Translation of Two Nested Genes in Bacteriophage P4 Controls Immunity-Specific Transcription Termination

Abstract

ABSTRACT In phage P4, transcription of the left operon may occur from both the constitutive P LE promoter and the regulated P LL promoter, about 400 nucleotides upstream of P LE . A strong Rho-dependent termination site, t imm , is located downstream of both promoters. When P4 immunity is expressed, transcription starting at P LE is efficiently terminated at t imm , whereas transcription from P LL is immunity insensitive and reads through t imm . We report the identification of two nested genes, kil and eta , located in the P4 left operon. The P4 kil gene, which encodes a 65-amino-acid polypeptide, is the first translated gene downstream of the P LE promoter, and its expression is controlled by P4 immunity. Overexpression of kil causes cell killing. This gene is the terminal part of a longer open reading frame, eta , which begins upstream of P LE . The eta gene is expressed when transcription starts from the P LL promoter. Three likely start codons predict a size between 197 and 199 amino acids for the Eta gene product. Both kil and eta overlap the t imm site. By cloning kil upstream of a tRNA reporter gene, we demonstrated that translation of the kil region prevents premature transcription termination at t imm . This suggests that P4 immunity might negatively control kil translation, thus enabling transcription termination at t imm . Transcription starting from P LL proceeds through t imm . Mutations that create nonsense codons in eta caused premature termination of transcription starting from P LL . Suppression of the nonsense mutation restored transcription readthrough at t imm . Thus, termination of transcription from P LL is prevented by translation of eta .