Department of Anatomy and Program in Immunology, University of California, San Francisco, Ca 94143-0452, USA
Received on June 30, 2002; revised on September 18, 2002; accepted on September 25, 2002
![]() |
Abstract |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Key words: GlcNAc-6-O-sulfotransferase / homing / L-selectin / mucin / sulfation
![]() |
Introduction |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
GlcNAc6ST-2 is also essential for the formation of the epitope of MECA-79, a monoclonal antibody that stains HEV and blocks L-selectin-dependent adherence of lymphocytes (Berg et al., 1991; Hemmerich et al., 1994
, 2001a
; Yeh et al., 2001
). The closest relatives of GlcNAc6ST-2 are GlcNAc6ST-3 and GlcNAc6ST-5, also known as I-GlcNAc6ST (Lee et al., 1999
) and C-GlcNAc6ST (Akama et al., 2000
; Hemmerich et al., 2001b
), respectively, because of their prominent expression in the intestine and cornea, respectively. These three sulfotransferases map to the same region of chromosome 16 and show the highest degree of sequence homology within the GlcNAc6ST subfamily (Hemmerich et al., 2001b
). Mutations in the gene for GlcNAc6ST-5 underlie macular corneal dystrophy in humans, which is attributable to the normal role of this enzyme in transferring sulfate to C-6 of GlcNAc within keratan sulfate (Akama et al., 2000
, 2001
). The remaining member of the family, GlcNAc6ST-4, like GlcNAC6ST-1, shows a broad expression pattern in human tissues (Bhakta et al., 2000
; Kitagawa et al., 2000
; Uchimura et al., 2000
). This enzyme is also reported to be a chondroitin 6-O-sulfotransferase that transfers sulfate to C-6 of GalNAc (Kitagawa et al., 2000
).
The present study focuses on GlcNAc6ST-3 because of the limited functional and biochemical information available about this enzyme. In comparing GlcNAc6ST-3 with GlcNAc6ST-1 and GlcNAc6ST-2, we find striking differences with respect to the acceptor specificities and the sulfated products that can be formed. Moreover, we have expanded the survey of tissue expression for GlcNAc6ST-3 by using a more extensive panel of tissue cDNAs and including isolated cell populations. The pertinence of our findings to L-selectin ligand synthesis is discussed.
![]() |
Results |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
COS-7 cells were transfected with a cDNA for each of these sulfotransferases together with a cDNA encoding one of the acceptor glycoproteins in the form of an Fc chimera. The Fc region itself (denoted as IgG) contains one potential site for N-linked glycosylation. During the transfection, the cells were labeled with [35S]-SO4. The chimeric proteins were purified on protein Aagarose resins and analyzed by sodium dodecyl sulfatepolyacrylamide gel electrophoresis (SDSPAGE). We verified by Coomassie blue staining (Figure 1B) that expression levels of the different acceptor proteins were similar. Differences in sulfation levels could therefore be taken to be due to differential activity of the enzymes. As documented previously and confirmed in Figure 1A, COS cells possess endogenous sulfotransferase activity that resulted in detectable levels of sulfation on several of the acceptors in the absence of exogenously provided activity (Tangemann et al., 1999). The GlcNAc6STs imparted only trace levels of sulfate to the IgG component of the chimeras as judged by the weak labeling seen for IgG construct. KSGal6ST, GlcNAc6ST-1, and GlcNAc6ST-2 catalyzed strong sulfation in all acceptors of both classes. In contrast, GlcNAc6ST-3 demonstrated a marked preference for the mucin containing acceptors in that only very weak (relative to the mock transfection) label was incorporated into NCAM-1, ICAM-1, or IgG.
|
|
|
|
As reviewed, GlcNAc6ST-2 has a demonstrated function in the elaboration of L-selectin ligands in HECs of lymph nodes. We next asked whether GlcNAc6ST-3, like GlcNAc6ST-2, was expressed in isolated HECs. Here we took advantage of the MECA-79 monoclonal antibody, which selectively stains HEVs in secondary lymphoid organs. HECs were isolated from collagenase-dissociated human tonsils by affinity to MECA-79 and a magnetic bead immunoselection (see Materials and methods). In parallel, T and B cells (CD3+ and CD19+ cells, respectively) were isolated from tonsillar cell suspensions using fluorescence-activated cell sorting. Semi-quantitative PCR was performed on cDNAs generated from these specimens. As shown in Figure 5B, we observed strong expression of GlcNAc6ST-2 in the isolated HECs, consistent with our previous observations (Bistrup et al., 1999). Interestingly, low levels of GlcNAc6ST-2 transcripts were also detected in the cDNAs derived from the populations of B and T cells, which were estimated to be >99% pure by flow cytometry analysis. A converse expression pattern was observed for GlcNAc6ST-3 (Figure 5A): B and T cells showed significant expression, whereas only a trace level was detectable in HECs. This minimal expression was very likely attributable to contamination of the HECs with adherent lymphocytes. We also found only barely detectable levels of GlcNAc6ST-3 transcripts in HECs isolated from mouse lymph node or Peyer's patches (data not shown).
|
|
![]() |
Discussion |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
GlcNAc-6-O-sulfotransferase activities have been demonstrated in extracts from a variety of tissues and tumors (Carter et al., 1988; Goso and Hotta, 1993
; Spiro et al., 1996
; Bowman et al., 1998
; Nakazawa et al., 1998
; Hasegawa et al., 2000
; Seko et al., 2000
; Delmotte et al., 2001
). Underlying these activities are members of the GlcNAc6ST subfamily of carbohydrate sulfotransferases, of which five have been cloned to date. This subfamily is distinct with respect to sequence homology and activities from the heparan sulfate GlcNSO3-6-O-sulfotransferase (HS6ST) subfamily of three enzymes that are responsible for addition of sulfate to the 6-position of GlcN or GlcNAc in heparan sulfate chains (reviewed in Fukuda et al., 2001
). It is strongly suspected that the existence of multiple sulfotransferases for a given modification underlies higher-order specificities, involving the elaboration of sulfate modifications within the context of different oligosaccharides (Bowman and Bertozzi, 1999
; Fukuda et al., 2001
). Further acceptor specificity may derive from features of the protein scaffold. An emerging view is that different sulfotransferases acting in concert with specific glycosyltransferases provide for a huge diversity of sulfate-based determinants ("sulfotopes") that enable a broad range of biological recognition events (Bowman and Bertozzi, 1999
).
Among the members of the GlcNAc6ST subfamily, two members have been extensively characterized with respect to their function, as reviewed in the Introduction. GlcNAc6ST-2 is strongly implicated as one of the contributing enzymes in the synthesis of the 6-sulfo sLex determinant on L-selectin ligands, as well as the formation of the MECA-79 epitope carried by these ligands. Akama et al. (2000, 2001
) have demonstrated the involvement of GlcNAc6ST-5 in the elaboration of the glycosaminoglycan chains of keratan sulfate. Although GlcNAc6ST-3 is highly homologous to GlcNAc6ST-5, and it is likely that their genes arose by gene duplication, GlcNAc6ST-3 is not able to generate highly sulfated keratan sulfate (Akama et al., 2001
). Also, when we compared the tissue distributions of GlcNAc6ST-3 and GlcNAc6ST-5, notable differences were found. The former was strongly expressed in tissues of the gut, whereas the latter was not. A commonality was their expression in peripheral blood mononuclear cells and brain.
To address the issue of the acceptor specificity for GlcNAc6ST-3, we analyzed the ability of several members of the GlcNAc6ST subfamily to sulfate N-linked versus O-linked chains in a series of glycoproteins. Compared to its closest relatives in the GlcNAc6ST family, GlcNAc6ST-3 was much more selective for O-linked chains of mucin-type acceptors with minimal activity in sulfating glycoproteins with predominantly N-linked glycosylation. No such preference was found for GlcNAc6ST-2, despite the fact that the functionally relevant HEV-expressed acceptors for this enzyme are mucin-type glycoproteins (Puri et al., 1995; Hemmerich et al., 2001a
). As expected, the sulfation of O-linked chains by GlcNAc6ST-3 within CHO cells required the presence of the core 2 branch. Our findings are compatible with the recent biochemical analyses in which cell-free sulfotransferase assays were performed with a series of recombinantly expressed GlcNAc6STs and defined oligosaccharides acceptors (Bowman et al., 2001
; Seko et al., 2002; Uchimura et al., 2002
). GlcNAc6ST-1 and GlcNAc6ST-2 were able to sulfate GlcNAc on a core 2 acceptor (Bowman et al., 2001
; Uchimura et al., 2002
), but both were also active on GlcNAc-Man structures found in N-Linked chains. In contrast, GlcNAc6ST-3 was found to act on the core 2 acceptor but exhibited very limited activity on GlcNAc-Man structures, consistent with our observation that this enzyme preferred mucin-type over N-linked acceptors in cotransfection assays. Our findings with respect to the acceptor preference for GlcNAc6ST-2 appear to be at variance with those of Hiraoka et al. (1999)
who studied the mouse homolog of this enzyme. They, like us, found that this enzyme could direct the sulfation of IgG chimeras of GlyCAM-1, MAdCAM-1, and CD34, but they observed no detectable activity above background on NCAM-1/IgG. Whether this discrepancy is attributable to the species difference or to technical issues (e.g., level of enzyme or acceptor glycoprotein) remains to be determined.
Our previous analysis of GlcNAc6ST-2 genetargeted mice has established a clear role for this enzyme in the elaboration of HEV ligands for L-selectin and in the generation of the MECA-79 epitope on these ligands (Hemmerich et al., 2001a). However, the phenotype of the null mice was only partial in that L-selectin ligands and MECA-79 reactivity were still present in HEVs, albeit to a greatly reduced extent and restricted to the abluminal aspects of the HEVs. Also, we have observed a reduced but still substantial level of sulfation in HEV-expressed ligands from GlcNAc6ST-2 null mice (Van Zante and Rosen, unpublished data). One of the candidates for these residual activities has been GlcNAc6ST-3. Two of our present findings render this possibility remote. First, we found only a trace level of GlcNAc6ST-3 transcripts in isolated HECs, which was very likely attributable to contaminating lymphocytes in the HEC preparation. Second, our transfection experiments indicate that GlcNAc6ST-3 is unable to direct the synthesis of the 6-sulfo sLex determinant, a critical structure for L-selectin ligands. A more likely candidate to account for the residual ligand and MECA-79 reactivity in GlcNAc6ST-2 null mice is GlcNAc6ST-1, which is known to be expressed in HEVs (Uchimura et al., 1998a
). This enzyme can participate in the generation of functional L-selectin ligands, the 6-sulfo sLex determinant, and the MECA-79 epitope (Kimura et al., 1999
; Kanamori et al., 2002
; Uchimura et al., 2002
). It should be noted that GlcNAc6ST-1 is also implicated in the generation of the 6-sulfo sLex determinant in the mouse embryo (Fan et al., 1999
).
We found evidence for GlcNAc6ST-2 expression in B and T cells, although at a markedly lower level than in HECs (Figure 5B). This low level of expression likely explains the failure to detect this transcript by in situ hybridization in the lymphocyte-rich regions of lymph nodes (Bistrup et al., 1999; Hiraoka et al., 1999
). Ohmori et al. (2000)
reported the presence of transcripts for GlcNAc6ST-2 in the human lymphoid leukemia line Nawalma. However, our results provide the first direct evidence of its presence in primary lymphocytes. The enzyme may account for the occurrence of the 6-sulfo sLex epitope on subpopulations of lymphocytes (Kannagi and Kanamori, 1999
).
A novel finding of the present study is the expression of GlcNAc6ST-3 in mononuclear cells. This was established in sets of cDNAs derived by three independent means (Figures 4A, C, and 5A). The preference of GlcNAc6ST-3 to sulfate mucin acceptors focuses attention on leukocyte cell surface molecules with mucin domains. In fact, several such molecules (CD43, CD44, and CD45) are reported to be sulfated (Giordanengo et al., 1995; Maiti et al., 1998
; Brown et al., 2001
), although the nature of the sulfated moieties has not been reported. In the case of CD44, ligand binding activity (i.e., hyaluronic acid binding) depends on its sulfation (Maiti et al., 1998
; Brown et al., 2001
). PSGL-1, another sialomucin, is broadly distributed on leukocytes (reviewed in McEver and Cummings, 1997
) and carries tyrosine sulfation. On NK subpopulations, PSGL-1 is further decorated in a cell type-specific manner with the PEN5 epitope, which is thought to involve a GlcNAc-6-sulfate modification (Andre et al., 2000
). This epitope is of functional interest because it is implicated in L-selectin ligand activity on these cells. The potential involvement of GlcNAc6ST-3 in modulating the function of cell surface mucins on leukocytes deserves further attention.
The present study confirms and extends the original finding that GlcNAc6ST-3 is strongly expressed in the gastrointestinal tract. Studies by Seko et al. (2000, 2002) indicate that GlcNAc6ST-3 corresponds to the GlcNAc-6-O-transferase activity detected in normal human colon mucosa. In view of the preference of GlcNAc6ST-3 for mucin-type acceptors, demonstrated herein, a role in the modification of gut-associated mucins should be considered. A variety of secreted and membrane-bound mucins are found in the gut (reviewed in Kim and Gum, 1995
). Normal functions and pathological roles ascribed to mucins are varied, including mechanical protection, lubrication, facilitating metastasis of carcinoma cells, and providing attachment sites for microbes. Most recently, a gene targeting approach has implicated the Muc2 mucin in the suppression of colorectal cancer (Velcich et al., 2002
). Mucins exhibit an enormous diversity of sulfated O-linked chains (Lo-Guidice et al., 1994
; Capon et al., 1997
). Mucin function undoubtedly will depend on these sulfation modifications whether through influence on overall physicochemical properties or via sulfotopes involved in specific recognition events. Further work on the contribution of GlcNAc6ST-3 to the structure and function of gut-associated mucins under both normal and pathological circumstances is clearly warranted.
![]() |
Materials and methods |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
For the N-glycanase experiments, Fc-fusion proteins were produced as above. Each 25 µl of protein Aagarose-bound protein was boiled for 2 min in 20 mM sodium phosphate, pH 7.5, 50 mM ethylenediamine tetra-acetic acid (EDTA), 0.02% sodium azide, 0.5% SDS, and 5% ß-mercaptoethanol. As a control, 25 µg of 1-acid glycoprotein was subjected to N-glycanase treatment. The denatured samples were treated with or without 10 U/ml of recombinant Peptide-NGlycosidase F (Glyko, Novato, CA) in the presence of 20 mM sodium phosphate, pH 7.5, 50 mM EDTA, 0.02% sodium azide, 1% Nondiet P-40 (Sigma, St. Louis, MO) and 1% ß-mercaptoethanol in a volume of 30 µl for 16 h at 37°C. The resulting samples were analyzed by SDSPAGE as mentioned.
For the experiments to determine the presence of the 6-sulfo sLex determinant, CHO cells were transfected with cDNAs encoding full-length CD34, Core2GlcNAcT-I (pCDNA1.1), FTVII (pcDNA3.1), and either GlcNAc-6ST-2 or -3 or the empty vector (pcDNA3.1). The transfected cells were cultured for 3 days in the presence of Na2[35S]SO4. Whole cell lysates (0.1% Triton-X 100 in PBS) were prepared and equalized for protein content. Equal aliquots of each sample were separated by 10% SDSPAGE and visualized by Coomassie blue staining and autoradiography. In parallel, equal aliquots were incubated with 8 µg G72 monoclonal antibody immobilized on anti-murine IgM Sepharose (Zymed, San Francisco, CA). Bound proteins were separated by 10% SDSPAGE and visualized by Coomassie staining and autoradiography. To determine the presence of the 6-sulfo sLex determinant by flow cytometry, CHO cells stably expressing Core2GlcNAcT-I and FTVII were transiently transfected by Lipofectamine as previously described (Bistrup et al., 1999) with cDNAs encoding Core2GlcNAcT-I and FTVII and either GlcNAc6ST-2 or -3 or the empty vector (pCDNA3.1). Two days after transfection, cells were analyzed by flow cytometry as previously described (Bistrup et al., 1999
) using the G152 monoclonal antibody (Mitsuoka et al., 1998
) or a mouse IgM (Pharmingen, San Diego, CA) isotype control.
For the core 2 dependency experiments, CHO cells were transfected with a cDNA for GlcNAc6ST-3 (or the empty vector), together with a plasmid encoding one of a series of the Ig fusion proteins (CD34/IgG, ICAM/IgG NCAM/IgG or fractalkine/IgG) with or without a plasmid encoding Core2GlcNAcT-I (pCDNA1.1). Transfection methods and analysis of the recombinant fusion proteins were as already described for the COS cell transfections.
Reverse transcriptase PCR experiments
Primers for each GlcNAc6ST were designed to avoid cross-reactivity among the GlcNAc6ST cDNAs. The following primers were used:
To verify the specificity of the PCR reactions, each of three primer pairs were tested against all three GlcNAc6ST cDNAs (20 ng of each cDNAs per reaction). Only the homologous combinations yielded PCR products of the predicted size. In addition the following PCR reactions were verified by directly sequencing the PCR product isolated from the agarose gel: GlcNAc6ST-3 product from CD19+ cells (B cells) cDNA; GlcNAC6ST-2 product from tonsillar B cell cDNA; and GlcNAC6ST-2 product from tonsillar T cell cDNA.
Tissue expression of GlcNAc6ST-3 and GlcNAc6ST-5 were determined using the Rapid-Scan Gene Expression Panel Human-24 (Origene Technologies, Rockville, MD). According to the manufacturer's description, poly A+ RNA was used to synthesize first-strand cDNA, employing oligo(dT) primers. The amount of first-strand cDNAs from each tissue was normalized to contain an equivalent concentration of ß-actin reverse transcripts. The cDNA pools from the 24 human tissues were diluted in water at four different concentrations in steps of 10-fold dilutions with the lowest concentration at approximately 1 pg/ml. Fragments were amplified by PCR using the primers for GlcNAc6ST-3 already given. PCR conditions were as follows: 1 cycle of 3 min at 94°C, 30 s at 68°C, 1 min at 68°C and 35 cycles of 30 s at 94°C, 30 s at 68°C, 1 min at 68°C, followed by 5 min at 68°C.
cDNAs derived from isolated purified populations of peripheral blood mononuclear cells were obtained from Clontech (K1428-1). Mononuclear cells were purified from peripheral blood on a Percoll gradient. Populations of CD4+ T cells, CD8+ T cells, and CD19+ cells (B cells) were obtained by immunomagnetic separation using antibody conjugated Dynabeads (Dynal, Lake Success, NY). According to the manufacturer, the purity of the populations was >95%, as evaluated by immunostaining the preparations. PCR was performed as described.
HECs were purified from human tonsils by immunomagnetic selection with MECA-79 by a modification of a previously described procedure (Girard and Springer, 1995; Sassetti et al., 2000
). Surgical specimens were digested with a combination of collagenase A (Boehringer-Mannheim, Indianapolis, IN) and dispase I (Roche Diagnostics, Indianapolis, IN), and the resulting cell suspension (2x108) was incubated with 10 µg MECA-79 in 1 ml staining buffer (phosphate buffered saline [PBS] containing 1% bovine serum albumin) at 4°C for 20 min. Cells were collected by centrifugation, washed with staining buffer, and incubated with 10 µg of biotinylated mouse anti-rat IgM (Caltag, Burlingame, CA) at 4°C for 20 min. Finally, cells were collected, washed, and incubated with 40 µl of streptavidin-conjugated beads (Dynal) at 4°C for 20 min. MECA-79 positive cells were selected using a Dynal cell separation magnet and extensive washing as directed by the manufacturer. The purity of the resulting HEC preparation was 95% as determined by microscopic examination of cellular morphology with the major contaminant being adherent lymphocytes.
To purify T and B lymphocytes, tonsillar lymphocytes were prepared by mincing surgical specimens of human tonsil and flushing the loose lymphocytes through a 100 µm cell strainer with cold RPMI 1640 medium. The filtered cells were pelleted for 5 min at 1000 rpm and washed with PBS. T and B lymphocytes were purified by sorting after incubation with anti-CD3-FITC (Pharmingen) or anti-CD19-PE (Caltag). The purity of the sorted populations was >99%. Total RNA was isolated from each purified cell population (3 x 106 T cells and 1 x 107 B cells) by lysis and extraction with RNAzol (Tel-Test, Friendswood, TX). To avoid genomic DNA contamination, the purified RNA was digested with RNase-free Dnase I (Gibco BRL) followed by ethanol precipitation. First strand cDNA was synthesized from 5 µg of total RNA primed with random hexamers using AMV reverse transcriptase (Gibco BRL). PCR reactions were carried out as described.
![]() |
Acknowledgements |
---|
![]() |
Footnotes |
---|
1 To whom correspondence should be addressed; e-mail: sdr{at}itsa.ucsf.edu
![]() |
Abbreviations |
---|
![]() |
References |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Akama, T.O., Nakayama, J., Nishida, K., Hiraoka, N., Suzuki, M., McAuliffe, J., Hindsgaul, O., Fukuda, M., and Fukuda, M.N. (2001) Human corneal GlcNac 6-O-sulfotransferase and mouse intestinal GlcNac6-O-sulfotransferase both produce keratan sulfate. J. Biol. Chem., 276, 1627116278.
Andre, P., Spertini, O., Guia, S., Rihet, P., Dignat-George, F., Brailly, H., Sampol, J., Anderson, P.J., and Vivier, E. (2000) Modification of P-selectin glycoprotein ligand-1 with a natural killer cell-restricted sulfated lactosamine creates an alternate ligand for L-selectin. Proc. Natl Acad. Sci. USA, 97, 34003405.
Bazan, J.F., Bacon, K.B., Harsiman, G., Wang, W., Soo, K., Rossi, D., Greaves, D.R., Zlotnik, A., and Schall, T.J. (1997) A new class of membrane-bound chemokine with a CX3C motif. Nature, 385, 640644.[CrossRef][ISI][Medline]
Berg, E.L., Robinson, M.K., Warnock, R.A., and Butcher, E.C. (1991) The human peripheral lymph node vascular addressin is a ligand for LECAM-1, the peripheral lymph node homing receptor. J. Cell Biol., 114, 343349.[Abstract]
Bhakta, S., Bartes, A., Bowman, K.G., Kao, W.M., Polsky, I., Lee, J.K., Cook, B.N., Bruehl, R.E., Rosen, S.D., Bertozzi, C.R., and Hemmerich, S. (2000) Sulfation of N-acetylglucosamine by chondroitin 6-sulfotransferase 2 (GST-5). J. Biol. Chem., 275, 4022640234.
Bierhuizen, M.F. and Fukuda, M. (1992) Expression cloning of a cDNA encoding UDP-GlcNAc:Gal beta 1-3-GalNAc-R (GlcNAc to GalNAc) beta 1-6GlcNAc transferase by gene transfer into CHO cells expressing polyoma large tumor antigen. Proc. Natl Acad. Sci. USA, 89, 93269330.[Abstract]
Bistrup, A., Bhakta, S., Lee, J.K., Belov, Y.C., Gunn, M.D., Zuo, F.-R., Huang, C.-C., Kannagi, R., Rosen, S.D., and Hemmerich, S. (1999) Sulfotransferases of two specificities function in the reconstitution of high-endothelial-cell ligands for L-selectin. J. Cell Biol., 145, 899910.
Bowman, K.G. and Bertozzi, C.R. (1999) Carbohydrate sulfotransferases: mediators of extracellular communication. Chem. Biol., 6, R9R22.[CrossRef][ISI][Medline]
Bowman, K.G., Hemmerich, S., Bakhta, S., Singer, M.S., Bistrup, A., Rosen, S.D., and Bertozzi, C.R. (1998) Identification of an N-acetylglucamine-6-O-sulfotransferase activity specific to lymphoid tissue: an enzyme with a possible role in lymphocyte homing. Chem. Biol., 5, 447460.[ISI][Medline]
Bowman, K.G., Cook, B.N., de Graffenried, C.L., and Bertozzi, C.R. (2001) Biosynthesis of L-selectin ligands: sulfation of sialyl Lewis x-related oligosaccharides by a family of GlcNAc-6-sulfotransferases. Biochemistry, 40, 53825391.[CrossRef][ISI][Medline]
Brown, K.L., Maiti, A., and Johnson, P. (2001) Role of sulfation in CD44-mediated hyaluronan binding induced by inflammatory mediators in human CD14(+) peripheral blood monocytes. J. Immunol., 167, 53675374.
Capon, C., Wieruszeski, J.M., Lemoine, J., Byrd, J.C., Leffler, H., and Kim, Y.S. (1997) Sulfated lewis x determinants as a major structural motif in glycans from LS174T-HM7 human colon carcinoma mucin. J. Biol. Chem., 272, 3195731968.
Carter, S.R., Slomiany, A., Gwozdzinski, K., Liau, Y.H., and Slomiany, B.L. (1988) Enzymatic sulfation of mucus glycoprotein in gastric mucosa. Effect of ethanol. J. Biol. Chem., 263, 1197711984.
Delmotte, P., Degroote, S., Lafitte, J.J., Lamblin, G., Perini, J.M., and Roussel, P. (2001) TNF{a} increases the expression of glycosyltransferases and sulfotransferases responsible for the biosynthesis of sialylated and/or sulfated Lewis x epitopes in the human bronchial mucosa. J. Biol. Chem., 277, 424431.
Fan, Q.W., Uchimura, K., Yuzawa, Y., Matsuo, S., Mitsuoka, C., Kannagi, R., Muramatsu, H., Kadomatsu, K., and Muramatsu, T. (1999) Spatially and temporally regulated expression of N-acetylglucosamine-6-O-sulfotransferase during mouse embryogenesis. Glycobiology, 9, 947955.
Fukuda, M., Hiraoka, N., Akama, T.O., and Fukuda, M.N. (2001) Carbohydrate-modifying sulfotransferases: structure, function, and pathophysiology. J. Biol. Chem., 276, 4774747750.
Fukuta, M., Inazawa, J., Torii, T., Tsuzuki, K., Shimada, E., and Habuchi, O. (1997) Molecular cloning and characterization of human keratan sulfate Gal-6-sulfotransferase. J. Biol. Chem., 272, 3232132328.
Giordanengo, V., Limouse, M., Peyron, J.F., and Lefebvre, J.C. (1995) Lymphocytic CD43 and CD45 bear sulfate residues potentially implicated in cell to cell interactions. Eur. J. Immunol., 25, 274278.[ISI][Medline]
Girard, J.-P. and Springer, T.A. (1995) Cloning from purified high endothelial venule cells of hevin, a close relative of the antiadhesive extracellular matrix protein SPARC. Immunity, 2, 113123.[ISI][Medline]
Goso, Y. and Hotta, K. (1993) Regional differences in sulfated oligosaccharides of rat gastrointestinal mucin as detected by two-dimensional chromatography. Arch. Biochem. Biophys., 302, 212217.[CrossRef][ISI][Medline]
Habuchi, O. (2000) Diversity and functions of glycosaminoglycan sulfotransferases. Biochim. Biophys. Acta., 1474, 115127.[ISI][Medline]
Hasegawa, N., Torii, T., Kato, T., Miyajima, H., Furuhata, A., Nakayasu, K., Kanai, A., and Habuchi, O. (2000) Decreased GlcNAc 6-O-sulfotransferase activity in the cornea with macular corneal dystrophy. Invest. Ophthalmol. Vis. Sci., 41, 36703677.
Hemmerich, S. and Rosen, S.D. (2000) Carbohydrate sulfotransferases in lymphocyte homing. Glycobiology, 10, 849856.
Hemmerich, S., Butcher, E.C., and Rosen, S.D. (1994) Sulfation-dependent recognition of HEV-ligands by L-selectin and MECA 79, an adhesion-blocking mAb. J. Exp. Med., 180, 22192226.[Abstract]
Hemmerich, S., Leffler, H., and Rosen, S.D. (1995) Structure of the O-glycans in GlyCAM-1, an endothelial-derived ligand for L-selectin. J. Biol. Chem., 270, 1203512047.
Hemmerich, S., Bistrup, A., Singer, M.S., Zante, A.v., Lee, J.K., Tsay, D., Peters, M., Carminati, J.L., Brennan, T.J., Carver-Moore, K., and others. (2001a) Sulfation of L-selectin ligands by an HEV-restricted sulfotransferase regulates lymphocyte homing to lymph nodes. Immunity, 15, 237247.[ISI][Medline]
Hemmerich, S., Lee, J.K., Bhakta, S., Bistrup., A., Ruddle, N., and Rosen, S.D. (2001b) Chromosomal localization and genomic organization for the galactose/N-acetylgalactosamine/N-acetylglucosamine 6-O-sulfotransferase gene family. Glycobiology, 11, 7587.
Hiraoka, N., Petryniak, B., Nakayama, J., Tsuboi, S., Suzuki, M., Yeh, J.-C., Izawa, D., Tanaka, T., Miyasaka, M., Lowe, J.B., and Fukuda, M. (1999) A novel, high endothelial venule-specific sulfotransferase expresses 6-sulfo sialyl lewis x, an L-selectin ligand displayed by CD34. Immunity, 11, 7989.[ISI][Medline]
Hooper, L.V., Manzella, S.M., and Baenziger, J.U. (1996) From legumes to leukocytes: biological roles for sulfated carbohydrates. FASEB J., 10, 11371146.
Kanamori, A., Kojima, N., Uchimura, K., Muramatsu, T., Tamatani, T., Berndt, M.C., Kansas, G.S., and Kannagi, R. (2002) Distinct sulfation requirements of selectins disclosed using cells which support rolling mediated by all three selectins under shear flow. J. Biol. Chem., 277, 3257832586.
Kannagi, R. and Kanamori, A. (1999) Glycobiology of sialyl 6-sulfo lewis x, a new carbohydrate ligand for selectins. Trends Glycosci. Glycotechnol., 11, 329344.[ISI]
Kim, Y.S. and Gum, J.R. Jr. (1995) Diversity of mucin genes, structure, function, and expression. Gastroenterology, 109, 9991001.[ISI][Medline]
Kimura, N., Mitsuoka, C., Kanamori, A., Hiraiwa, N., Uchimura, K., Muramatsu, T., Tamatani, T., Kansas, G.S., and Kannagi, R. (1999) Reconstitution of functional L-selectin ligands on a cultured human endothelial cell line by cotransfection of alpha13 fucosyltransferase VII and newly cloned GlcNAcbeta:6-sulfotransferase cDNA. Proc. Natl Acad. Sci. USA, 96, 45304535.
Kitagawa, H., Fujita, M., Ito, N., and Sugahara, K. (2000) Molecular cloning and expression of a novel chondroitin 6-O-sulfotransferase. J. Biol. Chem., 275, 2107521080.
Lee, J.K., Bhakta, S., Rosen, S.D., and Hemmerich, S. (1999) Cloning and characterization of a mammalian N-acetylglucosamine-6-sulfotransferase that is highly restricted to intestinal tissue. Biochem. Biophys. Res. Commun., 263, 543549.[CrossRef][ISI][Medline]
Li, X. and Tedder, T.F. (1999) CHST1 and CHST2 sulfotransferases expressed by human vascular endothelial cells: cDNA cloning, expression, and chromosomal localization. Genomics, 55, 345347.[CrossRef][ISI][Medline]
Lo-Guidice, J.M., Wieruszeski, J.M., Lemoine, J., Verbert, A., Roussel, P., and Lamblin, G. (1994) Sialylation and sulfation of the carbohydrate chains in respiratory mucins from a patient with cystic fibrosis. J. Biol. Chem., 269, 1879418813.
Maiti, A., Maki, G., and Johnson, P. (1998) TNF-alpha induction of CD44-mediated leukocyte adhesion by sulfation. Science, 282, 941943.
McEver, R.P. and Cummings, R.D. (1997) Role of PSGL-1 binding to selectins in leukocyte recruitment. J. Clin. Invest., 100, S97S103.[ISI][Medline]
Mitsuoka, C., Sawada-Kasugai, M., Ando-Furui, K., Izawa, M., Nakanishi, H., Nakamura, S., Ishida, H., Kiso, M., and Kannagi, R. (1998) Identification of a major carbohydrate capping group of the L-selectin ligand on high endothelial venules in human lymph nodes as 6-sulfo sialyl Lewis X. J. Biol. Chem., 273, 1122511233.
Nakazawa, K., Takahashi, I., and Yamamoto, Y. (1998) Glycosyltransferase and sulfotransferase activities in chick corneal stromal cells before and after in vitro culture. Arch. Biochem. Biophys., 359, 269282.[CrossRef][ISI][Medline]
Ohmori, K., Kanda, K., Mitsuoka, C., Kanamori, A., Kurata-Miura, K., Sasaki, K., Nishi, T., Tamatani, T., and Kannagi, R. (2000) P- and E-selectins recognize sialyl 6-sulfo lewis X, the recently identified L-selectin ligand. Biochem. Biophys. Res. Commun., 278, 9096.[CrossRef][ISI][Medline]
Puri, K.D., Finger, E.B., Gaudernack, G., and Springer, T.A. (1995) Sialomucin CD34 is the major L-selectin ligand in human tonsil high endothelial venules. J. Cell Biol., 131, 261270.[Abstract]
Rosen, S.D. (1999) Endothelial ligands for L-selectin: from lymphocyte recirculation to allograft rejection. Am. J. Pathol., 155, 10131020.
Sassetti, C., Van Zante, M., and Rosen, S.D. (2000) Identification of endoglycan, a member of the CD34/podocalyxin family of sialomucins. J. Biol. Chem., 275, 90019010.
Seko, A., Sumiya, J., Yonezawa, S., Nagata, K., and Yamashita, K. (2000) Biochemical differences between two types of N-acetylglucosamine:6sulfotransferases in human colonic adenocarcinomas and the adjacent normal mucosa: specific expression of a GlcNAc:
6sulfotransferase in mucinous adenocarcinoma. Glycobiology, 10, 919929.
Simmons, D.L. (1993) Cloning cell surface molecules by transient expression in mammalian cells. In D.A. Hartley (ed.), Cellular interactions in development. A practical approach. IRL Press, Oxford, pp. 93127.
Spiro, R.G., Yasumoto, Y., and Bhoyroo, V. (1996) Characterization of a rat liver Golgi sulphotransferase responsible for the 6-O-sulphation of N-acetylglucosamine residues in beta-linkage to mannose: role in assembly of sialyl-galactosyl-N-acetylglucosamine 6-sulphate sequence of N-linked oligosaccharides. Biochem. J., 319, 209216.[ISI][Medline]
Tangemann, K., Bistrup, A., Hemmerich, S., and Rosen, S.D. (1999) Sulfation of an HEV-expressed ligand for L-selectin: effects on tethering and rolling of lymphocytes. J. Exp. Med., 190, 935941.
Uchimura, K., Muramatsu, H., Kadomatsu, K., Fan, Q.W., Kurosawa, N., Mitsuoka, C., Kannagi, R., Habuchi, O., and Muramatsu, T. (1998a) Molecular cloning and characterization of an N-acetylglucosamine-6-O-sulfotransferase. J. Biol. Chem., 273, 2257722583.
Uchimura, K., Muramatsu, H., Kaname, T., Ogawa, H., Yamakawa, T., Fan, Q.W., Mitsuoka, C., Kannagi, R., Habuchi, O., Yokoyama, I., and others. (1998b) Human N-acetylglucosamine-6-O-sulfotransferase involved in the biosynthesis of 6-sulfo sialyl Lewis X: molecular cloning, chromosomal mapping, and expression in various organs and tumor cells. J. Biochem. (Tokyo), 124, 670678.[Abstract]
Uchimura, K., Fasakhany, F., Kadomatsu, K., Matsukawa, T., Yamakawa, T., Kurosawa, N., and Muramatsu, T. (2000) Diversity of N-acetylglucosamine-6-O-sulfotransferases: molecular cloning of a novel enzyme with different distribution and specificities. Biochem. Biophys. Res. Commun., 274, 291296.[CrossRef][ISI][Medline]
Uchimura, K., El-Fasakhany, F.M., Hori, M., Hemmerich, S., Blink, S.E., Kansas, G.S., Kanamori, A., Kumamoto, K., Kannagi, R., and Muramatsu, T. (2002) Specificities of N-acetylglucosamine-6-O-sulfotransferases in relation to L-selectin ligand synthesis and tumor-associated enzyme expression. J. Biol. Chem., 277, 39793984.
Velcich, A., Yang, W., Heyer, J., Fragale, A., Nicholas, C., Viani, S., Kucherlapati, R., Lipkin, M., Yang, K., and Augenlicht, L. (2002) Colorectal cancer in mice genetically deficient in the mucin Muc2. Science, 295, 17261729.
Watson, S.R., Imai, Y., Fennie, C., Geoffroy, J.S., Rosen, S.D., and Lasky, L.A. (1990) A homing receptor-IgG chimera as a probe for adhesive ligands of lymph node high endothelial venules. J. Cell Biol., 110, 22212229.[Abstract]
Yeh, J.C., Hiraoka, N., Petryniak, B., Nakayama, J., Ellies, L.G., Rabuka, D., Hindsgaul, O., Marth, J.D., Lowe, J.B., and Fukuda, M. (2001) Novel sulfated lymphocyte homing receptors and their control by a Core 1 extension beta 1,3-N-acetylglucosaminyltransferase. Cell, 105, 957969.[CrossRef][ISI][Medline]