title: E4orf6 variants with separate abilities to augment adenovirus replication and direct nuclear localization of the E1B 55-kilodalton protein.
abstract: The E4orf6 protein of group C adenovirus is an oncoprotein that, in association with the E1B 55-kDa protein and by E1B-independent means, promotes virus replication. An arginine-faced amphipathic alpha-helix in the E4orf6 protein is required for the E4orf6 protein to direct nuclear localization of the E1B 55-kDa protein and to enhance replication of an E4 deletion virus. In this study, E4orf6 protein variants containing arginine substitutions in the amphipathic alpha-helix were analyzed. Two of the six arginine residues within the alpha-helix, arginine-241 and arginine-243, were critical for directing nuclear localization of the E1B 55-kDa protein. The four remaining arginine residues appear to provide a net positive charge for the E4orf6 protein to direct nuclear localization of the E1B 55-kDa protein. The molecular determinants of the arginine-faced amphipathic alpha-helix that were required for the functional interaction between the E4orf6 and E1B 55-kDa proteins seen in the transfected cell differed from those required to support a productive infection. Several E4orf6 protein variants with arginine-to-glutamic acid substitutions that failed to direct nuclear localization of the E1B 55-kDa protein restored replication of an E4 deletion virus. Additionally, a variant containing an arginine-to-alanine substitution at position 243 that directed nuclear localization of the E1B 55-kDa protein failed to enhance virus replication. These results indicate that the ability of the E4orf6 protein to relocalize the E1B 55-kDa protein to the nucleus can be separated from the ability of the E4orf6 protein to support a productive infection.
authors.affiliation: Department of Microbiology and Immunology, School of Medicine, Wake Forest University, Winston-Salem, North Carolina 27157-1064, USA.
authors.forename: Joseph S
authors.initials: JS
authors.lastname: Orlando
authors.forename: David A
authors.initials: DA
authors.lastname: Ornelles
mesh_terms: D017501:Adenovirus E1B Proteins / Q000235:genetics / Q000378:metabolism*; D017504:Adenovirus E4 Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000260:Adenoviruses, Human / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine / Q000235:genetics / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D017124:Conserved Sequence; D014644:Genetic Variation; D006367:HeLa Cells; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D017386:Sequence Homology, Amino Acid; D014162:Transfection; D014779:Virus Replication / Q000502:physiology*
chemical_list: D017501:Adenovirus E1B Proteins; D017504:Adenovirus E4 Proteins; D001120:Arginine