Pyrimidine‐Ribonucleotide Pools and Their Turnover in Phage T4‐Infected Escherichia coli Cells

Abstract

Exogenous uracil was shown to be taken up efficiently into ribonucleotide pools a t all times during the cycle of T4 phage infection, in which stable host RNA synthesis is known to be turned off. This uptake was shown to be effected by ribonucleotide pool expansion and a net increase in mRNA content after phage infection. By comparing different phage mutants, the draining of ribonucleotide pools into DNA synthesis by ribotide reduction, was also found to be instrumental for pool labeling from exogenous uracil. The UDPG pool expanded as much as 3-fold during DNA negative mutant infection, presumably as a result of absent DNA glucosylation.From measurements of specific activities of nucleotides by double labeling and thin-layer chromatography, it was concluded that the incorporation into RNA of a radioactive, exogenous base can be reliably used for following T4 phage transcription.It was stated by Nierlich [1,2], that in bacteria a large synthesis of labile RNA could go undetected or be greatly underestimated by RNA base incorporation experiments under conditions of no synthesis of stable RNA. This statement was based on the demonstration that the entry of an exogenous radioactive base into the intracellular nucleotide pools was not facilitated by exchange reactions between nucleotides and free base or by nucleotide pool expansion, and that the labeling of the pools from the exogenous base was relatively slow [3]. The conclusion was that the entry of isotope is not faster than the removal of pool components for net RNA synthesis.In the case of stringent regulation of RNA synthesis in bacteria starved for amino acids, a ceasing synthesis of stable RNA could thus make synthesis of labile mRNA undetectable by incorporation experiments [a].

The bacteriophage T4 anti‐sigma factor AsiA is not necessary for the inhibition of early promoters in vivo

“…However, this is true only if the intracellular pool of UTP remains constant throughout phage development. That this is the case after infection with T4 wild type has been checked according to Fast and Sköld (1973). We do not know whether this is also the case after asiA mutant infections because middle and late transcription are abolished (Ouhammouch et al ., 1994; data not shown).…”

Induction of the heat shock regulon of Escherichia coli markedly increases production of bacterial viruses at high temperatures

“…Control experiments showed that this level of uracil was not limiting under these conditions. Incorporation of exogenous uracil into T4 RNA is a reliable measure of T4 transcription (17).…”