Bacteriophage P4 Vis protein is needed for prophage excision.
Upon infection of its host Escherichia coli, satellite bacteriophage P4 can integrate its genome into the bacterial chromosome by Int-mediated site-specific recombination between the attP and the attB sites. The opposite event, excision, may either occur spontaneously or be induced by a superinfecting P2 helper phage. In this work, we demonstrate that the product of the P4 vis gene, a regulator of the P4 late promoters P(LL) and P(sid), is needed for prophage excision. This conclusion is supported by the following evidence: (i) P4 mutants carrying either a frameshift mutation or a deletion of the vis gene were unable to excise both spontaneously or upon P2 phage superinfection; (ii) expression of the Vis protein from a plasmid induced P4 prophage excision; (iii) excision depended on a functional integrase (Int) protein, thus suggesting that Vis is involved in the formation of the excision complex, rather than in the excision recombination event per se; (iv) Vis protein bound P4 DNA in the attP region at two distinct boxes (Box I and Box II), located between the int gene and the attP core region, and caused bending of the bound DNA. Furthermore, we mapped by primer extension the 5' end of the int transcript and found that ectopic expression of Vis reduced its signal intensity, suggesting that Vis is also involved in negative regulation of the int promoter.
Calì
Simona
S
Dipartimento di Scienze Biomolecolari e Biotecnologie, Università degli Studi di Milano, 20133 Milan, Italy.
Spoldi
Elena
E
Piazzolla
Daniela
D
Dodd
Ian B
IB
Forti
Francesca
F
Dehò
Gianni
G
Ghisotti
Daniela
D
D000595:Amino Acid Sequence; D001287:Attachment Sites, Microbiological / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D003090:Coliphages / Q000502:physiology*; D016384:Consensus Sequence; D004279:DNA, Viral / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D019426:Integrases / Q000235:genetics; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D014764:Viral Proteins / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D014775:Virus Activation
D004279:DNA, Viral; D004268:DNA-Binding Proteins; D014764:Viral Proteins; C098826:Vis protein, Bacteriophage P4; D019426:Integrases