Dual role of the sequence-specific bacteriophage T4 endoribonuclease RegB. mRNA inactivation and mRNA destabilization

Abstract

Gene regB of bacteriophage T4 encodes a sequence-specific endoribonuclease that introduces cuts in early phage messenger RNAs. Cutting takes place specifically in the middle of the tetranucleotide GGAG, as soon as the first minute of infection. Out of the 20 processing sites so far identified, seven are in Shine-Dalgarno sequences. The others are localized in intercistronic regions or within coding sequences. In the latter case, cutting efficiency is much lower. regB-dependent cleavages can occur within AU-rich sequences downstream of processed GGAG motifs that are not in effective translation initiation sites. We looked for possible consequences of regB-dependent cuts on gene expression in two early regions of the T4 chromosome. In the comC alpha region, none of the three major RegB cleavage sites is in a Shine-Dalgarno sequence, and in the motA region the unique regB-dependent processing site is found within the Shine-Dalgarno sequence of the gene. We find that in the region of gene comC alpha, RegB decreases two- to threefold the chemical half-life of early transcripts, but does not change the functional half-life of mRNAs coding for protein ComC alpha. The amount of MotA protein synthesized by the wild-type is half that obtained in a regB mutant infection. We show that this is a direct consequence of mRNA processing by RegB at the Shine-Dalgarno sequence of motA. This regB-mediated translation inhibition is not accompanied by an important modification in motA mRNA chemical half-life. We show that rapid shut-off of MotA protein synthesis that occurs soon after infection results both from RegB processing within the translation initiation region of motA and from early transcription inhibition followed by regB-independent breakdown of the motA mRNA.