Encyclopedia of Signaling Molecules

2018 Edition
| Editors: Sangdun Choi

Galectin-9

Reference work entry
DOI: https://doi.org/10.1007/978-3-319-67199-4_101652

Synonyms

Historical Background

Galectin-9 (gal-9) is a member of the galectin protein family which consists of glycan-binding proteins that share a conserved carbohydrate recognition domain (CRD) and a binding affinity for β-galactoside sugars. Initially, galectins were designated as S-type lectins since it was found that the first discovered galectin, now referred to as galectin-1, depended on thiols (reducing conditions) to be active. Since this was not the case for many other galectins that were discovered later, the designation S-type lectin is now considered outdated. The galectin family name was defined in 1994 and consisted at that time of four members, i.e., galectin-1 to −4 (Barondes et al. 1994; Leffler et al. 2004). Galectin-9 was added to the protein family in 1997 when it was described by three independent groups as (i) a novel β-galactoside binding lectin from murine embryonic kidney (Wada and Kanwar 1997), (ii) a tumor antigen in Hodgkins disease (Tureci et al. 1997), and (iii) an urate transporter/channel (Leal-Pinto et al. 1997). Later, it was found that Ecalectin, an eosinophil chemoattractant that was described in 1998, was in fact also gal-9 (Matsushita et al. 2000). These early observations in different research fields are illustrative of the diverse functional activities of gal-9 that have been identified up to now.

Galectin-9 Gene Expression and Protein Isoforms

Galectin-9 is a tandem repeat galectin, i.e., it consists of two distinct and evolutionary conserved CRDs that are connected by a linker peptide. The protein is encoded by the LGALS9 gene which is located on chromosome 17q11.2. In close vicinity, two LGALS9-like genes can be found, i.e., LGALS9B and LGALS9C (Fig. 1a, b). While transcripts of these two genes have been identified, it is unclear whether this results in galectin-9-like proteins with distinct biological functions (Heusschen et al. 2013). The LGALS9 gene comprises of 11 exons which encode a protein of 355 amino acids. The primary transcript is subjected to extensive splicing involving exons 5, 6, and 10. Splicing of exons 5 and 6 affects the length of the linker peptide which results in three gal-9 protein isoforms, i.e., gal-9 L (all exons retained), gal-9 M (exon 5 spliced), and gal-9S (exons 5/6 spliced). Additional transcripts have been identified in which also exon 10 has been spliced. This does not affect the linker peptide but causes a frameshift and premature stop in the coding sequence of the C-terminal CRD (Fig. 1c). Whether stable and functionally active truncated gal-9 isoforms are actually transcribed in vivo is still unresolved. Nevertheless, different cell-type specific galectin-9 expression patterns that include exon 10 splice variants have been identified which is indicative of a specific role of each variant in cell biology (Spitzenberger et al. 2001; Heusschen et al. 2014).
Galectin-9, Fig. 1

(a) Location of galectin-9 (LGALS9) and galectin-9-like (LGALS9B and LGALS9C) genes on chromosome 17. (b) Homology of LGALS9, LGALS9B and LGALS9C proteins. Grey boxes depict differences with LGALS9. Sequences encoding the CRDs and linker region have green and blue bars above them respectively. (c) Galectin-9 (LGALS9) gene structure and confirmed splice variants. Boxes represent exons (white: untranslated region; green: CRD coding region; blue: Linker coding region; orange: frameshift in the coding region). So far, 6 splice variants have been identified that vary in the exclusion of exons 5, 6 and 10. As a result, linker length and the C-terminal CRD varies between splice variants. Gal-9FL: galectin-9 full length. (d) Protein localization of galectin-9. Galectin-9 can be found both intra- and extracellularly. Extracellularly galectin-9 can multimerize and is involved in adhesion of cells to the extracellular matrix and hetero- and homotypic cell adhesion via binding to varying carbohydrate epitopes (colored red, yellow and clear blue). In addition, galectin-9 can likely regulate receptor lattice formation and domain organization on the cell membrane. Galectin-9 is also found intracellularly, although its mode of action there is not clear yet (Reproduced with permission from Heusschen et al. 2013)

The regulation of gal-9 gene expression is still poorly understood. Several cytokines and Toll-like receptor ligands have been linked to the induction of gal-9 expression, including IFN-γ, IFN-β TNF-α, IL-1α, IL-1β, poly(I:C), LPS, and viral RNA. Many of the regulatory cytokines have also been associated with the expression of other galectins. Regarding the induction by IFN-γ, this appears to involve HDAC3 and IRF3 signaling (Gieseke et al. 2013; Thijssen et al. 2013). Of note, gal-9 expression is negatively regulated by miR-22 which targets the 3′ UTR of the galectin-9 transcript (Yang et al. 2015).

The Galectin-9 CRDs and Glycan Binding

Like most other galectin, the main functional activities of gal-9 are mediated through glycan binding. Since gal-9 consists of two CRDs, the gal-9 protein can bind two glycans. The binding valency can be affected by gal-9 dimerization – and possibly multimerization – through direct protein-protein interactions of the CRDs (Nagae et al. 2006; Miyanishi et al. 2007). The crystal structure of both gal-9 CRDs has been resolved (Nagae et al. 2006; Yoshida et al. 2010). In agreement with other galectins, each CRD is composed of a sandwich of two β-sheets composed of six (S1–S6) and five (F1–F5) antiparallel strands. The β-sheet sandwich is slightly bent thereby creating a groove for carbohydrate binding. Both CRDs also contain the conserved amino acids that are required for carbohydrate binding. There are also small structural differences between the CRDs which underlie the differences in glycan-binding specificity (Yoshida et al. 2010).

With regard to glycan binding, the galectin-9 CRDs can bind type I and type II N-acetyllactosamine (LacNAc) disaccharides. Binding affinity to complex N-glycans increases with increased branching. Increasing the number of repeats in poly-N-lactosamine also enhances binding affinity, especially of the N-terminal CRD. The N-terminal CRD also exhibits specific binding affinity for two glycolipid-type glycans, i.e., Forssman pentasaccharide and blood group A-hexasaccharide (Hirabayashi et al. 2002; Horlacher et al. 2010).

Galectin-9 in Physiology

Similar to other galectins, gal-9 protein can be found in the nucleus, in the cytoplasm, at the membrane, and in the extracellular environment (Fig. 1d). The protein lacks a secretion signal and the mechanism of gal-9 secretion is still unresolved. The exact functions of gal-9 in physiological processes are still poorly understood which is partly due to the complexicity of gal-9 protein expression. The initial studies that described galectin-9 have reported detectable expression in various tissues, most notably in tissues of endodermal origin and tissues related to the immune system. Gestational age-dependent regulation of gal-9 expression suggests a role in the development of different tissues (Wada et al. 1997). However, loss of gal-9 is not lethal and LGALS9 −/− mice are viable, similar as described for other galectins. The LGALS9 −/− mice do show increased levels of circulating neutrophils and lymphocytes as well as hyperproliferative B cells in germinal centers in splenic lymph nodes. Overall, larger cell numbers are observed in lymphoid organs (Orr et al. 2013). Together with the observed tissue expression, this supports the notion that galectin-9 is involved in shaping the immune response. Indeed, the initially identification of gal-9 as an eosinophil chemoattractant has linked the protein to the innate immune response. Additional studies have shown that gal-9 is also involved in the adaptive immune response by regulating T cell homeostasis and intracellular T cell signaling (for review see Wiersma et al. 2013). This activity has been linked to the interaction of gal-9 with T cell immunoglobulin mucin 3 (Tim-3) but also other gal-9 binding partners appear to be involved. For example, the interaction of gal-9 with protein disulfide isomerase can promote Th2 cell migration (Bi et al. 2011). The current view is that gal-9 has dual immunoregulatory activities, i.e., it stimulates innate immunity and suppresses adaptive immunity.

Additional functions/effects of gal-9 in/on (immune) cells involve regulation of cell survival, cell cycle control, cell adhesion, and cell differentiation (Hirashima et al. 2004; Wiersma et al. 2011; Heusschen et al. 2013). Moreover, gal-9 appears to be involved in the organization of different proteins in lipid rafts, as also described for other galectins. Finally, gal-9 was identified as a sugar-regulated urate transporter (Lipkowitz et al. 2004). In line with this functional diversity, multiple gal-9-binding glycoconjugates have been identified and this list continues to be expanded (Table 1).
Galectin-9, Table 1

Galectin-9 binding proteins

Name

Functional consequence of interaction

Carbohydrate dependent

CD40

Tim-3 independent control of CD40 signaling

Yes

CD44

Suppression of CD44-hyalunoric acid binding

Induction osteoblast differentiation

Increases iTreg cell stability

Yes

Galectin-9

Homomultimerization to increase valency

Yes

Galectin-8

Heteromultimerization to increase valency

Yes

Galectin-3

Heteromultimerization to increase valency

Yes

Glucagon receptor (Gcgr)

Reduces receptor mobility and enhances glucagon sensitivity

Yes

IgE/IgE glycopeptides

Prevention of mast cell degranulation

Yes

Latent membrane protein 1 (Lmp1)

ND

ND

NF-IL6 (C/EBPb)

Inflammatory cytokine production

ND

Protein disulfide isomerases

Regulation of T-cell migration

Yes

T cell immunoglobulin mucin-3 (Tim-3)

Modulation of various immune cells

Yes

Thrombin

Cleavage of gal-9 and decrease of eosinophil attraction

ND

Of note, as described previously, the gal-9 transcipt is extensively spliced which gives rise to multiple gal-9 protein isoforms. The processing and cellular distribution of these isoforms differs which can affect gal-9 function. For example, the length of the linker sequence affects the susceptible to proteolytic cleavage which influences the chemoattractive activity (Nishi et al. 2006). In addition, the isoforms in which exon 10 has been spliced appear to remain intracellular while other isoforms are found in the extracellular environment (Spitzenberger et al. 2001; Heusschen et al. 2014).

Galectin-9 in Pathology

Given the immunoregulatory functions of gal-9, the protein has been linked to different pathologies that involve an inadequate response of the immune system, including autoimmune diseases, like rheumatoid arthritis and multiple sclerosis, as well as graft versus host disease and asthma. In addition, gal-9 has been associated with the immune response to microbial infections. Several pathogens, e.g., viruses and bacteria, induce gal-9 expression in the infected host cells. This generally results in the activation of an innate immune response by the gal-9-mediated recruitment of eosinophils. At the same time, the adaptive immune response is hampered since elevated gal-9 expression has a suppressive effect on T cell responses. Of note, an intestinal nematode found in dogs expresses a gal-9-like protein that can suppress an inflammatory response (Wiersma et al. 2013; Merani et al. 2015). Hence, pathogens can exploit the dual immunomodulatory activity of galectin-9. Tumors also seem to exploit the dual activity of gal-9 during disease progression. In tumor cells, the expression of gal-9 is frequently reduced as compared to their normal counterparts, e.g., in gastric, colon, prostate, cervix, and skin cancer. The decreased gal-9 expression can reduce cell apoptosis and increase cell proliferation. In addition, loss of gal-9 can reduce tissue integrity thereby promoting tumor cell metastasis. In line with this, reduced tumoral gal-9 expression is frequently associated with poor patient survival. At the same time, gal-9 expression is frequently elevated in endothelial cells that line the tumor vasculature. This endothelial expression can further contribute to tumor progresssion by suppressing an adequate antitumor response and by facilitating tumor angiogenesis (Heusschen et al. 2013; Heusschen et al. 2014).

Summary

Galectin-9 was first described almost two decades ago. In the past 20 years, it has become apparent that the biology of this tandem repeat galectin is as complex as it is diverse.

This complexicity starts at the genomic level with the existence of two gal-9-like genes, both of which is still unclear whether they contribute to gal-9 protein expression. In general, the regulation of LGALS9 gene transcription is still poorly understood and awaits further investigation. Especially, the regulatory mechanisms that direct the posttranscriptional splicing and posttranslational modifications should be further unraveled as these processes give rise to multiple gal-9 isoforms. Deciphering the role of these isoforms in cellular functions and biological processes remains another major future research challenge.

Similar to most other galectin family members, gal-9 exerts its main functions by interacting with specific glycans on glycoproteins and glycolipids. Thus far, only a few of such glycoconjugates have been identified and – based on other galectins – it can be anticipated that multiple other gal-9-binding partners exist. In addition, gal-9 also likely participates in noncarbohydrate-mediated direct protein-protein interactions as described for other galectins. The identification of both carbohydrate dependent and independent gal-9 binding partners will provide more insight in the diverse activities of gal-9. Regarding the latter, the list of cellular and biological processes that involve gal-9 is continously expanded. It is now clear that gal-9 is involved in shaping the immune response, possibly by regulating the balance between innate and adaptive immunity. However, the exact mechanisms as well as the role of the different isoforms in this immunomodulatory process require further studies. In particular, because the immunomodulatory activity of gal-9 has been linked to different pathologies. Thus, a better insight in the regulatory mechanisms could provide therapeutic opportunities based on gal-9 activity.

References

  1. Barondes SH, Castronovo V, Cooper DN, Cummings RD, Drickamer K, Feizi T, et al. Galectins: a family of animal beta-galactoside-binding lectins. Cell. 1994;76:597–8.PubMedCrossRefGoogle Scholar
  2. Bi S, Hong PW, Lee B, Baum LG. Galectin-9 binding to cell surface protein disulfide isomerase regulates the redox environment to enhance t-cell migration and HIV entry. PNAS. 2011;108:10650–5.PubMedCrossRefPubMedCentralGoogle Scholar
  3. Gieseke F, Kruchen A, Tzaribachev N, Bentzien F, Dominici M, Müller I. Proinflammatory stimuli induce galectin-9 in human mesenchymal stromal cells to suppress t-cell proliferation. Eur J Immunol. 2013;43:2741–9.PubMedCrossRefGoogle Scholar
  4. Heusschen R, Griffioen AW, Thijssen VL. Galectin-9 in tumor biology: a jack of multiple trades. Biochim Biophys Acta. 2013;1836:177–85.PubMedPubMedCentralGoogle Scholar
  5. Heusschen R, Schulkens IA, van Beijnum J, Griffioen AW, Thijssen VL. Endothelial LGALS9 splice variant expression in endothelial cell biology and angiogenesis. Biochim Biophys Acta. 2014;1842:284–92.PubMedCrossRefGoogle Scholar
  6. Hirabayashi J, Hashidate T, Arata Y, Nishi N, Nakamura T, Hirashima M, et al. Oligosaccharide specificity of galectins: a search by frontal affinity chromatography. Biochim Biophys Acta. 2002;1572:232–54.PubMedCrossRefGoogle Scholar
  7. Hirashima M, Kashio Y, Nishi N, Yamauchi A, Imaizumi TA, Kageshita T, et al. Galectin-9 in physiological and pathological conditions. Glycoconj J. 2004;19:593–600.CrossRefGoogle Scholar
  8. Horlacher T, Oberli MA, Werz DB, Kröck L, Bufali S, Mishra R, et al. Determination of carbohydrate-binding preferences of human galectins with carbohydrate microarrays. ChemBioChem. 2010;11:1563–73.PubMedCrossRefGoogle Scholar
  9. Leal-Pinto E, Tao W, Rappaport J, Richardson M, Knorr BA, Abramson RG. Molecular cloning and functional reconstitution of a urate transporter/channel. J Biol Chem. 1997;272:617–25.PubMedCrossRefGoogle Scholar
  10. Leffler H, Carlsson S, Hedlund M, Qian Y, Poirier F. Introduction to galectins. Glycoconj J. 2004;19:433–40.CrossRefGoogle Scholar
  11. Lipkowitz MS, Leal-Pinto E, Cohen BE, Abramson RG. Galectin 9 is the sugar-regulated urate transporter/channel UAT. Glycoconj J. 2004;19:491–8.CrossRefGoogle Scholar
  12. Matsushita N, Nishi N, Seki M, Matsumoto R, Kuwabara I, Liu FT, et al. Requirement of divalent galactoside-binding activity of ecalectin/galectin-9 for eosinophil chemoattraction. J Biol Chem. 2000;275:8355–60.PubMedCrossRefGoogle Scholar
  13. Merani S, Chen W, Elahi S. The bitter side of sweet: the role of galectin-9 in immunopathogenesis of viral infections. Rev Med Virol. 2015;25:175–86.PubMedCrossRefGoogle Scholar
  14. Miyanishi N, Nishi N, Abe H, Kashio Y, Shinonaga R, Nakakita S, et al. Carbohydrate-recognition domains of galectin-9 are involved in intermolecular interaction with galectin-9 itself and other members of the galectin family. Glycobiology. 2007;17:423–32.PubMedCrossRefGoogle Scholar
  15. Nagae M, Nishi N, Murata T, Usui T, Nakamura T, Wakatsuki S, et al. Crystal structure of the galectin-9 n-terminal carbohydrate recognition domain from Mus musculus reveals the basic mechanism of carbohydrate recognition. J Biol Chem. 2006;281:35884–93.PubMedCrossRefGoogle Scholar
  16. Nishi N, Itoh A, Shoji H, Miyanaka H, Nakamura T. Galectin-8 and galectin-9 are novel substrates for thrombin. Glycobiology. 2006;16:15C–20C.PubMedCrossRefGoogle Scholar
  17. Orr SL, Le D, Long JM, Sobieszczuk P, Ma B, Tian H, et al. A phenotype survey of 36 mutant mouse strains with gene-targeted defects in glycosyltransferases or glycan-binding proteins. Glycobiology. 2013;23:363–80.PubMedCrossRefGoogle Scholar
  18. Spitzenberger F, Graessler J, Schroeder HE. Molecular and functional characterization of galectin 9 mrna isoforms in porcine and human cells and tissues. Biochimie. 2001;83:851–62.PubMedCrossRefGoogle Scholar
  19. Thijssen VL, Rabinovich GA, Griffioen AW. Vascular galectins: regulators of tumor progression and targets for cancer therapy. Cytokine Growth Factor Rev. 2013;24:547–58.PubMedCrossRefGoogle Scholar
  20. Tureci O, Schmitt H, Fadle N, Pfreundschuh M, Sahin U. Molecular definition of a novel human galectin which is immunogenic in patients with hodgkin’s disease. J Biol Chem. 1997;272:6416–22.PubMedCrossRefGoogle Scholar
  21. Wada J, Kanwar YS. Identification and characterization of galectin-9, a novel beta-galactoside-binding mammalian lectin. J Biol Chem. 1997;272:6078–86.PubMedCrossRefGoogle Scholar
  22. Wada J, Ota K, Kumar A, Wallner EI, Kanwar YS. Developmental regulation, expression, and apoptotic potential of galectin-9, a beta-galactoside binding lectin. J Clin Invest. 1997;99:2452–61.PubMedCrossRefPubMedCentralGoogle Scholar
  23. Wiersma VR, de Bruyn M, Helfrich W, Bremer E. Therapeutic potential of galectin-9 in human disease. Med Res Rev. 2013;33:E102–26.PubMedCrossRefGoogle Scholar
  24. Yang Q, Jiang W, Zhuang C, Geng Z, Hou C, Huang D, et al. MicroRNA-22 downregulation of galectin-9 influences lymphocyte apoptosis and tumor cell proliferation in liver cancer. Oncol Rep. 2015;34:1771–8.PubMedCrossRefGoogle Scholar
  25. Yoshida H, Teraoka M, Nishi N, Nakakita S, Nakamura T, Hirashima M, et al. X-ray structures of human galectin-9 c-terminal domain in complexes with a biantennary oligosaccharide and sialyllactose. J Biol Chem. 2010;285:36969–76.PubMedCrossRefPubMedCentralGoogle Scholar

Copyright information

© Springer International Publishing AG 2018

Authors and Affiliations

  1. 1.Departments of Medical Oncology and Radiation OncologyVU University Medical CenterAmsterdamThe Netherlands