IDENTIFICATION OF A PHAGE-CODED DNA-BINDING PROTEIN THAT REGULATES TRANSCRIPTION FROM LATE PROMOTERS IN BACTERIOPHAGE-P4

The genetic element P4 can propagate as a temperate phage or as a mult icopy plasmid in its host Escherichia coli. Late in the lytic cycle an d in the plasmid condition, transcription of the P4 essential genes de pends on the activation of the late promoters P-LL and P-sid, which co ntrol the transcription of the left and right operons, respectively. B oth P4 late promoters are positively regulated by the product of the P 4 delta gene, which is transcribed from P-sid. We have identified a ne w P4 gene, vis, that appears to play a relevant role in P4 late transc ription control. vis is the first gene downstream of P-LL and codes fo r a basic 88 amino acid protein with a potential helix-turn-helix moti f. Expression of the cloned vis gene suppresses all the phenotypic tra its exhibited by P4 vir1, a mutant that carries a promoter-up mutation in the late promoter P-LL. By Northern hybridization analysis we show ed that vis negatively regulates transcription from P-LL and enhances transcription from P-sid. Thus, vis auto-regulates its expression by r epressing its own promoter and enhancing transcription of gamma, which is required for P-LL, activation. The vis gene was fused with the glu tathione S-transferase gene and the GST-Vis fusion protein was partial ly purified. By gel retardation assays and DNA footprinting we demonst rated that GST-Vis binds to a 32 bp long region immediately downstream of P-LL. We also showed, by gel retardation, that GST-Vis binds to th e P-sid region. A sequence present in both P-LL and P-sid regions may represent the Vis binding consensus sequence. The dual role of Vis on the control of P4 late transcription may be required for a regulated e xpression of the replication functions when P4 propagates in the plasm id state. (C) 1996 Academic Press limited.