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Coronaviruses pp 213-229 | Cite as

A Field-Proven Yeast Two-Hybrid Protocol Used to Identify Coronavirus–Host Protein–Protein Interactions

  • Pierre-Olivier Vidalain
  • Yves Jacob
  • Marne C. Hagemeijer
  • Louis M. Jones
  • Grégory Neveu
  • Jean-Pierre Roussarie
  • Peter J. M. Rottier
  • Frédéric Tangy
  • Cornelis A. M. de Haan
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1282)

Abstract

Over the last 2 decades, yeast two-hybrid became an invaluable technique to decipher protein–protein interaction networks. In the field of virology, it has proven instrumental to identify virus–host interactions that are involved in viral embezzlement of cellular functions and inhibition of immune mechanisms. Here, we present a yeast two-hybrid protocol that has been used in our laboratory since 2006 to search for cellular partners of more than 300 viral proteins. Our aim was to develop a robust and straightforward pipeline, which minimizes false-positive interactions with a decent coverage of target cDNA libraries, and only requires a minimum of equipment. We also discuss reasons that motivated our technical choices and compromises that had to be made. This protocol has been used to screen most non-structural proteins of murine hepatitis virus (MHV), a member of betacoronavirus genus, against a mouse brain cDNA library. Typical results were obtained and are presented in this report.

Key words

Murine hepatitis virus Host–pathogen interactions Yeast two-hybrid Interactomics Proteomics 

Notes

Acknowledgment

This work was supported by Institut Pasteur and CNRS (Centre National de la Recherche Scientifique).

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Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Pierre-Olivier Vidalain
    • 1
    • 2
  • Yves Jacob
    • 3
    • 7
    • 8
  • Marne C. Hagemeijer
    • 5
    • 9
  • Louis M. Jones
    • 6
  • Grégory Neveu
    • 12
    • 13
    • 14
    • 15
  • Jean-Pierre Roussarie
    • 4
    • 10
    • 11
  • Peter J. M. Rottier
    • 4
  • Frédéric Tangy
    • 1
    • 2
  • Cornelis A. M. de Haan
    • 4
  1. 1.Unité de Génomique Virale etVaccinationInstitut PasteurParisFrance
  2. 2.CNRS, UMR3569ParisFrance
  3. 3.Unité de Génétique Moléculaire des Virus à ARN, Département de VirologieInstitut PasteurParisFrance
  4. 4.Laboratory of Molecular and Cellular NeuroscienceThe Rockefeller UniversityNew York USA
  5. 5.Virology Division, Department of Infectious Diseases & Immunology, Faculty of Veterinary MedicineUtrecht UniversityUtrechtThe Netherlands
  6. 6.Centre d’Informatique pour laBiologieInstitut PasteurParisFrance
  7. 7.CNRS, UMR3569ParisFrance
  8. 8.EA302Université Paris Diderot, Sorbonne Paris CitéParisFrance
  9. 9.Laboratory of Host-Pathogen Dynamics, Cell Biology and Physiology Center (CBPC)National Heart, Lung, and Blood Institute (NHLBI), National Institutes of HealthBethesdaUSA
  10. 10.Unité de Génomique Virale etVaccinationInstitut PasteurParisFrance
  11. 11.CNRS, UMR3569ParisFrance
  12. 12.Division of Infectious Diseases and Geographic Medicine, Department of MedicineStanford University School of MedicineStanfordUSA
  13. 13.Unité de Génomique Virale etVaccinationInstitut PasteurParisFrance
  14. 14.CNRS, UMR3569ParisFrance
  15. 15.Department of Microbiology and ImmunologyStanford University School of MedicineStanfordUSA

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