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Apoptotic cells selectively uptake minor glycoforms of vitronectin from serum

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Abstract

Apoptosis profoundly alters the carbohydrate layer coating the membrane of eukaryotic cells. Previously we showed that apoptotic cells became reactive with the α2,6-sialyl-specific lectin from Sambucus nigra agglutinin (SNA), regardless of their histological origin and the nature of the apoptotic stimulus. Here we reveal the basis of the phenomenon by showing that in apoptotic cancer cell lines SNA reactivity was mainly associated with a 67 kDa glycoprotein which we identified by MALDI-TOF/TOF and immunoblot analysis as bovine vitronectin (bVN). bVN was neither present in non-apoptotic cells, nor in cells induced to apoptosis in serum-free medium, indicating that its uptake from the cell culture serum occurred only during apoptosis. The bVN molecules associated with apoptotic cancer cell lines represented minor isoforms, lacking the carboxyterminal sequence and paradoxically containing a few α2,6-linked sialic acid residues. Despite their poor α2,6-sialylation, these bVN molecules were sufficient to turn apoptotic cells to SNA reactivity, which is a late apoptotic event occurring in cells positive to both annexin-V and propidium iodide. Unlike in cancer cell lines, the major bVN form taken up by apoptotic neutrophils and mononuclear cells was a 80 kDa form. In apoptotic SW948 cells we also detected the α2,6-sialylated forms of the stress-70 mitochondrial precursor (mortalin) and of tubulin-β2C. These data indicate that the acquisition of vitronectin isoforms from the environment is a general, although cell specific phenomenon, potentially playing an important role in post-apoptotic events and that the α2,6-sialylation of intracellular proteins is a new kind of posttranslational modification associated with apoptosis.

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Abbreviations

An-V:

Annexin-V

bVN:

Bovine vitronectin

FCS:

Fetal calf serum

IEF:

Isoelectrofocusing

PARP:

Poly (ADP-ribose) polymerase

PBMC:

Peripheral blood mononuclear cells

PI:

Propidium iodide

PMN:

Polymorphonuclear neutrophils

PNGase F:

Peptide N-glycanase F

SNA:

Sambucus nigra agglutinin

TPEN:

N,N,N′,N′-tetrakis-(2-pyridylmethyl)ethylenediamine

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Acknowledgments

We thank Dr. M.C. Betts for the critical reading of the manuscript.

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Authors and Affiliations

  1. Department of Experimental, Diagnostic and Specialty Medicine, DIMES, University of Bologna, Via S. Giacomo 14, 40126, Bologna, Italy

    Nadia Malagolini, Mariangela Catera, Mariella Chiricolo & Fabio Dall’Olio

  2. Institute of Molecular Pathology and Immunology, University of Porto-IPATIMUP, Porto, Portugal

    Hugo Osorio & Celso A. Reis

  3. Faculty of Medicine, University of Porto, Porto, Portugal

    Hugo Osorio & Celso A. Reis

  4. Institute of Biomedical Sciences of Abel Salazar (ICBAS), 4200-465, Porto, Portugal

    Celso A. Reis

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  1. Nadia Malagolini
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  2. Mariangela Catera
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  3. Hugo Osorio
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  4. Celso A. Reis
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Correspondence to Fabio Dall’Olio.

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Malagolini, N., Catera, M., Osorio, H. et al. Apoptotic cells selectively uptake minor glycoforms of vitronectin from serum. Apoptosis 18, 373–384 (2013). https://doi.org/10.1007/s10495-013-0812-z

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