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Galectin-1 induced activation of the apoptotic death-receptor pathway in human Jurkat T lymphocytes

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Abstract

Galectin-1 (gal-1), a member of the family of β-galactoside binding proteins, participates in several biological processes such as immunomodulation, cell adhesion, regulation of cell growth and apoptosis. The aim of this study was to investigate whether gal-1 interferes with the Fas (Apo-1/CD95)-associated apoptosis cascade in the T-cell lines Jurkat and MOLT-4. Gal-1 and an Apo-1 monoclonal antibody (mAb) induced DNA-fragmentation in Jurkat T-cells whereas MOLT-4 cells were resistant. Gal-1 stimulated DNA-fragmentation could be efficiently inhibited by caspase-8 inhibitor II (Z-IETD-FMK) and a neutralizing Fas mAb. Fas could be identified as a target for gal-1 recognition as demonstrated by immunofluorescence staining, binding of the receptor glycoprotein to immobilized gal-1 and analyses by immunoblotting as well as by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Gal-1 stimulates the activation and proteolytic processing of procaspase-8 and downstream procaspase-3 in Jurkat-T cells. Inhibition of gal-1 induced procaspase-8 activation by a neutralizing Fas mAb strongly suggests that gal-1 recognition of Fas is associated with caspase-8 activation. Our data provide the first experimental evidence for targeting of gal-1 to glycotopes on Fas and the subsequent activation of the apoptotic death-receptor pathway.

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Abbreviations

Bp:

Base pair

BSA:

Bovine serum albumin

CD:

Cluster of differentiation

Chaps:

3-[(3-Cholamidopropyl)dimethylammonio]-1-propanesulfonate

DTT:

Dithiothreitol

ECL:

Enhanced chemiluminescence

EDTA:

Ethylene-diaminetetraacetic acid

EGTA:

Ethylene glycol-bis(2-aminoethylether)-N,N,N′,N′-tetraacetic acid

FITC:

Fluorescein isothiocyanate

gal-1:

Galectin-1

HEPES:

N-2-hydroxyethyl-piperazine-N-2-ethanesulfonic acid

IgG:

Immunoglobulin G

HRP:

Horseradish peroxidase

kDa:

Kilo Dalton

LC-MS/MS:

Liquid chromatography-tandem mass spectrometry

mAb:

Monoclonal antibody

NHS:

N-hydroxysuccinimide

NP-40:

Nonidet P-40

pAb:

Polyclonal antibody

PIPES:

Piperazine-N,N´-bis(2-ethanesulfonic) acid

PAGE:

Polyacrylamide gel electrophoresis

PBS:

Phosphate-buffered saline

PI:

Propidium iodide

PtdSer:

Phosphatidylserine

SDS:

Sodium dodecyl sulphate

TBS:

Tris-buffered saline

Tris:

Tris(hydroxymethyl) aminomethane

T:

Tween 20

TUNEL:

Terminal deoxynucleotidyl transferase dUTP nick end labelling

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Acknowledgements

This work was supported by a grant from the Deutsche Forschungsgemeinschaft (DFG, WA 1771/1-1 and JE 181/7-1). We gratefully acknowledge the excellent technical assistance of Mrs. G. Gaede. We thank M. Schümann for help with the MS measurements.

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Authors and Affiliations

  1. Medical Faculty, Institute of Medical Biochemistry and Molecular Biology, University of Rostock, Schillingallee 70, 18057, Rostock, Germany

    Bettina Brandt, Tom Büchse, Ehab Fathi Abou-Eladab, Markus Tiedge & Hermann Walzel

  2. Faculty of Specific Education, Mansoura University, Mansoura, 35516, New Damietta City, Egypt

    Ehab Fathi Abou-Eladab

  3. Leibniz Institute of Molecular Pharmacology, Robert-Rössle-Str. 10, 13125, Berlin, Germany

    Eberhard Krause

  4. First Department of Obstetrics and Gynaecology, Ludwig Maximilians University of Munich, Maistrasse 11, 80337, Munich, Germany

    Udo Jeschke

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  1. Bettina Brandt
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Correspondence to Udo Jeschke.

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Brandt, B., Büchse, T., Abou-Eladab, E.F. et al. Galectin-1 induced activation of the apoptotic death-receptor pathway in human Jurkat T lymphocytes. Histochem Cell Biol 129, 599–609 (2008). https://doi.org/10.1007/s00418-008-0395-x

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