Relative activities of the transcriptional regulatory sites in the rplKAJLrpoBC gene cluster of Escherichia coli

Abstract

The pattern of transcription of the rplKAJLrpoBC gene cluster of Escherichia coli appears to be complex. At least four different promoters and a transcriptional attenuator have been identified. To compare the relative effect of each of the putative promoters and the attenuator on transcription of these genes, we fused these regulatory sites to lacZ. These transcriptional fusions were constructed on lambda transducing phages so a single copy of each could be stably integrated into the chromosome. The level of ,B-galactosidase in a lysogen of each phage reflects the activity of the transcriptional regulatory site. We find that the promoters preceding rplK (rplKp) and rplJ (rplJp) are indeed the major promoters of this gene cluster. The minor promoter before rplL (rplLp) is much weaker and contributes little to the transcription of the downstream genes. Under these conditions, we find no evidence of a promoter (rpoBp) in the rplL-rpoB intercistronic region. The attenuator (atn) terminates ca. 70o of the transcripts initiated at the promoters preceding it. Although we cannot rule out that some transcripts from rplKp may read through into rplJLrpoBC, we find that rplJp alone is sufficient for high-level expression of these genes.The genes for the subunits of Escherichia coli RNA polymerase are cotranscribed with ribosomal protein genes. The genes for ,B and 13' (rpoB and rpoC) are cotranscribed with at least the L10 and L7/12 ribosomal protein genes (rplL and rplJ, respectively) (26,33,42), whereas the a gene (rpoA) is part of an operon containing four ribosomal protein genes (23). More recently, it has been shown that the gene for a (rpoD) is cotranscribed with the S21 gene (rpsU) and the gene for DNA primase (dnaG) (10).The transcription pattern of the rplKAJLrpoBC gene cluster located at 90 min on the E. coli chromosome is complicated. Initial experiments demonstrated that the genes for the 1B and 1' subunits of RNA polymerase are cotranscribed with at least the rplJ and rplL ribosomal protein genes from a promoter upstream of rplJ (rpUp) (26,33,42). A variety of experiments subsequently suggested that there are also two internal promoters within rplJLrpoBC, one preceding rplL (rplLp) and the other before rpoB (rpoBp) (20,25,26,28,33). In addition, a transcriptional attenuator (atn) was located between rplL and rpoB and accounts for the lower frequency of rpoBC transcription compared with that of rplJL (5,6,14,26). A promoter responsible for the cotranscription of rplK and rplA was identified upstream of rplK (rplKp) (2,15,26,43). Recent S1 nuclease mapping of in vivo transcripts suggests that transcription initiated at rplKp is not necessarily terminated after rplA, but can read through into the rplJL genes (9).To further understand the regulation of this complex operon, we have directly compared the relative effect of these different promoters and the attenuator on rplKAJLrpoBC transcription. These transcriptional regulatory sites were fused, both separately and in combination, to the lacZ gene carried on a lambd...

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Transcription frequency modulates the efficiency of an attenuator preceding therpoBCRNA polymerase genes ofEscherichia coli: possible autogenous control

“…Therefore the regulation of RNA polymerase synthesis would appear to be governed by controls acting on the expression of rpoB and rpoC. Transcription of the gene cluster containing rpoBC exhibits a complex pattern (19)(20)(21)(22)(23). A strong promoter located in front of rplJ (rplJp) directs the transcription of the rplJLrpoBC genes while initiation at another strong promoter preceding rplK (rplKp) leads to transcription of not only the rplKA genes but also the downstream rpULrpoBC genes.…”

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“…The splits we detect are mostly between the rplKA and rplJL-rpoBC transcription units. The genes in this operon have multiple promotor and attenuator sites ( Ralling and Linn, 1984 ), and have been shown to be governed by a complex set of signals ( Steward and Linn, 1991 ). It appears that a complete tetracistronic product is transcribed from rplKAJL with less abundant bicistronic products of rplK-rplA and rplJ-rplL ( Downing and Dennis, 1987 ), which may explain the strong conservation of these four genes in this gene block.…”