Transcription activation by phage phi29 protein p4 is mediated by interaction with the alpha subunit of Bacillus subtilis RNA polymerase.
Regulatory protein p4 from Bacillus subtilis phage phi29 activates transcription from the viral late A3 promoter by stabilizing sigmaA-RNA polymerase at the promoter as a closed complex. Activation requires an interaction between protein p4 and RNA polymerase mediated by the protein p4 carboxyl-end, mainly through residue Arg-120. We have obtained derivatives of B. subtilis RNA polymerase alpha subunit with serial deletions at the carboxyl-end and reconstituted RNA polymerase holoenzymes harboring the mutant alpha subunits. Protein p4 promoted the binding of purified B. subtilis RNA polymerase alpha subunit to the A3 promoter in a cooperative way. Binding was abolished by deletion of the last 15 amino acids of the alpha subunit. Reconstituted RNA polymerases with deletions of 15 to 59 residues at the alpha subunit carboxyl-end could recognize and transcribe viral promoters not activated by protein p4, but they had lost their ability to recognize the A3 promoter in the presence of protein p4. In addition, these mutant reconstituted RNA polymerases could not interact with protein p4. We conclude that protein p4 activation of the viral A3 promoter requires an interaction between the carboxyl-end of protein p4 and the carboxyl-end of the alpha subunit of B. subtilis RNA polymerase that stabilizes the RNA polymerase at the promoter.
Mencía
M
M
Centro de Biología Molecular "Severo Ochoa," Consejo Superior de Investigaciones Científicas, Universidad Autónoma, Madrid, Spain.
Monsalve
M
M
Rojo
F
F
Salas
M
M
D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000201:enzymology*; D018983:DNA Footprinting; D012321:DNA-Directed RNA Polymerases / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D016415:Sequence Alignment; D017384:Sequence Deletion; D012808:Sigma Factor / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*; D015533:Transcriptional Activation; D014764:Viral Proteins / Q000378:metabolism*
D012808:Sigma Factor; D014157:Transcription Factors; D014764:Viral Proteins; C053987:p4 protein, Bacteriophage phi 29; C063505:RNA polymerase sigma A; D012321:DNA-Directed RNA Polymerases