SummaryIn this paper we show that RNase R is a cold shock protein that is induced seven-to eightfold by cold shock and that its expression is tightly regulated by temperature. Transcriptional studies reveal that the rnr gene is co-transcribed with flanking genes as an operon induced under cold shock. The induction of RNase R levels is mainly a result of the stabilization of the rnr transcripts. The transient stability of the rnr transcripts is shown to be regulated by PNPase at the end of the acclimation phase. Studies with an rnr mutant revealed a cold-shock phenotype showing that RNase R contributes to growth at low temperatures. We have shown that RNase R can be involved in the maturation of SsrA/tmRNA, an important small stable RNA involved in protein tagging and ribosome rescue. The wide biological significance of RNase R regarding adaptation to cold shock and its involvement in RNA surveillance, protein quality control and pathogenesis is discussed.
SummaryIn nature, bacteria remain mostly in the stationary phase of the life cycle. Although mRNA is a major determinant of gene expression, little is known about mRNA decay in the stationary phase. The results presented herein demonstrate that RNase R is induced in stationary phase and is involved in the posttranscriptional regulation of omp A mRNA. This work is the first report of RNase R activity on a full length mRNA. In the absence of RNase R in a single rnr mutant, higher levels of omp A mRNA are found as a consequence of the stabilization of omp A full transcript. This effect is growth-phase-specific and not a growth-rate-dependent event. These higher levels of omp A mRNA were correlated with increases in the amounts of OmpA protein. We have also analysed the role of other factors that could affect ompA mRNA stability in stationary phase. RNase E was found to have the most important role, followed by polyadenylation. PNPase also affected the decay of the ompA transcript but RNase II did not seem to contribute much to this degradation process. The participation of RNase R in poly(A)-dependent pathways of decay in stationary phase of growth is discussed. The results show that RNase R can be a modulator of gene expression in stationary phase cells.
SUMMARY
The Unfolded Protein Response (UPR) is composed by homeostatic signaling pathways that are activated by excessive protein misfolding in the endoplasmic reticulum (ER). Ire1 signaling is an important mediator of the UPR, leading to the activation of the transcription factor Xbp1. Here, we show that Drosophila Ire1 mutant photoreceptors have defects in the delivery of Rhodopsin-1 to the rhabdomere and in the secretion of Spacemaker/Eyes shut into the inter-rhabdomeral space. However, these defects are not observed in Xbp1 mutant photoreceptors. Ire1 mutant retinas have higher mRNA levels for targets of regulated Ire1-dependent decay (RIDD), including for the fatty acid transport protein (fatp). Importantly, downregulation of fatp by RNA interference rescues the Rhodopsin-1 delivery defects observed in Ire1 mutant photoreceptors. Our results show that the role of Ire1 during photoreceptor differentiation is independent of Xbp1 function and demonstrate the physiological relevance of the RIDD mechanism in this specific paradigm.
SummaryIn Escherichia coli, ribonucleases are effectors that rapidly modulate the levels of mRNAs for adaptation to a changing environment. Factors involved in the regulation of these ribonucleases can be relevant for mRNA stability. RNase II is one of the main ribonucleases responsible for exonucleolytic activity in E. coli extracts. We have identified and characterized a new E. coli gene, which was named gmr (gene modulating RNase II). The results demonstrate that a deletion of gmr can be associated with changes in RNase II levels and activity. Western analysis and exoribonuclease activity assays showed a threefold increase in RNase II in the gmr deletion strain. Gmr does not affect RNase II mRNA, but modulates RNase II at the level of protein stability. RNase II protein turnover is slower in the gmr deletion strain. We also show that RNase II levels change in different media, and that this regulation is abolished in a strain lacking gmr. The data presented here show that the regulation of ribonucleolytic activity can depend on growth conditions, and this regulation can be mediated by factors that are not RNases.
PNPase and RNase II are the key regulatory exonucleases controlling mRNA decay in Escherichia coli. The rnb transcripts were found to proceed through the terminator and PNPase was found to be involved in the 3' to 5' degradation of rnb mRNA. Analysis of these longer 3' termini revealed that they are located in UA-rich regions. Comparison of single and double mutants suggested that PNPase and RNase II could have different roles in the degradation of these unstructured regions. We have shown that RNase II levels can vary over a fivefold range in haploid cells and that its expression depends on PNPase levels. PNPase-deficient strains were found to have a 2-2.5-fold increase in RNase II activity, while PNPase-overproducing strains reduced the rnb message and RNase II levels. Conversely, the amount of PNPase in the rnb deletion strain was approximately twofold higher than that in the wild-type strain. These observations suggest that the two main exonucleases are inter-regulated through a fine tuning mechanism. We discuss the implications of these results with regard to mRNA degradation and cell metabolism.
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