Evolutionary cheating in Escherichia coli stationary phase cultures

Starved cultures of Escherichia coli are highly dynamic, undergoing frequent population shifts. The shifts result from the spread of mutants able to grow under conditions that impose growth arrest on the ancestral population. To analyze competitive interactions underlying this dynamic we measured the survival of a typical mutant and the wild type during such population shifts. Here we show that the survival advantage of the mutant at any given time during a takeover is inversely dependent on its frequency in the population, its growth adversely affects the survival of the wild type, and its ability to survive in stationary phase at fixation is lower than that of its ancestor. These mutants do not enter, or exit early, the nondividing stationary-phase state, cooperatively maintained by the wild type. Thus they end up overrepresented as compared to their initial frequency at the onset of the stationary phase, and subsequently they increase disproportionately their contribution in terms of progeny to the succeeding generation in the next growth cycle, which is a case of evolutionary cheating. If analyzed through the game theory framework, these results might be explained by the prisoner's dilemma type of conflict, which predicts that selfish defection is favored over cooperation.     

surA, an Escherichia coli gene essential for survival in stationary phase.

Mutations in genes not required for exponential growth but essential for survival in stationary phase were isolated in an effort to understand the ability of wild-type Escherichia coli cells to remain viable during prolonged periods of nutritional deprivation. The phenotype of these mutations is referred to as Sur- (survival) and the genes are designated sur. The detailed analysis of one of these mutations is presented here. The mutation (surA1) caused by insertion of a mini-Tn10 element defined a new gene located near 1 min on the E. coli chromosome. It was located directly upstream of pdxA and formed part of a complex operon. Evidence is presented supporting the interpretation that cells harboring the surA1 mutation die during stationary phase while similar insertion mutations in other genes of the operon do not lead to a Sur- phenotype. Strains harboring surA1 had a normal doubling time in both rich and minimal medium, but cultures lost viability after several days in stationary phase. Analysis of revertants and suppressors of surA1, which arose after prolonged incubation in stationary phase, indicates that DNA rearrangements (excisions and duplications) occurred in cultures of this strain even when the viable-cell counts were below 10(2) cells per ml. Cells containing suppressing mutations then grew in the same culture to 10(8) cells per ml, taking over the population. The implications of these observations to our understanding of stationary-phase mutagenesis are discussed.     

Identification and characterization of stationary phase inducible genes in Escherichia coli

During transition into stationary phase a large set of proteins is induced in Escherichia coli. Only a minority of the corresponding genes has been identified so far. Using the λplacMu system and a plate screen for carbon starvation-induced fusion activity, a series of chromosomal lacZ fusions (csi::lacZ) was isolated. In complex medium these fusions were induced either during late exponential phase or during entry into stationary phase. csi::lacZ expression in minimal media in response to starvation for carbon, nitrogen and phosphate sources and the roles of global regulators such as the alternative sigma factor sigma;^S (encoded by rpoS), cAMP/CRP and the relA gene product were investigated. The results show that almost every fusion exhibits its own characteristic pattern of expression, suggesting a complex control of stationary phase-inducible genes that involves various combinations of regulatory mechanisms for different genes. All fusions were mapped to the E. coli chromosome. Using fine mapping by Southern hybridization, cloning, sequencing and/or phenotypic analysis, csi-5, csi-17, and csi-18 could be localized in osmY (encoding a periplasmic protein), glpD (aerobic glycerol-3-phosphate dehydrogenase) and glgA (glycogen synthase), respectively. The other fusions seem to specify novel genes now designated csiA through to csiF. csi-17(glpD)::lacZ was shown to produce its own glucose-starvation induction, thus illustrating the Intricacies of gene-fusion technology when applied to the study of gene regulation.     

Escherichia coli produces linoleic acid during late stationary phase.

Escherichia coli produces linoleic acid in the late stationary phase. This was the case whether the cultures were grown aerobically or anaerobically on a supplemented glucose-salts medium. The linoleic acid was detected by thin-layer chromatography and was measured as the methyl ester by gas chromatography. The linoleic acid methyl ester was identified by its mass spectrum. Lipids extracted from late-stationary-phase cells generated thiobarbituric acid-reactive carbonyl products when incubated with a free radical initiator. In contrast, extracts from log-phase or early-stationary-phase cells failed to do so, in accordance with the presence of polyunsaturated fatty acid only in the stationary-phase cells.     

外源基因在大肠杆菌中的高效表达

为了提高外源蛋白在大杨杆菌中的表达量,人们对大肠杆菌表达系统进行了许多研究。作者综述了有关外源基因在大肠杆菌中高效表达的研究进展。     

Escherichia coli deltafur mutant displays low HPII catalase activity in stationary phase

Iron is among the most important micronutrients used by bacteria. As a partner of the Fenton reaction, however, iron potentiates oxygen toxicity. Strict regulation of iron metabolism, and its coupling with regulation of defenses against oxidative stress, is an essential factor for life in the presence of oxygen. In Escherichia coli, iron metabolism is regulated by the Fur protein. A Fur-deficient mutant, in stationary phase, displayed about 30y-fold lower HPII activity than the respective, Fur-proficient parental strain. Deletion of fur seems to affect HPII catalase specifically, since the mutant was capable of inducing HPI catalase when challenged with H(2)O(2). Low HPII catalase activity appears to be among the reasons for hydrogen peroxide hypersensitivity of the deltafur mutant.     

Global gene expression in Escherichia coli biofilms

It is now apparent that microorganisms undergo significant changes during the transition from planktonic to biofilm growth. These changes result in phenotypic adaptations that allow the formation of highly organized and structured sessile communities, which possess enhanced resistance to antimicrobial treatments and host immune defence responses. Escherichia coli has been used as a model organism to study the mechanisms of growth within adhered communities. In this study, we use DNA microarray technology to examine the global gene expression profile of E. coli during sessile growth compared with planktonic growth. Genes encoding proteins involved in adhesion (type 1 fimbriae) and, in particular, autoaggregation (Antigen 43) were highly expressed in the adhered population in a manner that is consistent with current models of sessile community development. Several novel gene clusters were induced upon the transition to biofilm growth, and these included genes expressed under oxygen-limiting conditions, genes encoding (putative) transport proteins, putative oxidoreductases and genes associated with enhanced heavy metal resistance. Of particular interest was the observation that many of the genes altered in expression have no current defined function. These genes, as well as those induced by stresses relevant to biofilm growth such as oxygen and nutrient limitation, may be important factors that trigger enhanced resistance mechanisms of sessile communities to antibiotics and hydrodynamic shear forces.     

Effect of sigmaS on sigmaE-directed cell lysis in Escherichia coli early stationary phase

The sigmaE regulon has been shown to perform a novel function that causes dead-cell lysis specific to the early stationary phase in addition to its well-known role in the extracytoplasmic stress response in Escherichia coli. Here, the effect of sigmaS as a general stress-responsive sigma factor on sigmaE-directed cell lysis was investigated. The lysis phenomena were observed in both rpoS mutant and parental strains constitutively expressing active sigmaE, but the former lysis occurred at a relatively early stage compared to the latter. Based on these results and experiments with hydrogen peroxide, we propose that some stresses generate living but non-culturable cells, which are subject to sigmaE-directed cell lysis.     

Direct expression of urogastrone gene in Escherichia coli

Human epidermal growth factor (urogastrone; UG) is a 53-amino acid polypeptide hormone. A 192-bp DNA fragment containing the coding sequence for methionyl UG (Met-UG) and the ribosome-binding site (RBS) was chemically synthesized and placed downstream from the promotor for the Escherichia coli outer-membrane lipoprotein gene (lpp) on a plasmid. E. coli cells harboring the plasmid directed the synthesis of Met-UG at 10(2)-10(3) molecules per cell. Next, the coding sequence for Met-UG was inserted in a runaway-replication plasmid and expressed under the control of the lpp promoter and the RBS derived from bacteriophage Mu cII gene. Upon heat induction, the cells harboring the recombinant plasmid synthesized 10(5) molecules of Met-UG per cell.     

Processivity errors of gene expression in Escherichia coli

Not all ribosomes that initiate translation of an mRNA sequence will successfully complete it and produce a full-length protein product. By comparing the amounts of lacZ monomer and lacZ dimer protein expressed from a plasmid in a strictly controlled assay, we calculate a dimer to monomer ratio of 0.76. We interpret this to mean that ribosomes have a 76% chance of completing the synthesis of a beta-galactosidase polypeptide. The remaining 24% of the initiated chains end in processivity accidents. For the wild-type, premature RNA polymerase termination is found to account for roughly one-third of the processivity accidents. For the hyperaccurate SmP mutant, we observe a processivity of 0.28, but the presence of streptomycin improves this to 0.50. Thus, the hyperaccuracy with respect to missense substitutions for this mutant is accompanied by a reduced processivity. Addition of streptomycin increase the first error class and reduces the second one. This finding is relevant to the optimization of ribosome function and the growth performance of ribosome mutants.     

Escherichia coli segments its controls on carbon-dependent gene expression into global and specific regulations

How bacteria adjust gene expression to cope with variable environments remains open to question. Here, we investigated the way global gene expression changes in E. coli correlated with the metabolism of seven carbon substrates chosen to trigger a large panel of metabolic pathways. Coarse-grained analysis of gene co-expression identified a novel regulation pattern: we established that the gene expression trend following immediately the reduction of growth rate (GR) was correlated to its initial expression level. Subsequent fine-grained analysis of co-expression demonstrated that the Crp regulator, coupled with a change in GR, governed the response of most GR-dependent genes. By contrast, the Cra, Mlc and Fur regulators governed the expression of genes responding to non-glycolytic substrates, glycolytic substrates or phosphotransferase system transported sugars following an idiosyncratic way. This work allowed us to expand additional genes in the panel of gene complement regulated by each regulator and to elucidate the regulatory functions of each regulator comprehensively. Interestingly, the bulk of genes controlled by Cra and Mlc were, respectively, co-regulated by Crp- or GR-related effect and our quantitative analysis showed that each factor took turns to work as the primary one or contributed equally depending on the conditions.     

Causes and consequences of DNA repair activity modulation during stationary phase in Escherichia coli

Escherichia coli responds to nutrient exhaustion by entering a state commonly referred to as the stationary phase. Cells entering the stationary phase redirect metabolic circuits to scavenge any available nutrients and become resistant to different stresses. However, many DNA repair pathways are downregulated in stationary-phase cells, which results in increased mutation rates. DNA repair activity generally depends on consumption of energy and often requires de novo proteins synthesis. Consequently, unless stringently regulated during stationary phase, DNA repair activities may lead to an irreversible depletion of energy sources and, therefore to cell death. Most stationary phase morphological and physiological modifications are regulated by an alternative RNA polymerase sigma factor RpoS. However, nutrient availability, and the frequency and nature of stresses, are different in distinct environmental niches, which impose conflicting choices that result in selection of the loss or of the modification of RpoS function. Consequently, DNA repair activity, which is partially controlled by RpoS, is differently modulated in different environments. This results in the variable mutation rates among different E. coli ecotypes. Hence, the polymorphism of mutation rates in natural E. coli populations can be viewed as a byproduct of the selection for improved fitness.