Summary
In order to characterize the tandem rrnB promoters transcribing one of the ribosomal RNA operons in E. coli we subcloned the basic promoter unit. This 185 bp fragment extends from -64 to +121 counted from the transcription start site of upstream promoter P1. The start site of downstream promoter P2 is also included in the promoter cartridge. S1 mapping experiments show that both promoters on this fragment are active in vivo. BAL-31 deletion mutations generated at the start site for promoter P2 were also tested by S1 mapping. Transcription from P2 remained active in all cases with the exception of one construction which lacks the -10 region. This demonstrates that the sequences downstream from the -10 region of P2 are not essential for basic promoter function.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Berk AJ, Sharp PA (1977) Sizing and mapping of early adenovirus mRNAs by gel electrophoresis of S1 nuclease-digested hybrids. Cell 12:721–732
Birnboim HC, Doly J (1979) A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res 7:1513–1523
Bolivar F, Rodriguez RL, Greene PJ, Betlach MC, Heynecker HL, Boyer HW, Crosa JH, Falcow S (1977) Construction and characterization of new cloning vehicles, II. A multipurpose cloning system. Gene 2:95–113
Boyer HW, Roulland-Dussoix D (1969) A complementation analysis of the restriction and modification of DNA in Escherichia coli. J Mol Biol 41:459–472
Brosius J, Dull TD, Sleeter DD, Noller HF (1981a) Gene organization and primary structure of a ribosomal RNA operon from Escherichia coli. J Mol Biol 148:107–127
Brosius J, Ullrich A, Raker MA, Gray A, Dull TD, Gutell RR, Noller HF (1981b) Construction and fine mapping of recombinant plasmids containing the rrnB ribosomal RNA operon from E. coli. Plasmid 6:112–118
Brosius J, Cate RL, Perlmutter AP (1982) Precise location of two promoters for the β-lactamase gene of pBR322. S1 mapping of ribonucleic acid isolated from Escherichia coli or synthesized in vitro. J Biol Chem 257:9205–9210
Brosius J (1984a) Plasmid vectors for the selection of promoters. Gene 27:151–160
Brosius J (1984b) Toxicity of an overproduced foreign gene product in Escherichia coli and its use in plasmid vectors for the selection of transcription terminators. Gene 27:161–172
Brosius J, Holy A (1984) Regulation of ribosomal RNA promoters with a synthetic lac operator. Proc Natl Acad Sci USA 81:6929–6933
Csordás-Tóth E, Boros I, Venetianer P (1979) Structure of the promoter region for the rrnB gene in E. coli. Nucleic Acids Res 7:2189–2197
De Boer HA, Gilbert SF, Nomura M (1979) DNA sequences of promoter regions for rRNA operons rrnE and rrnA in E. coli. Cell 17:201–209
Glaser G, Sarmientos P, Cashel M (1983) Functional interrelationship between two tandem E. coli ribosomal RNA promoters. Nature 302:74–76
Gourse RL, Stark MJR, Dahlberg AE (1983) Regions of DNA involved in the stringent control of plasmid-encoded rRNA in vivo. Cell 32:1347–1354
Gray HB, Ostrander DA, Hodnett JL, Legerski RJ, Robberson DL (1975) Extracellular nucleases of Pseudomonas BAL31, I. Characterization of single-strand-specific deoxyribonuclease and double stranded deoxyribonuclease activities. Nucleic Acids Res 2:1459–1492
Jinks-Robertson S, Gourse RL, Nomura M (1983) Expression of rRNA and tRNA genes in Escherichia coli: Evidence for feedback regulation by products of rRNA operons. Cell 33:865–876
Kajitani M, Ishihama A (1984) Promoter selectivity of Escherichia coli RNA polymerase. Differential stringent control of the multiple promoters from ribosomal RNA and protein operons. J Biol Chem 259:1951–1957
Kingston RE (1983) Effects of deletions near Escherichia coli rrnB promoter P2 on inhibition of in vitro transcription by guanosine tetraphosphate. Biochemistry 22:5249–5254
Legerski RJ, Hodnett JL, Gray HB (1978) Extracellular nucleases of Pseudomonas BAL31. III. Use of the double-strand deoxyribonuclease activity as the basis of a convenient method for the mapping of fragments of DNA produced by cleavage with restriction enzymes. Nucleic Acids Res 5:1445–1464
Maxam AM, Gilbert W (1980) Sequencing end-labeled DNA with base-specific chemical cleavage. In Grossmann L, Moldave K (eds), Methods in enzymology, vol 65. Academic Press, New York, pp 499–559
Miller JH (1972) Experiments in molecular genetics, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, p 433
Nomura M, Gourse RL, Baughman G (1984) Regulation of synthesis of ribosomes and ribosomal components. Annu Rev Biochem 53:75–117
Sarminetos P, Cashel M (1983) Carbon starvation and growth rate-dependent regulation of the Escherichia coli ribosomal RNA promoters: Differential control of dual promoters. Proc Natl Acad Sci USA 80:7010–7013
Sarmientos P, Sylvester JE, Contente S, Cashel M (1983) Differential stringent control of the tandem E. coli ribosomal RNA promoters from the rrnA operon expressed in vivo on multicopy plasmids. Cell 32:1337–1346
Shen WF, Squires C, Squires CL (1982) Nucleotide sequence of the rrnG promoter region of E. coli. Nucleic Acids Res 10:3303–3313
Sutcliffe JG (1979) Complete nucleotide sequence of the Escherichia coli plasmid pBR322. Cold Spring Harbor Symp. Quant Biol 43:77–90
Szeberényi J, Apirion D (1983) Initiation, processing and termination of ribosomal RNA from a hybrid 5S ribosomal RNA gene in a plasmid. J Mol Biol 168:525–561
Szeberényi J, Apirion D (1984) Synthesis and processing of 5S rRNA from an rrnB minigene in a plasmid. Biochim Biophys Acta 783:15–25
Talmadge K, Gilbert W (1980) Construction of plasmid vectors with unique PstI cloning sites in a signal sequence coding region. Gene 12:235–241
Talmadge K, Brosius J, Gilbert W (1981) An ‘internal’ signal sequence directs secretion and processing of proinsulin in bacteria. Nature 294:176–178
Weaver RF, Weissman C (1979) Mapping of RNA by a modification of the Berk-Sharp procedure: the 5′ termini of 15S β-globin mRNA precursor and mature 10S β-globin mRNA have identical map coordinates. Nucleic Acids Res 7:1175–1193
Young RA, Steitz JA (1979) Tandem promoters direct E. coli ribosomal RNA synthesis. Cell 17:225–234
Author information
Authors and Affiliations
Additional information
Communicated by K. Isono
Rights and permissions
About this article
Cite this article
Thayer, G.C., Brosius, J. In vivo transcription from deletion mutations introduced near Escherichia coli ribosomal RNA promoter P2 . Molec Gen Genet 199, 55–58 (1985). https://doi.org/10.1007/BF00327509
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF00327509