title: A field-proven yeast two-hybrid protocol used to identify coronavirus-host protein-protein interactions.
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.
authors.affiliation: Unité de Génomique Virale etVaccination, Institut Pasteur, Paris, France, pierre-olivier.vidalain@pasteur.fr.
authors.forename: Pierre-Olivier
authors.initials: PO
authors.lastname: Vidalain
authors.forename: Yves
authors.initials: Y
authors.lastname: Jacob
authors.forename: Marne C
authors.initials: MC
authors.lastname: Hagemeijer
authors.forename: Louis M
authors.initials: LM
authors.lastname: Jones
authors.forename: Grégory
authors.initials: G
authors.lastname: Neveu
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Roussarie
authors.forename: Peter J M
authors.initials: PJ
authors.lastname: Rottier
authors.forename: Frédéric
authors.initials: F
authors.lastname: Tangy
authors.forename: Cornelis A M
authors.initials: CA
authors.lastname: de Haan
mesh_terms: D000818:Animals; D054884:Host-Pathogen Interactions; D051379:Mice; D006517:Murine hepatitis virus / Q000502:physiology*; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D020798:Two-Hybrid System Techniques; D014764:Viral Proteins / Q000378:metabolism*; D053585:Virus Attachment
chemical_list: D009419:Nerve Tissue Proteins; D014764:Viral Proteins