Induction of a φC31 prophage inhibits rRNA transcription in Streptomyces coelicolor A3(2)

Abstract

A lysogen of Streptomyces coelicolor A3(2) containing a thermoinducible mutant of the temperate phage phi C31 (phi C31 cts1) was used to obtain synchronous phage development. Filter hybridization experiments indicated a marked reduction in rRNA synthesis after prophage induction. S1 nuclease mapping showed that transcription from each of the four promoters of one rRNA gene set (rrnD) was reduced to approximately the same extent, and that inhibition required protein synthesis. Crude preparations of RNA polymerase from induced lysogens had enhanced transcribing activity for phi C31 DNA which was lost upon further purification. The purified preparations were unimpaired in their ability to transcribe from the rrnD promoters in vitro and apparently unchanged in polypeptide composition. The factor(s) responsible for stimulating phage transcription, and possibly for inhibiting rRNA synthesis, may have been separated from the enzyme during purification.

Global transcription pattern of C31 after induction of a Streptomyces coelicolor lysogen at different growth stages

“…Two independently constructed lysogens of S . coelicolor A3(2) derivatives were used: S. coelicolor 01 (hisAI uraAI strAI Pgl-SCP1-; Rodriguez et al, 1986) and S. coelicolor TMCl (wild-type : Clayton & Bibb, 1990). Both contain the thermoinducible (pC31 ctsl prophage (Novikova et al, 1973).…”

“…This phage has a 1-72 kb deletion (AM) in its EcoRI-C fragment (Chater et al, 1981). Conditions for growth and thermal induction of Streptomyces cultures were as described by Rodriguez et al (1986) and Clayton & Bibb (1990). Standard recombinant DNA techniques were used to subclone fragments of (pC31 cl DNA into pBR322, pBR329, and M13mp19 and to prepare plasmid and replicative-form DNA (Sambrook et al, 1989).…”

“…For cultures of S. coelicolor 01, the extraction procedures were as in Rodriguez et al (1986); for S. coelicolor TMC1, procedures were as in Clayton & Bibb (1990).…”

“…After precipitation, alkaline denaturation and electrophoresis in alkaline 1 % (w/v) agarose gels, the protected DNA was visualized by probing Southern blots with suitable 32P-nick translated DNA probes (Sambrook et al, 1989). S1 mapping analysis of rrnD transcription was done with a 32P-end-labelled 570 bp DNA probe, as described by Clayton & Bibb (1990). Dot-blot analysis of RNA.…”

“…The ability of 4C31 to inhibit transcription of host rRNA genes is lost with increasing culture age Induction of a 4C31 prophage leads to inhibition of host rRNA transcription (Rodriguez et al, 1986;Clayton & Bibb, 1990). The effect of culture age on this inhibition was analysed by S1 mapping of transcripts from the rrnD rRNA gene cluster using the TMCl RNA samples, isolated at different stages of culture, with and without prophage induction.…”