Galectin-9

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Galectin-9 is the best-studied of the tandem-repeat galectins and the crystal structure of the N-terminal carbohydrate recognition domain (CRD) is known. In addition, Galectin-9...

  • uniquely binds poly-N-acetyllactosamine sequences by recognizing internal N-acetyllactosamine repeats[1]
  • binds distinct ligands from Galectin-1[2]
  • has three well-characterized linker domains between the CRDs, generated by alternative splicing, that regulate cellular localization and function of the protein
  • is the only tandem-repeat galectin that has been administered in animal models of disease to assess therapeutic potential[3][4][5]
  • null mice have increased susceptibility to autoimmune disease
  • binds to a unique glycoprotein ligand Tim-3 expressed in Th1 and Th17 cells[4][6][7][8]

Contents

CFG Participating Investigators contributing to the understanding of this paradigm

CFG Participating Investigators (PIs) contributing to the understanding of Galectin-9 include: Linda Baum, Richard Cummings, Gabriel Rabinovich, Sachiko Sato

Progress toward understanding this GBP paradigm

This section documents what is currently known about Galectin-9, its carbohydrate ligand(s), and how they interact to mediate cell communication. Further information can be found in the GBP Molecule Page for human and mouse Galectin-9 in the CFG database.

Carbohydrate ligands

Human galectin-9 binding to glycans has been studied by a variety of techniques including glycan microarray analysis and frontal affinity chromatography.

On the CFG glycan microarray, the individual N- and C-terminal domains of recombinant dog (Canis lupus) galectin-9, generated as GST (glutathione-S-transferase) chimeras, showed similarities in glycan recognition, but also distinct differences[9]. While both domains bound well to short sulfated glycans, such as 3-O-sulfated galactose in short LacNAc structures, only the N-terminal domain bound well to many glycans expressing blood group A-related sequences and to the Forssman glycolipid-like glycans, whereas the C-terminal domain bound less well to the blood group related structures, but showed binding to a linear sialylated poly-N-acetyllactosamine pentasaccharide.

In frontal affinity chromatography, recombinant human galectin-9 was found to preferentially bind to both branched N-glycans (Kd = 0.16 μM toward tetraantennary N-glycans terminating in galactose) and glycans with poly-N-acetyllactosamine sequences (Kd = 0.09 μM toward octasaccharides with 4 repeating LacNAc groups, and this was found for both the N- and C-terminal domains. By contrast, the N-terminal, but not the C-terminal domain, showed significant binding in the low μM range to Forssman glycolipid-derived pentasaccharides and to blood group A hexasaccharide[10].

Glycan microarray analyses in microarrays with relatively short glycan species[11], showed that both the recombinant full-length human galectin-9 and the N-terminal domain displayed very similar binding patterns, and both bound to LacNAc sequences and even better to short fucosylated glycans with terminal blood group A and B trisaccharide sequences.

Cellular expression of GBP and ligands

Galectin-9 is widely expressed in various tissues (heart, lung, liver, kidney, spleen, muscle, intestine, and thymus), but weakly expressed in brain[12]. Interestingly, the rat urate transporter was reported to be 99% identical to the sequence reported for rat galectin-9 [13], suggesting that these two proteins are the same[14][15], and suggest that galectin-9 may have multiple functions, occurring as a polytopic transmembrane protein to function as the urate transporter, and as a soluble protein in its signaling and cell-binding forms.

Biosynthesis of ligands


Structure

Galectin-9 (long isoform in humans) has 355 amino acids and behaves as an ~35 kDa protein; short isoforms differ in the linker peptide length and have lower apparent sizes compared to the full-length long isoform. The crystal structure of the N-terminal carbohydrate recognition domain (CRD) been defined.[16][17][18] The GBP shows strong interactions in a metal-free manner with poly-N-acetyllactosamine sequences comprised of repeating (-3Galβ1-4GlcNAcβ1-)n by recognizing internal N-acetyllactosamine repeats [16]. Generally, it binds distinct glycan ligands from Galectin-1 [2]). There are has three well-characterized linker domains between the CRDs, generated by alternative splicing [19], that may regulate cellular localization and function of the protein. Truncation of linker domain between CRDs in recombinant forms of galectin-9 stabilize the protein to proteolysis [20].

Biological roles of GBP-ligand interaction


CFG resources used in investigations

The best examples of CFG contributions to this paradigm are described below, with links to specific data sets. For a complete list of CFG data and resources relating to this paradigm, see the CFG database search results for Galectin-9.

Glycan profiling


Glycogene microarray

Probes for human galectin-9 have been included in all versions of the CFG glycogene chip, and probes for mouse galectin-9 are included on versions 2, 3, and 4.

Knockout mouse lines

CFG-generated Galectin-9 knockout mice have been used to study the biological functions of this paradigm GBP. (CFG PI data)

Glycan array

Investigators have used CFG carbohydrate compounds and glycan array screening to study ligand binding specificity of Galectin-9 (for example, click here). To see all glycan array results for Galectin-9, click here.

Related GBPs

Galectin-4 (CFG data), galectin-6, galectin-8 (CFG data), and galectin-12 (CFG data).

References

  1. Nagae, M. et al. Structural analysis of the recognition mechanism of poly-N-acetyllactosamine by the human galectin-9 N-terminal carbohydrate recognition domain. Glycobiology 19, 112-117 (2009).
  2. 2.0 2.1 Bi, S., Earl, L.A., Jacobs, L. & Baum, L.G. Structural features of galectin-9 and galectin-1 that determine distinct T cell death pathways. J Biol Chem 283, 12248-12258 (2008).
  3. Baba, M. et al. Galectin-9 inhibits glomerular hypertrophy in db/db diabetic mice via cell-cycle-dependent mechanisms. J Am Soc Nephrol 16, 3222-3234 (2005).
  4. 4.0 4.1 Seki, M. et al. Galectin-9 suppresses the generation of Th17, promotes the induction of regulatory T cells, and regulates experimental autoimmune arthritis. Clin Immunol 127, 78-88 (2008).
  5. Tsuchiyama, Y. et al. Efficacy of galectins in the amelioration of nephrotoxic serum nephritis in Wistar Kyoto rats. Kidney Int 58, 1941-1952 (2000).
  6. Naka, E.L., Ponciano, V.C., Cenedeze, M.A., Pacheco-Silva, A. & Camara, N.O. Detection of the Tim-3 ligand, galectin-9, inside the allograft during a rejection episode. Int Immunopharmacol 9, 658-662 (2009).
  7. Niwa, H. et al. Stable form of galectin-9, a Tim-3 ligand, inhibits contact hypersensitivity and psoriatic reactions: a potent therapeutic tool for Th1- and/or Th17-mediated skin inflammation. Clin Immunol 132, 184-194 (2009).
  8. Anderson, D.E. TIM-3 as a therapeutic target in human inflammatory diseases. Expert Opin Ther Targets 11, 1005-1009 (2007).
  9. Poland PA, Rondanino C, Kinlough CL, Heimburg-Molinaro J, Arthur CM, Stowell SR, Smith DF, Hughey RP. Identification and characterization of endogenous galectins expressed in Madin Darby canine kidney cells. J Biol Chem. 2011;286(8):6780-90
  10. Hirabayashi J, Hashidate T, Arata Y, Nishi N, Nakamura T, Hirashima M, Urashima T, Oka T, Futai M, Muller WE, Yagi F, Kasai K. Oligosaccharide specificity of galectins: a search by frontal affinity chromatography. Biochim Biophys Acta. 2002;1572(2-3):232-54
  11. Horlacher T, Oberli MA, Werz DB, Krock L, Bufali S, Mishra R, Sobek J, Simons K, Hirashima M, Niki T, Seeberger PH. Determination of carbohydrate-binding preferences of human galectins with carbohydrate microarrays. Chembiochem. 2010;11(11):1563-73
  12. 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(10):2452-61
  13. 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(1):617-25
  14. Lipkowitz MS, Leal-Pinto E, Rappoport JZ, Najfeld V, Abramson RG. Functional reconstitution, membrane targeting, genomic structure, and chromosomal localization of a human urate transporter. J Clin Invest. 2001;107(9):1103-15.
  15. Lipkowitz MS, Leal-Pinto E, Cohen BE, Abramson RG. Galectin 9 is the sugar-regulated urate transporter/channel UAT. Glycoconj J. 2004;19(7-9):491-8
  16. 16.0 16.1 Nagae M, Nishi N, Nakamura-Tsuruta S, Hirabayashi J, Wakatsuki S, Kato R. Structural analysis of the human galectin-9 N-terminal carbohydrate recognition domain reveals unexpected properties that differ from the mouse orthologue. J Mol Biol. 2008;375(1):119-35.
  17. Yoshida H, Teraoka M, Nishi N, Nakakita S, Nakamura T, Hirashima M, Kamitori S. X-ray structures of human galectin-9 C-terminal domain in complexes with a biantennary oligosaccharide and sialyllactose. J Biol Chem. 2010;285(47):36969-76.
  18. Nagae M, Nishi N, Murata T, Usui T, Nakamura T, Wakatsuki S, Kato R. 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(47):35884-93.
  19. Nishi N, Itoh A, Shoji H, Miyanaka H, Nakamura T. Galectin-8 and galectin-9 are novel substrates for thrombin. Glycobiology. 2006;16(11):15C-20C
  20. Nishi N, Itoh A, Fujiyama A, Yoshida N, Araya S, Hirashima M, Shoji H, Nakamura T. Development of highly stable galectins: truncation of the linker peptide confers protease-resistance on tandem-repeat type galectins. FEBS Lett. 2005;579(10):2058-64

Acknowledgements

The CFG is grateful to the following PIs for their contributions to this wiki page: Linda Baum, Richard Cummings

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