Accumulation of Glucose 6-Phosphate or Fructose 6-Phosphate Is Responsible for Destabilization of Glucose Transporter mRNA inEscherichia coli

Abstract

Previously we found that a mutation in either pgi or pfkA, encoding phosphoglucose isomerase or phosphofructokinase A, respectively, facilitates degradation of the ptsG mRNA in an RNase E-dependent manner in Escherichia coli (1). In this study, we examined the effects of a series of glycolytic genes on the degradation of ptsG mRNA and how the mutations destabilize the ptsG mRNA. The conditional lethal mutation ts8 in fda, encoding fructose-1,6-P 2 aldolase just downstream of pfkA in the glycolytic pathway, caused the destabilization of ptsG mRNA at the nonpermissive temperature. Mutations in any other gene did not destabilize the ptsG mRNA; rather, they reduced the ptsG transcription mainly by affecting the cAMP level. The rapid degradation of ptsG mRNA in mutant strains was completely dependent upon the presence of glucose or any one of its compounds, which enter the Embden-Meyerhof glycolytic pathway before the block points. A significant increase in the intracellular glucose-6-P level was observed in the presence of glucose in the pgi strain. An overexpression of glucose-6-phosphate dehydrogenase eliminated both the accumulation and the degradation of ptsG mRNA in the pgi strain. In addition, accumulation of fructose-6-P led to the rapid degradation of ptsG mRNA in a pgi pfkA mutant strain lacking glucose-6-P. We conclude that the RNase E-dependent destabilization of ptsG mRNA occurs in response to accumulation of glucose-6-P or fructose-6-P.In bacteria, a number of sugars represented by glucose are transported into the cells coupled with their phosphorylation by the phosphoenolpyruvate-dependent sugar phosphotransferase system (PTS) 1 (2-4), whereas the translocation of some other sugars such as lactose is catalyzed by non-PTS transport systems. In either case, the incorporated sugars are metabolized primarily by the Embden-Meyerhof glycolytic pathway and by the pentose phosphate pathway to produce numerous intermediary metabolites as well as energy in cells (5). The PTS in Escherichia coli consists of two common cytoplasmic proteins, enzyme I and HPr (histidine-containing protein of the PTS), as well as an array of sugar-specific enzyme II complexes (EIIs). The glucose-specific EII (glucose transporter) consists of cytoplasmic protein IIA Glc and membrane receptor IICB Glc encoded by crr and ptsG, respectively. The phosphoryl group from phosphoenolpyruvate is transferred sequentially to enzyme I, HPr, the EIIs, and finally glucose as it is translocated across the membrane.In addition to sugar transport and phosphorylation, the PTS plays important regulatory roles in a variety of cellular activities. This is particularly evident for the glucose-specific PTS. For example, IIA Glc regulates both the transport of non-PTS sugars and the activity of adenylate cyclase depending on its phosphorylation state (2-4). The former process, called inducer exclusion, is fully responsible for the glucose-lactose diauxie that is a prototype of catabolite repression (6, 7). A striking recent discovery regarding the regulatory...

Experimental design-aided systematic pathway optimization of glucose uptake and deoxyxylulose phosphate pathway for improved amorphadiene production

“…4). The decreased cell density at high induction level is likely to be due to the stress of high expression of galactose permease and glucose kinase (Lu et al 2012) or sugar phosphate stress due to accumulation of glucose-6-phosphate or fructose-6-phosphate (Morita et al 2003), and this requires further investigation. Nevertheless, this limitation can be mitigated with tight control of the expressions of galP and glk using the combinatorial modular expression system described herein.…”

A dual function for a bacterial small RNA: SgrS performs base pairing-dependent regulation and encodes a functional polypeptide

“…An analogous condition occurs in wild-type strains exposed to the nonmetabolizable glucose analog ␣-methyl glucoside (␣MG), resulting in accumulation of ␣MG-6-phosphate. In both situations, cell growth is inhibited and there is a specific destabilization of the ptsG mRNA, which encodes the major glucose transporter of the phosphoenolpyruvate phosphotransferase system (PTS) in E. coli (PtsG, IICB Glc ) (4,5). This posttranscriptional regulation of the ptsG mRNA under conditions of G6P accumulation (glucose-phosphate stress) suggested the existence of at least one specific regulatory response to deal with such stresses.…”

“…However, until recently, little was known about the mechanisms used by bacterial cells to deal with metabolic stress associated with intracellular phosphosugar accumulation. One form of phosphosugar stress occurs in pgi (phosphoglucose isomerase) mutant strains where stress is associated with accumulation of glucose-6-phosphate (G6P) when cells are exposed to glucose (4)(5)(6). An analogous condition occurs in wild-type strains exposed to the nonmetabolizable glucose analog ␣-methyl glucoside (␣MG), resulting in accumulation of ␣MG-6-phosphate.…”

Small RNAs making a small protein

“…1). When the glycolytic flux is blocked, for example by a mutation in pgi encoding phosphoglucose isomerase, the intracellular level of G6P greatly increases (4). The accumulation of G6P represents a metabolic stress because it is somehow toxic for cells (5).…”