N-mediated transcription antitermination in lambdoid phage H-19B is characterized by alternative NUT RNA structures and a reduced requirement for host factors.

Abstract

Gene expression in lambdoid phages in part is controlled by transcription antitermination. For most lambdoid phages, maximal expression of delayed early genes requires an RNA polymerase modified by the phage N and host Nus proteins at RNA NUT sites. The NUT sites (NUTL and NUTR) are made up of three elements: BOXA, BOXB and an intervening spacer sequence. We report on N antitermination in H-19B, a lambdoid phage carrying shiga toxin 1 genes. H-19B N requires NusA, but not two other host factors required by lambda N, NusB and ribosomal protein S10. The H-19B NUT site BOXA is not required, whereas the BOXB is required for N action. H-19B nut sites have dyad symmetries in the spacer regions that are not in other nut sites. Changes in one arm of the dyad symmetry inactivate the NUT RNA. Compensating changes increasing the number of mutant nucleotides but restoring dyad symmetry restore activity. Deletion of the sequences encoding the dyad symmetry has little effect. Thus, the specific nucleotides composing the dyad symmetry seem relatively unimportant. We propose that the RNA stem-loop structure, called the 'reducer', by sequestering nucleotides from the linear RNA brings into proximity sites on either side of the dyad symmetry that contribute to forming an active NUT site.