Genome-wide Transcriptional Response Research Articles

Genome-wide transcriptional responses of two metal-tolerant symbiotic Mesorhizobium isolates to Zinc and Cadmium exposure

BackgroundMesorhizobium metallidurans STM 2683T and Mesorhizobium sp. strain STM 4661 were isolated from nodules of the metallicolous legume Anthyllis vulneraria from distant mining spoils. They tolerate unusually high Zinc and Cadmium concentrations as compared to other mesorhizobia. This work aims to study the gene expression profiles associated with Zinc or Cadmium exposure and to identify genes involved in metal tolerance in these two metallicolous Mesorhizobium strains of interest for mine phytostabilization purposes.ResultsThe draft genomes of the two Mezorhizobium strains were sequenced and used to map RNAseq data obtained after Zinc or Cadmium stresses. Comparative genomics and transcriptomics allowed the rapid discovery of metal-specific or/and strain-specific genes. Respectively 1.05% (72/6,844) and 0.97% (68/6,994) predicted Coding DNA Sequences (CDS) for STM 2683 and STM 4661 were significantly differentially expressed upon metal exposure. Among these, a significant number of CDS involved in transport (13/72 and 13/68 for STM 2683 and STM 4661, respectively) and sequestration (15/72 and 16/68 for STM 2683 and STM 4661, respectively) were identified. Thirteen CDS presented homologs in both strains and were differentially regulated by Zinc and/or Cadmium. For instance, several PIB-type ATPases and genes likely to participate in metal sequestration were identified. Among the conserved CDS that showed differential regulation in the two isolates, we also found znuABC homologs encoding for a high affinity ABC-type Zinc import system probably involved in Zinc homeostasis. Additionally, global analyses suggested that both metals also repressed significantly the translational machinery.ConclusionsThe comparative RNAseq-based approach revealed a relatively low number of genes significantly regulated in the two Mesorhizobium strains. Very few of them were involved in the non-specific metal response, indicating that the approach was well suited for identifying genes that specifically respond to Zinc and Cadmium. Among significantly up-regulated genes, several encode metal efflux and sequestration systems which can be considered as the most widely represented mechanisms of rhizobial metal tolerance. Downstream functional studies will increase successful phytostabilization strategies by selecting appropriate metallicolous rhizobial partners.

Sa1969 Inter-Ethnic Differences in Genome-Wide Transcriptional Response to Vitamin D in the Colon

Sa1969 Inter-Ethnic Differences in Genome-Wide Transcriptional Response to Vitamin D in the Colon

Transcriptomic Response of Porcine PBMCs to Vaccination with Tetanus Toxoid as a Model Antigen

The aim of the present study was to characterize in vivo genome-wide transcriptional responses to immune stimulation in order to get insight into the resulting changes of allocation of resources. Vaccination with tetanus toxoid was used as a model for a mixed Th1 and Th2 immune response in pig. Expression profiles of PBMCs (peripheral blood mononuclear cells) before and at 12 time points over a period of four weeks after initial and booster vaccination at day 14 were studied by use of Affymetrix GeneChip microarrays and Ingenuity Pathway Analysis (IPA). The transcriptome data in total comprised more than 5000 genes with different transcript abundances (DE-genes). Within the single time stages the numbers of DE-genes were between several hundred and more than 1000. Ingenuity Pathway Analysis mainly revealed canonical pathways of cellular immune response and cytokine signaling as well as a broad range of processes in cellular and organismal growth, proliferation and development, cell signaling, biosynthesis and metabolism. Significant changes in the expression profiles of PBMCs already occurred very early after immune stimulation. At two hours after the first vaccination 679 DE-genes corresponding to 110 canonical pathways of cytokine signaling, cellular immune response and other multiple cellular functions were found. Immune competence and global disease resistance are heritable but difficult to measure and to address by breeding. Besides QTL mapping of immune traits gene expression profiling facilitates the detection of functional gene networks and thus functional candidate genes.

Effect of combined oxidative and nitrosative stresses on Staphylococcus aureus transcriptome

Staphylococcus aureus is a pathogen responsible for severe community- and nosocomially acquired infections. To fight pathogen intrusion, the innate immune system uses a plethora of weapons, with the generation of oxidative and nitrosative stresses among the most efficient. In this work, the S. aureus genome-wide transcriptional responses to oxidative stress generated by hydrogen peroxide, to nitrosative stress imposed by S-nitrosoglutathione (GSNO), and to the combination of the two were investigated using microarray analysis. The results showed that these stresses have a significant impact on the transcriptome of S. aureus. Hydrogen peroxide modified mainly the mRNA abundance of genes involved in oxidative detoxification and DNA metabolism, which together represent 14 % of the total number of upregulated genes. GSNO caused significant alteration of the expression of gene products with regulatory function. However, the simultaneous addition of GSNO and hydrogen peroxide was found to cause the more significant transcriptomic alteration, affecting ∼10 % of the total transcriptome. In particular, exposure of S. aureus to GSNO plus hydrogen peroxide modified the transcription of genes associated with cell envelope and iron metabolism, including induction of ftnA and dps genes that encode iron-storage and oxidative-protecting proteins. Further studies revealed that when exposed to combined GSNO-hydrogen peroxide stresses, S. aureus has decreased viability, which is enhanced in the presence of iron, and low siderophore activity. Altogether, this study revealed, for the first time, how the combined oxidative and nitrosative stresses inflicted during phagocytosis interfere at the transcriptional level with the S. aureus cellular metabolism.

Cytokine Response Is Determined by Duration of Receptor and Signal Transducers and Activators of Transcription 3 (STAT3) Activation

Paradoxically, the pro-inflammatory cytokine IL-6 and the anti-inflammatory cytokine IL-10 both activate STAT3, yet generate nearly opposing cellular responses. Here, we show that the temporal pattern of STAT3 activation codes for the specific cytokine response. A computational model of IL-6 and IL-10 signaling predicted that IL-6 stimulation results in transient activation of STAT3, with a rapid decline in phosphorylation and nuclear localization. In contrast, simulated IL-10 signaling resulted in sustained STAT3 activation. The predicted STAT3 patterns produced by each cytokine were confirmed experimentally in human dendritic cells. Time course microarray studies further showed that the dynamic genome-wide transcriptional responses were nearly identical at early time points following stimulation (when STAT3 is active in response to both IL-6 and IL-10) but divergent at later times (when STAT3 is active only in response to IL-10). Truncating STAT3 activation after IL-10 stimulation caused IL-10 to elicit an IL-6-like transcriptional and secretory response. That the duration of IL-10 receptor and STAT3 activation can direct distinct responses reveals a complex cellular information-coding mechanism that may be relevant to improving the prediction of the effects of drug candidates using this mechanism.

Multiphasic adaptation of the transcriptome of Saccharomyces cerevisiae to heat stress

The genome-wide transcriptional response of Saccharomyces cerevisiae to a temperature shift from 28°C to 41°C – a temperature at which cell viability was not impaired, but growth rate was – was determined over a period of 6h under well-controlled conditions. Care was taken not to introduce other perturbations. The data shows that the cells respond by rapidly changing their gene expression. Two phases were observed in this response: A short term transitional response during the first hour in which cell growth ceased, followed by a transcriptionally stable phase that was growth permissive. The initial response was broadest, involving almost half of all the ORFs being induced or repressed to a statistically significant level (here 1.5 fold). Functional analysis showed that genes involved in energy transduction, including trehalose metabolism, and molecular-chaperone encoding genes were induced most prominently. The latter set of genes presumably functions to counteract the effect of the temperature increase on protein denaturation. Furthermore, genes encoding parts of the translation- and transcription systems were repressed temporarily, in line with the observed lag phase in growth. After the initial adaptation phase a new stable state of gene expression was attained, compatible with cell growth. This new transcriptome was characterised by a continued induction of stress response genes, except for those involved in trehalose biosynthesis and a normalisation of the expression of genes coding for the transcription and translation machinery. Finally, our data, together with information available from the literature, show that the adaptation of specific cellular processes to severe temperature stress can occur at different hierarchical control levels. While trehalose metabolism seems to be adjusted mainly at the transcriptional level, the metabolic or post-transcriptional level seems to be more significant for the adaptation of parts of glycolysis and respiration.

A map of cell type-specific auxin responses.

In plants, changes in local auxin concentrations can trigger a range of developmental processes as distinct tissues respond differently to the same auxin stimulus. However, little is known about how auxin is interpreted by individual cell types. We performed a transcriptomic analysis of responses to auxin within four distinct tissues of the Arabidopsis thaliana root and demonstrate that different cell types show competence for discrete responses. The majority of auxin-responsive genes displayed a spatial bias in their induction or repression. The novel data set was used to examine how auxin influences tissue-specific transcriptional regulation of cell-identity markers. Additionally, the data were used in combination with spatial expression maps of the root to plot a transcriptomic auxin-response gradient across the apical and basal meristem. The readout revealed a strong correlation for thousands of genes between the relative response to auxin and expression along the longitudinal axis of the root. This data set and comparative analysis provide a transcriptome-level spatial breakdown of the response to auxin within an organ where this hormone mediates many aspects of development.

Genome-wide transcriptional responses of Escherichia coli to glyphosate, a potent inhibitor of the shikimate pathway enzyme 5-enolpyruvylshikimate-3-phosphate synthase

The shikimate pathway enzymes offer attractive targets for the development of antimetabolites. Glyphosate is an effective antimetabolite that inhibits 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase in the shikimate pathway, thereby resulting in a shortage of the chorismate-derived essential aromatic amino acids. However, little is known about the genome-wide transcriptional responses of bacteria to glyphosate shock. In the current study, a transcriptome analysis of Escherichia coli (E. coli) exposed to glyphosate identified the differential expression of 1040 genes, which represent 23.2% of the genome. The differentially expressed genes are primarily involved in amino acid metabolism, cell motility, and central carbon metabolism, indicating that the impact of glyphosate on the shikimate pathway also extends to other metabolic pathways. Expectedly, almost all genes encoding the proteins for the shikimate and specific aromatic amino acid pathways were downregulated after the addition of glyphosate. Furthermore, the expression of many energy- and metabolism-related genes was repressed. In contrast, glyphosate treatment induced the coordinated upregulation of at least 50 genes related to cell motility and chemotaxis. The reverse transcription-quantitative real-time PCR (RT-qPCR) data showed that the expression profiles of selected genes from the referred pathways were found to be consistent with the microarray data. The results suggest that the presence of glyphosate during growth induces metabolic starvation, an energy drain and other non-target effects.

Modeling Transcriptome Dynamics in a Complex World

An accurate prediction of how extrinsic stimuli influence changes in gene expression has been challenging. In this issue, Nagano and colleagues successfully model genome-wide mRNA expression changes under variable environmental conditions in rice, raising hopes that scientists will soon be able to predict genome-wide transcriptional responses in a variety of organisms in uncontrolled real-world settings.

Examination of the genome-wide transcriptional response of Escherichia coli O157:H7 to cinnamaldehyde exposure.

Cinnamaldehyde is a natural antimicrobial that has been found to be effective against many food-borne pathogens, including Escherichia coli O157:H7. Although its antimicrobial effects have been well investigated, limited information is available on its effects at the molecular level. Sublethal treatment at 200 mg/liter cinnamaldehyde inhibited growth of E. coli O157:H7 at 37°C and for ≤2 h caused cell elongation, but from 2 to 4 h growth resumed and cells reverted to normal length. To understand this transient behavior, genome-wide transcriptional analysis of E. coli O157:H7 was performed at 2 and 4 h of exposure to cinnamaldehyde in conjunction with reverse-phase high-performance liquid chromatography (RP-HPLC) analysis for cinnamaldehyde and other cinnamic compounds. Drastically different gene expression profiles were obtained at 2 and 4 h. RP-HPLC analysis showed that cinnamaldehyde was structurally stable for at least 2 h. At 2 h of exposure, cinnamaldehyde induced expression of many oxidative stress-related genes and repressed expression of DNA, protein, O-antigen, and fimbrial synthetic genes. At 4 h, many cinnamaldehyde-induced repressive effects on E. coli O157:H7 gene expression were reversed, and cells became more motile and grew at a slightly higher rate. Data indicated that by 4 h, E. coli O157:H7 was able to convert cinnamaldehyde into the less toxic cinnamic alcohol using dehydrogenase/reductase enzymes (YqhD and DkgA). This is the first study to characterize the ability of E. coli O157:H7 to convert cinnamaldehyde into cinnamic alcohol which, in turn, showed that the antimicrobial activity of cinnamaldehyde is mainly attributable to its carbonyl aldehyde group.

Cellular Plasticity Enables Adaptation to Unforeseen Cell-Cycle Rewiring Challenges

The fundamental dynamics of the cell cycle, underlying cell growth and reproduction, were previously found to be robust under a wide range of environmental and internal perturbations. This property was commonly attributed to its network structure, which enables the coordinated interactions among hundreds of proteins. Despite significant advances in deciphering the components and autonomous interactions of this network, understanding the interfaces of the cell cycle with other major cellular processes is still lacking. To gain insight into these interfaces, we used the process of genome-rewiring in yeast by placing an essential metabolic gene HIS3 from the histidine biosynthesis pathway, under the exclusive regulation of different cell-cycle promoters. In a medium lacking histidine and under partial inhibition of the HIS3p, the rewired cells encountered an unforeseen multitasking challenge; the cell-cycle regulatory genes were required to regulate the essential histidine-pathway gene in concert with the other metabolic demands, while simultaneously driving the cell cycle through its proper temporal phases. We show here that chemostat cell populations with rewired cell-cycle promoters adapted within a short time to accommodate the inhibition of HIS3p and stabilized a new phenotypic state. Furthermore, a significant fraction of the population was able to adapt and grow into mature colonies on plates under such inhibiting conditions. The adapted state was shown to be stably inherited across generations. These adaptation dynamics were accompanied by a non-specific and irreproducible genome-wide transcriptional response. Adaptation of the cell-cycle attests to its multitasking capabilities and flexible interface with cellular metabolic processes and requirements. Similar adaptation features were found in our previous work when rewiring HIS3 to the GAL system and switching cells from galactose to glucose. Thus, at the basis of cellular plasticity is the emergence of a yet-unknown general, non-specific mechanism allowing fast inherited adaptation to unforeseen challenges.

Genome-wide transcriptional response of Populus euphratica to long-term drought stress

Populus euphratica is native to semi-arid regions of the Xinjiang Uyghur Autonomous Region of China, and studying its drought responses will greatly increase the understanding of how trees acclimate to drought. Water was withheld for seven weeks in four different drought stress treatments, with regime 1 being the least drought stressed and regime 4 being the most, and the poplar's transcriptional profiles examined with Affymetrix Poplar GeneChip microarrays. The number of significantly up or down transcriptional changes increased with the severity of drought stress, with regime 1, 2, 3 and 4 showing 952, 1354, 2138 and 2360 altered transcripts, respectively. Only 277 of these were found in common across all four regimes, while 1938 transcripts were found to be unique to the individual treatments. Genes with altered transcript abundance included members of the transcription factor families AP2/EREPB, bZIP, NAC, NF-Y, WRKY, MYB and Homeobox, as well as genes for the small HSP, HSP70 and HSP90 heat shock protein families. Analysis of the transcript data from these experiments indicated that P. euphratica activates specific regulatory pathways according to the degree of drought stress it receives. These results provide important insights into the molecular mechanisms underpinning the drought stress responses of poplar, as well as providing candidates for future experimentation.

Insecticide resistance mechanisms in the green peach aphid Myzus persicae (Hemiptera: Aphididae) I: A transcriptomic survey.

BackgroundInsecticide resistance is one of the best examples of rapid micro-evolution found in nature. Since the development of the first synthetic insecticide in 1939, humans have invested considerable effort to stay ahead of resistance phenotypes that repeatedly develop in insects. Aphids are a group of insects that have become global pests in agriculture and frequently exhibit insecticide resistance. The green peach aphid, Myzus persicae, has developed resistance to at least seventy different synthetic compounds, and different insecticide resistance mechanisms have been reported worldwide.Methodology/Principal FindingsTo further characterize this resistance, we analyzed genome-wide transcriptional responses in three genotypes of M. persicae, each exhibiting different resistance mechanisms, in response to an anti-cholinesterase insecticide. The sensitive genotype (exhibiting no resistance mechanism) responded to the insecticide by up-regulating 183 genes primarily ones related to energy metabolism, detoxifying enzymes, proteins of extracellular transport, peptidases and cuticular proteins. The second genotype (resistant through a kdr sodium channel mutation), up-regulated 17 genes coding for detoxifying enzymes, peptidase and cuticular proteins. Finally, a multiply resistant genotype (carrying kdr and a modified acetylcholinesterase), up-regulated only 7 genes, appears not to require induced insecticide detoxification, and instead down-regulated many genes.Conclusions/SignificanceThis study suggests strongly that insecticide resistance in M. persicae is more complex that has been described, with the participation of a broad array of resistance mechanisms. The sensitive genotype exhibited the highest transcriptional plasticity, accounting for the wide range of potential adaptations to insecticides that this species can evolve. In contrast, the multiply resistant genotype exhibited a low transcriptional plasticity, even for the expression of genes encoding enzymes involved in insecticide detoxification. Our results emphasize the value of microarray studies to search for regulated genes in insects, but also highlights the many ways those different genotypes can assemble resistant phenotypes depending on the environmental pressure.

Gene expression profiling of Bangpungtongseong-san (Bofutsushosan) and Bangkihwangki-tang (Boiogito) administered individuals

Bangpungtongseong-san (Bofutsushosan, BTS) and Bangkihwangki-tang (Boiogito, BOT) are widely prescribed Korean, Japanese and Chinese traditional herbal medicines used to clinically prevent or improve obesity. In this study, we investigated genomewide transcriptional response to administration of either BTS or BOT to the Korean obese adults. From the blood samples, peripheral blood mononuclear cells (PBMCs) were isolated and RNA samples extracted from PBMCs were processed to conduct DNA microarray. Analysis of DNA microarray data revealed that administration of the both BTS and BOT evokes gene expression profile changes of PBMCs. In addition, comparative analysis between the drugs and placebo identified sets of specific genes regulated by BTS or BOT. Interestingly, analysis of gene list using functional annotation tool revealed that PMBCs of BOT-prescribed subjects have increased expression level of genes which are localized to mitochondria. The fact that mitochondrion is the main organelle of regulating cellular energy metabolism suggests that induced expressions of mitochondrial genes by BOT administration might be responsible for the anti-obesity effect of the medicine. We expect that further study based on our finding will provide molecular mechanisms of BTS and BOT effects on obesity.

Transcriptome profiling of Zymomonas mobilis under furfural stress

Furfural from lignocellulosic hydrolysates is the prevalent inhibitor to microorganisms during cellulosic ethanol production, but the molecular mechanisms of tolerance to this inhibitor in Zymomonas mobilis are still unclear. In this study, genome-wide transcriptional responses to furfural were investigated in Z. mobilis using microarray analysis. We found that 433 genes were differentially expressed in response to furfural. Furfural up- or down-regulated genes related to cell wall/membrane biogenesis, metabolism, and transcription. However, furfural has a subtle negative effect on Entner-Doudoroff pathway mRNAs. Our results revealed that furfural had effects on multiple aspects of cellular metabolism at the transcriptional level and that membrane might play important roles in response to furfural. This research has provided insights into the molecular response to furfural in Z. mobilis, and it will be helpful to construct more furfural-resistant strains for cellulosic ethanol production.

Transcriptome profiling of Zymomonas mobilis under ethanol stress

BackgroundHigh tolerance to ethanol is a desirable characteristics for ethanologenic strains used in industrial ethanol fermentation. A deeper understanding of the molecular mechanisms underlying ethanologenic strains tolerance of ethanol stress may guide the design of rational strategies to increase process performance in industrial alcoholic production. Many extensive studies have been performed in Saccharomyces cerevisiae and Escherichia coli. However, the physiological basis and genetic mechanisms involved in ethanol tolerance for Zymomonas mobilis are poorly understood on genomic level. To identify the genes required for tolerance to ethanol, microarray technology was used to investigate the transcriptome profiling of the ethanologenic Z. mobilis in response to ethanol stress.ResultsWe successfully identified 127 genes which were differentially expressed in response to ethanol. Ethanol up- or down-regulated genes related to cell wall/membrane biogenesis, metabolism, and transcription. These genes were classified as being involved in a wide range of cellular processes including carbohydrate metabolism, cell wall/membrane biogenesis, respiratory chain, terpenoid biosynthesis, DNA replication, DNA recombination, DNA repair, transport, transcriptional regulation, some universal stress response, etc.ConclusionIn this study, genome-wide transcriptional responses to ethanol were investigated for the first time in Z. mobilis using microarray analysis.Our results revealed that ethanol had effects on multiple aspects of cellular metabolism at the transcriptional level and that membrane might play important roles in response to ethanol. Although the molecular mechanism involved in tolerance and adaptation of ethanologenic strains to ethanol is still unclear, this research has provided insights into molecular response to ethanol in Z. mobilis. These data will also be helpful to construct more ethanol resistant strains for cellulosic ethanol production in the future.

Influence of oxygen limitation, absence of the cytochrome bc1 complex and low pH on global gene expression in Gluconobacter oxydans 621H using DNA microarray technology

The genome-wide transcriptional responses of the strictly aerobic α-proteobacterium Gluconobacter oxydans 621H to oxygen limitation, to the absence of the cytochrome bc(1) complex, and to low pH were studied using DNA microarray analyses. Oxygen limitation caused expression changes of 486 genes, representing 20% of the chromosomal genes. Genes with an increased mRNA level included those for terminal oxidases, the cytochrome bc(1) complex, transhydrogenase, two alcohol dehydrogenases, heme biosynthesis, PTS proteins, proteins involved in cyclic diGMP synthesis and degradation, two sigma factors, flagella and chemotaxis proteins, several stress proteins, and a putative exporter protein. The downregulated genes comprised those for respiratory dehydrogenases, enzymes of central metabolism, PQQ biosynthesis, outer membrane receptors, Sec proteins, and proteins involved in transcription and translation. A ΔqrcABC mutant of G. oxydans showed a growth defect during cultivation on mannitol at pH 4 under oxygen saturation. Comparison of the transcriptomes of this mutant versus the wild type under these conditions revealed 51 differentially expressed genes. Interestingly, almost all of the 45 genes with increased expression in the ΔqrcABC mutant at pH 4 were also upregulated in the wild type grown at pH 6 under oxygen limitation. These results support an active role of the cytochrome bc(1) complex in G. oxydans respiration. The transcriptome comparison of G. oxydans wild type at pH 4 versus pH 6 in mannitol medium under oxygen-saturated conditions uncovered only 72 differentially expressed genes. The 35 upregulated genes included those for cytochrome bd oxidase, major polyol dehydrogenase, iron storage and oxidative stress proteins. Among the 37 downregulated genes were some encoding enzymes dealing with carbon dioxide, such as biotin carboxylase, biotin carboxyl carrier protein, and carboanhydrase. These results give first insights into global transcriptional responses of G. oxydans.

The Global Transcriptional Response of Fission Yeast to Hydrogen Sulfide

BackgroundHydrogen sulfide (H2S) is a newly identified member of the small family of gasotransmitters that are endogenous gaseous signaling molecules that have a fundamental role in human biology and disease. Although it is a relatively recent discovery and the mechanism of H2S activity is not completely understood, it is known to be involved in a number of cellular processes; H2S can affect ion channels, transcription factors and protein kinases in mammals.Methodology/Principal FindingsIn this paper, we have used fission yeast as a model organism to study the global gene expression profile in response to H2S by microarray. We initially measured the genome-wide transcriptional response of fission yeast to H2S. Through the functional classification of genes whose expression profile changed in response to H2S, we found that H2S mainly influences genes that encode putative or known stress proteins, membrane transporters, cell cycle/meiotic proteins, transcription factors and respiration protein in the mitochondrion. Our analysis showed that there was a significant overlap between the genes affected by H2S and the stress response. We identified that the target genes of the MAPK pathway respond to H2S; we also identified that a number of transporters respond to H2S, these include sugar/carbohydrate transporters, ion transporters, and amino acid transporters. We found many mitochondrial genes to be down regulated upon H2S treatment and that H2S can reduce mitochondrial oxygen consumption.Conclusion/SignificanceThis study identifies potential molecular targets of the signaling molecule H2S in fission yeast and provides clues about the identity of homologues human proteins and will further the understanding of the cellular role of H2S in human diseases.

Molecular analysis of melatonin-induced changes in breast cancer cells: microarray study of anti-cancer effect of melatonin

Melatonin is a pineal secretory product that acts as a chemical messenger for the dark/light cycle and plays a central role in synchronization of circadian rhythms. Melatonin has been reported to exhibit a variety of therapeutic effects, including antiaging, anti-oxidative, anti-inflammation, and anticancer (such as induction of tumor suppressor genes and suppression of the metabolism of tumor cells, including breast cancer cells). In this study, we investigated the genome-wide transcriptional responses of MCF-7 human breast cancer cells exposed to melatonin by microarray gene expression profiling. We identified 1,946 and 983 genes that were 2-fold up- or down-regulated within 72 h of 1 nM and 100 nM melatonin treatment. Gene Ontology (GO) enrichment analysis was performed for identification of the biological functions and biological processes affected by the differential expression of genes in melatonin-stimulated cells. The differentially expressed genes that were dysregulated in many biological functions included cell proliferation, immune responses, translation, cell adhesion, apoptosis, and cell cycle. Our findings support the view that melatonin-stimulated changes in gene expression contribute to the anticancer effect of melatonin in breast cancer cells.

Ligand-Independent Phosphorylation of the Glucocorticoid Receptor Integrates Cellular Stress Pathways with Nuclear Receptor Signaling

Glucocorticoids are stress hormones that maintain homeostasis through gene regulation mediated by nuclear receptors. We have discovered that other cellular stressors are integrated with glucocorticoid signaling through a new hormone-independent phosphorylation site, Ser134, on the human glucocorticoid receptor (GR). Ser134 phosphorylation is induced by a variety of stress-activating stimuli in a p38 mitogen-activated protein kinase (MAPK)-dependent manner. Cells expressing a mutant glucocorticoid receptor incapable of phosphorylation at Ser134 (S134A-GR) had significantly altered hormone-dependent genome-wide transcriptional responses and associated hormone-mediated cellular functions. The phosphorylation of Ser134 significantly increased the association of the GR with the zeta isoform of the 14-3-3 class of signaling proteins (14-3-3zeta) on chromatin promoter regions, resulting in a blunted hormone-dependent transcriptional response of select genes. These data argue that the phosphorylation of Ser134 acts as a molecular sensor on the GR, monitoring the level of cellular stress to redirect glucocorticoid-regulated signaling through altered 14-3-3zeta cofactor binding and promoter recruitment. This posttranslational modification allows prior cellular stress signals to dictate the transcriptional response to glucocorticoids.