Relaxed mutants of Escherichia coli RNA polymerase

When Escherichia coli cells are treated with either polymixin or gramicidin at concentrations that block protein and RNA synthesis, they accumulate a significant amount of guanosine tetraphosphate ppGpp. Such accumulation occurs in stringent (relA+) as well as in relaxed (relA) strains and no guanosine pentaphosphate pppGpp is then detected within the cells. These observations suggest that polypeptide antibiotics elicit ppGpp formation through a mechanism different from the stringent control system triggered by amino acid starvation of bacteria. Experiments based on tetracycline action indicate, moreover, that the accumulation of ppGpp under polymixin or gramicidin treatment is connected with a strong restriction of the degradation rate of this nucleotide.     

Initiation of RNA molecules by purified Escherichia coli RNA polymerase

Summary The kinesties of appearance of, and the distribution among, the four bases of the chain-initial nucleotides (5-termini) of RNA chains synthesized in vitro under various conditions have been investigated. The results of this study have shown that when native T4 bacteriophage DNA is the template for the RNA polymerase, most chains start with a purine nucleotide. The ratio of ATP termini to GTP termini is independent of the reaction time and of the template/enzyme ratio in the reaction mixture. A similar preferential purine initiation was observed when denatured T4 is the template, but the ratio of ATP to GTP termini is reduced. All 5-termini of poly-AU chains synthesized on poly-dAT templates are ATP.The determination of the kinetics of initiation of RNA chains has allowed direct confirmation of some conclusions which had been inferred previously from sedimentation analyses of the RNA product. (1) Most RNA chains are initiated during a short period at the outset of the reaction. (2) Low concentrations of native DNA templates limit the number of RNA chains synthesized, not the rate of RNA chain growth. (3) The maximum number of RNA molecules which can be synthesized on denatured DNA templates is severalfold larger than the maximum number which can be synthesized on the same weight of native DNA.     

Degradation of RNA in Escherichia coli. A hypothesis

Summary A hypothesis to explain RNA degradation in Escherichia coli is proposed. In this hypothesis all classes of RNA are potentially degradable unless they are protected. The proposed mechanism for mRNA degradation requires a combination of endonuclease(s) and exonuclease(s) which degrades RNA in the 3 to 5 direction. Ribosomes attached to the newly synthesized 5 end of an mRNA molecule protect it from being attacked endonucleolytically; a delay in attachment of ribosomes to this end exposes it to endonucleolytic cleavage, followed by exonucleolytic digestion from the newly exposed 3 end to the 5 end. This mechanism is consistent with an overall 5 to 3 direction of degradation for mRNA. Exoribonucleases that degrade polyribonucleotides from the 5 end to 3 end are not required. The 3 end of the messenger is protected by its association with DNA. In order to enable the mRNA to remain anchored to the DNA while serving as an efficient template for protein synthesis, a special region near the 3 end of the mRNA is envisaged. This hypothetical region would not be translated.