scholarly journals The Escherichia coli RNA polymerase alpha subunit and transcriptional activation by bacteriophage lambda CII protein.

1998 ◽  
Vol 45 (1) ◽  
pp. 271-280 ◽  
Author(s):  
M Gabig ◽  
M Obuchowski ◽  
A Ciesielska ◽  
B Latała ◽  
A Wegrzyn ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Rna Polymerase ◽  
Transcriptional Activation ◽  
Bacteriophage Lambda ◽  
Alpha Subunit ◽  
Single Amino Acid ◽  
Amino Acid Residues ◽  
A Subunit ◽  
Rpoa Gene

Bacteriophage lambda is not able to lysogenise the Escherichia coli rpoA341 mutant. This mutation causes a single amino acid substitution Lys271Glu in the C-terminal domain of the RNA polymerase alpha subunit (alphaCTD). Our previous studies indicated that the impaired lysogenisation of the rpoA341 host is due to a defect in transcriptional activation by the phage CII protein and suggested a role for alphaCTD in this process. Here we used a series of truncation and point mutants in the rpoA gene placed on a plasmid to investigate the process of transcriptional activation by the cII gene product. Our results indicate that amino-acid residues 265, 268 and 271 in the a subunit may play an important role in the CII-mediated activation of the pE promoter (most probably residue 271) or may be involved in putative interactions between alphaCTD and an UP-like element near pE (most probably residues 265 and 268). Measurement of the activity of pE-lacZ, pI-lacZ and p(aQ)-lacZ fusions in the rpoA+ and rpoA341 hosts demonstrated that the mechanism of activation of these CII-dependent promoters may be in each case different.

scholarly journals Inhibition of transcription starting from bacteriophage lambda pR promoter during the stringent response in Escherichia coli: implications for lambda DNA replication.

1995 ◽  
Vol 42 (2) ◽  
pp. 233-239 ◽  
Author(s):  
A Szalewska-Pałasz ◽  
G Wegrzyn
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Dna Replication ◽  
Transcriptional Activation ◽  
Plasmid Dna ◽  
Bacteriophage Lambda ◽  
Stringent Response ◽  
Lacz Gene ◽  
Relative Level ◽  
Lambda Dna

Replication of lambda plasmid DNA is halted in amino acid-starved wild type (stringent) strains whereas it proceeds in relA (relaxed) mutants. The only transcription which could be important in lambda plasmid DNA replication in amino acid-starved Escherichia coli cells is that starting from the pR promoter. Using a fusion which consists of the lacZ gene under the control of bacteriophage lambda pR promoter we found that transcription starting from this promoter was inhibited during the stringent, but not the relaxed, response in E. coli. We confirmed our conclusion by estimating the relative level of the pR transcript by RNA-DNA hybridization. We propose that decreased transcription from the pR promoter which serves as transcriptional activation of ori lambda is responsible for inhibition of lambda plasmid replication during the stringent response. The results presented in this paper, combined with our recent findings (published elsewhere), indicate that the transcriptional activation of ori lambda may be a main regulatory process controlling lambda DNA replication not only during the relaxed response but also in normal growth conditions.

scholarly journals Simultaneous gain and loss of functions caused by a single amino acid substitution in the beta subunit of Escherichia coli RNA polymerase: suppression of nusA and rho mutations and conditional lethality.

Genetics ◽  
1992 ◽  
Vol 130 (3) ◽  
pp. 411-428 ◽  
Author(s):  
J Sparkowski ◽  
A Das
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Rna Polymerase ◽  
Amino Acid Substitution ◽  
Beta Subunit ◽  
Single Amino Acid Substitution ◽  
Single Amino Acid ◽  
N Gene ◽  
Allele Specific ◽  
Gain And Loss

Abstract Transcript elongation and termination in Escherichia coli is modulated, in part, by the nusA gene product, an acidic protein that interacts not only with RNA polymerase itself but also with ancillary factors, namely the host termination protein Rho and phage lambda antitermination protein, N. The E. coli nusA1 mutant fails to support lambda development due to a specific defect in N-mediated antitermination. Certain rifampicin-resistant (rifR) variants of the nusA1 host support lambda growth. We report here the isolation and pleiotropic properties of one such rifR mutant, ts8, resulting from a single amino acid substitution mutation in rpoB, the structural gene for polymerase beta subunit. ts8 is a recessive lethal mutation that blocks cell growth at 42 degrees. Pulse-labeling and analysis of newly synthesized proteins indicate that the mutant cell is proficient in RNA synthesis at high temperature. Apparently, ts8 causes a loss of some specialized function of RNA polymerase without a gross defect in general transcription activities. ts8 is an allele-specific suppressor of nusA1. It does not suppress nusAsal, nusB5 and nusE71 mutations nor does it bypass the requirement for a functional N gene and the nut site for antitermination and lambda growth. A mutation in the N gene, punA1, that restores lambda growth in the nusA1 mutant host but not in the nusAsal host, compensates for the nusAsal allele in the ts8 mutant. This combined effect of two allele-specific suppressors suggests that they enhance some aspect of polymerase-NusA-N interaction and function. ts8 suppresses the rho15 mutation, but not the rho112 mutation, indicating that it might render RNA polymerase susceptible to the action of a defective Rho protein. Marker rescue analysis has localized ts8 to a 910-bp internal segment of rpoB that encodes the Rif domain. By amplification, cloning and sequencing of this segment of the mutant chromosome we have determined that ts8 contains Phe in place of Ser522, caused by a C to T transition. By gene conversion, we have established that the simultaneous gain and loss of three functions of polymerase is caused by this single amino acid substitution. Clearly, a site in the beta subunit critical for the functioning of both termination and antitermination factors is altered by ts8. The alteration, we imagine, might make this site on polymerase receptive to some factors but repulsive to others.

Sign in / Sign up

Export Citation Format

Share Document