Induction of 6-sulfated glycans with cell adhesion activity via T-bet and GATA-3 in human helper T cells

Keiichiro Sakuma, Guoyun Chen, Masahiro Aoki, Reiji Kannagi

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

Background: Cell surface 6-sulfated glycans play important roles in various immunological events through cell-to-cell interactions. The 6-sulfation process is mediated by 6-sulfotransferase family isoenzymes. We previously demonstrated that GlcNAc6ST-1, one of the isoenzyme genes, is induced by GATA-3 and NF-κB in human helper T (Th) cells. However, transcriptional regulation of HEC-GlcNAc6ST, another isoenzyme important in Th cells, remains unclear. Methods: 5′-RACE analysis, chromatin immunoprecipitation, and reporter assays were performed to reveal transcriptional regulation of HEC-GlcNAc6ST. RNA-knockdown and forced expression experiments were performed to demonstrate the contribution of HEC-GlcNAc6ST to the 6-sulfated glycan expression. Results: We identified potential binding sites of Sp1, T-bet, and GATA-3 in the HEC-GlcNAc6ST promoter. Reporter assays indicated that transfection of Sp1 enhanced the activity, whereas mithramycin A, an Sp1-specific inhibitor, repressed it. Transfection of T-bet increased the activity, which was inhibited by introducing a mutation into the potential T-bet binding site. GATA-3 alone could not elevate the activity, although co-transfection of protein kinase A, which is known to enhance IL-5 transcription in Th2 cells through phosphorylation of GATA-3, caused elevation. RNA-knockdown and forced expression of HEC-GlcNAc6ST in Jurkat cells down- and up-regulated α2,6-sialylated 6-sulfo N-acetyllactosamine, a preferential ligand for B-cell-specific CD22 antigen, respectively. From these results, we concluded that T-bet and GATA-3 as well as Sp1 control the expression of glycan with cell-adhesion activity by regulating HEC-GlcNAc6ST transcription in Th cells. General significance: These results may provide a clue to biological regulation of Th-cell interaction with selectins and other carbohydrate-recognition molecules by T-bet and GATA-3.

Original languageEnglish (US)
Pages (from-to)841-848
Number of pages8
JournalBiochimica et Biophysica Acta - General Subjects
Volume1820
Issue number7
DOIs
StatePublished - Jul 1 2012

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T-cells
Cell adhesion
Helper-Inducer T-Lymphocytes
Cell Adhesion
Isoenzymes
Transfection
Polysaccharides
Transcription
Cell Communication
Sialic Acid Binding Ig-like Lectin 2
Assays
Binding Sites
RNA
Sulfotransferases
Selectins
Th2 Cells
Phosphorylation
Jurkat Cells
Chromatin Immunoprecipitation
Interleukin-5

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Induction of 6-sulfated glycans with cell adhesion activity via T-bet and GATA-3 in human helper T cells. / Sakuma, Keiichiro; Chen, Guoyun; Aoki, Masahiro; Kannagi, Reiji.

In: Biochimica et Biophysica Acta - General Subjects, Vol. 1820, No. 7, 01.07.2012, p. 841-848.

Research output: Contribution to journalArticle

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T1 - Induction of 6-sulfated glycans with cell adhesion activity via T-bet and GATA-3 in human helper T cells

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AU - Aoki, Masahiro

AU - Kannagi, Reiji

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N2 - Background: Cell surface 6-sulfated glycans play important roles in various immunological events through cell-to-cell interactions. The 6-sulfation process is mediated by 6-sulfotransferase family isoenzymes. We previously demonstrated that GlcNAc6ST-1, one of the isoenzyme genes, is induced by GATA-3 and NF-κB in human helper T (Th) cells. However, transcriptional regulation of HEC-GlcNAc6ST, another isoenzyme important in Th cells, remains unclear. Methods: 5′-RACE analysis, chromatin immunoprecipitation, and reporter assays were performed to reveal transcriptional regulation of HEC-GlcNAc6ST. RNA-knockdown and forced expression experiments were performed to demonstrate the contribution of HEC-GlcNAc6ST to the 6-sulfated glycan expression. Results: We identified potential binding sites of Sp1, T-bet, and GATA-3 in the HEC-GlcNAc6ST promoter. Reporter assays indicated that transfection of Sp1 enhanced the activity, whereas mithramycin A, an Sp1-specific inhibitor, repressed it. Transfection of T-bet increased the activity, which was inhibited by introducing a mutation into the potential T-bet binding site. GATA-3 alone could not elevate the activity, although co-transfection of protein kinase A, which is known to enhance IL-5 transcription in Th2 cells through phosphorylation of GATA-3, caused elevation. RNA-knockdown and forced expression of HEC-GlcNAc6ST in Jurkat cells down- and up-regulated α2,6-sialylated 6-sulfo N-acetyllactosamine, a preferential ligand for B-cell-specific CD22 antigen, respectively. From these results, we concluded that T-bet and GATA-3 as well as Sp1 control the expression of glycan with cell-adhesion activity by regulating HEC-GlcNAc6ST transcription in Th cells. General significance: These results may provide a clue to biological regulation of Th-cell interaction with selectins and other carbohydrate-recognition molecules by T-bet and GATA-3.

AB - Background: Cell surface 6-sulfated glycans play important roles in various immunological events through cell-to-cell interactions. The 6-sulfation process is mediated by 6-sulfotransferase family isoenzymes. We previously demonstrated that GlcNAc6ST-1, one of the isoenzyme genes, is induced by GATA-3 and NF-κB in human helper T (Th) cells. However, transcriptional regulation of HEC-GlcNAc6ST, another isoenzyme important in Th cells, remains unclear. Methods: 5′-RACE analysis, chromatin immunoprecipitation, and reporter assays were performed to reveal transcriptional regulation of HEC-GlcNAc6ST. RNA-knockdown and forced expression experiments were performed to demonstrate the contribution of HEC-GlcNAc6ST to the 6-sulfated glycan expression. Results: We identified potential binding sites of Sp1, T-bet, and GATA-3 in the HEC-GlcNAc6ST promoter. Reporter assays indicated that transfection of Sp1 enhanced the activity, whereas mithramycin A, an Sp1-specific inhibitor, repressed it. Transfection of T-bet increased the activity, which was inhibited by introducing a mutation into the potential T-bet binding site. GATA-3 alone could not elevate the activity, although co-transfection of protein kinase A, which is known to enhance IL-5 transcription in Th2 cells through phosphorylation of GATA-3, caused elevation. RNA-knockdown and forced expression of HEC-GlcNAc6ST in Jurkat cells down- and up-regulated α2,6-sialylated 6-sulfo N-acetyllactosamine, a preferential ligand for B-cell-specific CD22 antigen, respectively. From these results, we concluded that T-bet and GATA-3 as well as Sp1 control the expression of glycan with cell-adhesion activity by regulating HEC-GlcNAc6ST transcription in Th cells. General significance: These results may provide a clue to biological regulation of Th-cell interaction with selectins and other carbohydrate-recognition molecules by T-bet and GATA-3.

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