Expression of Mycobacteriophage Ms6 lysis genes is driven by two sigma(70)-like promoters and is dependent on a transcription termination signal present in the leader RNA.
A mycobacteriophage Ms6 strong promoter region (P(lys)) was isolated by using transcriptional fusions with the lacZ reporter gene. Two tandem sigma(70)-like promoter sequences (P1 and P2) were found in this region. DNA sequencing of the promoter downstream region revealed a 214-bp leader sequence followed by five adjacent coding regions of 231 bp (ORF1), 1,152 bp (ORF2), 996 bp (ORF3), 231 bp (ORF4), and 372 (ORF5). ORF1 has the potential to encode a 77-amino-acid protein which revealed similarity to mycobacteriophage TM4 gp90, a predicted protein with unknown function. ORF2 encodes a 384-amino-acid protein which is related to several bacteriophage amidases. This protein induced cell lysis upon addition of chloroform, confirming its mureinolytic activity. ORF3 encodes a 332-amino-acid protein which is related to TM4 gp30, a protein with sequence similarity to amidases. ORF4 encodes a 77-amino-acid holin-like protein with significant similarity to the holin of Lactococcus lactis r1t bacteriophage. ORF5 encodes a 124-amino-acid protein which is related to mycobacteriophage L5 gp30, a protein with unknown function. These data indicate that the promoter region P(lys) drives the transcription of the Ms6 lysis genes. An intrinsic transcription termination signal was identified in the leader sequence. Experiments using lacZ fusions showed that beta-galactosidase synthesis is inhibited when this transcription termination signal is present in the leader sequence. In conclusion, mycobacteriophage Ms6 cell lysis genes are expressed by their own promoter region, independently of virion structure and assembly protein genes. Moreover, an antitermination mechanism might be involved in their transcription regulation.
Garcia
Miguel
M
Laboratório de Microbiologia Molecular, Centro de Patogénese Molecular, Faculdade de Farmácia, Universidade de Lisboa, Lisbon, Portugal.
Pimentel
Madalena
M
Moniz-Pereira
José
J
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics; D002262:Carboxy-Lyases; D012321:DNA-Directed RNA Polymerases / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins; D015967:Gene Expression Regulation, Viral; D005814:Genes, Viral; D008565:Membrane Proteins / Q000235:genetics; D008969:Molecular Sequence Data; D009160:Mycobacteriophages / Q000235:genetics*; D020102:Mycobacterium smegmatis / Q000821:virology*; D009690:Nucleic Acid Conformation; D016366:Open Reading Frames; D011401:Promoter Regions, Genetic; D016415:Sequence Alignment; D012808:Sigma Factor / Q000235:genetics*; D013728:Terminator Regions, Genetic; D014158:Transcription, Genetic; D014169:Transformation, Bacterial; D014764:Viral Proteins / Q000235:genetics
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D012808:Sigma Factor; D014764:Viral Proteins; C081775:RNA polymerase sigma 70; D012321:DNA-Directed RNA Polymerases; D002262:Carboxy-Lyases; C079912:LysA protein, Bacteria; C489812:LysA protein, E coli