Regulation of pyrBI operon expression in Escherichia coli by UTP-sensitive reiterative RNA synthesis during transcriptional initiation.

Abstract

In Escherichia coli K-12, the pyrBI operon encodes the two nonidentical subunits of the pyrimidine biosynthetic enzyme aspartate transcarbamylase (ATCase). Expression of the pyrBI operon is regulated over an -300-fold range by pyrimidine availability. Most of this regulation (i.e., 50-fold) occurs through a UTP-sensitive attenuation control mechanism in which low intracellular levels of UTP cause transcriptional pausing within a segment of the pyrBI leader region that specifies a 44-codon open reading frame. This pausing permits close coupling of transcription and translation within the leader region, which suppresses transcriptional termination at an attenuator (a p-independent transcriptional terminator) located,in front of the pyrB structural gene (Turnbough et al. 1983;Roland et al. 1988;Liu and Turnbough 1989;Landick and Turnbough 1992; Donahue and Turnbough 1994).Pyrimidine-mediated regulation of pyrBI expression also occurs at the level of transcriptional initiation (Don1Corresponding author. ahue and Turnbough 1990;Liu et al. 1993). Recent studies have suggested that this regulation involves a run of three T-A base pairs (specifying UUU) in the initially transcribed region of the pyrBI promoter (Donahue and Turnbough 1990). The pyrBI promoter region contains the sequence 5'-TATAATGCCGGACAATTTGCCG; the -10 region and two adenine residues at which transcriptional initiation can occur in vitro are underlined, and the more downstream adenine residue appears to be the predominant transcriptional start site in vivo (Donahue and Turnbough 1990). It was discovered that RNA polymerase formed heparin-resistant, transcriptionally competent initiation complexes at the pyrBI promoter in the presence of ATP and CTP and, to a small extent, with ATP alone, indicating stabilization by the nascent transcripts ACAA and AA, respectively. In contrast, in the presence of ATP and UTP, RNA polymerase did not form initiation complexes that were capable of synthesizing pyrBI transcripts following a heparin challenge.This result suggested that the synthesis of a nascent transcript with the sequence AAUUU did not stabilize the initiation complex or perhaps interfered with pro-

The Escherichia coli fis Promoter Is Regulated by Changes in the Levels of Its Transcription Initiation Nucleotide CTP

“…These studies uncovered a variety of fascinating regulatory strategies resulting from changes in intracellular NTP pools, including alterations in the rate of transcription so as to control the coupling of translation and transcription attenuation (45), alterations in the predominant transcription start site position so as to regulate the synthesis mRNAs with differential capacity to control translation (30,49,50), and stimulation of high rates of reiterative transcription so as to control productive transcript synthesis (39,(51)(52)(53). An effect of the transcription initiation NTP concentration on the kinetics of transcription initiation complex formation was demonstrated in the ribosomal RNA promoter rrnB P1 (54).…”

Changes in Conserved Region 3 of Escherichia coliς70 Reduce Abortive Transcription and Enhance Promoter Escape

“…In addition, at high UTP concentrations polymerase tends to "stutter" at the run of three U residues encoded in the initial sequence ATTTC of gal P2 after initiating at the normal A residue start site (36). Stuttering refers to the nonproductive mode of reiterative incorporation of UMP producing poly(U) RNA chains that are released at a variety of lengths and can grow to hundreds of nucleotide residues (37). At low UTP concentrations, this stuttering does not occur.…”

The rpoB mutants destabilizing initiation complexes at stringently controlled promoters behave like “stringent” RNA polymerases in Escherichia coli

“…One such example is pyrBI, a pyrimidine biosynthetic operon, whose expression is inhibited during the stringent response (3). Thus, the expression of operons encoding rRNA, such as rrnB P1, and those encoding nucleotide biosynthetic enzymes, such as pyrBI, are coordinately regulated during the stringent response, even through each of these operons also has its own unique regulatory feature(s) (4)(5)(6). The stringent response serves as a global regulatory mechanism, coordinating the transcriptional activity of RNAP with the growth conditions of the cell.…”