Proc Natl Acad Sci U S A 95(8): 4504-4509

Immune regulation by the ST6Gal sialyltransferase

^{Howard Hughes Medical Institute, Division of Cellular and Molecular Medicine, and the Glycobiology Program, 9500 Gilman Drive 0625, University of California at San Diego, La Jolla, CA 92093; and}^{CYTEL Corporation, 3525 John Hopkins Court, San Diego, CA 92121}

^{Present address: Institute of Physiology, University of Zurich, 8057 Zurich, Switzerland.}

^{To whom correspondence should be addressed at: Howard Hughes Medical Institute, 9500 Gilman Drive 0625, University of California at San Diego, La Jolla, CA 92093.}

Communicated by Robert L. Hill, Duke University Medical Center, Durham, NC

Received 1997 Aug 5; Accepted 1998 Feb 4.

Abstract

The ST6Gal sialyltransferase controls production of the Siaα2-6Galβ1-4GlcNAc (Sia6LacNAc) trisaccharide, which is the ligand for the lectin CD22. Binding of CD22 to Sia6LacNAc is implicated in regulating lymphocyte adhesion and activation. We have investigated mice that lack ST6Gal and report that they are viable, yet exhibit hallmarks of severe immunosuppression unlike CD22-deficient mice. Notably, Sia6LacNAc-deficient mice display reduced serum IgM levels, impaired B cell proliferation in response to IgM and CD40 crosslinking, and attenuated antibody production to T-independent and T-dependent antigens. Deficiency of ST6Gal was further found to alter phosphotyrosine accumulation during signal transduction from the B lymphocyte antigen receptor. These studies reveal that the ST6Gal sialyltransferase and corresponding production of the Sia6LacNAc oligosaccharide are essential in promoting B lymphocyte activation and immune function.

Abstract

Sialyltransferases are a family of glycosyltransferase enzymes that add sialic acid residues during oligosaccharide diversification (reviewed in ref. 1). Sialic acid addition occurs in the Golgi apparatus and generally terminates further oligosaccharide chain elongation. The outer position of sialic acid linkages places these residues in a location to provide key structural determinants in ligand formation for endogenous and pathogenic lectins. Three sialic acid linkage types commonly exist among vertebrates and the corresponding sialyltransferase genes have been previously isolated. The most abundant sialic acid linkage found among mammalian cell surface oligosaccharides is of the α2-3 variety and can be produced independently by four sialyltransferases that each, nonetheless, bear unique substrate preferences among glycolipids, asparagine (N)-linked glycans, and serine/threonine (O)-linked glycans (2). Sialyltransferases have also been found to be developmentally regulated and differentially expressed among various cell types (1–6). For example, expression of α2-8 linked sialic acids is much less common than α2-3 linkages and appears restricted to a small subset of glycoproteins (7–10).

α2-6-linked sialic acids are also less abundant than α2-3-linked forms and are generated by at least four distinct gene products. However, the ST6Gal sialyltransferase appears solely responsible for producing the Siaα2-6Galβ1-4GlcNAc (Sia6LacNAc) terminus on various N glycans, and perhaps on some O glycans (1, 11). High levels of ST6Gal RNA have been found to preferentially accumulate in hematopoietic cells, as well as in the liver (3–5). Moreover, ST6Gal gene transcription is regulated by multiple promoters and altered by glucocorticoids and cytokines (12–14). Although the physiologic role of the ST6Gal sialyltransferase has not been defined previously by available genetic approaches, it has been shown to be unique in producing the ligand for the CD22 lectin molecule expressed on B lymphocytes.

CD22 is a transmembrane glycoprotein lectin found exclusively on B lymphocytes and is known to play a role in the immunologic activation of these cells (15–17). CD22 has been found associated with the antigen receptor and is a target for tyrosine kinase phosphorylation on the cytoplasmic domain, which thereby recruits various signal transduction molecules (18, 19). The extracellular domain of CD22 specifically binds the Sia6LacNAc trisaccharide (20–22). This trisaccharide ligand exists on several lymphoid molecules. Lymphocyte interactions involving CD22 binding to CD45 have been reported (23). As CD22 itself carries Sia6LacNAc, homotypic binding interactions have been shown to occur and may play a regulatory role in immune function (24, 25). These results suggest that CD22 and Sia6LacNAc are a lectin–ligand pair with the potential to control immune cell surface interactions. However, a relatively simple model for CD22 function has not developed from analyses of CD22 null mice by several laboratories (26–29). Results obtained have inferred both positive and negative roles for CD22 in B lymphocyte immune function, suggesting that CD22 may modulate threshold signaling responses from the antigen-receptor complex.

To investigate ST6Gal-dependent physiology we have chosen a complementary approach involving the generation of mice deficient in the carbohydrate ligand for CD22 by inactivating the ST6Gal sialyltransferase gene implicated in it’s synthesis. We report that such mice develop normally but harbor an immunodeficient phenotype that is distinct from CD22 null mice. These studies describe an essential role for the ST6Gal sialyltransferase in B lymphocyte immune responses.

Results are mean ± SD determined using three distinct mice of the indicated genotypes. ST6Gal activity is defined as detailed in Materials and Methods. Activity observed in Δ/Δ samples is not significantly distinct from background and is at the level of assay sensitivity.

Acknowledgments

We thank Ajit Varki for helpful comments on the manuscript and the generous gift of CD22–Ig chimera and Mary-Ann Campbell for the anti-CD22 antibody. This research was supported by the Howard Hughes Medical Institute (J.D.M.). T.H. was supported by scholarships from the Swiss National Science Foundation and the Swiss Foundation for Medical and Biological Fellowships.

Acknowledgments

ABBREVIATIONS

Sia6LacNAc	Siaα2-6Galβ1-4GlcNAc
FACS	fluorescence-activated cell sorting
ES	embryonic stem
wt	wild type
KLH	keyhole limpet hemocyanin
DNP	dinitrophenyl
LPS	lipopolysaccharide
IL-4	interleukin 4

ABBREVIATIONS

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