Posttranslational modification of the glycosylation inhibiting factor (GIF) gene product generates bioactive GIF.

Proceedings of the National Academy of Sciences of the United States of America ·Vol. 97 ·No. 24 ·2000-11-21 ·Pages 13251-6

Watarai H, Nozawa R, Tokunaga A, Yuyama N, Tomas M, Hinohara A, Ishizaka K, Ishii Y

Abstract

Glycosylation inhibiting factor (GIF) and macrophage migration inhibitory factor (MIF) share an identical structure gene. Here we unravel two steps of posttranslational modifications in GIF/MIF molecules in human suppressor T (Ts) cell hybridomas. Peptide mapping and MS analysis of the affinity-purified GIF from the Ts cells revealed that one modification is cysteinylation at Cys-60, and the other is phosphorylation at Ser-91. Cysteinylated GIF, but not the wild-type GIF/MIF, possessed immunosuppressive effects on the in vitro IgE antibody response and had high affinity for GIF receptors on the T helper hybridoma cells. In vitro treatment of wild-type recombinant human GIF/MIF with cystine resulted in preferential cysteinylation of Cys-60 in the molecules. The cysteinylated recombinant human GIF and the Ts hybridoma-derived cysteinylated GIF were comparable both in the affinity for the receptors and in the immunosuppressive activity. Polyclonal antibodies specific for a stretch of the amino acid sequence in alpha2-helix of GIF bound bioactive cysteinylated GIF but failed to bind wild-type GIF/MIF. These results strongly suggest that cysteinylation of Cys-60 and consequent conformational changes in the GIF/MIF molecules are responsible for the generation of GIF bioactivity.

MeSH Terms

Amino Acid Sequence Animals Cysteine/metabolism Glycosylation Humans Hybridomas Immunoglobulin E/immunology Lymphokines/chemistry,genetics,metabolism Mice Mice, Inbred BALB C Mice, Transgenic Models, Molecular Molecular Sequence Data Prostatic Secretory Proteins Protein Processing, Post-Translational Protein Structure, Secondary Receptors, Antigen, T-Cell, alpha-beta/genetics Recombinant Proteins/chemistry,metabolism Spleen/immunology T-Lymphocytes/immunology T-Lymphocytes, Regulatory/immunology

Chemicals

Lymphokines Prostatic Secretory Proteins Receptors, Antigen, T-Cell, alpha-beta Recombinant Proteins beta-microseminoprotein immunoglobulin-binding factors Immunoglobulin E Cysteine

Authors & Affiliations

8 authors, click to expand affiliations / ORCID

Watarai H

Pharmaceutical Research Laboratory, Kirin Brewery Company, Ltd., 3. Miyahara-cho, Takasaki 370-1295, Gunma, Japan.

Nozawa R

Tokunaga A

Yuyama N

Tomas M

Hinohara A

Ishizaka K

Ishii Y

References (26)

26 references, click to expand

IgE-binding factors from mouse T lymphocytes. III. Role of antigen-specific suppressor T cells in the formation of IgE-suppressive factor.
J Immunol. 1984 Dec;133(6):3266-73 PMID: 6208275
Formation of IgE-binding factors by human T-cell hybridomas.
Proc Natl Acad Sci U S A. 1984 Mar;81(5):1514-8 PMID: 6231643
Immunosuppressive effects of glycosylation inhibiting factor on the IgE and IgG antibody response.
J Immunol. 1986 May 1;136(9):3172-9 PMID: 2937840
Studies on murine IgE with monoclonal antibodies. I. Characterization of rat monoclonal anti-IgE antibodies and the use of these antibodies for determinations of serum IgE levels and for anaphylactic reactions.
Int Arch Allergy Appl Immunol. 1988;85(1):47-54 PMID: 3257463
Antigen-specific T cells that form IgE-potentiating factor, IgG-potentiating factor, and antigen-specific glycosylation-enhancing factor on antigenic stimulation.
J Immunol. 1988 Apr 15;140(8):2534-42 PMID: 3258613
I-Ad-binding peptides derived from unrelated protein antigens share a common structural motif.
J Immunol. 1988 Jul 1;141(1):45-8 PMID: 2837512
Molecular cloning of a cDNA encoding a human macrophage migration inhibitory factor.
Proc Natl Acad Sci U S A. 1989 Oct;86(19):7522-6 PMID: 2552447
Oxidized redox state of glutathione in the endoplasmic reticulum.
Science. 1992 Sep 11;257(5076):1496-502 PMID: 1523409
Molecular cloning and functional expression of a cDNA encoding glycosylation-inhibiting factor.
Proc Natl Acad Sci U S A. 1993 Nov 1;90(21):10056-60 PMID: 8234256
Cloning the human gene for macrophage migration inhibitory factor (MIF).
Genomics. 1994 Jan 1;19(1):48-51 PMID: 8188240
The macrophage is an important and previously unrecognized source of macrophage migration inhibitory factor.
J Exp Med. 1994 Jun 1;179(6):1895-902 PMID: 8195715
Naive T cells can mediate delayed-type hypersensitivity response in T cell receptor transgenic mice.
Eur J Immunol. 1994 Jul;24(7):1512-6 PMID: 8026513
Purification, bioactivity, and secondary structure analysis of mouse and human macrophage migration inhibitory factor (MIF).
Biochemistry. 1994 Nov 29;33(47):14144-55 PMID: 7947826
Requirement of posttranslational modifications for the generation of biologic activity of glycosylation-inhibiting factor.
Proc Natl Acad Sci U S A. 1994 Nov 8;91(23):11227-31 PMID: 7972039
Association of the "major histocompatibility complex subregion" I-J determinant with bioactive glycosylation-inhibiting factor.
Proc Natl Acad Sci U S A. 1995 Sep 26;92(20):9196-200 PMID: 7568100
Biochemical characterization of antigen-specific glycosylation-inhibiting factor from antigen-specific suppressor T cells. I. Identification of a 55-kilodalton glycosylation-inhibiting factor peptide with TCR alpha-chain determinant.
J Immunol. 1996 Mar 1;156(5):1728-34 PMID: 8596020
Biochemical characterization of antigen-specific glycosylation-inhibiting factor from antigen-specific suppressor T cells. II. The 55-kDa glycosylation-inhibiting factor peptide is a derivative of TCR alpha-chain and a subunit of antigen-specific glycosylation-inhibiting factor.
J Immunol. 1996 Mar 1;156(5):1735-42 PMID: 8596021
The crystal structure of human glycosylation-inhibiting factor is a trimeric barrel with three 6-stranded beta-sheets.
Proc Natl Acad Sci U S A. 1996 Apr 2;93(7):3007-10 PMID: 8610159
Conversion of inactive glycosylation inhibiting factor to bioactive derivatives by modification of a SH group.
Proc Natl Acad Sci U S A. 1997 Jan 7;94(1):202-7 PMID: 8990186
High-affinity binding of bioactive glycosylation-inhibiting factor to antigen-primed T cells and natural killer cells.
Proc Natl Acad Sci U S A. 1997 May 13;94(10):5278-83 PMID: 9144228
Immunosuppressive activities of recombinant glycosylation-inhibiting factor mutants.
J Immunol. 1999 Jan 1;162(1):195-202 PMID: 9886386
Target cells for an immunosuppressive cytokine, glycosylation-inhibiting factor.
Int Immunol. 1999 Jul;11(7):1149-56 PMID: 10383948
Definition and transfer of a serological epitope specific for peptide-empty forms of MHC class I.
Int Immunol. 1999 Dec;11(12):1897-906 PMID: 10590255
Stimulus-specific interaction between activator-coactivator cognates revealed with a novel complex-specific antiserum.
J Biol Chem. 2000 Mar 24;275(12):8263-6 PMID: 10722651
Monoclonal dinitrophenyl-specific murine IgE antibody: preparation, isolation, and characterization.
J Immunol. 1980 Jun;124(6):2728-37 PMID: 7373045
Regulation of IgE synthesis.
Annu Rev Immunol. 1984;2:159-82 PMID: 6085750

Article Info

Journal

Proceedings of the National Academy of Sciences of the United States of America

Abbr.

Proc Natl Acad Sci U S A

ISSN

0027-8424

DOI

10.1073/pnas.230445397

Published

2000-11-21

Pages

13251-6

Language

English

Region

United States

NLM ID

7505876

PMCID

PMC27211

Subset

External Links

PubMed source DOI: 10.1073/pnas.230445397 PMC Full Text View journal details

Analysis Services

Posttranslational modification of the glycosylation inhibiting factor (GIF) gene product generates bioactive GIF.

Contact

WeChat Official Account