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Frontiers in Endocrinology 2018During differentiation transient, inducers produce permanent changes in gene expression. A similar phenomenon, transcriptional hysteresis, produced by transient or... (Review)
Review
During differentiation transient, inducers produce permanent changes in gene expression. A similar phenomenon, transcriptional hysteresis, produced by transient or prolonged exposure to glucose, leads to cumulative, persistent, and largely irreversible effects on glucose-regulated gene expression, and may drive key aspects of metabolic memory, obesity, diabetes, and aging, and explain the protective effects of dietary restriction during aging. The most relevant effects of glucose-induced transcriptional hysteresis are the persistent effects of elevated glucose on genes that control glucose metabolism itself. A key observation is that, as with the lac operon, glucose induces genes that promote glycolysis and inhibits gene expression of alternative metabolic pathways including the pentose pathway, beta oxidation, and the TCA cycle. A similar pattern of metabolic gene expression is observed during aging, suggesting that cumulative exposure to glucose during aging produces this metabolic shift. Conversely, dietary restriction, which increases lifespan and delays age-related impairments, produces the opposite metabolic profile, leading to a shift away from glycolysis and toward the use of alternative substrates, including lipid and ketone metabolisms. The effect of glucose on gene expression leads to a positive feedback loop that leads to metastable persistent expression of genes that promote glycolysis and inhibit alternative pathways, a phenomenon first observed in the regulation of the lac operon. On the other hand, this pattern of gene expression can also be inhibited by activation of peroxisome proliferator activating receptor transcription factors that promote beta oxidation and inhibit metabolism of glucose-derived carbon bonds in the TCA cycle. Several pathological consequences may arise from glucose-induced transcriptional hysteresis. First, elevated glucose induces glycolytic genes in pancreatic beta cells, which induces a semi-stable persistent increase in insulin secretion, which could drive obesity and insulin resistance, and also due to glucose toxicity could eventually lead to beta-cell decompensation and diabetes. Diabetic complications persist even after complete normalization of glucose, a phenomenon known as metabolic memory. This too can be explained by persistent bistable expression of glucose-induced glycolytic genes.
PubMed: 29892261
DOI: 10.3389/fendo.2018.00232 -
CBE Life Sciences Education 2016Concept inventories (CIs) are valuable tools for educators that assess student achievement and identify misconceptions held by students. Results of student responses can...
Concept inventories (CIs) are valuable tools for educators that assess student achievement and identify misconceptions held by students. Results of student responses can be used to adjust or develop new instructional methods for a given topic. The regulation of gene expression in both prokaryotes and eukaryotes is an important concept in genetics and one that is particularly challenging for undergraduate students. As part of a larger study examining instructional methods related to gene regulation, the authors developed a 12-item CI assessing student knowledge of the lac operon. Using an established protocol, the authors wrote open-ended questions and conducted in-class testing with undergraduate microbiology and genetics students to discover common errors made by students about the lac operon and to determine aspects of item validity. Using these results, we constructed a 12-item multiple-choice lac operon CI called the Lac Operon Concept Inventory (LOCI), The LOCI was reviewed by two experts in the field for content validity. The LOCI underwent item analysis and was assessed for reliability with a sample of undergraduate genetics students (n = 115). The data obtained were found to be valid and reliable (coefficient alpha = 0.994) with adequate discriminatory power and item difficulty.
Topics: Educational Measurement; Female; Humans; Lac Operon; Male; Pilot Projects; Reproducibility of Results; Students; Thinking
PubMed: 27252300
DOI: 10.1187/cbe.15-07-0162 -
Journal of Microbiology & Biology... 2020Undergraduate biology courses rely heavily on visual representation of information. Students view images of plants, animals, and microbes, interpret data presented in...
Undergraduate biology courses rely heavily on visual representation of information. Students view images of plants, animals, and microbes, interpret data presented in graphs, and use drawings to understand how cells and molecules interact in three dimensions. Traditional teaching approaches exclude students with visual impairments and disadvantage students with disabilities that affect their interpretation and processing of visual and spatial information, and also students who simply do not identify as "visual learners." By using new technologies to develop tactile teaching tools (TTTs) that can be employed in classrooms, we aim to create inclusive learning environments and more effectively instruct diverse learners. The advent of affordable and accessible 3D printing technology makes it possible to create tactile models that represent molecules, cells, and entire organisms more accurately than traditional visual representations. We describe the assessment of a 3D gene expression puzzle as a guided inquiry learning activity in which students must correctly assemble a series of components in order to achieve an output. Upon completion of the puzzle, the TTT provides tactile feedback through vibration to signal transcriptional activation. Analysis of pre- and postassessment performance demonstrated statistically significant increases in individual students' paired assessment scores in two different classroom implementations, with a greater effect size at a rural minority-serving institution than an urban R1 university. These encouraging preliminary data suggest that TTTs with guided-inquiry learning disproportionately benefit disadvantaged student populations and could serve as a tool in leveling the playing field when teaching abstract biological concepts in diverse educational settings.
PubMed: 32341727
DOI: 10.1128/jmbe.v21i1.2091 -
BMC Microbiology Aug 2015Classical bacteriological characteristics of Salmonella enterica indicate that the members of this species are unable to utilize lactose as a carbon source. However,...
BACKGROUND
Classical bacteriological characteristics of Salmonella enterica indicate that the members of this species are unable to utilize lactose as a carbon source. However, lactose-fermenting (Lac+) strains of several Salmonella serovars have been isolated from different foodborne outbreaks as well as different geographical regions worldwide. In the present study, we sequenced the genomes of 13 Lac + S. enterica isolates and characterized the lac region, comparing it to the lac region in other enteric bacterial species.
RESULTS
Genetic analysis of the lac operons in the S. enterica genomes revealed that they all contain intact lacI, lacZ, and lacY genes. However, lacA was truncated in all of the S. enterica subsp. enterica isolates, encoding a 56 amino acid peptide rather than the full length 220 amino acid LacA protein. Molecular analyses of the 13 isolates revealed that the lac operon resided on a plasmid in some strains and in others was integrated into the bacterial chromosome. In most cases, an insertion sequence flanked at least one end of the operon. Interestingly, the S. enterica Montevideo and S. enterica Senftenberg isolates were found to harbor a plasmid with a high degree of sequence similarity to a plasmid from Klebsiella pneumoniae strain NK29 that also harbors the lac operon. In addition, two S. enterica Tennessee isolates carried two copies of the lac operon. Phylogenetic analysis based on lacIZY gene sequences determines distinct clusters, and reveals a greater correlation between lacIZY sequence and flanking organization than with either bacterial species or genomic location.
CONCLUSIONS
Our results indicate that the lac region is highly mobile among Enterobacteriaceae and demonstrate that the Lac + S. enterica subsp. enterica serovars acquired the lac region through parallel events. The acquisition of the lac operon by several S. enterica serovars may be indicative of environmental adaptation by these bacteria.
Topics: Chromosomes, Bacterial; Cluster Analysis; DNA Transposable Elements; DNA, Bacterial; Gene Order; Genome, Bacterial; Lac Operon; Molecular Sequence Data; Phylogeny; Plasmids; Salmonella enterica; Sequence Analysis, DNA; Sequence Homology
PubMed: 26303940
DOI: 10.1186/s12866-015-0511-8 -
Frontiers in Microbiology 2022is an important human pathogen that can infect almost every organ system, resulting in a high incidence of morbidity and mortality. The operon is an important...
is an important human pathogen that can infect almost every organ system, resulting in a high incidence of morbidity and mortality. The operon is an important regulator of several staphylococcal phenotypes, including biofilm development, cell wall crosslinking, antibiotic resistance, oxidative stress, and acute and chronic implant-associated osteomyelitis. Our previous study showed that, by modulating murein hydrolase activity, the operon negatively regulates the proteases that govern cell death. Here, we report further elucidation of the mechanism of cell death, which is regulated by the operon at the molecular level in the USA300 LAC strain. We showed that deletion of enhances weak-acid-dependent cell death, because, in the biofilm microenvironment, this mutant strain consumes glucose and produces acetate and acetoin at higher rates than wild-type USA300 LAC strain. We proposed the increased intracellular acidification leads to increased cell death. MsaB, a dual-function transcription factor and RNA chaperone, is a negative regulator of the regulon, which has been shown to play an important role in overflow metabolism and programmed cell death during biofilm development in . We found that MsaB binds directly to the promoter, which represses expression of the regulon and prevents transcription of the and operons. In addition, we observed that pyruvate induced expression of the operon (MsaB). The results reported here have enabled us to decipher the role of the operon in staphylococcal metabolic adaption during biofilm development.
PubMed: 35722290
DOI: 10.3389/fmicb.2022.914512 -
Frontiers in Microbiology 2022is used extensively in the fermented milk-beverage industry as a starter culture. Acid production capacity during fermentation is the main criterion for evaluating...
is used extensively in the fermented milk-beverage industry as a starter culture. Acid production capacity during fermentation is the main criterion for evaluating starters although it is strain-dependent. In this study, the acid production rates of 114 strains were determined and then classified into high acid (HC), medium acid (MC), and low acid (LC) groups. Comparative genomics analysis found that the operon genes encoding the phosphoenolpyruvate-lactose phosphotransferase system (PTS) were located on plasmids in the HC strains; however, it is notable that the corresponding operons were located on the chromosome in LC strains. Real-time PCR analysis showed that the copy numbers of lac operon genes in HC strains were between 3.1 and 9.3. To investigate the relationship between copy number and acid production rate, the operon cluster of the HC group was constitutively expressed in LC strains. The resulting copy numbers of operon genes were between 15.8 and 18.1; phospho-β-galactosidase activity increased by 1.68-1.99-fold; and the acid production rates increased by 1.24-1.40-fold, which enhanced the utilization rate of lactose from 17.5 to 42.6% in the recombinant strains. The markedly increased expression of operon genes increased lactose catabolism and thereby increased the acid production rate of .
PubMed: 36386630
DOI: 10.3389/fmicb.2022.1016904 -
ACS Synthetic Biology Feb 2023Reverse genetics (RG) systems have been instrumental for determining the molecular aspects of viral replication, pathogenesis, and for the development of therapeutics....
Reverse genetics (RG) systems have been instrumental for determining the molecular aspects of viral replication, pathogenesis, and for the development of therapeutics. Here, we demonstrate that genes encoding the influenza surface antigens hemagglutinin and neuraminidase have varying stability when cloned into a common RG plasmid and transformed into . Using GFP as a reporter, we demonstrate that expresses the target genes in the RG plasmid at low levels. Incorporating operators or a transcriptional terminator into the plasmid reduced expression and stabilized the viral genes to varying degrees. Sandwiching the viral gene between two operators provided the largest contribution to stability and we confirmed the stabilization is Lac repressor-dependent and crucial for subsequent plasmid propagations in . Viruses rescued from the operator-stabilized plasmid displayed similar kinetics and titers to the original plasmid in two different viral backbones. Together, these results indicate that silencing transcription from the plasmid in helps to maintain the correct influenza gene sequence and that the operator addition does not impair virus production. It is envisaged that sandwiching DNA segments between operators can be used for reducing DNA segment instability in any plasmid that is propagated in which express the Lac repressor.
Topics: Humans; Escherichia coli; Lac Repressors; Influenza, Human; Reverse Genetics; Plasmids; Lac Operon
PubMed: 36716395
DOI: 10.1021/acssynbio.2c00358 -
Biophysical Journal Jan 2022The lactose uptake pathway of E. coli is a paradigmatic example of multistability in gene regulatory circuits. In the induced state of the lac pathway, the genes...
The lactose uptake pathway of E. coli is a paradigmatic example of multistability in gene regulatory circuits. In the induced state of the lac pathway, the genes comprising the lac operon are transcribed, leading to the production of proteins that import and metabolize lactose. In the uninduced state, a stable repressor-DNA loop frequently blocks the transcription of the lac genes. Transitions from one phenotypic state to the other are driven by fluctuations, which arise from the random timing of the binding of ligands and proteins. This stochasticity affects transcription and translation, and ultimately molecular copy numbers. Our aim is to understand the transition from the induced to the uninduced state of the lac operon. We use a detailed computational model to show that repressor-operator binding and unbinding, fluctuations in the total number of repressors, and inducer-repressor binding and unbinding all play a role in this transition. Based on the timescales on which these processes operate, we construct a minimal model of the transition to the uninduced state and compare the results with simulations and experimental observations. The induced state turns out to be very stable, with a transition rate to the uninduced state lower than 2×10 per minute. In contrast to the transition to the induced state, the transition to the uninduced state is well described in terms of a 2D diffusive system crossing a barrier, with the diffusion rates emerging from a model of repressor unbinding.
Topics: Escherichia coli; Escherichia coli Proteins; Lac Operon; Lac Repressors; Lactose
PubMed: 34953812
DOI: 10.1016/j.bpj.2021.12.027 -
Biophysical Journal Mar 2022The expression of the lac operon of E. coli is subject to positive feedback during growth in the presence of gratuitous inducers, but its existence in the presence of...
The expression of the lac operon of E. coli is subject to positive feedback during growth in the presence of gratuitous inducers, but its existence in the presence of lactose remains controversial. The key question in this debate is: Do the lactose enzymes, Lac permease and β-galactosidase, promote accumulation of allolactose? If so, positive feedback exists since allolactose does stimulate synthesis of the lactose enzymes. Here, we addressed the above question by developing methods for determining the intracellular allolactose concentration as well as the kinetics of enzyme induction and dilution. We show that, during lac induction in the presence of lactose, the intracellular allolactose concentration increases with the lactose enzyme level, which implies that lactose enzymes promote allolactose accumulation, and positive feedback exists. We also show that, during lac repression in the presence of lactose + glucose, the intracellular allolactose concentration decreases with the lactose enzyme levels, which suggests that, under these conditions, the positive feedback loop turns in the reverse direction. The induction and dilution rates derived from the transient data show that the positive feedback loop is reversed due to a radical shift of the steady-state induction level. This is formally identical to the mechanism driving catabolite repression in the presence of TMG + glucose.
Topics: Catabolite Repression; Escherichia coli; Feedback; Glucose; Lactose
PubMed: 35065914
DOI: 10.1016/j.bpj.2022.01.017 -
Frontiers in Cell and Developmental... 2023Since Jacob and Monod's discovery of the lac operon ∼1960, the explanations offered for most metabolic adaptations have been genetic. The focus has been on the...
Since Jacob and Monod's discovery of the lac operon ∼1960, the explanations offered for most metabolic adaptations have been genetic. The focus has been on the adaptive changes in gene expression that occur, which are often referred to as "metabolic reprogramming." The contributions metabolism makes to adaptation have been largely ignored. Here we point out that metabolic adaptations, including the associated changes in gene expression, are highly dependent on the metabolic state of an organism prior to the environmental change to which it is adapting, and on the plasticity of that state. In support of this hypothesis, we examine the paradigmatic example of a genetically driven adaptation, the adaptation of to growth on lactose, and the paradigmatic example of a metabolic driven adaptation, the Crabtree effect in yeast. Using a framework based on metabolic control analysis, we have reevaluated what is known about both adaptations, and conclude that knowledge of the metabolic properties of these organisms prior to environmental change is critical for understanding not only how they survive long enough to adapt, but also how the ensuing changes in gene expression occur, and their phenotypes post-adaptation. It would be useful if future explanations for metabolic adaptations acknowledged the contributions made to them by metabolism, and described the complex interplay between metabolic systems and genetic systems that make these adaptations possible.
PubMed: 37377740
DOI: 10.3389/fcell.2023.1197226