

Siglecs are Type I transmembrane proteins where the NH3+-terminus is in the extracellular space and the COO−-terminus is cytosolic.[7] Each Siglec contains an N-terminal V-type immunoglobulin domain (Ig domain) which acts as the binding receptor for sialic acid. These lectins are placed into the group of I-type lectins because the lectin domain is an immunoglobulin fold. All Siglecs are extended from the cell surface by C2-type Ig domains which have no binding activity. Siglecs differ in the number of these C2-type domains.[6] As these proteins contain Ig domains, they are members of the Immunoglobulin superfamily (IgSF).

Most Siglecs, such as CD22 and the CD33-related family, contain ITIMs (Immunoreceptor tyrosine-based inhibitory motifs) in their cytosolic region.[7] These act to down-regulate signaling pathways involving phosphorylation, such as those induced by ITAMs (Immunoreceptor tyrosine-based activation motifs).[8] Some, however, like Siglec-14, contain positive amino acid residues that help dock ITAM-containing adaptor proteins such as DAP12.[1]

Ligand binding

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Due to the acidic nature of sialic acid, Siglec active sites contain a conserved arginine residue which is positively charged at physiological pH. This amino acid forms salt bridges with the carboxyl group of the sugar residue.[6] This is best seen in Sialoadhesin, where arginine at position 97 forms salt bridges with the COO− group of the sialic acid, producing a stable interaction.[9] Each lectin domain is specific for the linkage that connects sialic acid to the glycan. Sialic acid contains numerous hydroxyl groups which can be involved in the formation of glycosidic linkages, which are observed at carbons number 2, 3, 6, and 8 of the sugar backbone. The binding specificity of each Siglec is due to different chemical interactions between the sugar ligand and the Siglec amino acids. The position in space of the individual groups on the sugar and the protein amino acids affects the sialic acid linkage to which each Siglec binds. For example, Sialoadhesin preferentially binds α2,3 linkages over α2,6 linkages.[9]

Function

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Schematic representation of the CD22 and B-cell receptor signalling process, showing the domain structure of CD22
Simplified schematic representation of the CD22 and B-cell receptor signalling process. pTyr refers to phosphotyrosine. The blocked line represents inhibition.

The primary function of Siglecs is to bind glycans containing sialic acids. These receptor-glycan interactions can be used in cell adhesion, cell signalling and others. The function of Siglecs is limited to their cellular distribution. For example, MAG is found only on oligodendrocytes and schwann cells whereas Sialoadhesin is localised to macrophages.

Most Siglecs are short and do not extend far from the cell surface. This prevents most Siglecs from binding to other cells as mammalian cells are covered in sialic acid-containing glycans. This means that the majority of Siglecs only bind ligands on the surface of the same cell, so called cis -ligands, as they are "swamped" by glycans on the same cell. One exception is Sialoadhesin which contains 16 C2-Ig domains, producing a long, extended protein allowing it to bind trans-ligands, i.e. ligands found on other cells. Others, such as MAG, have also been shown to bind trans-ligands.

Signalling

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The members of the siglec family are paired receptors with opposing intracellular signaling functions.[10][11] Due to their ITIM-containing cytoplasmic regions, most Siglecs interfere with cellular signalling, inhibiting immune cell activation. Once bound to their ligands, Siglecs recruit inhibitory proteins such as SHP phosphatases via their ITIM domains.[12] The tyrosine contained within the ITIM is phosphorylated after ligand binding and acts as a docking site for SH2 domain-containing proteins like SHP phosphatases. This leads to de-phosphorylation of cellular proteins, down-regulating activating signalling pathways.

Examples of negative signalling:

  • CD22 is found on B cells. B cells become active when the B-cell receptor (BCR) binds to its cognate ligand. Once the BCR is bound to its ligand, the receptor auto-phosphorylates its cytoplasmic region (cytoplasmic tail). This leads to phosphorylation of the three ITIMs in CD22's cytoplasmic tail, leading to the recruitment of SHP-1 which negatively regulates BCR-based cellular activation. This creates an activation threshold for B cell activation whereby transient activation of B cells is prevented.[13] CD22 inhibition of BCR signalling was originally thought to be sialic acid-binding-independent, but evidence suggests α2,6 sialic acid ligands are required for inhibition.[14]
  • Siglec-7 is found on Natural Killer cells (NK cells). Siglec-7 leads to cellular inactivation once bound to its sialic acid-containing cognate ligand and is found in high levels on NK cell surfaces. It is used in cell-cell contacts, binding to sialylated glycans on target cells leading to inhibition of NK cell-dependent killing of the target cell. Mammalian cells contain high levels of sialic acid and so when NK cells bind so called "self-cells", they are not activated and do not kill host cells.

Siglec-14 contains an arginine residue in its transmembrane region.[15] This binds to the ITAM-containing DAP10 and DAP12 proteins. When bound to its ligand, Siglec-14 leads to activation of cellular signalling pathways via the DAP10 and DAP12 proteins.[7] These proteins up-regulate phosphorylation cascades involving numerous cellular proteins, leading to cellular activation. Siglec-14 appears to co-localise with Siglec-5, and as this protein inhibits cellular signalling pathways, co-ordinate opposing functions within immune cells.[15]

Phagocytosis and adhesion

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Siglecs that can bind trans-ligands, such as Sialoadhesin, allow cell-cell interactions to take place. These glycan-Siglec interactions allow cells to bind one another, allowing signalling in some cases, or in the case of Sialoadhesin, pathogen uptake. Sialoadhesin's function was originally thought to be important in binding to red blood cells. Sialoadhesin lacks a cytosolic ITIM or a positive residue to bind ITAM-containing adaptors and so is thought not to influence signalling. Studies show that this protein is involved in phagocytosis of bacteria that contain highly sialylated glycan structures such as the lipopolysaccharide of Neisseria meningitidis.[16] Binding to these structures allows the macrophage to phagocytose these bacteria, clearing the system of pathogens.

Siglec-7 is also used in binding to pathogens such as Campylobacter jejuni. This occurs in a sialic acid-dependent manner and brings NK cells and monocytes, on which Siglec-7 is expressed, into contact with these bacteria.[17] The NK cell is then able to kill these foreign pathogens.

Knock-out studies

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Knock-out studies are often used to uncover the function proteins have within a cell. Mice are often used as they express orthologous proteins of ours, or extremely similar homologues.

Some examples of knock-out Siglecs include:

  • CD22: Walker & Smith conducted experiments with CD22 knock-outs and deletion mutants to discern CD22's function.[18] These mutant B cells did not infer any autoimmune disease, but they did see an increased production of autoantibodies due to the lack of BCR signalling inhibition, usually conducted by CD22. Autoantibodies are specific for self proteins and can harm the host. CD22 is normally up-regulated by lipopolysaccharide binding to Toll-like receptors. The mutant B cells can not up-regulate the mutant protein and so become hyper-sensitive in the presence of lipopolysaccharide. This means that the B cells overproduce antibodies when antibodies would not normally have been produced.
  • MAG (Myelin-associated glycoprotein) is expressed on cells that form myelin sheaths (schwann cells and oligodendrocytes) around neurons. MAG binds to sialylated ligands on the neuron. Knock-out of MAG in the peripheral nervous system leads to decreased myelination of neurons. Knock-out of MAG in the central nervous system of mice does not appear to affect myelination, but the interaction between the myelin and the neuron does deteriorate with age. This leads to neurological defects as the action potential can not pass so rapidly down the length of the axon during neural stimulation. Removing the ligand for MAG, by knocking-out the GalNAc transferase gene required for ligand formation, has similar effects to that of the MAG knock-out mice[19]

Human/Primate Siglecs

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Name Cellular distribution[7] Sialic acid linkage specificity[6] No. of C2-Ig domains[7] ITIM or positive residue[7]
Siglec-1 (Sialoadhesin) Macrophages α2,3>α2,6 16 None
Siglec-2 (CD22) B cells α2,6 6 ITIM
Siglec-3 (CD33) Myeloid progenitors, Monocytes α2,6>α2,3[20] 1 ITIM
Siglec-4 (MAG) Myelin α2,3>α2,6 4 None
Siglec-5 (CD170) Neutrophils, Monocytes α2,3 3 ITIM
Siglec-6 (CD327) Trophoblasts, Mast cells α2,6 2 ITIM
Siglec-7 (CD328) NK cells α2,8>α2,6>α2,3 2 ITIM
Siglec-8 Eosinophils, Mast cells α2,3>α2,6 2 ITIM
Siglec-9 (CD329) Monocytes, Neutrophils, Dendritic cells α2,3=α2,6 (prefers sulfated residues) 2 ITIM
Siglec-10 B cells α2,3=α2,6 4 ITIM
Siglec-11 B cells α2,8 4 ITIM
Siglec-12[21] Macrophages No binding[6] 2 ITIM
Siglec-13[22] Chimpanzee monocytes
Siglec-14 Unknown α2,6[15] 2 Arginine[15]
Siglec-15[4] Osteoclasts, Macrophages, DCs Siglec-15[4][23][24] α2,6[25] 1 Lysine[25]
Siglec-16[26] Tissue macrophages
Siglec-17 [22] NK cells

This table briefly summarises the cellular distribution of each human/primate Siglec; the linkage specificity each has for sialic acid binding; the number of C2-Ig domains it contains; and whether it contains an ITIM or a positive residue to bind ITAM-containing adaptor proteins. References in the column headings correspond to all information displayed in that column, unless other references are shown. Siglec-12 information is referenced by[21] only, excluding the linkage specificity.

Mimetics

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Many pathologies have been linked to the spontaneous interactions between sialic acid and the immunosuppressive sialic acid-binding immunoglobulin-like lectin (Siglec) receptors on immune cells such as cancer,[27] HIV-1[28] and Group B Strep Infection.[29][30] The sialic acid family branches from glycans, sugar chains comprising various monosaccharides that cover the membrane of every living cell and display a staggering structural diversity. Sialic acids function in protein folding, neural development, cellular interactions, among many other physiological processes. As sialic acids are abundantly expressed in vertebrates and not in microorganisms, they are considered self-antigens or self-structures that play major role in inhibiting harmful immune system activity by regulating neutrophils and B cell tolerance.[31]

Within the immune system, Siglecs, especially those related to CD33, sialic acid and Siglec-binding pathogens are subjected to the runaway Red Queen co-evolution phenomenon by a selection pressure that maintains the innate immune system's capacity for self-recognition and ensures prevention of autoimmunity diseases.[32][33] This evolutionary chain and incessant mutations have made Siglecs one of the most rapidly evolving gene, evidenced by both intra- and inter-species differences.[33] The polymorphism of human-unique Siglec-12, -14 and -16 suggests that the selection pressure is ongoing.[32]

As Siglecs feature distinct binding preferences for the sialic acid and its modifications, several attempts have been made to chemically modify natural sialic acid ligands and eventually led to the creation of sialic acid mimetics (SAMs) with enhanced binding capacity and selectivity towards Siglecs.[34]

Synthesis

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SAMs can be used to target Siglecs and modulate Siglec-expressing cells by modifying the sialic acid backbone at various positions, from C-2 to C-9.[34][35][36] The carboxylic acid, however, must be left intact.[34] The first attempts were made to develop high-affinity sialic acid mimetics for Siglec-2, which led to the discovery that with increased binding affinity came hydrogen bonding and lipophilic interactions between SAMs and Siglec-2.[34] Several separate modifications have been made at the C-2, C-5 and C-9 positions, leading Mesch et al. to hypothesizing that the simultaneous modification at all three positions could lead to optimization of binding.[37]

Success in drastically enhanced binding of SAMs to Siglec 2 suggests that a similar approach can work on other members of the family. Some modifications have included an additional simultaneous modification at the C-4 position on the sialic acid backbone.[34] The development of (copper) I-catalyzed azide alkyne cycloaddition (CuAAC) click chemistry has expedited the identification of new SAMs and allowed for the creation of novel SAMs with high binding to Siglec-3, -5, -6, -7 and -10.[38] As of 2017, SAMs for most Siglecs have been reported, except for Siglec -6, -8, -11, -14, -15 and -16.[34]

Clustering of receptors and high-avidity binding, collectively known as multivalent binding, can enhance the effectiveness of SAMs in the human body. Currently, advancements in glycoengineering have made use of SAM-decorated nanoparticles, SAM-decorated polymers and on-cell synthesis of SAMs to present SAMs to Siglecs.[34] Liposomes crosslinked with SAMs also have been shown to aid in presenting antigens to antigen-presenting cells via the Siglec-1 or -7 pathways.[39] Moreover, human cells, engineered with sialic acids carrying Ac5NeuNPoc incorporated into its sialoglycans and 3-bromo-benzyl azide, showed hyperactivity towards Siglec-2.[34][40]

References

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Siglec

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Siglecs, or sialic acid-binding immunoglobulin-like , constitute a family of cell surface receptors that specifically recognize sialic acid-containing glycans on glycoproteins and glycolipids, primarily expressed on hematopoietic cells of the to regulate innate and adaptive immune responses. In humans, there are 14 functional Siglec genes, encoding proteins such as Siglec-1 through Siglec-4 and Siglec-5 through Siglec-11, as well as Siglec-14 through Siglec-16, with additional pseudogenes reflecting evolutionary dynamics. Structurally, Siglecs are type I transmembrane glycoproteins belonging to the , featuring an extracellular region with a variable number of immunoglobulin-like domains—typically a membrane-distal V-set domain responsible for binding, followed by varying numbers of C2-set domains for interaction and . Intracellularly, most Siglecs contain immunoreceptor tyrosine-based inhibitory motifs (ITIMs) that recruit phosphatases like SHP-1 and SHP-2 upon binding, delivering inhibitory signals to dampen immune cell ; however, a subset associates with DAP12, an adaptor protein bearing an (ITAM), enabling activating functions. This dual inhibitory-activating potential allows Siglecs to fine-tune immune signaling in response to self-associated molecular patterns (SAMPs) on host cells versus pathogen-associated molecular patterns. Siglecs are classified into two main subsets based on evolutionary conservation and sequence similarity: the conserved Siglecs (Siglec-1/sialoadhesin, , Siglec-4/MAG, and Siglec-15), which are ancient and found across tetrapods with relatively stable structures, and the rapidly evolving CD33-related Siglecs ( and Siglec-5 to -11, -14, and -16), which exhibit species-specific duplications, deletions, and polymorphisms driven by -host coevolution. Expression patterns are cell-type specific; for instance, Siglec-1 is predominantly on macrophages and dendritic cells, on B lymphocytes, on myeloid cells including monocytes and , and Siglec-7 and -9 on natural killer cells and subsets of T cells. These patterns underscore their roles in diverse immune contexts, from self/non-self to recognition. Functionally, Siglecs primarily mediate immune inhibition to promote tolerance and prevent by recognizing sialylated glycans on healthy cells, thereby setting thresholds for immune ; for example, Siglec-2 on B cells attenuates BCR signaling to control production, while Siglec-3 on myeloid cells modulates in tissues like . Activating Siglecs, such as paired like Siglec-14 (counterpart to inhibitory Siglec-5), can enhance responses against pathogens by promoting or release. Beyond immunity, Siglecs influence , migration, and survival, with implications in diseases including , cancer, neurodegeneration, and infections, where dysregulated sialic acid-Siglec interactions contribute to immune evasion or chronic .

Discovery and History

Initial Identification

The discovery of the first Siglec family member, sialoadhesin (Siglec-1), occurred in when it was identified as a sialic acid-dependent receptor on resident in , specifically recognizing glycoconjugates on sheep erythrocytes. This receptor was characterized as a large, macrophage-restricted glycoprotein that mediated sialic acid-specific hemagglutination, marking it as a novel involved in interactions with sialylated cells. Initial studies highlighted its expression on stromal macrophages in lymphohematopoietic tissues, distinguishing it from other known macrophage markers. In the early 1990s, (Siglec-2) was identified as a B-cell-specific surface that mediated homotypic and heterotypic , particularly to monocytes and erythrocytes. Cloned from a B-lymphocyte , was noted for its seven immunoglobulin-like domains and its role in facilitating cell-cell interactions during B-cell activation. Subsequent investigations in the mid-1990s revealed its function as an inhibitory receptor that modulates B-cell signaling, with studies demonstrating hyperresponsive B cells lacking this regulatory control. CD33 (Siglec-3) was recognized in 1988 through the cloning of its cDNA from a myeloid cell line, establishing it as a differentiation expressed on myeloid progenitor cells and maturing monocytes but absent on mature granulocytes and non-myeloid lineages. This marker was particularly valuable for identifying early myeloid commitment in hematopoiesis, with monoclonal antibodies like My9 confirming its restricted expression pattern on leukemic blasts in . By the early 1990s, further characterization emphasized its two immunoglobulin-like domains and potential involvement in myeloid , though its precise ligand interactions remained under exploration at the time. Initial studies in the late 1990s linked sialoadhesin, , and through shared structural features, including N-terminal V-set immunoglobulin-like domains that bind sialic acids and multiple C2-set domains in their extracellular regions. These commonalities, along with their expression on hematopoietic cells and roles in sialic acid-dependent recognition, prompted Paul Crocker to propose the unified "Siglec" in 1998, denoting sialic acid-binding immunoglobulin-like as a distinct subfamily of I-type . This classification laid the groundwork for recognizing Siglecs as a cohesive group prior to broader family expansion.

Evolution of the Siglec Family

The Siglec family emerged in the common ancestor of jawed vertebrates, with orthologs of conserved members such as Siglec-4 (myelin-associated glycoprotein, MAG) identified in cartilaginous and , indicating an ancient role in vertebrate immunity and neural function. This phylogenetic origin traces back over 500 million years, predating the diversification of tetrapods, and suggests that Siglecs arose as sialic acid-binding receptors to modulate early immune responses in vertebrates. An ancestral gene is proposed as the progenitor for the rapidly evolving CD33-related (CD33r) subfamily, forming an initial through duplication events that allowed to pressures and host glycan changes. The Siglec family divides into two main subfamilies based on evolutionary conservation and sequence similarity: the conserved group, including Siglec-1 (sialoadhesin), Siglec-2 (), Siglec-4 (MAG), and Siglec-15, which are present across most mammals with stable orthology and consistent sialic acid-binding preferences; and the CD33r subfamily (Siglec-3/ and Siglec-5 through -14 and -16), characterized by rapid evolution, high interspecies variability, and species-specific expansions or contractions. The conserved subfamily maintains core functions in and signaling, while the CD33r group exhibits dynamic changes driven by gene duplications, conversions, and pseudogenizations, reflecting an with pathogens that mimic host sialic acids. For instance, placental mammals display 5–20 CD33r Siglecs, with mice possessing approximately 7 (Siglec-D, -E, -F, -G, -H, -I) compared to 10–13 in s (Siglec-3, -5 to -12, -14, -16), highlighting lineage-specific diversification. These duplication events, often occurring in tandem clusters on s (e.g., chromosome 19q13), have generated functional diversity, including paired activating and inhibitory receptors that fine-tune immune activation. The evolution of Siglecs has been intimately linked to the co-evolution of sialic acids, the primary ligands that enable self-recognition in the immune system. Sialic acids, enriched on vertebrate cell surfaces, serve as "self-associated molecular patterns" (SAMPs) that Siglecs detect to inhibit overzealous immune responses and prevent autoimmunity, a mechanism that likely originated in early jawed vertebrates to distinguish host glycans from microbial mimics. Pathogen exploitation of sialic acid-Siglec interactions has driven selective pressures, leading to expansions in the CD33r subfamily and adaptations like human-specific loss of Neu5Gc (N-glycolylneuraminic acid), which altered ligand availability and Siglec binding specificities to enhance self/non-self discrimination. This co-evolutionary dynamic underscores Siglecs' role in balancing tolerance and defense across vertebrate lineages.

Structure and Classification

Overall Molecular Architecture

Siglecs constitute a family of type I transmembrane glycoproteins within the , characterized by a modular extracellular domain that facilitates glycan recognition and cell-cell interactions. The extracellular region typically comprises an N-terminal V-set immunoglobulin-like domain, responsible for binding, followed by 1 to 16 C2-set immunoglobulin-like domains that provide and extend the receptor to its s. This domain organization varies across family members; for instance, the CD33-related Siglecs generally feature 1 V-set and 1-4 C2-set domains, while sialoadhesin (Siglec-1) possesses 1 V-set and 16 C2-set domains. The V-set domain shares conserved features with other I-type lectins, including a characteristic residue essential for interaction. Anchoring the protein to the plasma membrane is a single-pass , typically a hydrophobic alpha-helix of approximately 20-25 , located membrane-proximally to the extracellular Ig-like domains. This region often includes a basic residue in certain activating Siglecs, such as Siglec-14, Siglec-15, and Siglec-16, which enables non-covalent association with adaptor proteins like DAP12 for . The cytoplasmic tail, generally short (40-50 ), harbors key signaling motifs: most Siglecs contain one or two immunoreceptor tyrosine-based inhibitory motifs (ITIMs) with the (I/V/L/S)-x-Tyr-x-x-(L/V/I), which, upon , recruit Src homology 2 domain-containing phosphatases such as SHP-1 and SHP-2 to dampen immune responses. Notably, Siglec-16 deviates from this inhibitory paradigm, lacking functional ITIMs and instead relying on DAP12 association to transduce activating signals via the ITAM motif in the adaptor. Siglecs are extensively post-translationally modified by N-linked at multiple residues within the extracellular Ig-like domains, contributing to , stability, and protection from . These glycosylation patterns, which include both sialylated and non-sialylated glycans, modulate the overall conformation of the extracellular region, thereby influencing ligand accessibility and preventing unwanted cis-interactions with self-glycans on the same cell surface.

Ligand Binding Mechanism

Siglecs primarily recognize and bind -containing glycans through their amino-terminal V-set immunoglobulin-like (Ig) domain, which contains a conserved residue that forms a with the carboxylate group of , such as (Neu5Ac). This interaction anchors the sialic acid in a binding pocket, with additional hydrophobic and polar contacts stabilizing the glycan . The V-set domain's specificity arises from variations in surrounding residues, allowing differential recognition of sialic acid modifications and linkages. Different Siglecs exhibit preferences for specific linkages to underlying glycans, influencing their binding affinities. For instance, Siglec-1 (sialoadhesin) shows a strong preference for α2-3-linked over α2-6-linked ones, enabling selective interactions with certain cell surface glycoconjugates. In contrast, Siglec-2 () favors α2-6 linkages, while other family members like Siglec-7 display affinity for both α2-6 and α2-8 disialylated structures. These linkage preferences, often with dissociation constants in the millimolar range for monovalent , can be enhanced by multivalency in natural glycan contexts. Binding affinity is further modulated by cis and trans interactions, where cis refers to Siglec-ligand engagement on the same cell surface and trans involves interactions between different cells. Cis ligands, abundant on immune cells expressing Siglecs, can occupy binding sites and reduce trans affinity, establishing a threshold for intercellular . This regulatory mechanism helps maintain immune by preventing excessive self-recognition while allowing activation upon encountering hypersialylated trans ligands, such as on pathogens or tumors. Crystallographic studies have elucidated the structural basis of these interactions, particularly for Siglec-7 bound to Neu5Ac derivatives. High-resolution structures reveal that the sialic acid's forms the key with the conserved (Arg107 in Siglec-7), while the acetamido group at C5 and at C6-9 engage in bonds and van der Waals contacts within the V-set pocket. For example, in complexes with α2-8-linked disialic acids, an additional sialic residue extends into a secondary , enhancing affinity through cooperative interactions. These insights from underscore the molecular determinants of glycan specificity across the Siglec family.

Family Classification

Siglecs are classified into two primary subfamilies based on evolutionary conservation, , and structural characteristics: the conserved subfamily and the CD33-related (CD33r) subfamily. This division reflects distinct phylogenetic branches, with the conserved group representing ancient origins stable across mammals and the CD33r group showing rapid diversification. The conserved subfamily includes Siglec-1 (sialoadhesin), Siglec-2 (), Siglec-4 (myelin-associated glycoprotein), and Siglec-15, which exhibit high sequence similarity to sialoadhesin and are preserved in most mammalian lineages without major expansions or losses. These members feature sialoadhesin-like extracellular architectures, including a prominent N-terminal V-set immunoglobulin domain essential for recognition, and are distinguished by their limited variability across species. The CD33r subfamily encompasses Siglec-3 (), Siglec-5 to Siglec-11, and Siglec-14 to Siglec-16, forming a larger, dynamically evolving cluster marked by interspecies diversity, including pseudogenes (such as human and ), gene duplications, and species-specific variants that contribute to immune adaptation. This subfamily's expansion is particularly evident in , where additional members like Siglec-14 and Siglec-16 arose through recent duplications of Siglec-5 and Siglec-11, respectively. Key classification criteria include the number of extracellular immunoglobulin-like domains, with examples in conserved Siglecs ranging from 2 (Siglec-15) to 17 (Siglec-1), while CD33r members have 2–5 domains; the composition of cytoplasmic tails, where CD33r Siglecs often harbor immunoreceptor tyrosine-based inhibitory motifs (ITIMs) or ITIM-like sequences for potential signaling; and differential preferences for sialic acid α2,3- versus α2,6-linkages in their V-set binding sites. In human nomenclature, SIGLEC genes are designated with numerical identifiers (SIGLEC1–SIGLEC16), predominantly clustered on 19q13.3–q13.4 for the CD33r subfamily and Siglec-2/Siglec-4, while SIGLEC1 localizes to 20p13 and SIGLEC15 to 18q12.3.

Biological Functions

Signaling Pathways

Siglecs primarily exert their regulatory effects through intracellular signaling pathways that can be inhibitory or activating, depending on their cytoplasmic motifs. The majority of Siglecs, such as those in the CD33-related subfamily, contain immunoreceptor tyrosine-based inhibitory motifs (ITIMs) in their cytoplasmic tails. Upon ligand binding and tyrosine phosphorylation by Src family kinases, these ITIMs recruit Src homology 2 (SH2) domain-containing protein tyrosine phosphatases, including SHP-1 and SHP-2. This recruitment leads to of key signaling molecules, such as immunoreceptor tyrosine-based motifs (ITAMs) on associated receptors, thereby dampening immune cell . A prominent example is (Siglec-2) on B cells, where ITIM-mediated SHP-1 recruitment dephosphorylates the (BCR) complex, inhibiting downstream pathways like calcium mobilization and proliferation. In contrast, a of Siglecs lacks ITIMs but features a charged residue in the , enabling association with the adaptor protein DAP12, which contains an ITAM. engagement phosphorylates the DAP12 ITAM, recruiting and activating spleen (Syk), which initiates proinflammatory signaling cascades, including MAPK and activation. Siglec-15 exemplifies this activating mode, pairing with DAP12 to enhance Syk-dependent responses in macrophages and monocytes, such as increased TGF-β production upon recognition. Siglec signaling often involves crosstalk with other immune receptors to fine-tune responses. For instance, CD22 interacts with β7 integrin on B cells, where SHP-1 recruitment inhibits integrin endocytosis and modulates cell homing. Similarly, Siglec-9 on macrophages attenuates Toll-like receptor (TLR) signaling by reducing LPS-induced MAPK phosphorylation, thereby limiting excessive inflammation. Cis-interactions with sialylated glycans on the same cell surface further modulate Siglec signaling by masking receptor availability and dampening trans-ligand responses. In B cells, cis-ligands bind to maintain it in a clustered, inactive state, requiring high-avidity trans-interactions to trigger inhibitory signaling and prevent . This mechanism ensures balanced immune thresholds across Siglec family members.

Phagocytosis and Cell Adhesion

Siglec-1, also known as sialoadhesin or CD169, is prominently expressed on macrophages and plays a key role in by binding to sialic acid-containing glycans on host cells and pathogens, facilitating tethering and initial capture. This interaction enables macrophages to adhere to sialylated surfaces, such as those on apoptotic cells or microbial pathogens like group B Streptococcus, promoting subsequent engulfment without triggering strong inflammatory responses. For instance, Siglec-1 enhances the and of sialylated particles in cooperation with other receptors like Fcγ receptors and TIM-4 on alveolar macrophages, thereby supporting efficient pathogen clearance in the lungs. In phagocytosis, specific Siglecs exhibit both facilitatory and inhibitory functions depending on the cellular context. On neutrophils, Siglec-5 and the paired activating receptor Siglec-14 interact with sialic acids on target cells, inhibiting activation (e.g., CD11b/CD18) and thereby suppressing neutrophil-mediated and of apoptotic or opsonized targets. This inhibitory mechanism limits excessive phagocytic activity, as demonstrated by enhanced upon blockade of Siglec-5/14 with sialidase treatment or antibodies, which significantly increases neutrophil-tumor cell conjugate formation in donor studies. Conversely, Siglec-E on murine macrophages and dendritic cells modulates apoptotic cell clearance by suppressing (ROS) production during engulfment, which prevents oxidative damage and promotes efficient while maintaining anti-inflammatory homeostasis; deficiency in Siglec-E leads to impaired ROS regulation and reduced clearance efficiency in models of neurodegeneration and . Counter-receptors such as and MUC1 serve as cis-ligands that modulate Siglec-mediated strength on the same cell surface, fine-tuning immune cell interactions. , a glycosylphosphatidylinositol-anchored protein rich in sialic acids, engages Siglec-10 in cis to dampen activation and enhance tolerance during , reducing pro-inflammatory release in response to damaged tissues. Similarly, MUC1, a with extensive O-linked sialylation, acts as a cis-ligand for Siglec-9 on myeloid cells, altering dynamics and promoting tumor-associated differentiation while inhibiting trans-interactions that could drive strong . Siglecs contribute to immune synapse formation by localizing to the contact interface between immune cells and targets, stabilizing and modulating activation thresholds. For example, inhibitory Siglecs like Siglec-G and (Siglec-2) are recruited to the B cell via sialylated ligands on antigen-presenting cells, inhibiting signaling and inducing BIM-dependent of self-reactive s, which is essential for as evidenced by reduced B cell depletion in double-knockout models. In natural killer cells, Siglec-7 similarly clusters at the with tumor targets, where cis-ligand masking limits activation, but trans-engagement with hypersialylated surfaces can suppress cytotoxic granule release, thereby regulating stability and preventing overactivation.

Insights from Knockout Studies

Knockout studies in mice have provided key insights into the roles of Siglecs, particularly in regulating immune and preventing pathological responses. In Siglec-1-deficient mice, macrophages exhibit reduced of sialylated pathogens, leading to impaired clearance of such as Campylobacter jejuni and group B Streptococcus, which highlights Siglec-1's function in facilitating uptake of sialylated microbes by myeloid cells. These mice show attenuated severity in experimental autoimmune encephalomyelitis (EAE), with increased regulatory T cells and reduced Th17 cells, suggesting Siglec-1 may promote pro-inflammatory responses in certain autoimmune contexts. CD22 knockout mice demonstrate hyperactive s with enhanced BCR signaling, characterized by increased calcium influx and proliferation in response to antigens, underscoring CD22's role as a negative regulator of activation. These mice develop elevated autoantibodies and lupus-like symptoms upon aging or in autoimmune-prone backgrounds, including high-affinity anti-DNA antibodies and immune complex-mediated , indicating CD22's essential function in maintaining tolerance and preventing systemic . Siglec-G-deficient mice, the murine ortholog of human Siglec-10, exhibit expanded populations with heightened responsiveness, elevated serum IgM levels, and increased B cells and plasma cells in aging animals, reflecting Siglec-G's inhibitory control over innate-like B cell expansion and . These changes contribute to enhanced formation and potential amplification of antibody-mediated responses, though single knockouts do not typically develop spontaneous on standard backgrounds. Studies of double knockouts reveal functional redundancy among CD33rSiglecs in B cell regulation; CD22/Siglec-G double-deficient mice show massively expanded , reduced B2 cells, hyperproliferative responses to TLR ligands, and overt systemic with anti-nuclear and anti-DNA , far exceeding phenotypes in single knockouts. This redundancy emphasizes the compensatory inhibitory roles of and Siglec-G in suppressing aberrant B cell activation and production across the CD33r subfamily.

Siglecs Across Mammals

Human and Primate Siglecs

express 14 functional Siglecs, which are primarily expressed on cells of the and a few other cell types, such as neurons and glial cells. These include Siglec-1 (also known as sialoadhesin or CD169), which is predominantly found on macrophages; Siglec-2 (), restricted to B cells; and Siglec-3 (), expressed on myeloid progenitors, monocytes, macrophages, dendritic cells, mast cells, and . Other notable examples are Siglec-7 and Siglec-9, both present on natural killer (NK) cells, monocytes, macrophages, neutrophils, and subsets of T cells; Siglec-8, specific to , mast cells, and ; Siglec-10, on B cells and monocytes; Siglec-11, on macrophages and ; Siglec-14, on neutrophils, monocytes, and macrophages; Siglec-15, primarily on osteoclasts and some macrophages; and Siglec-16, on macrophages. Additionally, Siglec-4 (myelin-associated glycoprotein or MAG) is expressed on and Schwann cells in the , while Siglec-5 is found on neutrophils and monocytes, and Siglec-6 on placental trophoblasts and B cells. The Siglec family in , particularly great apes, exhibits rapid , with notable expansions and variations in the CD33-related (CD33r) subgroup, which includes Siglec-3, -5 through -11, and -14 through -16. This expansion is thought to reflect pathogen-driven selective pressures, leading to increased gene diversity in great apes compared to humans. A key human-specific change is the complete loss of Siglec-13 through an Alu-mediated deletion, rendering it absent in modern humans but functional in chimpanzees and baboons, where it is expressed on monocytes and epithelial cells. Furthermore, Siglec-12 exists as a in humans due to a abolishing binding, a feature fixed in both modern and archaic humans but retained in great apes. Siglec-16 is primate-specific, while Siglec-11 shows human-specific upregulation in brain . These variations highlight the dynamic of Siglecs in , potentially influencing immune recognition and self-tolerance. Human Siglecs display preferences for specific linkages, primarily α2-3, α2-6, or α2-8, and vary in the number of extracellular immunoglobulin-like (Ig) domains, which contribute to ligand avidity. The following table summarizes these features for the 14 functional human Siglecs:
SiglecExtracellular Ig DomainsPreferred Sialic Acid Linkages
Siglec-117α2-3
Siglec-27α2-6
Siglec-32α2-3, α2-6
Siglec-45α2-3
Siglec-54α2-3, α2-6
Siglec-64α2-6, α2-3 (with )
Siglec-73α2-8, branched α2-6
Siglec-83α2-3 (with 6'-sulfate)
Siglec-93α2-3, α2-6, α2-8
Siglec-105α2-3, α2-6 (prefers α2-6)
Siglec-114α2-8
Siglec-143α2-3, α2-6 (similar to Siglec-5)
Siglec-152α2-3
Siglec-163α2-8 (similar to Siglec-11)
These binding preferences influence cellular interactions, with most Siglecs featuring an N-terminal V-set domain for sialic acid recognition.

Siglecs in Non-Primate Mammals

In non-primate mammals, the Siglec family exhibits notable diversity, particularly through expansions in the CD33-related (CD33r) subgroup, which contrasts with the more conserved repertoire in . Rodents, such as mice, possess nine Siglec genes in total, including five CD33r members: (CD33), Siglec-E, Siglec-F, Siglec-G, and Siglec-H. These expansions, with Siglec-E, -F, -G, and -H being unique to , likely arose from events that enhanced immune regulation in these species. This larger repertoire allows to fine-tune sialic acid-mediated interactions in innate immunity, making them valuable model organisms for studying Siglec functions despite differences from human orthologs. A prominent example of functional divergence is mouse Siglec-F, which serves as an inhibitory receptor on eosinophils, promoting their apoptosis upon ligand engagement and thereby dampening allergic inflammation. This parallels human Siglec-8, its closest functional analog, but Siglec-F lacks the exact sequence motifs for certain antibody-induced signaling pathways observed in humans, highlighting species-specific adaptations in eosinophil regulation. Similarly, mouse Siglec-E modulates macrophage and neutrophil responses to bacterial infections, suppressing excessive inflammation, while Siglec-G on B cells influences self-nonself discrimination in adaptive immunity. These rodent-specific Siglecs underscore the role of evolutionary expansions in tailoring immune homeostasis to environmental pressures like pathogens. In species, Siglec expression supports surveillance and immune modulation, with implications for veterinary health. In , Siglec-1 (sialoadhesin) is expressed on macrophages and dendritic cells, where it facilitates recognition and uptake of sialylated pathogens, such as viruses, enhancing in immunity. Bovine dendritic cells preferentially utilize Siglec-1 over Siglec-10 for binding enveloped viruses like , illustrating its role in bridging innate recognition to adaptive responses. Porcine CD33r Siglecs, including Siglec-3, Siglec-5, and Siglec-10, are expressed on myeloid cells and contribute to sialic acid-dependent interactions that influence viral tropism and immune evasion. These receptors can bind sialylated viral envelopes, potentially increasing host cell susceptibility to infections like porcine reproductive and respiratory virus (PRRSV), where engagement modulates activation and viral entry. In pigs, variations in CD33r Siglec expression may underlie breed-specific differences in outcomes, informing strategies for health management. The diversity of Siglecs in non-primate mammals highlights their adaptability in veterinary , where models elucidate basic mechanisms, and studies reveal applications in disease resistance breeding and design. For instance, targeting Siglec-F in mice has informed eosinophil-targeted therapies, while bovine and porcine Siglecs offer insights into zoonotic pathogen control. This comparative perspective emphasizes Siglecs' conserved yet expanded roles across mammals, beyond primate-centric views.

Therapeutic Potential

Siglec Mimetics and Synthesis

mimetics (SAMs) are synthetic analogs of (Neu5Ac), the primary in mammalian glycans, engineered to bind Siglecs with greater potency and specificity than natural ligands. Key modifications target the side chain at the C6 position of Neu5Ac, often replacing it with aromatic groups such as benzyl or benzoyl moieties to enhance hydrophobic interactions within the Siglec V-set domain. These alterations exploit structural insights from Siglec-ligand complexes to optimize binding while maintaining the core scaffold responsible for recognition. The production of SAMs relies on two principal synthesis strategies: chemoenzymatic and . Chemoenzymatic approaches leverage sialyltransferases to incorporate modified onto acceptors, combining of the sialic acid analog with enzymatic for scalable assembly of multivalent ligands. involves iterative protection-deprotection cycles in chemistry to build the ring and substituents from non-carbohydrate starting materials, enabling precise control over and substitution patterns. Notable examples of SAMs include GT1b-lactone derivatives tailored for Siglec-2 (), which mimic polysialylated motifs to achieve selective B-cell targeting, and benzoyl-modified sialic acids optimized for Siglec-7 engagement on natural killer cells. These compounds demonstrate binding enhancements of up to 1000-fold over unmodified Neu5Ac, as measured by and cell-based assays, primarily due to stabilized interactions at the C9 position and multivalent presentation.

Clinical Applications and Targeting

Siglecs have emerged as promising therapeutic targets in various diseases due to their roles in modulating immune responses, with (Siglec-3) being the most clinically advanced example. In (AML), is overexpressed on leukemic blasts, making it a key target for antibody-drug conjugates. (GO), a -targeted immunoconjugate approved by the FDA in 2017 for CD33-positive AML, delivers the cytotoxic agent to CD33-expressing cells, leading to improved overall survival when combined with standard in newly diagnosed patients. Siglec-15 has been implicated in promoting , particularly in , where its elevated expression in the metastatic niche enhances tumor-induced osteoclastogenesis and suppresses T-cell activity. A 2024 study demonstrated that blocking Siglec-15 with a reduced osteoclast formation and tumor burden in preclinical models of , highlighting its potential as a glyco-immune checkpoint. Recent preclinical as of 2025 has explored Siglec-15 –GM-CSF chimeras, which suppress tumor progression in models of Siglec-15-overexpressing cancers by enhancing immune activation. Therapeutic strategies targeting Siglecs include blocking antibodies to inhibit immunosuppressive functions in cancer and agonists to dampen hyperinflammation in . For Siglec-3 in cancer, non-conjugate blocking antibodies have shown preclinical efficacy in disrupting CD33-mediated inhibition of immune cells, enhancing antitumor responses when combined with checkpoint inhibitors. In , Siglec-9 agonists, such as synthetic derivatives, inhibit activation and NETosis, reducing inflammatory damage in models of hyperinflammation akin to bacterial . Challenges in Siglec targeting arise from functional among family members and cis-inhibition, where Siglecs bind endogenous sialic acids on the same cell, limiting accessibility for therapeutics. To overcome these, bispecific antibodies that simultaneously engage multiple Siglecs or pair Siglec with immune activators have demonstrated superior in preclinical cancer models by bypassing redundancy and enhancing immune engagement. Emerging applications focus on Siglec-8 for disorders, where antagonists like lirentelimab () induce and reduce tissue infiltration.

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