Autogenous regulation of Escherichia coli polynucleotide phosphorylase expression revisited

The Escherichia coli polynucleotide phosphorylase (PNPase, encoded by pnp), a phosphorolytic exoribonuclease, post-transcriptionally regulates its own expression at the level of mRNA stability and translation. Its primary transcript is very efficiently processed by RNase III, an endonuclease that makes a staggered double strand cleavage about in the middle of a long stem-loop in the 5'-untranslated region. The processed pnp mRNA is then rapidly degraded in a PNPase-dependent manner. Two non-mutually exclusive models have been proposed to explain PNPase autogenous regulation. The earlier one suggested that PNPase impedes translation of the RNase III processed pnp mRNA thus exposing the transcript to degradative pathways. More recently this has been replaced by the current model, which maintains that PNPase would simply degrade the promoter proximal small RNA generated by the RNase III endonucleolytic cleavage thus destroying the double stranded structure at the 5'-end that otherwise stabilizes the pnp mRNA. In our opinion, however, the first model was not completely ruled out. Moreover, the RNA decay pathway acting upon the pnp mRNA after disruption of the 5' double stranded structure remained to be determined. Here we provide additional support to the current model and show that the RNase III-processed pnp mRNA devoid of the double stranded structure at its 5'-end is not translatable and is degraded by RNase E in a PNPase-independent manner. Thus the role of PNPase in autoregulation is simply to remove, in concert with RNase III, the 5'-fragment of the cleaved structure that both allows translation and prevents the RNase E-mediated PNPase-independent degradation of the pnp transcript

Piovesello - Structure I - 3D models

the two folders contain the three-dimensional models of each lithic artifact of Stucture I (SI), a lithic concentration found in the gravettian archaeological site of Piovesello, Cassimoreno, Piacenza, IT

Autogenous regulation of Escherichia coli polynucleotide phosphorylase during cold acclimation by transcription termination and antitermination

Adaptation of Escherichia coli at low temperature implicates a drastic reprogramming of gene expression patterns. Mechanisms operating downstream of transcription initiation, such as control of transcription termination, mRNA stability and translatability, play a major role in controlling gene expression in the cold acclimation phase. It was previously shown that Rho-dependent transcription termination within pnp, the gene encoding polynucleotide phosphorylase (PNPase), was suppressed in pnp nonsense mutants, whereas it was restored by complementation with wild type allele. Using a tRNA gene as a reporter and the strong Rho-dependent transcription terminator t (imm) of bacteriophage P4 as a tester, here we show that specific sites in the 5'-untranslated region of pnp mRNA are required for PNPase-sensitive cold-induced suppression of Rho-dependent transcription termination. We suggest that suppression of Rho-dependent transcription termination within pnp and its restoration by PNPase is an autogenous regulatory circuit that modulates pnp expression during cold acclimation

Transcriptional and post-transcriptional control of polynucleotide phosphorylase during cold acclimation in Escherichia coli

Polynucleotide phosphorylase (PNPase, polyribonucleotide nucleotidyltransferase, EC 2.7.7.8) is one of the cold shock-induced proteins in Escherichia coli and pnp, the gene encoding it, is essential for growth at low temperatures. We have analysed the expression of pnp upon cold shock and found a dramatic transient variation of pnp transcription profile: within the first hour after temperature downshift the amount of pnp transcripts detectable by Northern blotting increased more than 10-fold and new mRNA species that cover pnp and the downstream region, including the cold shock gene deaD appeared; 2 h after temperature downshift the transcription profile reverted to a preshift-like pattern in a PNPase-independent manner. The higher amount of pnp transcripts appeared to be mainly due to an increased stability of the RNAs. The abundance of pnp transcripts was not paralleled by comparable variation of the protein: PNPase steadily increased about twofold during the first 3 h at low temperature, as determined both by Western blotting and enzymatic activity assay, suggesting that PNPase, unlike other known cold shock proteins, is not efficiently translated in the acclimation phase. In experiments aimed at assessing the role of PNPase in autogenous control during cold shock, we detected a Rho-dependent termination site within pnp. In the cold acclimation phase, termination at this site depended upon the presence of PNPase, suggesting that during cold shock pnp is autogenously regulated at the level of transcription elongation