Galectins are among the most widely expressed class of lectins in all organisms. They typically bind β-galactose-containing glycoconjugates and share primary structural homology in their carbohydrate-recognition domains (CRDs). Galectins have many biological functions, including roles in development, regulation of immune cell activities, and microbial recognition as part of the innate immune system. This chapter describes the diversity of the galectin family and presents an overview of what is known about their biosynthesis, secretion, and biological roles.

HISTORICAL BACKGROUND ON DISCOVERY OF GALECTINS

Following the discovery of the Ashwell–Morell asialoglycoprotein receptor in the liver, many investigators sought other such receptors by affinity chromatography with immobilized asialoglycoproteins. In 1975, a novel lectin of ∼15 kDa was isolated from electric organs of the electric eel, and named “electrolectin.” This noncovalently linked homodimer displayed hemagglutinating activity toward trypsinized rabbit erythrocytes that was inhibitable by β-galactosides; inclusion of β-mercaptoethanol in the isolation buffers was required to maintain the binding activity. Soon thereafter, in 1976, similar β-galactoside-binding lectins were isolated from chicken muscle and from extracts of calf heart and lung (∼15 kDa; now designated as galectin-1). These proteins were initially referred to as S-type lectins to denote their sulfhydryl dependency, presence of free cysteine residues, and solubility, along with shared primary sequence. Interestingly, and unlike most galectins, electrolectin does not contain free Cys residues, but a key Trp in the binding site of its CRD can be oxidized, causing loss of activity. In the early 1980s, a 35 kDa protein termed CBP35 that also bound to β-galactosides was identified in mouse fibroblasts. The same protein was studied by other groups under the names IgE-binding protein, L-29, and L-31 and is currently known as galectin-3.

The nomenclature for galectins was systematized in 1994 and the first galectin type found (∼15 kDa, discussed above) was renamed galectin-1. All other members of this family were numbered consecutively by order of discovery. Multiple distinct galectins are usually expressed in a single species (up to 15 in the mouse). A list of the vertebrate galectins is shown in Figure 36.1A.

FIGURE 36.1.

Different types of galectins in vertebrates and invertebrates and their organization and sequences. (A) Fifteen types of vertebrate galectins are known. (B) Examples of sequence alignments of some representative human galectins are shown. The amino acid (more...)

THE GALECTIN FAMILY

In 1982, a shared amino acid sequence motif was identified in the CRD of all galectins. The canonical CRD of galectins has ∼130 amino acids, although only a small number of residues within the CRD directly contact glycan ligands. A comparison of more than 100 galectin sequences from many different sources reveals that eight residues, which have been shown to be involved in glycan binding by X-ray crystallographic analyses, are mostly invariant. In addition, another dozen residues appear to be highly conserved. Part of the conserved galectin sequence motif is shown in Figure 36.1b, along with a comparison of several human galectins.

Based on their CRD number and organization, galectins have been classified into three major groups, termed prototypical galectins (galectin-1, -2, -7, -10, -11, -13, -14, and -15) that contain a single CRD that may associate as homodimers, chimera-type galectins (galectin-3), which are characterized by having a single CRD and an amino-terminal polypeptide “tail” rich in proline, glycine, and tyrosine residues, through which it can form oligmers, and tandem-repeat galectins (galectin-4, -8, -9, and -12, which comprise proteins that carry two CRDs connected by a peptide linker that can range from five to >50 amino acids in length. Galectin transcripts may be differentially spliced to generate multiple isoforms. Invertebrates have galectins with one, two, or four tandemly arrayed CRDs.

Some galectins appear to be species specific, such as galectin-5 (prototype) and galectin-6 (tandem repeat) that are found in rodents but not humans, and galectin-11 (ovagal11; prototype) that has only been reported in sheep, and galectin-15 has only been found in sheep and goat. The amino-terminal domain of galectin-12 has significant homologies with other galectins, whereas the carboxy-terminal domain displays greater divergence from other galectins.

Several proteins with sequence homology with galectins, which do not bind typical β-galactosides, have been described as galectin-related proteins (GRPs). For example, galectin-10, also known as the Charcot–Leyden crystal protein, is highly abundant and is present in eosinophil granules and preferentially binds to β-mannosides. Several GRPs, including GRIFIN (galectin-related interfiber protein), PP-13, and PPL-13 that lack glycan-binding activity, are related to galectin-10. It is noteworthy that the GRIFIN homolog in the zebrafish (Danio rerio DrGRIFIN) is a functional β-galactoside-binding protein and, like its mammalian counterpart, is also highly expressed in the eye lens.

TAXONOMIC DISTRIBUTION AND EVOLUTION OF GALECTINS

Hallmarks of the galectin family are the extensive taxonomic distribution of its members, together with the evolutionary conservation of their primary structures, gene organization, and structure of the CRD. The early emergence and structural conservation of the galectin family in eukaryotic evolution is revealed in the fungus Coprinopsis cinerea in the sponge Geodia cydonium, and in a protein that displays the galectin fold in the protozoan parasite Toxoplasma gondii. Galectin-like sequences have also been identified in plants (Arabidopsis thaliana) and some viruses that infect pigs and fish, such as the porcine adenovirus and lymphocystis disease virus. However, galectin-like proteins or sequence motifs in the latter should be interpreted with caution as it could result from horizontal gene transfer.

The evolution of the galectin family has been inferred from the rigorous analysis of primary structures, gene organization, and taxonomic distribution of family members identified in a variety of organisms, including fungi, invertebrates, and vertebrates. Among the arthropods and nematodes, galectins are abundantly present in Drosophila melanogaster (six candidate genes) and Caenorhabditis elegans (26 candidate genes), respectively. At least 14 of these candidate galectin genes in C. elegans produce transcripts encoding proteins. Generally, the primary structures and domain organizations of galectins from invertebrates do not fit the canonical features of vertebrate galectins described above (Figure 36.1C).

Galectins are ubiquitous in vertebrates. In fish, galectins have been identified and characterized in numerous species. The zebrafish (D. rerio) genome has ortholog genes for all three galectin types (proto-, chimera-, and tandem-repeat) identified in mammals. Among amphibians, several galectin subtypes have been found in various tissues, mucus, and eggs from salamanders (the axolotl Ambystoma mexicanum), toads (Bufo arenarum), and frogs (Rana catesbeiana and Xenopus laevis).

The evolution of galectins along the vertebrate taxa leading to the mammals has been rationalized as the duplication of an early single-CRD galectin gene that would have led to a bi-CRD galectin gene, in which the amino- and carboxy-terminal CRDs subsequently diverged into two different subtypes, defined by an exon–intron structure (F4-CRD and F3-CRD) (F refers to the β-sandwich strand structure discussed below). All vertebrate single-CRD galectins belong to either the F3- (e.g., galectin-1, -2, -3, -5) or F4- (e.g., galectin-7, -10, -13, -14) subtype, whereas the mammalian tandem-repeat galectins such as galectin-4, -6, -8, -9, and -12 contain both F4 and F3 subtypes. However, how the multiple CRD galectins from invertebrates relate to the vertebrate tandem-repeat galectins remains to be fully understood, but a preliminary phylogenetic analysis of the oyster galectin CvGal1 revealed that the individual CRDs cluster with the mammalian single-CRD galectins rather than with the tandem-repeat galectins, suggesting that the CvGal1 gene is the product of two consecutive gene duplications of a single-CRD galectin gene ancestral to the early invertebrate taxa that was conserved along the chordate lineages. The striking structural and evolutionary conservation of the CRD architecture and arrangements supports the key roles of galectins in essential biological functions related to β-galactoside recognition.

STRUCTURE OF GALECTINS

The crystal structures of several galectins in complexes with glycan ligands are known, including galectins-1, -2, and -7, the carboxy-terminal domain of galectin-3, and individual domains of galectin-4, -8, and -9. In all cases, the galectin CRD is comprised of five- and six-stranded antiparallel β sheets arranged in a β-sandwich or jelly-roll configuration that lacks an α-helix (Figure 36.2A). In the dimeric proteins, such as galectin-1, -2, and -7, the subunits are related by a twofold rotational axis perpendicular to the plane of the β sheets. The glycan-binding sites in the CRD are located at opposite ends of the dimer, although the orientation of the subunits in the galectin-7 dimer is different from the other canonical galectin dimers. The compactly arranged structure of the CRD partly explains the protease resistance of the galectin CRD and the high degree of conservation and requirement for the 130 amino acids in the CRD.

FIGURE 36.2.

(A) Ribbon diagram of the crystal structure of human galectin-1 complexed with lactose. The homodimer is shown with each monomer colored differently and orthogonal views are presented. The subunit interface is based on interactions between the carboxy- (more...)

The galectin-1 CRD displays highly specific interactions with Gal and GlcNAc residues in typical glycans. Interactions with glycans are generally through hydrogen bonding, electrostatic interactions, and van der Waals interactions through ring stacking with Gal and the highly conserved Trp residue (Figures 36.2B,C). In general, the open-ended structure of the glycan-binding site is predicted to allow access to extended Gal-containing glycans, such as the poly-N-acetyllactosamines and blood group–related structures (Chapter 14).

The binding preference of invertebrate galectins for blood group glycans can be illustrated by a unique structural feature of the binding site of the C. elegans and oyster galectins. They show a significantly shorter loop 4 that accommodates the 2´-fucosyl moiety that is a common feature of both the A and B blood groups (Figures 36.3A,B). The α(1-3)-linked GalNAc or Gal moieties of the A and B blood group tetrasaccharides, respectively, are recognized by the -NH at position 5 of the conserved Trp, and a hydrophobic pocket at the external side of the β3-strand (amino acids 30–38) recognizes the methyl of the -NAc group in α(1-3)GalNAc of the A1/2 antigens.

FIGURE 36.3.

Structural aspects of galectins from mammals and invertebrates. (A) Sequence alignment of the carbohydrate-recognition domain (CRD) regions from bovine galectin-1, zebrafish Drgal1-L2, C. elegans N16, and the CRDs 1–4 of CvGal1. (B) Homology modeling (more...)

There are several known and predicted subsites on galectins near the carbohydrate-binding site that could serve to enhance affinity for more extended glycans. Some of these subsites have been identified by cocrystal structural analysis with glycans and by glycan-binding studies. The crystal structure of bovine galectin-1 was derived in a complex with a biantennary N-glycan containing two terminal β-Gal residues. The N-glycan is bridged between two galectin-1 dimers, thus effectively creating a crystal latticework. This type of crystal latticework may be unique among vertebrate galectins and may be critical for their signaling and adhesive functions (Figures 36.4 and 36.5).

FIGURE 36.4.

Possible biosynthetic routes for galectins in animal cells, using galectin-1 as an example. The mRNA for the protein is translated on free polysomes in the cytoplasm and the newly synthesized protein there is capable of binding glycan ligands or interacting (more...)

FIGURE 36.5.

Functional interactions of galectins with cell-surface glycoconjugates and to extracellular glycoconjugates leads to cell adhesion and cell signaling. Interactions of galectins with intracellular ligands may contribute to regulating intracellular pathways. (more...)

GLYCAN LIGANDS FOR GALECTINS

The interactions of galectins with glycans are complex and several factors contribute to high-affinity binding, including their natural multivalency and oligomeric state, as well as the multivalency of their natural ligands. Although most galectins bind weakly to simple β-galactosides, such as disaccharides or trisaccharides (K_d in the high micromolar to low millimolar range), they typically bind much stronger to natural glycoconjugate ligands (apparent K_d in the submicromolar range). Studies utilizing large libraries of glycans reveal that each galectin CRD recognizes different glycan ligands and shows highest affinity binding to different structures. For example, galectin-3 binds tightly to glycans with repeating [-3Galβ1-4GlcNAcβ-]_n or poly-N-acetyllactosamine sequences containing 3-4 repeating units, regardless of the presence of terminal β-Gal residue, and binding is further enhanced if the penultimate β-Gal residues are substituted with Galα1-3, GalNAcα1-3, or Fucα1-2 residues. In contrast, human galectin-1 binds well only to glycans with terminal β-Gal residues, and does not bind blood group antigens. Galectin-8 has two CRDs within its single polypeptide and the two CRDs bind different glycans. For example, the amino-terminal CRD of human galectin-8 binds α2-3-sialylated glycans with high affinity (K_d ∼ 50 nm), whereas the carboxy-terminal CRD binds to the blood group A determinant on a LacNAc core, and does not bind sialylated glycans.

Cocrystallization of galectins with simple β-Gal-containing disaccharides has revealed that many galectins have H-bonds to the C4 and C6 hydroxyl of Gal and the C3 hydroxyl of GlcNAc (Figure 36.3). The binding sites of galectins may be viewed as containing several subsites: one for Gal, another for GlcNAc, and other subsites that may be filled by other sugars and the aglycone moiety, e.g., protein or lipid. Thus, other sugar residues linked to the disaccharide Gal-GlcNAc can enhance or decrease binding affinity, depending on the galectin. Some galectins from fungi, invertebrates, and mammals (such as galectin-4 and -8) preferentially bind selected ABO(H) blood group glycans as discussed above.

Galectins bind selectively to only some cell-surface and extracellular matrix ligands at low lectin concentrations but at higher concentrations they more often show broad binding to many different glycoproteins. However, the precise physiological ligands for each galectin and the physiological concentrations of galectins at sites of interactions are not well understood.

Galectins can also bind to exogenous ligands, such as exposed glycans on the surface of viruses, bacteria, fungi, and parasites, displaying a remarkable diversity of recognition. For example, galectin-1 can bind to complex type N-glycans on the HIV-1 gp120 envelope glycoprotein and to Trichomonas vaginalis lipophosphoglycan, whereas galectin-3 recognizes both terminal and internal N-acetyllactosamine units in lipopolysaccharides from meningococcus (Neisseria meningitidis), LacdiNAc from Schistosoma mansoni, and oligomannans from Candida albicans. The lipophosphoglycan from Leishmania major is recognized by both galectin-3 and -9, whereas galectin-4 recognizes Escherichia coli strains that display blood group B glycans.

BIOSYNTHESIS AND EXPORT OF GALECTINS

All members of the galectin family lack a classical signal sequence or membrane-anchoring domains, and are synthesized on free polysomes in the cytoplasm and accumulate there before secretion. Galectins are unusual among all types of animal lectins in that they can be found in the nucleus, cytosol, outer plasma membrane, and extracellular matrix. One rather common modification of galectins in animal cells is blockage of the amino terminus, although galectin-3 has been shown to also have two serine residues in its unique amino-terminal region that can be phosphorylated. Newly synthesized galectins isolated directly from the cytosol of cells are functional in binding β-galactosides.

Interestingly, there is compelling evidence for galectins having non-glycan-binding partners in the cytosol. The complexity of galectin biosynthesis, secretion, and oligomerization is illustrated schematically for galectin-1 in Figure 36.4. Curiously, the export of galectins from cells does not involve direct movement through the secretory pathway. It appears the secretion and availability of glycan ligands in the cells generating the galectin may also regulate export, activity, and stability of galectins.

BIOLOGICAL ROLES OF GALECTINS

Members of the galectin family display a striking functional diversification that ranges from key roles in early development, tissue regeneration, and cancer, to immune homeostasis and recognition/effector functions against potential pathogens. By binding to endogenous glycans, galectins can contribute to cell–cell and cell–matrix interactions, and galectin signaling at the cell surface can also modulate cellular functions. In addition, intracellular galectins may, as in the case of galectins-3, regulate cellular activities and may contribute to some fundamental processes such as pre-mRNA splicing (Figure 36.5). The galectin-mediated recognition of glycans on the surface of potential pathogens and parasites discovered in the past few years has revealed their role(s) as pattern recognition receptors and effector factors in innate immune functions. Much information about the biological functions of galectins has been revealed largely through using galectin-deficient mouse models. Such studies, which have been performed primarily in mice deficient in galectin-1, -3, -7, and -9, have revealed roles of galectins in a wide range of physiological pathways (Chapter 41).

FURTHER READING

• Drickamer K. 1988. Two distinct classes of carbohydrate-recognition domains in animal lectins. J Biol Chem 263: 9557–9560. [PubMed: 3290208]
• Barondes SH, Castronovo V, Cooper DN, Cummings RD, Drickamer K, Feizi T, Gitt MA, Hirabayashi J, Hughes C, Kasai K, et al. 1994. Galectins: A family of animal β-galactoside-binding lectins. Cell 76: 597–598. [PubMed: 8124704]
• Houzelstein D, Gonçalves IR, Fadden AJ, Sidhu SS, Cooper DNW, Drickamer K, Leffler H, Poirier F. 2004. Phylogenetic analysis of the vertebrate galectin family. Mol Biol Evol 21: 1177–1187. [PubMed: 14963092]
• Liu FT, Rabinovich GA. 2005. Galectins as modulators of tumour progression. Nat Rev Cancer 5: 29–41. [PubMed: 15630413]
• van Die I, Cummings RD. 2006. Glycans modulate immune responses in helminth infections and allergy. Chem Immunol Allergy 90: 91–112. [PubMed: 16210905]
• Elola MT, Wolfenstein-Todel C, Troncoso MF, Vasta GR, Rabinovich GA. 2007. Galectins: Matricellular glycan-binding proteins linking cell adhesion, migration, and survival. Cell Mol Life Sci 64: 1679–1700. [PubMed: 17497244]
• Mercier S, St-Pierre C, Pelletier I, Ouellet M, Tremblay MJ, Sato S. 2008. Galectin-1 promotes HIV-1 infectivity in macrophages through stabilization of viral adsorption. Virology 371: 121–129. [PubMed: 18028978]
• Stowell SR, Cummings RD. 2008. Interactions of galectins with leukocytes. In Animal lectins: A functional view (ed. Vasta GR, Ahmed H.). CRC Press, Boca Raton, FL.
• Vasta GR. 2009. Roles of galectins in infection. Nat Rev Microbiol 7: 424–438. [PMC free article: PMC3759161] [PubMed: 19444247]
• Dam TK, Brewer CF. 2010. Maintenance of cell surface glycan density by lectin–glycan interactions: A homeostatic and innate immune regulatory mechanism. Glycobiology 20: 1061–1064. [PubMed: 20548106]
• Stowell SR, Arthur CM, Dias-Baruffi M, Rodrigues LC, Gourdine JP, Heimburg-Molinaro J, Ju T, Molinaro RJ, Rivera-Marrero C, Xia B, et al. 2010. Innate immune lectins kill bacteria expressing blood group antigen. Nat Med 16: 295–301. [PMC free article: PMC2853181] [PubMed: 20154696]
• DiLella S, Sundblad V, Cerliani JP, Guardia CM, Estrin DA, Vasta GR, Rabinovich GA. 2011. When galectins recognize glycans: From biochemistry to physiology and back again. Biochemistry 50: 7842–7857. [PMC free article: PMC3429939] [PubMed: 21848324]
• Liu FT, Yang RY, Hsu DK. 2012. Galectins in acute and chronic inflammation. Ann NY Acad Sci 1253: 80–91. [PubMed: 22329844]
• Rabinovich GA, van Kooyk Y, Cobb BA. 2012. Glycobiology of immune responses. Ann NY Acad Sci 1253: 1–15. [PMC free article: PMC3884643] [PubMed: 22524422]
• Thurston TL, Wandel MP, vonMuhlinen N, Foeglein A, Randow F. 2012. Galectin 8 targets damaged vesicles for autophagy to defend cells against bacterial invasion. Nature 482: 414–418. [PMC free article: PMC3343631] [PubMed: 22246324]
• Vasta GR, Ahmed H, Nita-Lazar M, Banerjee A, Pasek M, Shridhar S, Guha P, Fernández-Robledo JA. 2012. Galectins as self/non-self recognition receptors in innate and adaptive immunity: An unresolved paradox. Front Immunol 3: 1–14. [PMC free article: PMC3396283] [PubMed: 22811679]
• Yang RY, Havel PJ, Liu FT. 2012. Galectin-12: A protein associated with lipid droplets that regulates lipid metabolism and energy balance. Adipocyte 1: 96–100. [PMC free article: PMC3609087] [PubMed: 23700518]
• Chen HY, Weng IC, Hong MH, Liu FT. 2014. Galectins as bacterial sensors in the host innate response. Curr Opin Microbiol 17: 75–81. [PubMed: 24581696]
• Croci DO, Cerliani JP, Dalotto-Moreno T, Méndez-Huergo SP, Mascanfroni ID, Dergan-Dylon S, Toscano MA, Caramelo JJ, García-Vallejo JJ, Ouyang J, et al. 2014. Glycosylation-dependent lectin–receptor interactions preserve angiogenesis in anti-VEGF refractory tumors. Cell 156: 744–758. [PubMed: 24529377]
• Toledo KA, Fermino ML, AndradeCdel C, Riul TB, Alves RT, Muller VD, Russo RR, Stowell SR, Cummings RD, Aquino VH, et al. 2014. Galectin-1 exerts inhibitory effects during DENV-1 infection. PLoS One 9: e112474. [PMC free article: PMC4231055] [PubMed: 25392933]
• Vladoiu MC, Labrie M, St-Pierre Y. 2014. Intracellular galectins in cancer cells: Potential new targets for therapy. Int J Oncol 44: 1001–1014. [PubMed: 24452506]
• Arthur CM, Baruffi MD, Cummings RD, Stowell SR. 2015. Evolving mechanistic insights into galectin functions. Methods Mol Biol 1207: 1–35. [PMC free article: PMC5755386] [PubMed: 25253130]
• Blidner AG, Méndez-Huergo SP, Cagnoni AJ, Rabinovich GA. 2015. Re-wiring regulatory cell networks in immunity by galectin-glycan interactions. FEBS Lett 589: 3407–3418. [PubMed: 26352298]
• Than NG, Romero R, Balogh A, Karpati E, Mastrolia SA, Staretz-Chacham O, Hahn S, Erez O, Papp Z, Kim CJ. 2015. Galectins: Double-edged swords in the cross-roads of pregnancy complications and female reproductive tract inflammation and neoplasia. J Pathol Transl Med 49: 181–208. [PMC free article: PMC4440931] [PubMed: 26018511]
• Rabinovich GA, Conejo-Garcia JR. 2017. Shaping the immune landscape in cancer by galectin-driven regulatory pathways. J Mol Biol 428: 3266–3281. [PubMed: 27038510]
• Salvagno GL, Pavan C. 2017. Prognostic biomarkers in acute coronary syndrome. Ann Transl Med 4: 258. [PMC free article: PMC4958728] [PubMed: 27500159]