Cell propagation of cholera toxin CTA ADP-ribosylating factor by exosome mediated transfer

Cristiana Zanetti, Angelo Gallina, Alessia Fabbri, Sofia Parisi, Angela Palermo, Katia Fecchi, Zaira Boussadia, Maria Carollo, Mario Falchi, Luca Pasquini, Maria Luisa Fiani, Massimo Sargiacomo

Research output: Contribution to journalArticle

1 Citation (Scopus)

Abstract

In this study, we report how the cholera toxin (CT) A subunit (CTA), the enzyme moiety responsible for signaling alteration in host cells, enters the exosomal pathway, secretes extracellularly, transmits itself to a cell population. The first evidence for long-term transmission of CT’s toxic effect via extracellular vesicles was obtained in Chinese hamster ovary (CHO) cells. To follow the CT intracellular route towards exosome secretion, we used a novel strategy for generating metabolically-labeled fluorescent exosomes that can be counted by flow cytometry assay (FACS) and characterized. Our results clearly show the association of CT with exosomes, together with the heat shock protein 90 (HSP90) and Protein Disulfide Isomerase (PDI) molecules, proteins required for translocation of CTA across the ER membrane into the cytoplasm. Confocal microscopy showed direct internalization of CT containing fluorescent exo into CHO cells coupled with morphological changes in the recipient cells that are characteristic of CT action. Moreover, Me665 cells treated with CT-containing exosomes showed an increase in Adenosine 3’,5’-Cyclic Monophosphate (cAMP) level, reaching levels comparable to those seen in cells exposed directly to CT. Our results prompt the idea that CT can exploit an exosome-mediated cell communication pathway to extend its pathophysiological action beyond an initial host cell, into a multitude of cells. This finding could have implications for cholera disease pathogenesis and epidemiology.

Original languageEnglish
Article number1521
JournalInternational Journal of Molecular Sciences
Volume19
Issue number5
DOIs
Publication statusPublished - May 19 2018

Fingerprint

cholera
Exosomes
Administrative data processing
adenosine diphosphate
Cholera Toxin
Adenosine Diphosphate
Proteins
propagation
cells
Cells
Epidemiology
Flow cytometry
Confocal microscopy
Assays
ovaries
Enzymes
hamsters
Association reactions
Cricetulus
proteins

Keywords

  • Caveolin-1
  • Cholera toxin
  • Endocytic pathway
  • Exosomes
  • Monosialganglioside GM1

ASJC Scopus subject areas

  • Catalysis
  • Molecular Biology
  • Spectroscopy
  • Computer Science Applications
  • Physical and Theoretical Chemistry
  • Organic Chemistry
  • Inorganic Chemistry

Cite this

Cell propagation of cholera toxin CTA ADP-ribosylating factor by exosome mediated transfer. / Zanetti, Cristiana; Gallina, Angelo; Fabbri, Alessia; Parisi, Sofia; Palermo, Angela; Fecchi, Katia; Boussadia, Zaira; Carollo, Maria; Falchi, Mario; Pasquini, Luca; Fiani, Maria Luisa; Sargiacomo, Massimo.

In: International Journal of Molecular Sciences, Vol. 19, No. 5, 1521, 19.05.2018.

Research output: Contribution to journalArticle

@article{4d028152a29f4d5a9863648443f0f653,
title = "Cell propagation of cholera toxin CTA ADP-ribosylating factor by exosome mediated transfer",
abstract = "In this study, we report how the cholera toxin (CT) A subunit (CTA), the enzyme moiety responsible for signaling alteration in host cells, enters the exosomal pathway, secretes extracellularly, transmits itself to a cell population. The first evidence for long-term transmission of CT’s toxic effect via extracellular vesicles was obtained in Chinese hamster ovary (CHO) cells. To follow the CT intracellular route towards exosome secretion, we used a novel strategy for generating metabolically-labeled fluorescent exosomes that can be counted by flow cytometry assay (FACS) and characterized. Our results clearly show the association of CT with exosomes, together with the heat shock protein 90 (HSP90) and Protein Disulfide Isomerase (PDI) molecules, proteins required for translocation of CTA across the ER membrane into the cytoplasm. Confocal microscopy showed direct internalization of CT containing fluorescent exo into CHO cells coupled with morphological changes in the recipient cells that are characteristic of CT action. Moreover, Me665 cells treated with CT-containing exosomes showed an increase in Adenosine 3’,5’-Cyclic Monophosphate (cAMP) level, reaching levels comparable to those seen in cells exposed directly to CT. Our results prompt the idea that CT can exploit an exosome-mediated cell communication pathway to extend its pathophysiological action beyond an initial host cell, into a multitude of cells. This finding could have implications for cholera disease pathogenesis and epidemiology.",
keywords = "Caveolin-1, Cholera toxin, Endocytic pathway, Exosomes, Monosialganglioside GM1",
author = "Cristiana Zanetti and Angelo Gallina and Alessia Fabbri and Sofia Parisi and Angela Palermo and Katia Fecchi and Zaira Boussadia and Maria Carollo and Mario Falchi and Luca Pasquini and Fiani, {Maria Luisa} and Massimo Sargiacomo",
year = "2018",
month = "5",
day = "19",
doi = "10.3390/ijms19051521",
language = "English",
volume = "19",
journal = "International Journal of Molecular Sciences",
issn = "1661-6596",
publisher = "MDPI AG",
number = "5",

}

TY - JOUR

T1 - Cell propagation of cholera toxin CTA ADP-ribosylating factor by exosome mediated transfer

AU - Zanetti, Cristiana

AU - Gallina, Angelo

AU - Fabbri, Alessia

AU - Parisi, Sofia

AU - Palermo, Angela

AU - Fecchi, Katia

AU - Boussadia, Zaira

AU - Carollo, Maria

AU - Falchi, Mario

AU - Pasquini, Luca

AU - Fiani, Maria Luisa

AU - Sargiacomo, Massimo

PY - 2018/5/19

Y1 - 2018/5/19

N2 - In this study, we report how the cholera toxin (CT) A subunit (CTA), the enzyme moiety responsible for signaling alteration in host cells, enters the exosomal pathway, secretes extracellularly, transmits itself to a cell population. The first evidence for long-term transmission of CT’s toxic effect via extracellular vesicles was obtained in Chinese hamster ovary (CHO) cells. To follow the CT intracellular route towards exosome secretion, we used a novel strategy for generating metabolically-labeled fluorescent exosomes that can be counted by flow cytometry assay (FACS) and characterized. Our results clearly show the association of CT with exosomes, together with the heat shock protein 90 (HSP90) and Protein Disulfide Isomerase (PDI) molecules, proteins required for translocation of CTA across the ER membrane into the cytoplasm. Confocal microscopy showed direct internalization of CT containing fluorescent exo into CHO cells coupled with morphological changes in the recipient cells that are characteristic of CT action. Moreover, Me665 cells treated with CT-containing exosomes showed an increase in Adenosine 3’,5’-Cyclic Monophosphate (cAMP) level, reaching levels comparable to those seen in cells exposed directly to CT. Our results prompt the idea that CT can exploit an exosome-mediated cell communication pathway to extend its pathophysiological action beyond an initial host cell, into a multitude of cells. This finding could have implications for cholera disease pathogenesis and epidemiology.

AB - In this study, we report how the cholera toxin (CT) A subunit (CTA), the enzyme moiety responsible for signaling alteration in host cells, enters the exosomal pathway, secretes extracellularly, transmits itself to a cell population. The first evidence for long-term transmission of CT’s toxic effect via extracellular vesicles was obtained in Chinese hamster ovary (CHO) cells. To follow the CT intracellular route towards exosome secretion, we used a novel strategy for generating metabolically-labeled fluorescent exosomes that can be counted by flow cytometry assay (FACS) and characterized. Our results clearly show the association of CT with exosomes, together with the heat shock protein 90 (HSP90) and Protein Disulfide Isomerase (PDI) molecules, proteins required for translocation of CTA across the ER membrane into the cytoplasm. Confocal microscopy showed direct internalization of CT containing fluorescent exo into CHO cells coupled with morphological changes in the recipient cells that are characteristic of CT action. Moreover, Me665 cells treated with CT-containing exosomes showed an increase in Adenosine 3’,5’-Cyclic Monophosphate (cAMP) level, reaching levels comparable to those seen in cells exposed directly to CT. Our results prompt the idea that CT can exploit an exosome-mediated cell communication pathway to extend its pathophysiological action beyond an initial host cell, into a multitude of cells. This finding could have implications for cholera disease pathogenesis and epidemiology.

KW - Caveolin-1

KW - Cholera toxin

KW - Endocytic pathway

KW - Exosomes

KW - Monosialganglioside GM1

UR - http://www.scopus.com/inward/record.url?scp=85047331864&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85047331864&partnerID=8YFLogxK

U2 - 10.3390/ijms19051521

DO - 10.3390/ijms19051521

M3 - Article

C2 - 29783743

AN - SCOPUS:85047331864

VL - 19

JO - International Journal of Molecular Sciences

JF - International Journal of Molecular Sciences

SN - 1661-6596

IS - 5

M1 - 1521

ER -