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PubMed2001

Global and specific translational regulation in the genomic response of Saccharomyces cerevisiae to a rapid transfer from a fermentable to a nonfermentable carbon source.

The global gene expression program that accompanies the adaptation of Saccharomyces cerevisiae to an abrupt transfer from a fermentable to a nonfermentable carbon source was characterized by using a cDNA microarray to monitor the relative abundances and polysomal distributions of mRNAs. Features of the program included a transient reduction in global translational activity and a severe decrease in polysome size of transcripts encoding ribosomal proteins. While the overall translation initiation of newly synthesized and preexisting mRNAs was generally repressed after the carbon source shift, the mRNA encoded by YPL250C was an exception in that it selectively mobilized into polysomes, although its relative abundance remained unchanged. In addition, splicing of HAC1 transcripts, which has previously been reported to occur during accumulation of unfolded proteins in the endoplasmic reticulum, was observed after the carbon shift. This finding suggests that the nonconventional splicing complex, composed of the kinase-endonuclease Ire1p and the tRNA ligase Rlg1p, was activated. While spliced HAC1 transcripts mobilized into polysomes, the vast majority of unspliced HAC1 RNA accumulated in nonpolysomal fractions before and after the carbon source shift, indicating that translation of unspliced HAC1 RNA is blocked at the translation initiation step, in addition to the previously reported elongation step. These findings reveal that S. cerevisiae reacts to the carbon source shift with a remarkable variety of responses, including translational regulation of specific mRNAs and activation of specific enzymes involved in a nonconventional splicing mechanism.

Base Sequence↗

PubMed2004

[Effect of carbon source and nitrate concentration on denitrifying dephosphorus removal and variation of ORP].

Effect of added carbon source and nitrate concentration on the denitrifying phosphorus removal by SBR process was systematicaly studied, at the same time the variation of oxidation reductiun potential (ORP) was investigated. The results showed the phosphate release rate and the denitrifying and dephosphorus uptake rate in anoxic phase increased with the high carbon source concentration under anaerobic condition (100-300mg/L). However when the carbon source added in anaerobic phase was high to 300mg/L, the residual COD inhibited the succeed denitrifying dephosphorus uptake. High nitrate concentration (5, 15, 40mg/L) in anoxic phase increased the initial denitrifying dephosphorus rate. Once the nitrate depletes, phosphate uptake changed to phosphate release. Moreover, the time of the turning point occurred later with the higher nitrate addition. ORP can be used as a control parameter of phosphorus release, and it can also indicate the denitrificaiton react degree during the anoxic phosphorus removal but can't be used as control parameter of phosphorus uptake.

Bacteria, Anaerobic↗

PubMed2004

Regulation of expression of cellulosomes and noncellulosomal (hemi)cellulolytic enzymes in Clostridium cellulovorans during growth on different carbon sources.

Cellulosomes and noncellulosomal (hemi)cellulolytic enzymes are produced by Clostridium cellulovorans to degrade plant cell walls. To understand their synergistic relationship, changes in mRNA and protein expression in cellulosomes and noncellulosomal (hemi)cellulolytic enzymes (hereafter called noncellulosomal enzymes) of cultures grown on cellobiose, cellulose, pectin, xylan, and corn fiber or mixtures thereof were examined. Cellulase expression, favored particularly by the presence of Avicel, was found with all substrates. Comparison of cellulosome and noncellulosomal enzymes showed that expression profiles were strongly affected by the carbon source. High xylanase or pectate lyase expression was observed when C. cellulovorans was grown on xylan or pectin, respectively. Mixed carbon substrates (cellulose-pectin-xylan mixture or corn fiber) induced a wider variety of enzymes than a single carbon source, such as cellobiose, pectin, or xylan. Cellulosomal proteome profiles were more affected by the carbon source than the noncellulosomal enzymes. Transcription and protein analyses revealed that cellulosomes and noncellulosomal enzymes were expressed simultaneously on mixed carbon sources, but their degree of inducibility varied when the substrate was either cellulose or cellobiose. Cellulosomes and noncellulosomal enzymes had synergistic activity on various carbon substrates. These results indicated that expression of plant cell wall-degrading enzymes is highly influenced by the available carbon source and that synergy between cellulosomes and noncellulosomal enzymes contribute to plant cell wall degradation.

Cellulase↗

PubMed2005

Global transcriptional programs reveal a carbon source foraging strategy by Escherichia coli.

By exploring global gene expression of Escherichia coli growing on six different carbon sources, we discovered a striking genome transcription pattern: as carbon substrate quality declines, cells systematically increase the number of genes expressed. Gene induction occurs in a hierarchical manner and includes many factors for uptake and metabolism of better but currently unavailable carbon sources. Concomitantly, cells also increase their motility. Thus, as the growth potential of the environment decreases, cells appear to devote progressively more energy on the mere possibility of improving conditions. This adaptation is not what would be predicated by classic regulatory models alone. We also observe an inverse correlation between gene activation and rRNA synthesis suggesting that reapportioning RNA polymerase (RNAP) contributes to the expanded genome activation. Significant differences in RNAP distribution in vivo, monitored using an RNAP-green fluorescent protein fusion, from energy-rich and energy-poor carbon source cultures support this hypothesis. Together, these findings represent the integration of both substrate-specific and global regulatory systems, and may be a bacterial approximation to metazoan risk-prone foraging behavior.

Acetic Acid↗

PubMed2004

Roles of pyruvate kinase and malic enzyme in Corynebacterium glutamicum for growth on carbon sources requiring gluconeogenesis.

In many bacteria, pyruvate kinase serves a well-defined function in glycolysis, catalyzing an ATP-generating reaction. However, its role during growth on carbon sources requiring glucoeneogenesis is less well investigated. We analyzed a defined pyruvate kinase gene (pyk) deletion mutant of Corynebacterium glutamicum, which is unable to grow on ribose as sole carbon source. Unexpectedly, the pyk deletion mutant was also unable to grow on acetate or citrate as sole carbon sources unless low amounts of pyruvate were added to the growth medium. A spontaneous suppressor mutant of the pyk deletion strain that regained the ability to grow on acetate was isolated. DNA microarray experiments revealed increased expression of the malic enzyme gene malE. The point mutation upstream of malE identified in this mutant was responsible for the loss of carbon-source-dependent regulation, as revealed by transcriptional fusion analysis. Overexpression of malE was sufficient to restore growth of the pyk deletion strain on acetate or citrate. The requirement of increased malic enzyme levels to re-route the carbon flux at the interface between glycolysis, gluconeogenesis and the tricarboxylic acid cycle in order to compensate for the absence of pyruvate kinase indicates a metabolic flux bifurcation at the metabolic node phosphoenolpyruvate. Whereas during growth of C. glutamicum on acetate or citrate most of the phosphoenolpyruvate generated from oxaloacetate is metabolized in gluconeogenesis, a fraction is converted by pyruvate kinase in the glycolytic direction to sustain proper pyruvate availability for biomass synthesis.

Acetates↗

PubMed2006

Use of population genetics to derive nonrecombinant Saccharomyces cerevisiae strains that grow using xylose as a sole carbon source.

According to scientific dogma, Saccharomyces cerevisiae cannot grow utilizing xylose as a sole carbon source. Although recombinant DNA technology has overcome this deficiency to some degree, efficient utilization of xylose appears to require complex global changes in gene expression. This complexity provides a significant challenge to the development of yeasts suitable for the utilization of xylose-rich lignocellulosic substrates. In contrast to the dogma, we have found that native strains of S. cerevisiae can grow on xylose as a sole carbon source, albeit very slowly. This observation provided the basis for a new approach using natural selection to develop strains of S. cerevisiae with improved ability to utilize xylose. By applying natural selection and breeding over an extended period, we have developed S. cerevisiae strains that can double in less than 6 h using xylose as a sole carbon source. Strains with improved growth rate possessed increased xylose reductase and xylitol dehydrogenase activities, with the latter showing the greater improvement. This unique, completely nonrecombinant approach to developing xylose-utilizing strains of S. cerevisiae opens an alternative route to the development of yeast that can fully utilize lignocellulosic substrates.

Aldehyde Reductase↗

PubMed1997

Identification of a Saccharomyces cerevisiae mitochondrial-DNA which can act as a promoter tightly regulated by carbon source when placed in the nucleus.

Screening of a promoter probe gene bank for DNA sequences that could act as inducible promoters following growth on non-fermentable carbon sources led to the identification of the mitochondrially encoded cytochrome oxidase subunit 1 gene (COX1) as an active sequence. Carbon-source regulation of this promoter was confirmed by a beta-galactosidase assay which showed a 31- and 180-fold induction of expression after growth on ethanol or lactate-based media respectively. Two elements matching the CCAAT-binding-factor motif, which is involved in activating transcription on non-fermentable carbon sources, were identified in the putative promoter. Expression was found to be reduced to low levels in otherwise isogenic hap3 and hap4 mutant strains. Thus, this mitochondrial DNA when placed in the nucleus can act as a promoter that is subject to strict carbon-source regulation. These observations are discussed both with respect to the origin of the S. cerevisiae COX1 gene in particular and with respect to the origin of introns in general.

Blotting, Southern↗

PubMed1994

Carbon sources for D-lactate formation in rat liver.

Carbon sources for D-lactate formation were investigated in vitro using 6,000 x g supernatant of rat liver homogenate and by rat liver perfusion in situ. As carbon sources, L-threonine, glucose, glycerol, acetone, and acetoacetate were tested. Glycerol was the best substrate for D-lactate formation via methylglyoxal in rat liver. Glucose was the second most preferred substrate, while L-threonine, acetone, and acetoacetate were poor substrates for D-lactate formation. Glycerol was several times more effective than normal as a substrate of D-lactate in the supernatants of liver homogenates of diabetic and starved rats, while it was less effective as a substrate of L-lactate. The glycerol kinase [EC 2.7.1.30] activities in livers increased in the diabetic and starved states. These and other results can explain why the plasma concentration of D-lactate increases several-fold after running and why the D-lactate contents in plasma, liver, and skeletal muscle are markedly increases in diabetic and starved rats.

Acetoacetates↗

PubMed2003

The effect of carbohydrate carbon sources on the production of constitutive and inducible laccases by Botryosphaeria sp.

The influence of carbohydrates: glucose, fructose, galactose, galacturonic acid, xylose, lactose, sucrose, pectin and inulin, were evaluated as sole carbon source for the production of laccases by the ascomycete, Botryosphaeria sp. Veratryl alcohol, a laccase inducer, was added to culture media to study inducible laccase production on the same carbon sources. Inulinase and pectinase were also produced when Botryosphaeria sp. was grown on inulin, and galacturonic acid and pectin, respectively, and their levels were less in the presence of veratryl alcohol. Botryosphaeria sp. produced constitutive laccases on all carbon sources examined, and veratryl alcohol increased the laccase production on most of carbon sources studied except for inulin and galacturonic acid. Evidence is presented that Botryosphaeria sp. is also pectinolytic.

Ascomycota↗

PubMed2002

The influence of carbon sources and morphology on nystatin production by Streptomyces noursei.

Carbon source nutrition and morphology were examined during cell growth and production of nystatin by Streptomyces noursei ATCC 11455. This strain was able to utilise glucose, fructose, glycerol and soluble starch for cell growth, but failed to grow on media supplemented with galactose, xylose, maltose, sucrose, lactose and raffinose. Utilisation of glucose had a negative influence on production of nystatin independent of the specific growth rate when phosphate and ammonium was in excess. Consumption of carbon sources was related to the specific growth rate. S. noursei ATCC 11455 formed mainly mycelial clumps during cultivation, while pellet growth dominated the culture of the morphologically altered high producing mutant S. noursei NG7.19. When the pellet size increased above a critical size, cell growth and nystatin production terminated. Fluorescent staining of hyphae revealed that this coincided with loss of activity inside the core of the pellets, probably due to diffusion limitation of oxygen or other nutrients.

Anti-Bacterial Agents↗

PubMed2000

Carbon source regulation of a dextranase gene from the filamentous fungus Penicillium minioluteum.

The regulation of dextranase (dexA) gene expression in the filamentous fungus Penicillium minioluteum grown on different carbon sources was studied. Growth in the presence of dextran leads to high expression of the dextranase enzyme, but growth in starch, glucose, glycerol, lactose and sorbitol did not. Dextran induced dexA gene expression at the mRNA level. However, in cultures containing dextran plus glucose or glycerol, the transcript was detected 24 h later than in the case where dextran was the only carbon source. When the glucose or glycerol concentration in the dextran-containing medium was kept at about 1% (w/v), no dextranase-transcripts were detected. It was found that both glucose and glycerol inhibited enzyme synthesis, because 1% (w/v) addition of both carbon sources to dextran-growing cultures was able to abolish the inducing effect of dextran. Our results suggest that dextran utilization responds to both specific induction and to glucose and glycerol repression, providing evidence that P. minioluteum dexA expression is regulated by the carbon source at the transcriptional level.

Blotting, Northern↗

PubMedDate not supplied

[Influence of carbon sources on the biological activity and morphology of Bacillus thuringiensis parasporal crystals].

The influence of the carbon source on biological activity and morphological properties of the parasporal crystals of Bacillus thuringiensis subsp. kurstaki cultivated on three different media has been studied. Biological activity of delta-endotoxins contained in crystals was tested according to their impact on insects and microorganisms. Variations in carbon sources composition resulted in changes of biological activity and morphology of crystals, caused by alterations in synthesis rate of various delta-endotoxins contained in the crystals. Variations in the content of a certain carbon source (e.g. starch) in the medium produced no effect on the crystals properties, and the content variations of the delta-endotoxins content was proportional to the producer biomass. Control of the antibacterial effect of delta-endotoxins allows a valid evaluation of their biological activity under various growth conditions as well as comparison of the biological potential of entomopathogenic chemicals for plant protection containing delta-endotoxins with different specificity of insecticidal activity.

Bacillus thuringiensis↗

PubMed1977

Gluconeogenesis in Saccharomyces cerevisiae: determination of fructose-1,6-bisphosphatase activity in cells grown in the presence of glycolytic carbon sources.

The activity of fructose-1,6-bisphosphatase (FBP), a gluconeogenic enzyme, was determined in wild-type Saccharomyces cerevisiae X2180 grown in the presence of the glycolytic carbon sources, glucose, fructose, and galactose. The activities of phosphofructokinase (PFK), a glycolytic enzyme, and phosphoglucose isomerase (PGI), an enzyme functioning both in glycolysis and gluconeogenesis, were determined for purposes of comparison. A measurable amount of FBP activity was present in 20-h-old cells grown with moderate shaking in 1% glucose-nutrient or minimal medium. This activity increased significantly in 40 and 60-h-old cells. Similar levels of FBP activity were also present in 20-, 40-, and 60-h-old cells grown in 1% fructose-nutrient medium. A higher level of FBP activity was present in 20-h-old cells grown in 1% galactose-nutrient medium than in 20-h-old cells grown in 1% glucose- or fructose-nutrient medium. The FBP activity in glucose- or fructose-grown cells was higher than the corresponding activity in cells grown under similar conditions for 40 and 60 h in the presence of ethanol, a gluconeogenic carbon source. The PFK activity was significantly less in galactose- and ethanol-grown cells. The PGI activity was relatively constant in 20-, 40-, and 60-h-old cells grown in the presence of glucose, fructose, and galactose, but this activity was reduced approximately 50% in ethanol-grown cells. It is concluded from these results that, depending upon the concentration of carbon source and the time of incubation, FBP, a strictly gloconeogenic enzyme, is synthesized by S. cerevisiae grown in the presence of glycolytic carbon sources.

Ethanol↗

PubMed1998

Regulation of hly expression in Listeria monocytogenes by carbon sources and pH occurs through separate mechanisms mediated by PrfA.

Expression of the PrfA-controlled virulence gene hly (encoding the pore-forming cytolysin listeriolysin) is under negative regulation by readily metabolized carbon sources in Listeria monocytogenes. However, the hyperhemolytic strain NCTC 7973 exhibits deregulated hly expression in the presence of repressing sugars, raising the possibility that a defect in carbon source regulation is responsible for its anomalous behavior. We show here that the activity of a second glucose-repressed enzyme, alpha-glucosidase, is 10-fold higher in NCTC 7973 than in 10403S. Using hly-gus fusions, we show that the prfA allele from NCTC 7973 causes deregulated hly-gus expression in the presence of sugars in either the wild-type or the NCTC 7973 background, while the 10403S prfA allele restores carbon source regulation. However, the prfA genotype does not affect the regulation of alpha-glucosidase activity by repressing sugars. Of the two mutational differences in PrfA, only a Gly145Ser change is important for regulation of hly-gus. Therefore, NCTC 7973 and 10403S have genetic differences in at least two loci: one in prfA that affects carbon source regulation of virulence genes and another in an unidentified gene(s) that up-regulates alpha-glucosidase activity. We also show that the decrease in pH associated with utilization of sugars negatively regulates hly-gus expression, although sugars can affect hly-gus expression by another mechanism that is independent of pH.

Alleles↗

PubMed2005

Competition between polyphosphate and glycogen accumulating organisms in enhanced biological phosphorus removal systems with acetate and propionate as carbon sources.

Enhanced biological phosphorus removal (EBPR) is a widely used process for achieving phosphorus removal from wastewater. A potential reason for EBPR failure is the undesirable growth of glycogen accumulating organisms (GAOs), which can compete for carbon sources with the bacterial group responsible for phosphorus removal from wastewater: the polyphosphate accumulating organisms (PAOs). This study investigates the impact of carbon source on EBPR performance and the competition between PAOs and GAOs. Two sequencing batch reactors (SBRs) were operated during a 4-6 month period and fed with a media containing acetate or propionate, respectively, as the sole carbon source. It was found that the acetate fed SBR rarely achieved a high level of phosphorus removal, and that a large portion of the microbial community was comprised of "Candidatus Competibacter phosphatis", a known GAO. The propionate fed SBR, however, achieved stable phosphorus removal throughout the study, apart from one brief disturbance. The bacterial community of the propionate fed SBR was dominated by "Candidatus Accumulibacter phosphatis", a known PAO, and did not contain Competibacter. In a separate experiment, another SBR was seeded with a mixture of PAOs and a group of alphaproteobacterial GAOs, both enriched with propionate as the sole carbon source. Stable EBPR was achieved and the PAO population increased while the GAOs appeared to be out-competed. The results of this paper suggest that propionate may provide PAOs with a selective advantage over GAOs in the PAO-GAO competition, particularly through the minimisation of Competibacter. Propionate may be a more suitable substrate than acetate for enhancing phosphorus removal in EBPR systems.

Acetates↗

PubMed2005

Determination of microbial carbon sources in petroleum contaminated sediments using molecular 14C analysis.

Understanding microbial carbon sources is fundamental to elucidating the role of microbial communities in carbon cycling and in the biodegradation of organic contaminants. Because the majority of anthropogenic contaminants are either directly or indirectly derived from fossil fuels that are devoid of 14C, radiocarbon can be used as a natural inverse tracer of contaminant carbon in the contemporary environment. Here, 14C analysis of individual microbial phospholipid fatty acids (PLFA) was used to characterize the carbon sources utilized bythe active microbial community in salt marsh sediments contaminated by the Florida oil spill of 1969 in Wild Harbor, West Falmouth, MA. A specific goal was to determine whether this community is actively degrading petroleum residues that persist in these sediments. The delta14C values of microbial PLFA in all sediment horizons (contaminated and noncontaminated) matched the delta14C of the total sedimentary organic carbon after petroleum removal, indicating that no measurable metabolism of petroleum residues was occurring. This result agrees with ancillary data such as the delta13C content and distribution of PLFA, and the residual hydrocarbon composition determined by comprehensive two-dimensional gas chromatography (GCxGC) analysis. We hypothesize that microbes have chosen to respire the natural organic matter rather than the residual petroleum hydrocarbons because the former is more labile. Future efforts directed at determining indices of microbial degradation of petroleum hydrocarbons should consider competition with natural organic matter.

Accidents↗

PubMed2006

Polyhydroxyalkanoates (PHA) production using wastewater as carbon source and activated sludge as microorganisms.

Activated sludge from different full-scale wastewater treatment plants (municipal, pulp and paper industry, starch manufacturing and cheese manufacturing wastewaters) was used as a source of microorganisms to produce biodegradable plastics in shake flask experiments. Acetate, glucose and different wastewaters were used as carbon sources. Pulp and paper wastewater sludge was found to accumulate maximum concentration (43% of dry weight of suspended solids) of polyhydroxy alkanoates (PHA) with acetate as carbon source. Among the different wastewaters tested as a source of carbon, pulp and paper industry and starch industry wastewaters were found to be the best source of carbon while employing pulp and paper activated sludge for maximum accumulation of PHA. High concentration of volatile fatty acids in these wastewaters was the probable reason.

Acetates↗

PubMed1996

Influence of a supplementary carbon source on biodegradation of pyridine by freely suspended and immobilized Pimelobacter sp..

The effect of the presence of supplementary glucose or acetate on the growth and pyridine-degrading activity of freely suspended and calcium-alginate-immobilized Pimelobacter sp. was investigated. Although the supplementary carbon sources could be degraded simultaneously with pyridine, Pimelobacter sp. exhibited a preference for pyridine over supplementary carbon sources. Thus, the pyridine-degrading activity of the freely suspended cells was not decreased significantly by the addition of either glucose (1.5-6 mM) or acetate (6-24 mM) to the pyridine (6-24 mM). In the semi-continuous immobilized cell culture, immobilized cells also exhibited a preference for pyridine over supplementary carbon sources and did not switch their substrate preference throughout the culture. Owing to a high cell concentration, the volumetric pyridine degradation rate at 24 mM pyridine in the immobilized cell culture was approximately six times higher than that in the freely suspended cell culture. Furthermore, the immobilized cells could be reused 16 times without losing their pyridine-degrading activity during the culture period tested. Taken together, the use of immobilized Pimelobacter sp. for the degradation of pyridine is quite feasible because of the preference for pyridine over supplementary carbon sources, the high volumetric pyridine degradation rate, and the reusability of immobilized cells.

Acetates↗