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Sample records for airway gene expression

  1. Regulation of Airway Mucin Gene Expression

    PubMed Central

    Thai, Philip; Loukoianov, Artem; Wachi, Shinichiro; Wu, Reen

    2015-01-01

    Mucins are important components that exert a variety of functions in cell-cell interaction, epidermal growth factor receptor signaling, and airways protection. In the conducting airways of the lungs, mucins are the major contributor to the viscoelastic property of mucous secretion, which is the major barrier to trapping inhaled microbial organism, particulates, and oxidative pollutants. The homeostasis of mucin production is an important feature in conducting airways for the maintenance of mucociliary function. Aberrant mucin secretion and accumulation in airway lumen are clinical hallmarks associated with various lung diseases, such as asthma, chronic obstructive pulmonary disease, cystic fibrosis, emphysema, and lung cancer. Among 20 known mucin genes identified, 11 of them have been verified at either the mRNA and/or protein level in airways. The regulation of mucin genes is complicated, as are the mediators and signaling pathways. This review summarizes the current view on the mediators, the signaling pathways, and the transcriptional units that are involved in the regulation of airway mucin gene expression. In addition, we also point out essential features of epigenetic mechanisms for the regulation of these genes. PMID:17961085

  2. Altered Epithelial Gene Expression in Peripheral Airways of Severe Asthma

    PubMed Central

    Singhania, Akul; Rupani, Hitasha; Jayasekera, Nivenka; Lumb, Simon; Hales, Paul; Gozzard, Neil; Davies, Donna E.

    2017-01-01

    Management of severe asthma remains a challenge despite treatment with glucocorticosteroid therapy. The majority of studies investigating disease mechanisms in treatment-resistant severe asthma have previously focused on the large central airways, with very few utilizing transcriptomic approaches. The small peripheral airways, which comprise the majority of the airway surface area, remain an unexplored area in severe asthma and were targeted for global epithelial gene expression profiling in this study. Differences between central and peripheral airways were evaluated using transcriptomic analysis (Affymetrix HG U133 plus 2.0 GeneChips) of epithelial brushings obtained from severe asthma patients (N = 17) and healthy volunteers (N = 23). Results were validated in an independent cohort (N = 10) by real-time quantitative PCR. The IL-13 disease signature that is associated with an asthmatic phenotype was upregulated in severe asthmatics compared to healthy controls but was predominantly evident within the peripheral airways, as were genes related to mast cell presence. The gene expression response associated with glucocorticosteroid therapy (i.e. FKBP5) was also upregulated in severe asthmatics compared to healthy controls but, in contrast, was more pronounced in central airways. Moreover, an altered epithelial repair response (e.g. FGFBP1) was evident across both airway sites reflecting a significant aspect of disease in severe asthma unadressed by current therapies. A transcriptomic approach to understand epithelial activation in severe asthma has thus highlighted the need for better-targeted therapy to the peripheral airways in severe asthma, where the IL-13 disease signature persists despite treatment with currently available therapy. PMID:28045928

  3. Altered Epithelial Gene Expression in Peripheral Airways of Severe Asthma.

    PubMed

    Singhania, Akul; Rupani, Hitasha; Jayasekera, Nivenka; Lumb, Simon; Hales, Paul; Gozzard, Neil; Davies, Donna E; Woelk, Christopher H; Howarth, Peter H

    2017-01-01

    Management of severe asthma remains a challenge despite treatment with glucocorticosteroid therapy. The majority of studies investigating disease mechanisms in treatment-resistant severe asthma have previously focused on the large central airways, with very few utilizing transcriptomic approaches. The small peripheral airways, which comprise the majority of the airway surface area, remain an unexplored area in severe asthma and were targeted for global epithelial gene expression profiling in this study. Differences between central and peripheral airways were evaluated using transcriptomic analysis (Affymetrix HG U133 plus 2.0 GeneChips) of epithelial brushings obtained from severe asthma patients (N = 17) and healthy volunteers (N = 23). Results were validated in an independent cohort (N = 10) by real-time quantitative PCR. The IL-13 disease signature that is associated with an asthmatic phenotype was upregulated in severe asthmatics compared to healthy controls but was predominantly evident within the peripheral airways, as were genes related to mast cell presence. The gene expression response associated with glucocorticosteroid therapy (i.e. FKBP5) was also upregulated in severe asthmatics compared to healthy controls but, in contrast, was more pronounced in central airways. Moreover, an altered epithelial repair response (e.g. FGFBP1) was evident across both airway sites reflecting a significant aspect of disease in severe asthma unadressed by current therapies. A transcriptomic approach to understand epithelial activation in severe asthma has thus highlighted the need for better-targeted therapy to the peripheral airways in severe asthma, where the IL-13 disease signature persists despite treatment with currently available therapy.

  4. Molecular mechanisms controlling CFTR gene expression in the airway

    PubMed Central

    Zhang, Zhaolin; Ott, Christopher J; Lewandowska, Marzena A; Leir, Shih-Hsing; Harris, Ann

    2012-01-01

    Abstract The low levels of CFTR gene expression and paucity of CFTR protein in human airway epithelial cells are not easily reconciled with the pivotal role of the lung in cystic fibrosis pathology. Previous data suggested that the regulatory mechanisms controlling CFTR gene expression might be different in airway epithelium in comparison to intestinal epithelium where CFTR mRNA and protein is much more abundant. Here we examine chromatin structure and modification across the CFTR locus in primary human tracheal (HTE) and bronchial (NHBE) epithelial cells and airway cell lines including 16HBE14o- and Calu3. We identify regions of open chromatin that appear selective for primary airway epithelial cells and show that several of these are enriched for a histone modification (H3K4me1) that is characteristic of enhancers. Consistent with these observations, three of these sites encompass elements that have cooperative enhancer function in reporter gene assays in 16HBE14o- cells. Finally, we use chromosome conformation capture (3C) to examine the three-dimensional structure of nearly 800 kb of chromosome 7 encompassing CFTR and observe long-range interactions between the CFTR promoter and regions far outside the locus in cell types that express high levels of CFTR. PMID:21895967

  5. Airway Epithelial Expression Quantitative Trait Loci Reveal Genes Underlying Asthma and Other Airway Diseases

    PubMed Central

    Luo, Wei; Obeidat, Ma’en; Di Narzo, Antonio Fabio; Chen, Rong; Sin, Don D.; Paré, Peter D.

    2016-01-01

    Genome-wide association studies (GWASs) have identified loci that are robustly associated with asthma and related phenotypes; however, the molecular mechanisms underlying these associations need to be explored. The most relevant tissues to study the functional consequences of asthma are the airways. We used publically available data to derive expression quantitative trait loci (eQTLs) for human epithelial cells from small and large airways and applied the eQTLs in the interpretation of GWAS results of asthma and related phenotypes. For the small airways (n = 105), we discovered 660 eQTLs at a 10% false discovery rate (FDR), among which 315 eQTLs were not previously reported in a large-scale eQTL study of whole lung tissue. A large fraction of the identified eQTLs is supported by data from Encyclopedia of DNA Elements (ENCODE) showing that the eQTLs reside in regulatory elements (57.5 and 67.6% of cis- and trans-eQTLs, respectively). Published pulmonary GWAS hits were enriched as airway epithelial eQTLs (9.2-fold). Further, genes regulated by asthma GWAS loci in epithelium are significantly enriched in immune response pathways, such as IL-4 signaling (FDR, 5.2 × 10−4). The airway epithelial eQTLs described in this study are complementary to previously reported lung eQTLs and represent a powerful resource to link GWAS-associated variants to their regulatory function and thus elucidate the molecular mechanisms underlying asthma and airway-related conditions. PMID:26102239

  6. Airway gene expression of IL-1 pathway mediators predicts exacerbation risk in obstructive airway disease

    PubMed Central

    Baines, Katherine J; Fu, Juan-juan; McDonald, Vanessa M; Gibson, Peter G

    2017-01-01

    Background Exacerbations of asthma and COPD are a major cause of morbidity and mortality and are responsible for significant health care costs. This study further investigates interleukin (IL)-1 pathway activation and its relationship with exacerbations of asthma and COPD. Methods In this prospective cohort study, 95 participants with stable asthma (n=35) or COPD (n=60) were recruited and exacerbations recorded over the following 12 months. Gene expressions of IL-1 pathway biomarkers, including the IL-1 receptors (IL1R1, IL1R2, and IL1RN), and signaling molecules (IRAK2, IRAK3, and PELI1), were measured in sputum using real-time quantitative polymerase chain reaction. Mediators were compared between the frequent (≥2 exacerbations in the 12 months) and infrequent exacerbators, and the predictive relationships investigated using receiver operating characteristic curves and area under the curve (AUC) values. Results Of the 95 participants, 89 completed the exacerbation follow-up, where 30 participants (n=22 COPD, n=8 asthma) had two or more exacerbations. At the baseline visit, expressions of IRAK2, IRAK3, PELI1, and IL1R1 were elevated in participants with frequent exacerbations of both asthma and COPD combined and separately. In the combined population, sputum gene expression of IRAK3 (AUC=75.4%; P<0.001) was the best predictor of future frequent exacerbations, followed by IL1R1 (AUC=72.8%; P<0.001), PELI1 (AUC=71.2%; P<0.001), and IRAK2 (AUC=68.6; P=0.004). High IL-1 pathway gene expression was associated with frequent prior year exacerbations and correlated with the number and severity of exacerbations. Conclusion The upregulation of IL-1 pathway mediators is associated with frequent exacerbations of obstructive airway disease. Further studies should investigate these mediators as both potential diagnostic biomarkers predicting at-risk patients and novel treatment targets. PMID:28223794

  7. Identification of Genes Expressed by Human Airway Eosinophils after an In Vivo Allergen Challenge

    PubMed Central

    Esnault, Stephane; Kelly, Elizabeth A.; Schwantes, Elizabeth A.; Liu, Lin Ying; DeLain, Larissa P.; Hauer, Jami A.; Bochkov, Yury A.; Denlinger, Loren C.; Malter, James S.; Mathur, Sameer K.; Jarjour, Nizar N.

    2013-01-01

    Background The mechanism for the contribution of eosinophils (EOS) to asthma pathophysiology is not fully understood. Genome-wide expression analysis of airway EOS by microarrays has been limited by the ability to generate high quality RNA from sufficient numbers of airway EOS. Objective To identify, by genome-wide expression analyses, a compendium of expressed genes characteristic of airway EOS following an in vivo allergen challenge. Methods Atopic, mild asthmatic subjects were recruited for these studies. Induced sputum was obtained before and 48h after a whole lung allergen challenge (WLAC). Individuals also received a segmental bronchoprovocation with allergen (SBP-Ag) 1 month before and after administering a single dose of mepolizumab (anti-IL-5 monoclonal antibody) to reduce airway EOS. Bronchoalveolar lavage (BAL) was performed before and 48 h after SBP-Ag. Gene expression of sputum and BAL cells was analyzed by microarrays. The results were validated by qPCR in BAL cells and purified BAL EOS. Results A total of 299 transcripts were up-regulated by more than 2-fold in total BAL cells following SBP-Ag. Mepolizumab treatment resulted in a reduction of airway EOS by 54.5% and decreased expression of 99 of the 299 transcripts. 3 of 6 post-WLAC sputum samples showed increased expression of EOS-specific genes, along with the expression of 361 other genes. Finally, the intersection of the 3 groups of transcripts (increased in BAL post SBP-Ag (299), decreased after mepolizumab (99), and increased in sputum after WLAC (365)) was composed of 57 genes characterizing airway EOS gene expression. Conclusion We identified 57 genes that were highly expressed by BAL EOS compared to unseparated BAL cells after in vivo allergen challenge. 41 of these genes had not been previously described in EOS and are thus potential new candidates to elucidate EOS contribution to airway biology. PMID:23844029

  8. Airway epithelial gene expression in the diagnostic evaluation of smokers with suspect lung cancer.

    PubMed

    Spira, Avrum; Beane, Jennifer E; Shah, Vishal; Steiling, Katrina; Liu, Gang; Schembri, Frank; Gilman, Sean; Dumas, Yves-Martine; Calner, Paul; Sebastiani, Paola; Sridhar, Sriram; Beamis, John; Lamb, Carla; Anderson, Timothy; Gerry, Norman; Keane, Joseph; Lenburg, Marc E; Brody, Jerome S

    2007-03-01

    Lung cancer is the leading cause of death from cancer in the US and the world. The high mortality rate (80-85% within 5 years) results, in part, from a lack of effective tools to diagnose the disease at an early stage. Given that cigarette smoke creates a field of injury throughout the airway, we sought to determine if gene expression in histologically normal large-airway epithelial cells obtained at bronchoscopy from smokers with suspicion of lung cancer could be used as a lung cancer biomarker. Using a training set (n = 77) and gene-expression profiles from Affymetrix HG-U133A microarrays, we identified an 80-gene biomarker that distinguishes smokers with and without lung cancer. We tested the biomarker on an independent test set (n = 52), with an accuracy of 83% (80% sensitive, 84% specific), and on an additional validation set independently obtained from five medical centers (n = 35). Our biomarker had approximately 90% sensitivity for stage 1 cancer across all subjects. Combining cytopathology of lower airway cells obtained at bronchoscopy with the biomarker yielded 95% sensitivity and a 95% negative predictive value. These findings indicate that gene expression in cytologically normal large-airway epithelial cells can serve as a lung cancer biomarker, potentially owing to a cancer-specific airway-wide response to cigarette smoke.

  9. Gene Delivery to the Airway

    PubMed Central

    Keiser, Nicholas W.; Engelhardt, John F.

    2013-01-01

    This unit describes generation of and gene transfer to several commonly used airway models. Isolation and transduction of primary airway epithelial cells are first described. Next, the preparation of polarized airway epithelial monolayers is outlined. Transduction of these polarized cells is also described. Methods are presented for generation of tracheal xenografts as well as both ex vivo and in vivo gene transfer to these xenografts. Finally, a method for in vivo gene delivery to the lungs of rodents is included. Methods for evaluating transgene expression are given in the support protocols. PMID:23853081

  10. POU2AF1 Functions in the Human Airway Epithelium To Regulate Expression of Host Defense Genes.

    PubMed

    Zhou, Haixia; Brekman, Angelika; Zuo, Wu-Lin; Ou, Xuemei; Shaykhiev, Renat; Agosto-Perez, Francisco J; Wang, Rui; Walters, Matthew S; Salit, Jacqueline; Strulovici-Barel, Yael; Staudt, Michelle R; Kaner, Robert J; Mezey, Jason G; Crystal, Ronald G; Wang, Guoqing

    2016-04-01

    In the process of seeking novel lung host defense regulators by analyzing genome-wide RNA sequence data from normal human airway epithelium, we detected expression of POU domain class 2-associating factor 1 (POU2AF1), a known transcription cofactor previously thought to be expressed only in lymphocytes. Lymphocyte contamination of human airway epithelial samples obtained by bronchoscopy and brushing was excluded by immunohistochemistry staining, the observation of upregulation of POU2AF1 in purified airway basal stem/progenitor cells undergoing differentiation, and analysis of differentiating single basal cell clones. Lentivirus-mediated upregulation of POU2AF1 in airway basal cells induced upregulation of host defense genes, including MX1, IFIT3, IFITM, and known POU2AF1 downstream genes HLA-DRA, ID2, ID3, IL6, and BCL6. Interestingly, expression of these genes paralleled changes of POU2AF1 expression during airway epithelium differentiation in vitro, suggesting POU2AF1 helps to maintain a host defense tone even in pathogen-free condition. Cigarette smoke, a known risk factor for airway infection, suppressed POU2AF1 expression both in vivo in humans and in vitro in human airway epithelial cultures, accompanied by deregulation of POU2AF1 downstream genes. Finally, enhancing POU2AF1 expression in human airway epithelium attenuated the suppression of host defense genes by smoking. Together, these findings suggest a novel function of POU2AF1 as a potential regulator of host defense genes in the human airway epithelium.

  11. Minicircle DNA Provides Enhanced and Prolonged Transgene Expression Following Airway Gene Transfer

    PubMed Central

    Munye, Mustafa M.; Tagalakis, Aristides D.; Barnes, Josephine L.; Brown, Rachel E.; McAnulty, Robin J.; Howe, Steven J.; Hart, Stephen L.

    2016-01-01

    Gene therapy for cystic fibrosis using non-viral, plasmid-based formulations has been the subject of intensive research for over two decades but a clinically viable product has yet to materialise in large part due to inefficient transgene expression. Minicircle DNA give enhanced and more persistent transgene expression compared to plasmid DNA in a number of organ systems but has not been assessed in the lung. In this study we compared minicircle DNA with plasmid DNA in transfections of airway epithelial cells. In vitro, luciferase gene expression from minicircles was 5–10-fold higher than with plasmid DNA. In eGFP transfections in vitro both the mean fluorescence intensity and percentage of cells transfected was 2–4-fold higher with minicircle DNA. Administration of equimolar amounts of DNA to mouse lungs resulted in a reduced inflammatory response and more persistent transgene expression, with luciferase activity persisting for 2 weeks from minicircle DNA compared to plasmid formulations. Transfection of equal mass amounts of DNA in mouse lungs resulted in a 6-fold increase in transgene expression in addition to more persistent transgene expression. Our findings have clear implications for gene therapy of airway disorders where plasmid DNA transfections have so far proven inefficient in clinical trials. PMID:26975732

  12. The role of reactive oxygen and nitrogen species in airway epithelial gene expression.

    PubMed Central

    Martin, L D; Krunkosky, T M; Voynow, J A; Adler, K B

    1998-01-01

    The body first encounters deleterious inhaled substances, such as allergens, industrial particles, pollutants, and infectious agents, at the airway epithelium. When this occurs, the epithelium and its resident inflammatory cells respond defensively by increasing production of cytokines, mucus, and reactive oxygen and nitrogen species (ROS/RNS). As inflammation in the airway increases, additional infiltrating cells increase the level of these products. Recent interest has focused on ROS/RNS as potential modulators of the expression of inflammation-associated genes important to the pathogenesis of various respiratory diseases. ROS/RNS appear to play a variety of roles that lead to changes in expression of genes such as interleukin-6 and intercellular adhesion molecule 1. By controlling this regulation, the reactive species can serve as exogenous stimuli, as intercellular signaling molecules, and as modulators of the redox state in epithelial cells. Unraveling the molecular mechanisms affected by ROS/RNS acting in these capacities should aid in the understanding of how stimulated defense mechanisms within the airway can lead to disease. Images Figure 1 PMID:9788898

  13. Quality control in microarray assessment of gene expression in human airway epithelium

    PubMed Central

    Raman, Tina; O'Connor, Timothy P; Hackett, Neil R; Wang, Wei; Harvey, Ben-Gary; Attiyeh, Marc A; Dang, David T; Teater, Matthew; Crystal, Ronald G

    2009-01-01

    Background Microarray technology provides a powerful tool for defining gene expression profiles of airway epithelium that lend insight into the pathogenesis of human airway disorders. The focus of this study was to establish rigorous quality control parameters to ensure that microarray assessment of the airway epithelium is not confounded by experimental artifact. Samples (total n = 223) of trachea, large and small airway epithelium were collected by fiberoptic bronchoscopy of 144 individuals and hybridized to Affymetrix microarrays. The pre- and post-chip quality control (QC) criteria established, included: (1) RNA quality, assessed by RNA Integrity Number (RIN) ≥ 7.0; (2) cRNA transcript integrity, assessed by signal intensity ratio of GAPDH 3' to 5' probe sets ≤ 3.0; and (3) the multi-chip normalization scaling factor ≤ 10.0. Results Of the 223 samples, all three criteria were assessed in 191; of these 184 (96.3%) passed all three criteria. For the remaining 32 samples, the RIN was not available, and only the other two criteria were used; of these 29 (90.6%) passed these two criteria. Correlation coefficients for pairwise comparisons of expression levels for 100 maintenance genes in which at least one array failed the QC criteria (average Pearson r = 0.90 ± 0.04) were significantly lower (p < 0.0001) than correlation coefficients for pairwise comparisons between arrays that passed the QC criteria (average Pearson r = 0.97 ± 0.01). Inter-array variability was significantly decreased (p < 0.0001) among samples passing the QC criteria compared with samples failing the QC criteria. Conclusion Based on the aberrant maintenance gene data generated from samples failing the established QC criteria, we propose that the QC criteria outlined in this study can accurately distinguish high quality from low quality data, and can be used to delete poor quality microarray samples before proceeding to higher-order biological analyses and interpretation. PMID:19852842

  14. Smoking-induced gene expression changes in the bronchial airway are reflected in nasal and buccal epithelium

    PubMed Central

    Sridhar, Sriram; Schembri, Frank; Zeskind, Julie; Shah, Vishal; Gustafson, Adam M; Steiling, Katrina; Liu, Gang; Dumas, Yves-Martine; Zhang, Xiaohui; Brody, Jerome S; Lenburg, Marc E; Spira, Avrum

    2008-01-01

    Background Cigarette smoking is a leading cause of preventable death and a significant cause of lung cancer and chronic obstructive pulmonary disease. Prior studies have demonstrated that smoking creates a field of molecular injury throughout the airway epithelium exposed to cigarette smoke. We have previously characterized gene expression in the bronchial epithelium of never smokers and identified the gene expression changes that occur in the mainstem bronchus in response to smoking. In this study, we explored relationships in whole-genome gene expression between extrathorcic (buccal and nasal) and intrathoracic (bronchial) epithelium in healthy current and never smokers. Results Using genes that have been previously defined as being expressed in the bronchial airway of never smokers (the "normal airway transcriptome"), we found that bronchial and nasal epithelium from non-smokers were most similar in gene expression when compared to other epithelial and nonepithelial tissues, with several antioxidant, detoxification, and structural genes being highly expressed in both the bronchus and nose. Principle component analysis of previously defined smoking-induced genes from the bronchus suggested that smoking had a similar effect on gene expression in nasal epithelium. Gene set enrichment analysis demonstrated that this set of genes was also highly enriched among the genes most altered by smoking in both nasal and buccal epithelial samples. The expression of several detoxification genes was commonly altered by smoking in all three respiratory epithelial tissues, suggesting a common airway-wide response to tobacco exposure. Conclusion Our findings support a relationship between gene expression in extra- and intrathoracic airway epithelial cells and extend the concept of a smoking-induced field of injury to epithelial cells that line the mouth and nose. This relationship could potentially be utilized to develop a non-invasive biomarker for tobacco exposure as well as a

  15. Distinct microRNA Expression in Human Airway Cells of Asthmatic Donors Identifies a Novel Asthma-associated Gene

    EPA Science Inventory

    Airway inflammation is the hallmark of asthma and suggests a dysregulation of homeostatic mechanisms. MicroRNAs (miRNAs) are key regulators of gene expression, necessary for the proper function of cellular processes. Here, we tested the hypothesis that differences between healthy...

  16. Oxidative Stress Regulates CFTR Gene Expression in Human Airway Epithelial Cells through a Distal Antioxidant Response Element

    PubMed Central

    Zhang, Zhaolin; Leir, Shih-Hsing

    2015-01-01

    Cystic fibrosis transmembrane conductance regulator gene (CFTR) expression in human airway epithelial cells involves the recruitment of distal cis-regulatory elements, which are associated with airway-selective DNase hypersensitive sites at −44 kb and −35 kb from the gene. The −35-kb site encompasses an enhancer that is regulated by the immune mediators interferon regulatory factor 1 and 2 and by nuclear factor Y. Here we investigate the −44-kb element, which also has enhancer activity in vitro in airway epithelial cells but is inactive in intestinal epithelial cells. This site contains an antioxidant response element (ARE) that plays a critical role in its function in airway cell lines and primary human bronchial epithelial cells. The natural antioxidant sulforaphane (SFN) induces nuclear translocation of nuclear factor, erythroid 2-like 2 (Nrf2), a transcription factor that regulates genes with AREs in their promoters, many of which are involved in response to injury. Under normal conditions, the −44-kb ARE is occupied by the repressor BTB and CNC homology 1, basic leucine zipper transcription factor (Bach1), and v-Maf avian musculoaponeurotic fibrosarcoma oncogene homolog K (MafK) heterodimers. After 2 hours of SFN treatment, Nrf2 displaces these repressive factors and activates CFTR expression. Site-directed mutagenesis shows that both the ARE and an adjacent NF-κB binding site are required for activation of the –44-kb element in airway epithelial cells. Moreover, this element is functionally linked to the −35-kb enhancer in modulating CFTR expression in response to environmental stresses in the airway. PMID:25259561

  17. Microarray gene expression analysis of the human airway in patients exposed to sulfur mustard.

    PubMed

    Najafi, Ali; Masoudi-Nejad, Ali; Imani Fooladi, Abbas Ali; Ghanei, Mostafa; Nourani, Mohamad Reza

    2014-08-01

    There is much data about the acute effects of sulfur mustard gas on humans, animals and cells. But less is known regarding the molecular basics of chronic complications in humans. Basically, mustard gas, as an alkylating agent, causes several chronic problems in the eyes, skin and more importantly in the pulmonary system which is the main cause of death. Although recent proteomic research has been carried out on bronchoalveolar lavage (BAL) and serum, but high-throughput transcriptomics have not yet been applied to chronic airway remodeling. This is the first cDNA-microarray report on the chronic human mustard lung disease, 25 years after exposure during the Iran-Iraq war. Microarray transcriptional profiling indicated that a total of 122 genes were significantly dysregulated in tissues located in the airway of patients. These genes are associated with the extracellular matrix components, apoptosis, stress response, inflammation and mucus secretion.

  18. Changes in Expression of Genes Regulating Airway Inflammation Following a High-Fat Mixed Meal in Asthmatics

    PubMed Central

    Li, Qian; Baines, Katherine J.; Gibson, Peter G.; Wood, Lisa G.

    2016-01-01

    Consumption of a high fat meal can increase neutrophilic airway inflammation in asthma subjects. This study investigates the molecular mechanisms driving airway neutrophilia following a high fat meal in asthmatics. Subjects with asthma (n = 11) and healthy controls (n = 8) consumed a high-fat/energy meal, containing total energy (TE) of 3846 kJ and 48 g of total fat (20.5 g saturated). Sputum was induced at 0 and 4 h, and gene expression was examined by microarray and quantitative real-time PCR (qPCR). Following the high fat dietary challenge, 168 entities were significantly differentially expressed greater than >1.5 fold in subjects with asthma, whereas, in healthy controls, only 14 entities were differentially expressed. Of the 168 genes that were changed in asthma, several biological processes were overrepresented, with 25 genes involved in “immune system processes”. qPCR confirmed that S100P, S100A16, MAL and MUC1 were significantly increased in the asthma group post-meal. We also observed a strong correlation and a moderate correlation between the change in NLRP12 and S100A16 gene expression at 4 h compared to baseline, and the change in total and saturated non-esterified plasma fatty acid levels at 2 h compared to baseline. In summary, our data identifies differences in inflammatory gene expression that may contribute to increased airway neutrophilia following a high fat meal in subjects with asthma and may provide useful therapeutic targets for immunomodulation. This may be particularly relevant to obese asthmatics, who are habitually consuming diets with a high fat content. PMID:26751474

  19. Immunological ignorance allows long-term gene expression after perinatal recombinant adeno-associated virus-mediated gene transfer to murine airways.

    PubMed

    Carlon, Marianne S; Vidović, Dragana; Dooley, James; da Cunha, Marina Mori; Maris, Michael; Lampi, Youlia; Toelen, Jaan; Van den Haute, Chris; Baekelandt, Veerle; Deprest, Jan; Verbeken, Erik; Liston, Adrian; Gijsbers, Rik; Debyser, Zeger

    2014-06-01

    Gene therapy of the lung has the potential to treat life-threatening diseases such as cystic fibrosis and α(1)-antitrypsin or surfactant deficiencies. A major hurdle for successful gene therapy is the development of an immune response against the transgene and/or viral vector. We hypothesized that by targeting the airways in the perinatal period, induction of an immune response against the vector particle could be prevented because of immaturity of the immune system, in turn allowing repeated gene transfer later in adult life to ensure long-term gene expression. Therefore, we readministered recombinant adeno-associated viral vector serotype 5 (rAAV2/5) to mouse airways 3 and 6 months after initial perinatal gene transfer. Our findings demonstrate that perinatal rAAV2/5-mediated gene transfer to the airways avoids a strong immune response. This immunological ignorance allows the readministration of an autologous vector later in adult life, resulting in efficient and stable gene transfer up to 7 months, without evidence of a decrease in transgene expression. Together, these data provide a basis to further explore perinatal gene therapy for pulmonary conditions with adequate gene expression up to 7 months.

  20. Widespread and efficient marker gene expression in the airway epithelia of fetal sheep after minimally invasive tracheal application of recombinant adenovirus in utero.

    PubMed

    Peebles, D; Gregory, L G; David, A; Themis, M; Waddington, S N; Knapton, H J; Miah, M; Cook, T; Lawrence, L; Nivsarkar, M; Rodeck, C; Coutelle, C

    2004-01-01

    Cystic fibrosis is a common lethal genetic disease caused by functional absence of the cystic fibrosis transmembrane conductance regulator (CFTR). Although a candidate disease for in utero gene therapy, demonstration of potentially therapeutic levels of transgene expression in the fetal airways after minimally invasive gene delivery is a mandatory prerequisite before application of this approach in humans can be considered. We report here on the delivery of a beta-galactosidase expressing adenovirus directly to the airways of fetal sheep in utero using ultrasound-guided percutaneous injection of the trachea in the fetal chest. Injection of adenoviral particles to the fetal airways was not associated with mortality and resulted in low-level expression in the peripheral airways. However, complexation of the virus with DEAE dextran, which confers a positive charge to the virus, and pretreatment of the airways with Na-caprate, which opens tight junctions, increased transgene expression, and a combination of these two enhancers resulted in widespread and efficient gene transfer of the fetal trachea and bronchial tree. Using a percutaneous ultrasound-guided injection technique, we have clearly demonstrated proof of principle for substantial transgene delivery to the fetal airways providing levels of gene expression that could be relevant for a therapeutic application of CFTR expressing vectors.

  1. Coordinate Control of Expression of Nrf2-Modulated Genes in the Human Small Airway Epithelium Is Highly Responsive to Cigarette Smoking

    PubMed Central

    Hübner, Ralf-Harto; Schwartz, Jamie D; De Bishnu, P; Ferris, Barbara; Omberg, Larsson; Mezey, Jason G; Hackett, Neil R; Crystal, Ronald G

    2009-01-01

    Nuclear factor erythroid 2–related factor 2 (Nrf2) is an oxidant-responsive transcription factor known to induce detoxifying and antioxidant genes. Cigarette smoke, with its large oxidant content, is a major stress on the cells of small airway epithelium, which are vulnerable to oxidant damage. We assessed the role of cigarette smoke in activation of Nrf2 in the human small airway epithelium in vivo. Fiberoptic bronchoscopy was used to sample the small airway epithelium in healthy-nonsmoker and healthy-smoker, and gene expression was assessed using microarrays. Relative to nonsmokers, Nrf2 protein in the small airway epithelium of smokers was activated and localized in the nucleus. The human homologs of 201 known murine Nrf2-modulated genes were identified, and 13 highly smoking-responsive Nrf2-modulated genes were identified. Construction of an Nrf2 index to assess the expression levels of these 13 genes in the airway epithelium of smokers showed coordinate control, an observation confirmed by quantitative PCR. This coordinate level of expression of the 13 Nrf2-modulated genes was independent of smoking history or demographic parameters. The Nrf2 index was used to identify two novel Nrf2-modulated, smoking-responsive genes, pirin (PIR) and UDP glucuronosyltransferase 1-family polypeptide A4 (UGT1A4). Both genes were demonstrated to contain functional antioxidant response elements in the promoter region. These observations suggest that Nrf2 plays an important role in regulating cellular defenses against smoking in the highly vulnerable small airway epithelium cells, and that there is variability within the human population in the Nrf2 responsiveness to oxidant burden. PMID:19593404

  2. Effects of Lupenone, Lupeol, and Taraxerol Derived from Adenophora triphylla on the Gene Expression and Production of Airway MUC5AC Mucin

    PubMed Central

    Yoon, Yong Pill; Lee, Hyun Jae; Lee, Dong-Ung; Lee, Sang Kook; Hong, Jang-Hee

    2015-01-01

    Background Adenophora triphylla var. japonica is empirically used for controlling airway inflammatory diseases in folk medicine. We evaluated the gene expression and production of mucin from airway epithelial cells in response to lupenone, lupeol and taraxerol derived from Adenophora triphylla var. japonica. Methods Confluent NCI-H292 cells were pretreated with lupenone, lupeol or taraxerol for 30 minutes and then stimulated with tumor necrosis factor α (TNF-α) for 24 hours. The MUC5AC mucin gene expression and production were measured by reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. Additionally, we examined whether lupenone, lupeol or taraxerol affects MUC5AC mucin production induced by epidermal growth factor (EGF) and phorbol 12-myristate 13-acetate (PMA), the other 2 stimulators of airway mucin production. Results Lupenone, lupeol, and taraxerol inhibited the gene expression and production of MUC5AC mucin induced by TNF-α from NCI-H292 cells, respectively. The 3 compounds inhibited the EGF or PMA-induced production of MUC5AC mucin in NCI-H292 cells. Conclusion These results indicated that lupenone, lupeol and taraxerol derived from Adenophora triphylla var. japonica regulates the production and gene expression of mucin, by directly acting on airway epithelial cells. In addition, the results partly explain the mechanism of of Adenophora triphylla var. japonica as a traditional remedy for diverse inflammatory pulmonary diseases. PMID:26175774

  3. Differential Roles of Hydrogen Peroxide in Adaptive and Inflammatory Gene Expression Induced by Exposure of Human Airway Epithelial Cells to Zn2+

    EPA Science Inventory

    Oxidant stress is believed to play an important role in particulate matter (PM)–mediated toxicity in the respiratory tract. Zinc (Zn2+) is a ubiquitous component of PM that has been shown to induce adverse responses such as inflammatory and adaptive gene expression in airway epit...

  4. Effects of 10 cigarette smoke condensates on primary human airway epithelial cells by comparative gene and cytokine expression studies.

    PubMed

    Pickett, Gavin; Seagrave, Jeanclare; Boggs, Susan; Polzin, Gregory; Richter, Patricia; Tesfaigzi, Yohannes

    2010-03-01

    Cigarettes vary in tobacco blend, filter ventilation, additives, and other physical and chemical properties, but little is known regarding potential differences in toxicity to a smoker's airway epithelia. We compared changes in gene expression and cytokine production in primary normal human bronchial epithelial cells following treatment for 18 h with cigarette smoke condensates (CSCs) prepared from five commercial and four research cigarettes, at doses of approximately 4 microg/ml nicotine. Nine of the CSCs were produced under a standard International Organization for Standardization smoking machine regimen and one was produced by a more intense smoking machine regimen. Isolated messenger RNA (mRNA) was analyzed by microarray hybridization, and media was analyzed for secreted cytokines and chemokines. Twenty-one genes were differentially expressed by at least 9 of the 10 CSCs by more than twofold, including genes encoding detoxifying and antioxidant proteins. Cytochrome P450, family 1, subfamily A, polypeptide 1 (CYP1A1) and NAD(P)H dehydrogenase, quinone 1 (NQO-1) were selected for validation with quantitative real-time PCR (qRT-PCR) and Western blot analyses. NQO-1 expression determined with microarrays, qRT-PCR, and Western blotting differed among the CSC types, with good correlation among the different assays. CYP1A1 mRNA levels varied substantially, but there was little correlation with the protein levels. For each CSC, the three most induced and three most repressed genes were identified. These genes may be useful as markers of exposure to that particular cigarette type. Furthermore, differences in interleukin-8 secretion were observed. These studies lay the foundation for future investigations to analyze differences in the responses of in vivo systems to tobacco products marketed with claims of reduced exposure or reduced harm.

  5. Differential Gene Expression Profiles and Selected Cytokine Protein Analysis of Mediastinal Lymph Nodes of Horses with Chronic Recurrent Airway Obstruction (RAO) Support an Interleukin-17 Immune Response

    PubMed Central

    2015-01-01

    Recurrent airway obstruction (RAO) is a pulmonary inflammatory condition that afflicts certain mature horses exposed to organic dust particulates in hay. Its clinical and pathological features, manifested by reversible bronchoconstriction, excessive mucus production and airway neutrophilia, resemble the pulmonary alterations that occur in agricultural workers with occupational asthma. The immunological basis of RAO remains uncertain although its chronicity, its localization to a mucosal surface and its domination by a neutrophilic, non-septic inflammatory response, suggest involvement of Interleukin-17 (IL-17). We examined global gene expression profiles in mediastinal (pulmonary-draining) lymph nodes isolated from RAO-affected and control horses. Differential expression of > 200 genes, coupled with network analysis, supports an IL-17 response centered about NF-κB. Immunohistochemical analysis of mediastinal lymph node sections demonstrated increased IL-17 staining intensity in diseased horses. This result, along with the finding of increased IL-17 concentrations in lymph node homogenates from RAO-affected horses (P = 0.1) and a down-regulation of IL-4 gene and protein expression, provides additional evidence of the involvement of IL-17 in the chronic stages of RAO. Additional investigations are needed to ascertain the cellular source of IL-17 in this equine model of occupational asthma. Understanding the immunopathogenesis of this disorder likely will enhance the development of therapeutic interventions beneficial to human and animal pulmonary health. PMID:26561853

  6. Apigenin Inhibits Tumor Necrosis Factor-α-Induced Production and Gene Expression of Mucin through Regulating Nuclear Factor-Kappa B Signaling Pathway in Airway Epithelial Cells

    PubMed Central

    Seo, Hyo-Seok; Sikder, Mohamed Asaduzzaman; Lee, Hyun Jae; Ryu, Jiho; Lee, Choong Jae

    2014-01-01

    In the present study, we investigated whether apigenin significantly affects tumor necrosis factor-α (TNF-α)-induced production and gene expression of MUC5AC mucin in airway epithelial cells. Confluent NCI-H292 cells were pretreated with apigenin for 30 min and then stimulated with TNF-α for 24 h or the indicated periods. The MUC5AC mucin gene expression and mucin protein production were measured by reverse transcription - polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA), respectively. Apigenin significantly inhibited MUC5AC mucin production and down-regulated MUC5AC gene expression induced by TNF-α in NCI-H292 cells. To elucidate the action mechanism of apigenin, effect of apigenin on TNF-α-induced nuclear factor kappa B (NF-κB) signaling pathway was also investigated by western blot analysis. Apigenin inhibited NF-κB activation induced by TNF-α. Inhibition of inhibitory kappa B kinase (IKK) by apigenin led to the suppression of inhibitory kappa B alpha (IκBα) phosphorylation and degradation, p65 nuclear translocation. This, in turn, led to the down-regulation of MUC5AC protein production in NCI-H292 cells. Apigenin also has an influence on upstream signaling of IKK because it inhibited the expression of adaptor protein, receptor interacting protein 1 (RIP1). These results suggest that apigenin can regulate the production and gene expression of mucin through regulating NF-κB signaling pathway in airway epithelial cells. PMID:25489420

  7. Preliminary Study on Gene Expression of Chitinase-Like Cytokines in Human Airway Epithelial Cell Under Chitin and Chitosan Microparticles Treatment.

    PubMed

    Alimohammadi, Masumeh; Yeganeh, Farshid; Haji Molla Hoseini, Mostafa

    2016-06-01

    Small-sized chitin and chitosan microparticles (MPs) reduce allergic inflammation. We examined the capacity of these glycans to stimulate A549 human airway epithelial cells to determine the feasibility of using of these glycans as allergic therapeutic modality. A549 cells were treated with MPs and then expressions levels of chitinase domain-containing 1 (CHID1) and chitinase 3-like 1 (CHI3L1) genes were determined by quantitative real-time PCR. IL-6 production was measured by ELISA. Chitin MPs resulted in upregulation of CHI3L1 expression by 35.7-fold while mRNA expression did not change with chitosan MPs. Compared to the untreated group, production of IL-6 was significantly decreased in the chitosan MPs-treated group, but chitin MPs treatment cause elevation of IL-6 level. This study demonstrates that chitin potently induces CHI3L1 expression, but chitosan is relatively inert. This effect and inhibition of pro-inflammatory cytokine (IL-6) suggest that chitosan MPs may possess more potential for therapeutic uses in human airway allergic inflammation.

  8. ARSENITE ACTIVATES KB-DEPENDENT IL-8 GENE EXPRESSION IN AIRWAY EPITHELIM IN THE ABSENCE OF NUCLEAR TRANSLOCATION OF NF-KB

    EPA Science Inventory

    Airway epithelial cells respond to certain environmental stresses by mounting a proinflammatory response, which is characterized by enhanced synthesis and release of the neutrophil chemotactic and activating factor interleukin-8 (IL-8). IL-8 expression is regulated at the transcr...

  9. Lymphocyte Gene Expression Characteristic of Immediate Airway Responses (IAR) and Methacholine (MCH) Hyperresponsiveness in Mice Sensitized and Challenged with Isocyanates

    EPA Science Inventory

    Exposure to isocyanates has been associated with occupational airway diseases, including asthma. Previously we reported on respiratory and immune responses following dermal sensitization and intranasal challenge of BALB/c mice with 6 different isocyanates. The purpose of this st...

  10. Quercetin Blocks Airway Epithelial Cell Chemokine Expression

    PubMed Central

    Nanua, Suparna; Zick, Suzanna M.; Andrade, Juan E.; Sajjan, Umadevi S.; Burgess, John R.; Lukacs, Nicholas W.; Hershenson, Marc B.

    2006-01-01

    Quercetin (3,3′,4′,5,7-pentahydroxyflavone), a dietary flavonoid, is an inhibitor of phosphatidylinositol (PI) 3-kinase and potent antioxidant. We hypothesized that quercetin blocks airway epithelial cell chemokine expression via PI 3-kinase–dependent mechanisms. Pretreatment with quercetin and the PI 3–kinase inhibitor LY294002 each reduced TNF-α–induced IL-8 and monocyte chemoattractant protein (MCP)-1 (also called CCL2) expression in cultured human airway epithelial cells. Quercetin also inhibited TNF-α–induced PI 3-kinase activity, Akt phosphorylation, intracellular H2O2 production, NF-κB transactivation, IL-8 promoter activity, and steady-state mRNA levels, consistent with the notion that quercetin inhibits chemokine expression by attenuating NF-κB transactivation via a PI 3-kinase/Akt-dependent pathway. Quercetin also reduced TNF-α–induced chemokine secretion in the presence of the transcriptional inhibitor actinomycin D, while inducing phosphorylation of eukaryotic translation initiation factor (eIF)-2α, suggesting that quercetin attenuates chemokine expression by post-transcriptional as well as transcriptional mechanisms. Finally, we tested the effects of quercetin in cockroach antigen–sensitized and –challenged mice. These mice show MCP-1–dependent airways hyperresponsiveness and inflammation. Quercetin significantly reduced lung MCP-1 and methacholine responsiveness. We conclude that quercetin blocks airway cell chemokine expression via transcriptional and post-transcriptional pathways. PMID:16794257

  11. Induction of CFTR gene expression by 1,25(OH)2 vitamin D3, 25OH vitamin D3, and vitamin D3 in cultured human airway epithelial cells and in mouse airways.

    PubMed

    DiFranco, Kristina M; Mulligan, Jennifer K; Sumal, Aman S; Diamond, Gill

    2017-01-24

    Cystic fibrosis (CF) is an autosomal recessive disorder caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene, which often leads to protein misfolding and no CFTR surface localization. This then leads to chronic airway infections, inflammation, and tissue damage. Although vitamin D has been explored as a therapy to treat CF due to its antimicrobial-inducing and anti-inflammatory properties, the effect of 1,25-dihydroxyvitamin D3 (1α,25(OH)2D3) on CFTR directly has not been studied. We treated cultured healthy and diseased bronchial epithelial cells (BEC) with 10nM 1α,25(OH)2D3 for 6 and 24h and found that 1α,25(OH)2D3 increases both mRNA and protein CFTR levels using RT-qPCR, flow cytometry and fluorescence immunohistochemistry. Treatment of CF cells with 10nM 1α,25(OH)2D3 led to an increase in both total and surface CFTR expression, suggesting 1α,25(OH)2D3 could be used to increase properly localized CFTR in airway cells. To determine if BEC could convert the more clinically relevant cholecalciferol to 25OHD3, cultured non-CF and CF BECs were treated with a range of cholecalciferol concentrations, and 25OHD3 levels were quantified by ELISA. We found that 25OHD3 levels increased in a concentration-dependent manner. Treatment of BEC with 10μM cholecalciferol led to increases in both CYP24A1 and CFTR mRNA levels, even when added to the apical surface of cells grown in an air-liquid interface, suggesting that topical administration of vitamin D could be used therapeutically. To demonstrate this in vivo, we intranasally delivered 1μM 1α,25(OH)2D3 into mice. After 6h, we observed induction of both Cyp24A1 and CFTR expression in the tracheas of treated mice. The major findings of this study are that vitamin D can be converted to the active form when topically administered to the airway, and this could be used to increase CFTR levels in patients with CF. This could potentially be useful as an adjunctive therapy, together with

  12. Inhibition of beta-defensin gene expression in airway epithelial cells by low doses of residual oil fly ash is mediated by vanadium.

    PubMed

    Klein-Patel, Marcia E; Diamond, Gill; Boniotto, Michele; Saad, Sherif; Ryan, Lisa K

    2006-07-01

    Poor ambient air quality is associated with increased morbidity and mortality, including respiratory infections. However, its effects on various host-defense mechanisms are poorly understood. This study utilized an in vitro model to study the effect of particulate matter (PM(2.5)) on one antimicrobial mechanism of host defense in the airway, beta-defensin-2 and its bovine homologue, tracheal antimicrobial peptide (TAP) induction in response to lipopolysaccharide (LPS) and IL-1beta. Our model utilized cultured primary bovine tracheal epithelial (BTE) cells and the human alveolar type II epithelial cell line, A549, treated with 0-20 microg/cm(2) residual oil fly ash (ROFA) for 6 h. The cells were then washed and stimulated for 18 h with 100 ng/ml LPS or for 6 h with 100 ng/ml IL-1beta. ROFA inhibited the LPS-induced increase in TAP mRNA and protein without inducing significant cytotoxicity. As little as 2.5 microg/cm(2) of ROFA inhibited LPS-induced TAP gene expression by 30%. The inhibitory activity was associated with the soluble fraction and not the washed particle. The activity in the leachate was attributed to vanadium, but not nickel or iron. SiO(2) and TiO(2) were utilized as controls and did not inhibit LPS induction of TAP gene expression in BTE. ROFA also inhibited the increase of IL-1beta-induced human beta-defensin-2, a homologue of TAP, in A549 cells. The results show that ROFA, V(2)O(5), and VOSO(4) inhibit the ability of airway epithelial cells to respond to inflammatory stimuli at low, physiologically relevant doses and suggest that exposure to these agents could result in an impairment of defense against airborne pathogens.

  13. Cold weather exercise and airway cytokine expression.

    PubMed

    Davis, Michael S; Malayer, Jerry R; Vandeventer, Lori; Royer, Christopher M; McKenzie, Erica C; Williamson, Katherine K

    2005-06-01

    Athletes who perform repeated exercise while breathing cold air have a high prevalence of asthmalike chronic airway disease, but the mechanism linking such activity to airway inflammation is unknown. We used a novel animal model (exercising horses) to test the hypothesis that exercise-induced chronic airway disease is caused by exposure of intrapulmonary airways to unconditioned air, resulting in the upregulation of cytokine expression. Bronchoalveolar lavage fluid (BALF) was obtained from eight horses 5 h after submaximal exercise while they breathed room temperature or subfreezing air in a random crossover design. BALF total and differential nucleated cell counts were determined, and relative cytokine mRNA expression in BALF nucleated cells was quantified by real-time RT-PCR using primer and probe sequences specific for equine targets. There were no significant changes in total or differential cell concentrations between BALF recovered after warm and cold air exercise, although there was a strong trend toward increased concentrations of airway epithelial cells after cold air exercise (P = 0.0625). T(H)2 cytokines IL-4, IL-5, and IL-10 were preferentially upregulated after cold air exercise 12-, 9-, and 10-fold, respectively, compared with warm air exercise. Other cytokines (IL-2 and IL-6) were upregulated to a lesser extent (6- and 3-fold, respectively) or not at all (IL-1, IL-8, IFN-gamma, and TNF-alpha). These results suggest that cold weather exercise can lead to asthmalike airway disease through the local induction of cytokines typical of the T(H)2 phenotype.

  14. NDRG1 is important to maintain the integrity of airway epithelial barrier through claudin-9 expression.

    PubMed

    Gon, Yasuhiro; Maruoka, Shuichiro; Kishi, Hiroyuki; Kozu, Yutaka; Kazumichi, Kuroda; Nomura, Yasuyuki; Takeshita, Ikuko; Oshima, Takeshi; Hashimoto, Shu

    2017-02-13

    Impairment of epithelial barrier integrity caused by environmental triggers is associated with the pathogenesis of airway inflammation. Using human airway epithelial cells, we attempted to identify molecule(s) that promote airway epithelial barrier integrity. Microarray analyses were conducted using the Affimetrix human whole genome gene chip, and we identified the N-myc downstream-regulated gene 1 (NDRG1) gene, which was induced during the development of the epithelial cell barrier. Immunohistochemical analysis revealed strong NDRG1 expression in ciliated epithelial cells in nasal tissues sampled from patients with chronic rhinosinusitis (CRS), and the low expression of NDRG1 was observed in goblet cells or damaged epithelial cells. NDRG1 gene knockdown with its specific siRNA decreased the transepithelial electrical resistance and increased the dextran permeability. Immunocytochemistry revealed that NDRG1 knockdown disrupted tight junctions of airway epithelial cells. Next, we analyzed the effects of NDRG1 knockdown on the expression of tight and adhesion junction molecules. NDRG1 knockdown significantly decreased only claudin-9 expression, but did not decrease other claudin family molecules, such as E-cadherin, and ZO-1, -2, or -3. Knockdown of claudin-9 markedly impaired the barrier function in airway epithelial cells. These results suggest that NDRG1 is important for the barrier integrity in airway epithelial cells.

  15. Ozone enhances diesel exhaust particles (DEP)-induced interleukin-8 (IL-8) gene expression in human airway epithelial cells through activation of nuclear factors- kappaB (NF-kappaB) and IL-6 (NF-IL6).

    PubMed

    Kafoury, Ramzi M; Kelley, James

    2005-12-01

    Ozone, a highly reactive oxidant gas is a major component of photochemical smog. As an inhaled toxicant, ozone induces its adverse effects mainly on the lung. Inhalation of particulate matter has been reported to cause airway inflammation in humans and animals. Furthermore, epidemiological evidence has indicated that exposure to particulate matter (PM[2.5-10]), including diesel exhaust particles (DEP) has been correlated with increased acute and chronic respiratory morbidity and exacerbation of asthma. Previously, exposure to ozone or particulate matter and their effect on the lung have been addressed as separate environmental problems. Ozone and particulate matter may be chemically coupled in the ambient air. In the present study we determined whether ozone exposure enhances DEP effect on interleukin-8 (IL-8) gene expression in human airway epithelial cells. We report that ozone exposure (0.5 ppm x 1 hr) significantly increased DEP-induced IL-8 gene expression in A549 cells (117 +/- 19 pg/ml, n = 6, p < 0.05) as compared to cultures treated with DEP (100 microg/ml x 4 hr) alone (31 +/- 3 pg/ml, n = 6), or cultures exposed to purified air (24 +/- 6 pg/ml, n = 6). The increased DEP-induced IL-8 gene expression following ozone exposure was attributed to ozone-induced increase in the activity of the transcription factors NF-kappaB and NF-IL6. The results of the present study indicate that ozone exposure enhances the toxicity of DEP in human airway epithelial cells by augmenting IL-8 gene expression, a potent chemoattractant of neutrophils in the lung.

  16. Expression of ligands for Siglec-8 and Siglec-9 in human airways and airway cells

    PubMed Central

    Jia, Yi; Yu, Huifeng; Fernandes, Steve M.; Wei, Yadong; Gonzalez-Gil, Anabel; Motari, Mary G.; Vajn, Katarina; Stevens, Whitney W.; Peters, Anju T.; Bochner, Bruce S.; Kern, Robert C.; Schleimer, Robert P.; Schnaar, Ronald L.

    2015-01-01

    Background Balanced activation and inhibition of the immune system ensures pathogen clearance while avoiding hyperinflammation. Siglecs, sialic acid binding proteins found on subsets of immune cells, often inhibit inflammation: Siglec-8 on eosinophils and Siglec-9 on neutrophils engage sialoglycan ligands on airways to diminish ongoing inflammation. The identities of human siglec ligands and their expression during inflammation are largely unknown. Objective The histological distribution, expression and molecular characteristics of siglec ligands were explored in healthy and inflamed human upper airways and in a cellular model of airway inflammation. Methods Normal and chronically inflamed upper airway tissues were stained for siglec ligands. The ligands were extracted from normal and inflamed tissues and from human Calu-3 cells for quantitative analysis by siglec blotting and isolation by siglec capture. Results Siglec-8 ligands were expressed on a subpopulation of submucosal gland cells of human inferior turbinate, whereas Siglec-9 ligands were expressed more broadly (submucosal glands, epithelium, connective tissue); both were significantly upregulated in chronic rhinosinusitis patients. Human airway (Calu-3) cells expressed Siglec-9 ligands on mucin 5B under inflammatory control via the NF-κB pathway, and mucin 5B carried sialoglycan ligands of Siglec-9 on human upper airway tissue. Conclusion Inflammation results in upregulation of immune inhibitory Siglec-8 and Siglec-9 sialoglycan ligands on human airways. Siglec-9 ligands were upregulated via the NF-κB pathway resulting in their enhanced expression on mucin 5B. Siglec sialoglycan ligand expression in inflamed cells and tissues may contribute to the control of airway inflammation. PMID:25747723

  17. Expression of IL-4/IL-13 receptors in differentiating human airway epithelial cells

    PubMed Central

    Martin, Linda D.; Stern, Randi; Laxman, Bharathi; Marroquin, Bertha A.

    2010-01-01

    IL-4 and IL-13 elicit several important responses in airway epithelium including chemokine secretion and mucous secretion that may contribute to airway inflammation, cell migration, and differentiation. These cytokines have overlapping but not identical effector profiles likely due to shared subunits in their receptor complexes. These receptors are variably described in epithelial cells, and the relative expression, localization, and function of these receptors in differentiated and repairing epithelial cells are not clear. We examined IL-4/IL-13 receptor expression and localization in primary airway epithelial cells collected from normal human lungs and grown under conditions yielding both undifferentiated and differentiated cells inclusive of basal, goblet, and ciliated cell phenotypes. Gene expression of the IL-4Rα, IL-2Rγc, IL-13Rα1, and IL-13Rα2 receptor subunits increased with differentiation, but different patterns of localization and protein abundance were seen for each subunit based on both differentiation and the cell subtypes present. Increased expression of receptor subunits observed in more differentiated cells was associated with more substantial functional responses to IL-4 stimulation including increased eotaxin-3 expression and accelerated migration after injury. We demonstrate substantial differences in IL-4/IL-13 receptor subunit expression and responsiveness to IL-4 based on the extent of airway epithelial cell differentiation and suggest that these differences may have functional consequences in airway inflammation. PMID:20729386

  18. Smoking-induced CXCL14 expression in the human airway epithelium links chronic obstructive pulmonary disease to lung cancer.

    PubMed

    Shaykhiev, Renat; Sackrowitz, Rachel; Fukui, Tomoya; Zuo, Wu-Lin; Chao, Ion Wa; Strulovici-Barel, Yael; Downey, Robert J; Crystal, Ronald G

    2013-09-01

    CXCL14, a recently described epithelial cytokine, plays putative multiple roles in inflammation and carcinogenesis. In the context that chronic obstructive pulmonary disease (COPD) and lung cancer are both smoking-related disorders associated with airway epithelial disorder and inflammation, we hypothesized that the airway epithelium responds to cigarette smoking with altered CXCL14 gene expression, contributing to the disease-relevant phenotype. Using genome-wide microarrays with subsequent immunohistochemical analysis, the data demonstrate that the expression of CXCL14 is up-regulated in the airway epithelium of healthy smokers and further increased in COPD smokers, especially within hyperplastic/metaplastic lesions, in association with multiple genes relevant to epithelial structural integrity and cancer. In vitro experiments revealed that the expression of CXCL14 is induced in the differentiated airway epithelium by cigarette smoke extract, and that epidermal growth factor mediates CXCL14 up-regulation in the airway epithelium through its effects on the basal stem/progenitor cell population. Analyses of two independent lung cancer cohorts revealed a dramatic up-regulation of CXCL14 expression in adenocarcinoma and squamous-cell carcinoma. High expression of the COPD-associated CXCL14-correlating cluster of genes was linked in lung adenocarcinoma with poor survival. These data suggest that the smoking-induced expression of CXCL14 in the airway epithelium represents a novel potential molecular link between smoking-associated airway epithelial injury, COPD, and lung cancer.

  19. Thick airway surface liquid volume and weak mucin expression in pendrin-deficient human airway epithelia

    PubMed Central

    Lee, Hyun Jae; Yoo, Jee Eun; Namkung, Wan; Cho, Hyung-Ju; Kim, Kyubo; Kang, Joo Wan; Yoon, Joo-Heon; Choi, Jae Young

    2015-01-01

    Pendrin is an anion exchanger whose mutations are known to cause hearing loss. However, recent data support the linkage between pendrin expression and airway diseases, such as asthma. To evaluate the role of pendrin in the regulation of the airway surface liquid (ASL) volume and mucin expression, we investigated the function and expression of pendrin and ion channels and anion exchangers. Human nasal epithelial cells were cultured from 16 deaf patients carrying pendrin mutations (DFNB4) and 17 controls. The cells were treated with IL-13 to induce mucus hypersecretion. Airway surface liquid thickness was measured and real-time polymerase chain reaction was performed targeting various transporters and MUC5AC. Anion exchanger activity was measured using a pH-sensitive fluorescent probe. Periodic acid-Schiff staining was performed on the cultured cells and inferior turbinate tissues. The ASL layer of the nasal epithelia from DFNB4 subjects was thicker than the controls, and the difference became more prominent following IL-13 stimulation. There was no difference in anion exchange activity after IL-13 treatment in the cells from DFNB4 patients, while it increased in the controls. Goblet cell metaplasia induced by IL-13 treatment seen in the controls was not observed in the DFNB4 cells. Furthermore, the periodic acid-Schiff staining-positive area was lesser in the inferior turbinate tissues from DFNB4 patients that those from controls. Pendrin plays a critical role in ASL volume regulation and mucin expression as pendrin-deficient airway epithelial cells are refractory to stimulation with IL-13. Specific blockers targeting pendrin in the airways may therefore have therapeutic potential in the treatment of allergic airway diseases. PMID:26243215

  20. The Allergic Airway Inflammation Repository--a user-friendly, curated resource of mRNA expression levels in studies of allergic airways.

    PubMed

    Gawel, D R; Rani James, A; Benson, M; Liljenström, R; Muraro, A; Nestor, C E; Zhang, H; Gustafsson, M

    2014-08-01

    Public microarray databases allow analysis of expression levels of candidate genes in different contexts. However, finding relevant microarray data is complicated by the large number of available studies. We have compiled a user-friendly, open-access database of mRNA microarray experiments relevant to allergic airway inflammation, the Allergic Airway Inflammation Repository (AAIR, http://aair.cimed.ike.liu.se/). The aim is to allow allergy researchers to determine the expression profile of their genes of interest in multiple clinical data sets and several experimental systems quickly and intuitively. AAIR also provides quick links to other relevant information such as experimental protocols, related literature and raw data files.

  1. Human airway epithelia express catalytically active NEU3 sialidase.

    PubMed

    Lillehoj, Erik P; Hyun, Sang Won; Feng, Chiguang; Zhang, Lei; Liu, Anguo; Guang, Wei; Nguyen, Chinh; Sun, Wenji; Luzina, Irina G; Webb, Tonya J; Atamas, Sergei P; Passaniti, Antonino; Twaddell, William S; Puché, Adam C; Wang, Lai-Xi; Cross, Alan S; Goldblum, Simeon E

    2014-05-01

    Sialic acids on glycoconjugates play a pivotal role in many biological processes. In the airways, sialylated glycoproteins and glycolipids are strategically positioned on the plasma membranes of epithelia to regulate receptor-ligand, cell-cell, and host-pathogen interactions at the molecular level. We now demonstrate, for the first time, sialidase activity for ganglioside substrates in human airway epithelia. Of the four known mammalian sialidases, NEU3 has a substrate preference for gangliosides and is expressed at mRNA and protein levels at comparable abundance in epithelia derived from human trachea, bronchi, small airways, and alveoli. In small airway and alveolar epithelia, NEU3 protein was immunolocalized to the plasma membrane, cytosolic, and nuclear subcellular fractions. Small interfering RNA-induced silencing of NEU3 expression diminished sialidase activity for a ganglioside substrate by >70%. NEU3 immunostaining of intact human lung tissue could be localized to the superficial epithelia, including the ciliated brush border, as well as to nuclei. However, NEU3 was reduced in subepithelial tissues. These results indicate that human airway epithelia express catalytically active NEU3 sialidase.

  2. Creation and characterization of an airway epithelial cell line for stable expression of CFTR variants

    PubMed Central

    Gottschalk, Laura B.; Vecchio-Pagan, Briana; Sharma, Neeraj; Han, Sangwoo T.; Franca, Arianna; Wohler, Elizabeth S.; Batista, Denise A.S.; Goff, Loyal A.; Cutting, Garry R.

    2016-01-01

    Background Analysis of the functional consequences and treatment response of rare CFTR variants is challenging due to the limited availability of primary airways cells. Methods A Flp recombination target (FRT) site for stable expression of CFTR was incorporated into an immortalized CF bronchial epithelial cell line (CFBE41o−). CFTR cDNA was integrated into the FRT site. Expression was evaluated by western blotting and confocal microscopy and function measured by short circuit current. RNA sequencing was used to compare the transcriptional profile of the resulting CF8Flp cell line to primary cells and tissues. Results Functional CFTR was expressed from integrated cDNA at the FRT site of the CF8Flp cell line at levels comparable to that seen in native airway cells. CF8Flp cells expressing WT-CFTR have a stable transcriptome comparable to that of primary cultured airway epithelial cells, including genes that play key roles in CFTR pathways. Conclusion CF8Flp cells provide a viable substitute for primary CF airway cells for the analysis of CFTR variants in a native context. PMID:26694805

  3. Targeted expression of IL-11 in the murine airway causes lymphocytic inflammation, bronchial remodeling, and airways obstruction.

    PubMed Central

    Tang, W; Geba, G P; Zheng, T; Ray, P; Homer, R J; Kuhn, C; Flavell, R A; Elias, J A

    1996-01-01

    Interleukin-11 is a pleotropic cytokine produced by lung stromal cells in response to respiratory viruses, cytokines, and histamine. To further define its potential effector functions, the Clara cell 10-kD protein promoter was used to express IL-11 and the airways of the resulting transgene mice were characterized. In contrast to transgene (-) littermates, the airways of IL-11 transgene (+) animals manifest nodular peribronchiolar mononuclear cell infiltrates and impressive airways remodeling with subepithelial fibrosis. The inflammatory foci contained large numbers of B220(+) and MHC Class II(+) cells and lesser numbers of CD3(+), CD4(+), and CD8(+) cells. The fibrotic response contained increased amounts of types III and I collagen, increased numbers of alpha smooth muscle actin and desmin-containing cells and a spectrum of stromal elements including fibroblasts, myofibroblasts, and smooth muscle cells. Physiologic evaluation also demonstrated that 2-mo-old transgene (+) mice had increased airways resistance and non-specific airways hyperresponsiveness to methacholine when compared with their transgene (-) littermates. These studies demonstrate that the targeted expression of IL-11 in the mouse airway causes a B and T cell-predominant inflammatory response, airway remodeling with increased types III and I collagen, the local accumulation of fibroblasts, myofibroblasts, and myocytes, and obstructive physiologic dysregulation. IL-11 may play an important role in the inflammatory and fibrotic responses in viral and/or nonviral human airway disorders. PMID:8981933

  4. Expression of interleukin-18 by porcine airway and intestinal epithelium.

    PubMed

    Muneta, Yoshihiro; Goji, Noriko; Tsuji, Noriko M; Mikami, Osamu; Shimoji, Yoshihiro; Nakajima, Yasuyuki; Yokomizo, Yuichi; Mori, Yasuyuki

    2002-08-01

    In this study, we investigated the expression of interleukin-18 (IL-18) in porcine airway and intestinal epithelium. We found constitutive protein expression of precursor IL-18 in primary culture of porcine airway epithelium. Immunohistochemical staining revealed that porcine IL-18 was localized in the porcine airway epithelium and that it was significantly upregulated with experimental endotoxemia induced by Escherichia coli lipopolysaccharide (LPS) inoculation. We also confirmed by immunohistochemical staining that IL-18 was expressed in porcine intestinal epithelial cells. Moreover, the concentration of IL-18 in intestinal cell lysates of 1-day-old piglets was about 3-fold and 6-fold less than that in those of 1-month-old and 6-month-old piglets, respectively. Exogenous IL-18 was able to induce interferon-gamma (IFN-gamma) in the peripheral blood of 1-day-old piglets, whereas concanavalin A (ConA) was not able to induce IFN-gamma in the same condition. These results suggest that mucosal epithelial cells are among the major sources of IL-18 in pig and that IL-18 may be useful as a therapeutic agent for the enhancement of immune responses and as a vaccine adjuvant, especially in neonatal piglets.

  5. CRTH2 antagonism significantly ameliorates airway hyperreactivity and downregulates inflammation-induced genes in a mouse model of airway inflammation.

    PubMed

    Lukacs, Nicholas W; Berlin, Aaron A; Franz-Bacon, Karin; Sásik, Roman; Sprague, L James; Ly, Tai Wei; Hardiman, Gary; Boehme, Stefen A; Bacon, Kevin B

    2008-11-01

    Prostaglandin D(2), the ligand for the G protein-coupled receptors DP1 and CRTH2, has been implicated in the pathogenesis of the allergic response in diseases such as asthma, rhinitis, and atopic dermatitis. This prostanoid also fulfills a number of physiological, anti-inflammatory roles through its receptor DP1. We investigated the role of PGD(2) and CRTH2 in allergic pulmonary inflammation by using a highly potent and specific antagonist of CRTH2. Administration of this antagonist ameliorated inflammation caused by either acute or subchronic sensitization using the cockroach egg antigen. Gene expression and ELISA analysis revealed that there was reduced proinflammatory cytokine mRNA or protein produced, as well as a wide array of genes associated with the Th2-type proinflammatory response. Importantly, the CRTH2 antagonist reduced antigen-specific IgE, IgG1, and IgG2a antibody levels as well as decreased mucus deposition and leukocyte infiltration in the large airways. Collectively, these findings suggest that the PGD(2)-CRTH2 activation axis has a pivotal role in mediating the inflammation and the underlying immune response in a T cell-driven model of allergic airway inflammation.

  6. GS-5759, a Bifunctional β2-Adrenoceptor Agonist and Phosphodiesterase 4 Inhibitor for Chronic Obstructive Pulmonary Disease with a Unique Mode of Action: Effects on Gene Expression in Human Airway Epithelial Cells.

    PubMed

    Joshi, Taruna; Yan, Dong; Hamed, Omar; Tannheimer, Stacey L; Phillips, Gary B; Wright, Clifford D; Kim, Musong; Salmon, Michael; Newton, Robert; Giembycz, Mark A

    2017-02-01

    (R)-6-[(3-{[4-(5-{[2-hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)ethyl]amino}pent-1-yn-1-yl)phenyl] carbamoyl}phenyl)sulphonyl]-4-[(3-methoxyphenyl)amino]-8-methylquinoline-3-carboxamide trifluoroacetic acid (GS-5759) is a bifunctional ligand composed of a quinolinone-containing pharmacophore [β2-adrenoceptor agonist orthostere (β2A)] found in several β2-adrenoceptor agonists, including indacaterol, linked covalently to a phosphodiesterase 4 (PDE4) inhibitor related to 6-[3-(dimethylcarbamoyl)benzenesulphonyl]-4-[(3-methoxyphenyl)amino]-8-methylquinoline-3-carboxamide (GSK 256066) by a pent-1-yn-1-ylbenzene spacer. GS-5759 had a similar affinity for PDE4B1 and the native β2-adrenoceptor expressed on BEAS-2B human airway epithelial cells. However, compared with the monofunctional parent compound, β2A, the KA of GS-5759 for the β2-adrenoceptor was 35-fold lower. Schild analysis determined that the affinities of the β-adrenoceptor antagonists, (2R,3R)-1-[(2,3-dihydro-7-methyl-1H-inden-4-yl)oxy]-3-[(1-methylethyl) amino]-2-butanol (ICI 118551) and propranolol, were agonist-dependent, being significantly lower for GS-5759 than β2A. Collectively, these data can be explained by "forced proximity," bivalent binding where the pharmacophore in GS-5759 responsible for PDE4 inhibition also interacts with a nonallosteric domain within the β2-adrenoceptor that enhances the affinity of β2A for the orthosteric site. Microarray analyses revealed that, after 2-hour exposure, GS-5759 increased the expression of >3500 genes in BEAS-2B cells that were highly rank-order correlated with gene expression changes produced by indacaterol and GSK 256066 in combination (Ind/GSK). Moreover, the line of regression began close to the origin with a slope of 0.88, indicating that the magnitude of most gene expression changes produced by Ind/GSK was quantitatively replicated by GS-5759. Thus, GS-5759 is a novel compound exhibiting dual β2-adrenoceptor agonism and PDE4 inhibition

  7. Airway Epithelial miRNA Expression Is Altered in Asthma

    PubMed Central

    Solberg, Owen D.; Ostrin, Edwin J.; Love, Michael I.; Peng, Jeffrey C.; Bhakta, Nirav R.; Nguyen, Christine; Solon, Margaret; Nguyen, Cindy; Barczak, Andrea J.; Zlock, Lorna T.; Blagev, Denitza P.; Finkbeiner, Walter E.; Ansel, K. Mark; Arron, Joseph R.; Erle, David J.

    2012-01-01

    Rationale: Changes in airway epithelial cell differentiation, driven in part by IL-13, are important in asthma. Micro-RNAs (miRNAs) regulate cell differentiation in many systems and could contribute to epithelial abnormalities in asthma. Objectives: To determine whether airway epithelial miRNA expression is altered in asthma and identify IL-13–regulated miRNAs. Methods: We used miRNA microarrays to analyze bronchial epithelial brushings from 16 steroid-naive subjects with asthma before and after inhaled corticosteroids, 19 steroid-using subjects with asthma, and 12 healthy control subjects, and the effects of IL-13 and corticosteroids on cultured bronchial epithelial cells. We used quantitative polymerase chain reaction to confirm selected microarray results. Measurements and Main Results: Most (12 of 16) steroid-naive subjects with asthma had a markedly abnormal pattern of bronchial epithelial miRNA expression by microarray analysis. Compared with control subjects, 217 miRNAs were differentially expressed in steroid-naive subjects with asthma and 200 in steroid-using subjects with asthma (false discovery rate < 0.05). Treatment with inhaled corticosteroids had modest effects on miRNA expression in steroid-naive asthma, inducing a statistically significant (false discovery rate < 0.05) change for only nine miRNAs. qPCR analysis confirmed differential expression of 22 miRNAs that were highly differentially expressed by microarrays. IL-13 stimulation recapitulated changes in many differentially expressed miRNAs, including four members of the miR-34/449 family, and these changes in miR-34/449 family members were resistant to corticosteroids. Conclusions: Dramatic alterations of airway epithelial cell miRNA levels are a common feature of asthma. These alterations are only modestly corrected by inhaled corticosteroids. IL-13 effects may account for some of these alterations, including repression of miR-34/449 family members that have established roles in airway

  8. DNA methylation is globally disrupted and associated with expression changes in chronic obstructive pulmonary disease small airways.

    PubMed

    Vucic, Emily A; Chari, Raj; Thu, Kelsie L; Wilson, Ian M; Cotton, Allison M; Kennett, Jennifer Y; Zhang, May; Lonergan, Kim M; Steiling, Katrina; Brown, Carolyn J; McWilliams, Annette; Ohtani, Keishi; Lenburg, Marc E; Sin, Don D; Spira, Avrum; Macaulay, Calum E; Lam, Stephen; Lam, Wan L

    2014-05-01

    DNA methylation is an epigenetic modification that is highly disrupted in response to cigarette smoke and involved in a wide spectrum of malignant and nonmalignant diseases, but surprisingly not previously assessed in small airways of patients with chronic obstructive pulmonary disease (COPD). Small airways are the primary sites of airflow obstruction in COPD. We sought to determine whether DNA methylation patterns are disrupted in small airway epithelia of patients with COPD, and evaluate whether changes in gene expression are associated with these disruptions. Genome-wide methylation and gene expression analysis were performed on small airway epithelial DNA and RNA obtained from the same patient during bronchoscopy, using Illumina's Infinium HM27 and Affymetrix's Genechip Human Gene 1.0 ST arrays. To control for known effects of cigarette smoking on DNA methylation, methylation and gene expression profiles were compared between former smokers with and without COPD matched for age, pack-years, and years of smoking cessation. Our results indicate that aberrant DNA methylation is (1) a genome-wide phenomenon in small airways of patients with COPD, and (2) associated with altered expression of genes and pathways important to COPD, such as the NF-E2-related factor 2 oxidative response pathway. DNA methylation is likely an important mechanism contributing to modulation of genes important to COPD pathology. Because these methylation events may underlie disease-specific gene expression changes, their characterization is a critical first step toward the development of epigenetic markers and an opportunity for developing novel epigenetic therapeutic interventions for COPD.

  9. Identification of genes differentially regulated by vitamin D deficiency that alter lung pathophysiology and inflammation in allergic airways disease.

    PubMed

    Foong, Rachel E; Bosco, Anthony; Troy, Niamh M; Gorman, Shelley; Hart, Prue H; Kicic, Anthony; Zosky, Graeme R

    2016-09-01

    Vitamin D deficiency is associated with asthma risk. Vitamin D deficiency may enhance the inflammatory response, and we have previously shown that airway remodeling and airway hyperresponsiveness is increased in vitamin D-deficient mice. In this study, we hypothesize that vitamin D deficiency would exacerbate house dust mite (HDM)-induced inflammation and alterations in lung structure and function. A BALB/c mouse model of vitamin D deficiency was established by dietary manipulation. Responsiveness to methacholine, airway smooth muscle (ASM) mass, mucus cell metaplasia, lung and airway inflammation, and cytokines in bronchoalveolar lavage (BAL) fluid were assessed. Gene expression patterns in mouse lung samples were profiled by RNA-Seq. HDM exposure increased inflammation and inflammatory cytokines in BAL, baseline airway resistance, tissue elastance, and ASM mass. Vitamin D deficiency enhanced the HDM-induced influx of lymphocytes into BAL, ameliorated the HDM-induced increase in ASM mass, and protected against the HDM-induced increase in baseline airway resistance. RNA-Seq identified nine genes that were differentially regulated by vitamin D deficiency in the lungs of HDM-treated mice. Immunohistochemical staining confirmed that protein expression of midline 1 (MID1) and adrenomedullin was differentially regulated such that they promoted inflammation, while hypoxia-inducible lipid droplet-associated, which is associated with ASM remodeling, was downregulated. Protein expression studies in human bronchial epithelial cells also showed that addition of vitamin D decreased MID1 expression. Differential regulation of these genes by vitamin D deficiency could determine lung inflammation and pathophysiology and suggest that the effect of vitamin D deficiency on HDM-induced allergic airways disease is complex.

  10. Incorporation of adenovirus in calcium phosphate precipitates enhances gene transfer to airway epithelia in vitro and in vivo.

    PubMed Central

    Fasbender, A; Lee, J H; Walters, R W; Moninger, T O; Zabner, J; Welsh, M J

    1998-01-01

    Adenovirus (Ad)-mediated gene transfer to airway epithelia is inefficient because the apical membrane lacks the receptor activity to bind adenovirus fiber protein. Calcium phosphate (CaPi) precipitates have been used to deliver plasmid DNA to cultured cell lines. However, such precipitates are not effective in many primary cultures or in vivo. Here we show that incorporating recombinant adenovirus into a CaPi coprecipitate markedly enhances transgene expression in cells that are resistant to adenovirus infection. Enhancement requires that the virus be contained in the precipitate and viral proteins are required to increase expression. Ad: CaPi coprecipitates increase gene transfer by increasing fiber-independent binding of virus to cells. With differentiated cystic fibrosis (CF) airway epithelia in vitro, a 20-min application of Ad:CaPi coprecipitates that encode CF transmembrane conductance regulator produced as much CF transmembrane conductance regulator Cl- current as a 24-h application of adenovirus alone. We found that Ad:CaPi coprecipitates also increased transgene expression in mouse lung in vivo; importantly, expression was particularly prominent in airway epithelia. These results suggest a new mechanism for gene transfer that may be applicable to a number of different gene transfer applications and could be of value in gene transfer to CF airway epithelia in vivo. PMID:9649572

  11. Thixotropic solutions enhance viral-mediated gene transfer to airway epithelia.

    PubMed

    Seiler, Michael P; Luner, Paul; Moninger, Thomas O; Karp, Philip H; Keshavjee, Shaf; Zabner, Joseph

    2002-08-01

    Adenovirus-mediated gene transfer to airway epithelia is inefficient in part because its receptor is absent on the apical surface of the airways. Targeting adenovirus to other receptors, increasing the viral concentration, and even prolonging the incubation time with adenovirus vectors can partially overcome the lack of receptors and facilitate gene transfer. Unfortunately, mucociliary clearance would prevent prolonged incubation time in vivo. Thixotropic solutions (TS) are gels that upon a vigorous shearing force reversibly become liquid. We hypothesized that formulating recombinant adenoviruses in TS would decrease virus clearance and thus enhance gene transfer to the airway epithelia. We found that clearance of virus-sized fluorescent beads by human airway epithelia in vitro and by monkey trachea in vivo were markedly decreased when the beads were formulated in TS compared with phosphate-buffered saline (PBS). Adenovirus formulated in TS significantly increased adenovirus-mediated gene transfer of a reporter gene in human airway epithelia in vitro and in murine airway epithelia in vivo. Furthermore, an adenovirus encoding the cystic fibrosis transmembrane regulator (CFTR) gene (AdCFTR) formulated in TS was more efficient in correcting the chloride transport defect in cystic fibrosis airway epithelia than AdCFTR formulated in PBS. These data indicate a novel strategy to augment the efficiency of gene transfer to the airways that may be applicable to a number of different gene transfer vectors and could be of value in gene transfer to cystic fibrosis (CF) airway epithelia in vivo.

  12. [Expression and role of sugar chains on airway mucus during the exacerbation of airway inflammation].

    PubMed

    Ishibashi, Yuji; Inouye, Yoshio; Taniguchi, Akiyoshi

    2012-01-01

    Human bronchial mucins, such as MUC5AC, have traditionally been defined as a family of high-molecular weight glycoproteins. Changes in the contents of sugar chains on MUC5AC are among the fundamental features in inflammatory respiratory disease. The changes have been shown to lead to unfavorable alterations in the viscosity of mucus, resulting in impairment of mucociliary transport, vulnerability to viral/bacterial infection as sugar chains play an important role in adhesion of some viruses and bacteria to the epithelium, and finally inflammatory cell infiltration in the airway. Recently, we found that expression of some glycosyltransferases associated with the contents and structure of sugar chains is regulated by phosphatidylinositol-phospholipase (PI-PL) C signaling in cells. L-Carbocisteine, a mucoregulatory drug, normalized or balanced fucosylated and sialylated sugar chains, such as sialyl Lewis x through inhibition of PI-PL C signaling. We prepared MUC5AC fusion protein with tandem repeats associated with MUC5AC, and confirmed that L-carbocisteine inhibited the increases in viscosity associated with sialyl Lewis x expression levels. In addition, the clinical study (2008) noted that L-carbocisteine reduced the frequency of common colds and exacerbation of symptoms in patients with COPD. These favorable effects in patients may be due to normalization of sugar chain contents on mucins. We suggest that the inhibitory effect on infection of airway epithelial cells by rhinoviruses, respiratory syncytial virus, and influenza viruses by treatment with L-carbocisteine may also be based on the regulation of sugar chain contents or structures on mucins.

  13. Vitamin D deficiency causes airway hyperresponsiveness, increases airway smooth muscle mass, and reduces TGF-β expression in the lungs of female BALB/c mice.

    PubMed

    Foong, Rachel E; Shaw, Nicole C; Berry, Luke J; Hart, Prue H; Gorman, Shelley; Zosky, Graeme R

    2014-01-01

    Abstract Vitamin D deficiency is associated with disease severity in asthma. We tested whether there is a causal association between vitamin D deficiency, airway smooth muscle (ASM) mass, and the development of airway hyperresponsiveness (AHR). A physiologically relevant mouse model of vitamin D deficiency was developed by raising BALB/c mice on vitamin D-deficient or -replete diets. AHR was assessed by measuring lung function responses to increasing doses of inhaled methacholine. Five-micron sections from formalin-fixed lungs were used for ASM measurement and assessment of lung structure using stereological methods. Transforming growth factor (TGF)-β levels were measured in bronchoalveolar lavage fluid (BALF). Lungs were dissected from embryonic day (E) 17.5 vitamin D-deficient and -replete fetal mice for quantification of ASM density and relative gene expression of TGF-β signaling pathway molecules. Eight-week-old adult vitamin D-deficient female mice had significantly increased airway resistance and ASM in the large airways compared with controls. Vitamin D-deficient female mice had a smaller lung volume, volume of parenchyma, and alveolar septa. Both vitamin D-deficient male and female mice had reduced TGF-β levels in BALF. Vitamin D deficiency did not have an effect on ASM density in E17.5 mice, however, expression of TGF-β1 and TGF-β receptor I was downregulated in vitamin D-deficient female fetal mice. Decreased expression of TGF-β1 and TGF-β receptor I during early lung development in vitamin D-deficient mice may contribute to airway remodeling and AHR in vitamin D-deficient adult female mice. This study provides a link between vitamin D deficiency and respiratory symptoms in chronic lung disease.

  14. Cigarette smoke induces genetic instability in airway epithelial cells by suppressing FANCD2 expression

    PubMed Central

    Hays, L E; Zodrow, D M; Yates, J E; Deffebach, M E; Jacoby, D B; Olson, S B; Pankow, J F; Bagby, G C

    2008-01-01

    Chromosomal abnormalities are commonly found in bronchogenic carcinoma cells, but the molecular causes of chromosomal instability (CIN) and their relationship to cigarette smoke has not been defined. Because the Fanconi anaemia (FA)/BRCA pathway is essential for maintenance of chromosomal stability, we tested the hypothesis that cigarette smoke suppresses that activity of this pathway. Here, we show that cigarette smoke condensate (CSC) inhibited translation of FANCD2 mRNA (but not FANCC or FANCG) in normal airway epithelial cells and that this suppression of FANCD2 expression was sufficient to induce both genetic instability and programmed cell death in the exposed cell population. Cigarette smoke condensate also suppressed FANCD2 function and induced CIN in bronchogenic carcinoma cells, but these cells were resistant to CSC-induced apoptosis relative to normal airway epithelial cells. We, therefore, suggest that CSC exerts pressure on airway epithelial cells that results in selection and emergence of genetically unstable somatic mutant clones that may have lost the capacity to effectively execute an apoptotic programme. Carcinogen-mediated suppression of FANCD2 gene expression provides a plausible molecular mechanism for CIN in bronchogenic carcinogenesis. PMID:18475298

  15. Non-raft adenylyl cyclase 2 defines a cAMP signaling compartment that selectively regulates IL-6 expression in airway smooth muscle cells: differential regulation of gene expression by AC isoforms.

    PubMed

    Bogard, Amy S; Birg, Anna V; Ostrom, Rennolds S

    2014-04-01

    Adenylyl cyclase (AC) isoforms differ in their tissue distribution, cellular localization, regulation, and protein interactions. Most cell types express multiple AC isoforms. We hypothesized that cAMP produced by different AC isoforms regulates unique cellular responses in human bronchial smooth muscle cells (BSMC). Overexpression of AC2, AC3, or AC6 had distinct effects on forskolin (Fsk)-induced expression of a number of known cAMP-responsive genes. These data show that different AC isoforms can differentially regulate gene expression. Most notable, overexpression and activation of AC2 enhanced interleukin 6 (IL-6) expression, but overexpression of AC3 or AC6 had no effect. IL-6 production by BSMC was induced by Fsk and select G protein-coupled receptor (GPCR) agonists, though IL-6 levels did not directly correlate with global cAMP levels. Treatment with PKA selective 6-Bnz-cAMP or Epac selective 8-CPT-2Me-cAMP cAMP analogs revealed a predominant role for PKA in cAMP-mediated induction of IL-6. IL-6 promoter mutations demonstrated that AP-1 and CRE transcription sites were required for Fsk to stimulate IL-6 expression. Our present study defines an AC2 cAMP signaling compartment that specifically regulates IL-6 expression in BSMC via Epac and PKA and demonstrates that other AC isoforms are excluded from this pool.

  16. Mucus altering agents as adjuncts for nonviral gene transfer to airway epithelium.

    PubMed

    Ferrari, S; Kitson, C; Farley, R; Steel, R; Marriott, C; Parkins, D A; Scarpa, M; Wainwright, B; Evans, M J; Colledge, W H; Geddes, D M; Alton, E W

    2001-09-01

    Nonviral vectors have been shown to be a safe and valid alternative to recombinant viruses for gene therapy of cystic fibrosis (CF). Nevertheless, gene transfer efficiency needs to be increased before clinical efficacy is likely in man. One barrier to increased efficacy is normal airway mucus. Using an ex vivo model of sheep tracheal epithelium, we show that this barrier can, in part, be overcome by treatment with the mucolytic agents, Nacystelyn or N-acetylcysteine using either a cationic lipid or a cationic polymer as the gene transfer agent. Further, in vivo application of either Nacystelyn or the anticholinergic glycopyrrolate, both clinically used agents, resulted in increased reporter gene expression in the mouse lung, but no significant correction of the bioelectric defect in CF null mice. These results, whilst unlikely to be sufficient in themselves to achieve clinically relevant gene therapy, may be a further useful step in the attainment of this goal.

  17. Expression of cystic fibrosis transmembrane conductance regulator corrects defective chloride channel regulation in cystic fibrosis airway epithelial cells

    NASA Astrophysics Data System (ADS)

    Rich, Devra P.; Anderson, Matthew P.; Gregory, Richard J.; Cheng, Seng H.; Paul, Sucharita; Jefferson, Douglas M.; McCann, John D.; Klinger, Katherine W.; Smith, Alan E.; Welsh, Michael J.

    1990-09-01

    The cystic fibrosis transmembrane conductance regulator (CFTR) was expressed in cultured cystic fibrosis airway epithelial cells and Cl- channel activation assessed in single cells using a fluorescence microscopic assay and the patch-clamp technique. Expression of CFTR, but not of a mutant form of CFTR (ΔF508), corrected the Cl- channel defect. Correction of the phenotypic defect demonstrates a causal relationship between mutations in the CFTR gene and defective Cl- transport which is the hallmark of the disease.

  18. Identification of human metapneumovirus-induced gene networks in airway epithelial cells by microarray analysis

    SciTech Connect

    Bao, X.; Sinha, M. |; Liu, T.; Hong, C.; Luxon, B.A. |; Garofalo, R.P. ||; Casola, A. ||

    2008-04-25

    Human metapneumovirus (hMPV) is a major cause of lower respiratory tract infections in infants, elderly and immunocompromised patients. Little is known about the response to hMPV infection of airway epithelial cells, which play a pivotal role in initiating and shaping innate and adaptive immune responses. In this study, we analyzed the transcriptional profiles of airway epithelial cells infected with hMPV using high-density oligonucleotide microarrays. Of the 47,400 transcripts and variants represented on the Affimetrix GeneChip Human Genome HG-U133 plus 2 array, 1601 genes were significantly altered following hMPV infection. Altered genes were then assigned to functional categories and mapped to signaling pathways. Many up-regulated genes are involved in the initiation of pro-inflammatory and antiviral immune responses, including chemokines, cytokines, type I interferon and interferon-inducible proteins. Other important functional classes up-regulated by hMPV infection include cellular signaling, gene transcription and apoptosis. Notably, genes associated with antioxidant and membrane transport activity, several metabolic pathways and cell proliferation were down-regulated in response to hMPV infection. Real-time PCR and Western blot assays were used to confirm the expression of genes related to several of these functional groups. The overall result of this study provides novel information on host gene expression upon infection with hMPV and also serves as a foundation for future investigations of genes and pathways involved in the pathogenesis of this important viral infection. Furthermore, it can facilitate a comparative analysis of other paramyxoviral infections to determine the transcriptional changes that are conserved versus the one that are specific to individual pathogens.

  19. Spi2 gene polymorphism is not associated with recurrent airway obstruction and inflammatory airway disease in thoroughbred horses

    PubMed Central

    da Silva, Aline Correa; Brass, Karin Erica; da Silva Loreto, Elgion; Vinocur, Myriam Elizabeth; Pozzobon, Ricardo; da Silva Azevedo, Marcos

    2011-01-01

    The aim was to detect the presence of polymorphisms at exons 1, 2, 3 and 4 of the Spi2 gene, and evaluate a possible association between them and recurrent airway obstruction (RAO) or inflammatory airway disease (IAD) in thoroughbred horses, through single-strand conformational-polymorphism (SSCP) screening. Although polymorphism was not detected in exons 1, 2 and 3, three alleles and six genotypes were identified in exon 4. The frequencies of allele A (0.6388) and genotype AA (0.3888) were higher in horses affected by RAO, although no association was found between polymorphism and horses with either RAO or IAD. PMID:21931519

  20. Differential transcriptional regulation of IL-8 expression by human airway epithelial cells exposed to diesel exhaust particles

    SciTech Connect

    Tal, Tamara L.; Simmons, Steven O.; Silbajoris, Robert; Dailey, Lisa; Cho, Seung-Hyun; Ramabhadran, Ram; Linak, William; Reed, William; Bromberg, Philip A.; Samet, James M.

    2010-02-15

    Exposure to diesel exhaust particles (DEP) induces inflammatory signaling characterized by MAP kinase-mediated activation of NFkB and AP-1 in vitro and in bronchial biopsies obtained from human subjects exposed to DEP. NFkB and AP-1 activation results in the upregulation of genes involved in promoting inflammation in airway epithelial cells, a principal target of inhaled DEP. IL-8 is a proinflammatory chemokine expressed by the airway epithelium in response to environmental pollutants. The mechanism by which DEP exposure induces IL-8 expression is not well understood. In the current study, we sought to determine whether DEP with varying organic content induces IL-8 expression in lung epithelial cells, as well as, to develop a method to rapidly evaluate the upstream mechanism(s) by which DEP induces IL-8 expression. Exposure to DEP with varying organic content differentially induced IL-8 expression and IL-8 promoter activity human airway epithelial cells. Mutational analysis of the IL-8 promoter was also performed using recombinant human cell lines expressing reporters linked to the mutated promoters. Treatment with a low organic-containing DEP stimulated IL-8 expression by a mechanism that is predominantly NFkB-dependent. In contrast, exposure to high organic-containing DEP induced IL-8 expression independently of NFkB through a mechanism that requires AP-1 activity. Our study reveals that exposure to DEP of varying organic content induces proinflammatory gene expression through multiple specific mechanisms in human airway epithelial cells. The approaches used in the present study demonstrate the utility of a promoter-reporter assay ensemble for identifying transcriptional pathways activated by pollutant exposure.

  1. Airway basal cells of healthy smokers express an embryonic stem cell signature relevant to lung cancer.

    PubMed

    Shaykhiev, Renat; Wang, Rui; Zwick, Rachel K; Hackett, Neil R; Leung, Roland; Moore, Malcolm A S; Sima, Camelia S; Chao, Ion Wa; Downey, Robert J; Strulovici-Barel, Yael; Salit, Jacqueline; Crystal, Ronald G

    2013-09-01

    Activation of the human embryonic stem cell (hESC) signature genes has been observed in various epithelial cancers. In this study, we found that the hESC signature is selectively induced in the airway basal stem/progenitor cell population of healthy smokers (BC-S), with a pattern similar to that activated in all major types of human lung cancer. We further identified a subset of 6 BC-S hESC genes, whose coherent overexpression in lung adenocarcinoma (AdCa) was associated with reduced lung function, poorer differentiation grade, more advanced tumor stage, remarkably shorter survival, and higher frequency of TP53 mutations. BC-S shared with hESC and a considerable subset of lung carcinomas a common TP53 inactivation molecular pattern which strongly correlated with the BC-S hESC gene expression. These data provide transcriptome-based evidence that smoking-induced reprogramming of airway BC toward the hESC-like phenotype might represent a common early molecular event in the development of aggressive lung carcinomas in humans.

  2. High expression of midkine in the airways of patients with cystic fibrosis.

    PubMed

    Nordin, Sara L; Jovic, Sandra; Kurut, Anil; Andersson, Cecilia; Gela, Anele; Bjartell, Anders; Mörgelin, Matthias; Olin, Anders I; Lund, Mikael; Egesten, Arne

    2013-12-01

    Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene result in impaired host defense during cystic fibrosis (CF), where Pseudomonas aeruginosa becomes a key pathogen. We investigated the expression pattern of the antibacterial growth factor midkine (MK) in CF and the possible interference with its activity by the altered airway microenvironment. High MK expression was found in CF lung tissue compared with control samples, involving epithelia of the large and small airways, alveoli, and cells of the submucosa (i.e., neutrophils and mast cells). In CF sputum, MK was present at 100-fold higher levels, but was also subject to increased degradation, compared with MK in sputum from healthy control subjects. MK exerted a bactericidal effect on P. aeruginosa, but increasing salt concentrations and low pH impaired this activity. Molecular modeling suggested that the effects of salt and pH were attributable to electrostatic screening and a charge-neutralization of the membrane, respectively. Both the neutrophil elastase and elastase of P. aeruginosa cleaved MK to smaller fragments, resulting in impaired bactericidal activity. Thus, MK is highly expressed in CF, but its bactericidal properties may be impaired by the altered microenvironment, as reflected by the in vitro conditions used in this study.

  3. Grepafloxacin inhibits tumor necrosis factor-alpha-induced interleukin-8 expression in human airway epithelial cells.

    PubMed

    Hashimoto, S; Matsumoto, K; Gon, Y; Maruoka, S; Hayashi, S; Asai, Y; Machino, T; Horie, T

    2000-01-01

    We examined the effect of grepafloxacin (GPFX), a new fluoroquinolone antimicrobial agent, on interleukin-8 (IL-8) expression in tumor necrosis factor-alpha (TNF-alpha)-stimulated human airway epithelial cells (AEC). GPFX inhibited IL-8 protein production as well as mRNA expression in a concentration-dependent manner (2.5 - 25 micro g/ml), but the inhibition of IL-8 expression by corresponding concentrations of GPFX to serum and airway lining fluids was not complete. We discuss the modulatory effect of GPFX on IL-8 production in the context of its efficacy on controlling chronic airway inflammatory diseases.

  4. Cigarette smoke extract inhibits expression of peroxiredoxin V and increases airway epithelial permeability.

    PubMed

    Serikov, Vladimir B; Leutenegger, Christian; Krutilina, Raisa; Kropotov, Andrei; Pleskach, Nadezhda; Suh, Jung H; Tomilin, Nikolay V

    2006-01-01

    Inhaled cigarette smoke induces oxidative stress in the epithelium of airways. Peroxiredoxin V (PRXV) is a potent antioxidant protein, highly expressed in cells of the airway epithelium. The goal of our study was to determine whether cigarette smoke extract (CSE) influenced expression of this protein in airway epithelia in vivo and in vitro. In Sprague-Dawley rats, we determined effects of CSE on airway epithelial permeability, mRNA levels and expression of PRXV protein. Exposure of isolated tracheal segment in vitro to 20% CSE for 4 h resulted in development of increased permeability to albumin, significantly reduced mRNA levels for PRXV, and reduced amounts of PRXV protein in the epithelium. In cultures of the airway epithelial cell lines (Calu-3, JME), primary airway cell culture (cow), and alveolar epithelial cells A549, CSE also significantly decreased transepithelial electrical resistance and expression of PRXV protein, and induced glutathione and protein oxidation. To demonstrate functional importance of PRXV, we exposed clones of HeLa cells with siRNA-downregulated PRXV to hydrogen peroxide, which resulted in increased rate of cell death and protein oxidation. CSE directly downregulates expression of functionally important antioxidant enzyme PRXV in the epithelial cells of airways, which represents one pathophysiological mechanism of cigarette smoke toxicity.

  5. Gene-environment interactions in a mutant mouse kindred with native airway constrictor hyperresponsiveness.

    PubMed

    Pinto, Lawrence H; Eaton, Emily; Chen, Bohao; Fleisher, Jonah; Shuster, Dmitry; McCauley, Joel; Kedainis, Dalius; Siepka, Sandra M; Shimomura, Kazuhiro; Song, Eun-Joo; Husain, Aliya; Lakser, Oren J; Mitchell, Richard W; Dowell, Maria L; Brown, Melanie; Camoretti-Mercado, Blanca; Naclerio, Robert; Sperling, Anne I; Levin, Stephen I; Turek, Fred W; Solway, Julian

    2008-01-01

    We mutagenized male BTBR mice with N-ethyl-N-nitrosourea and screened 1315 of their G3 offspring for airway hyperresponsiveness. A phenovariant G3 mouse with exaggerated methacholine bronchoconstrictor response was identified and his progeny bred in a nonspecific-pathogen-free (SPF) facility where sentinels tested positive for minute virus of mice and mouse parvovirus and where softwood bedding was used. The mutant phenotype was inherited through G11 as a single autosomal semidominant mutation with marked gender restriction, with males exhibiting almost full penetrance and very few females phenotypically abnormal. Between G11 and G12, facility infection eradication was undertaken and bedding was changed to hardwood. We could no longer detect airway hyperresponsiveness in more than 37 G12 offspring of 26 hyperresponsive G11 males. Also, we could not identify the mutant phenotype among offspring of hyperresponsive G8-G10 sires rederived into an SPF facility despite 21 attempts. These two observations suggest that both genetic and environmental factors were needed for phenotype expression. We suspect that rederivation into an SPF facility or altered exposure to pathogens or other unidentified substances modified environmental interactions with the mutant allele, and so resulted in disappearance of the hyperresponsive phenotype. Our experience suggests that future searches for genes that confer susceptibility for airway hyperresponsiveness might not be able to identify some genes that confer susceptibility if the searches are performed in SPF facilities. Experimenters are advised to arrange for multigeneration constancy of mouse care in order to clone mutant genes. Indeed, we were not able to map the mutation before losing the phenotype.

  6. Immune Mediators Regulate CFTR Expression through a Bifunctional Airway-Selective Enhancer

    PubMed Central

    Zhang, Zhaolin; Leir, Shih-Hsing

    2013-01-01

    An airway-selective DNase-hypersensitive site (DHS) at kb −35 (DHS-35kb) 5′ to the cystic fibrosis transmembrane conductance regulator (CFTR) gene is evident in many lung cell lines and primary human tracheal epithelial cells but is absent from intestinal epithelia. The DHS-35kb contains an element with enhancer activity in 16HBE14o- airway epithelial cells and is enriched for monomethylated H3K4 histones (H3K4me1). We now define a 350-bp region within DHS-35kb which has full enhancer activity and binds interferon regulatory factor 1 (IRF1) and nuclear factor Y (NF-Y) in vitro and in vivo. Small interfering RNA (siRNA)-mediated depletion of IRF1 or overexpression of IRF2, an antagonist of IRF1, reduces CFTR expression in 16HBE14o- cells. NF-Y is critical for maintenance of H3K4me1 enrichment at DHS-35kb since depletion of NF-YA, a subunit of NF-Y, reduces H3K4me1 enrichment at this site. Moreover, depletion of SETD7, an H3K4 monomethyltransferase, reduces both H3K4me1 and NF-Y occupancy, suggesting a requirement of H3K4me1 for NF-Y binding. NF-Y depletion also represses Sin3A and reduces its occupancy across the CFTR locus, which is accompanied by an increase in p300 enrichment at multiple sites. Our results reveal that the DHS-35kb airway-selective enhancer element plays a pivotal role in regulation of CFTR expression by two independent regulatory mechanisms. PMID:23689137

  7. Effect of dexamethasone and ACC on bacteria-induced mucin expression in human airway mucosa.

    PubMed

    Hauber, Hans-Peter; Goldmann, Torsten; Vollmer, Ekkehard; Wollenberg, Barbara; Zabel, Peter

    2007-11-01

    Gram-negative bacteria can stimulate mucin production, but excessive mucus supports bacterial infection and consequently leads to airway obstruction. Therefore, the effect of dexamethasone (DEX) and the antioxidant acetyl-cysteine (ACC) on bacteria-induced mucus expression was investigated. Explanted human airway mucosa and mucoepidermoid cells (Calu-3) were stimulated with lipopolysaccharide (LPS) or PAM3 (a synthetic lipoprotein). DEX or ACC were added to either LPS- or PAM3-stimulated airway mucosa or Calu-3 cells. Mucin mRNA expression (MUC5AC) and total mucus glycoconjugates (mucin protein) were quantified using real-time PCR and periodic acid Schiff staining. LPS and PAM3 significantly increased mucin expression in airway mucosa and Calu-3 cells (P < 0.05). DEX alone had no significant effect on mucin expression in airway mucosa or Calu-3 cells (P > 0.05). In contrast, DEX significantly reduced LPS- and PAM3-induced mucin expression in explanted mucosal tissue and mucin expression in Calu-3 cells (P < 0.05). In explanted human airway mucosa ACC alone significantly increased mucin expression (P < 0.05). In contrast, ACC significantly decreased LPS- and PAM3-induced mucin expression (P < 0.05). In Calu-3 cells ACC alone had no significant effect on mucin expression (P > 0.05). ACC decreased LPS- and PAM3-induced mucin expression, but this effect was not significant (P > 0.05). These data suggest that DEX can effectively reduce bacteria-induced mucin expression in the airways. ACC alone may increase mucin expression in noninfected mucosa, but it decreased bacteria-induced mucin expression. Further studies are warranted to evaluate whether the effect of DEX or ACC is clinically relevant.

  8. Soluble Fibre Meal Challenge Reduces Airway Inflammation and Expression of GPR43 and GPR41 in Asthma

    PubMed Central

    Halnes, Isabel; Baines, Katherine J.; Berthon, Bronwyn S.; MacDonald-Wicks, Lesley K.; Gibson, Peter G.; Wood, Lisa G.

    2017-01-01

    Short chain fatty acids (SCFAs) are produced following the fermentation of soluble fibre by gut bacteria. In animal models, both dietary fibre and SCFAs have demonstrated anti-inflammatory effects via the activation of free fatty acid receptors, such as G protein-coupled receptor 41 and 43 (GPR41 and GPR43). This pilot study examined the acute effect of a single dose of soluble fibre on airway inflammation—including changes in gene expression of free fatty acid receptors—in asthma. Adults with stable asthma consumed a soluble fibre meal (n = 17) containing 3.5 g inulin and probiotics, or a control meal (n = 12) of simple carbohydrates. Exhaled nitric oxide (eNO) was measured and induced sputum was collected at 0 and 4 h for differential cell counts, measurement of interleukin-8 (IL-8) protein concentration, and GPR41 and GPR43 gene expression. At 4 h after meal consumption, airway inflammation biomarkers, including sputum total cell count, neutrophils, macrophages, lymphocytes, sputum IL-8, and eNO significantly decreased compared to baseline in the soluble fibre group only. This corresponded with upregulated GPR41 and GPR43 sputum gene expression and improved lung function in the soluble fibre group alone. Soluble fibre has acute anti-inflammatory effects in asthmatic airways. Long-term effects of soluble fibre as an anti-inflammatory therapy in asthma warrants further investigation. PMID:28075383

  9. SAFETY AND EFFICIENCY OF MODULATING PARACELLULAR PERMEABILITY TO ENHANCE AIRWAY EPITHELIAL GENE TRANSFER IN VIVO

    EPA Science Inventory


    ABSTRACT

    We evaluated the safety of agents that enhance gene transfer by modulating paracellular permeability. Lactate dehydrogenase (LDH) and cytokine release were measured in polarized primary human airway epithelial (HAE) cells after luminal application of vehicle, ...

  10. Nacystelyn enhances adenoviral vector-mediated gene delivery to mouse airways.

    PubMed

    Kushwah, R; Oliver, J R; Cao, H; Hu, J

    2007-08-01

    Adenoviral vector-mediated gene delivery has been vastly investigated for cystic fibrosis (CF) gene therapy; however, one of its drawbacks is the low efficiency of gene transfer, which is due to basolateral colocalization of viral receptors, immune responses to viral vectors and the presence of a thick mucus layer in the airways of CF patients. Therefore, enhancement of gene transfer can lead to reduction in the viral dosage, which could further reduce the acute toxicity associated with the use of adenoviral vectors. Nacystelyn (NAL) is a mucolytic agent with anti-inflammatory and antioxidant properties, and has been used clinically in CF patients to reduce mucus viscosity in the airways. In this study, we show that pretreatment of the airways with NAL followed by administration of adenoviral vectors in complex with DEAE-Dextran can significantly enhance gene delivery to the airways of mice without any harmful effects. Moreover, NAL pretreatment can reduce the airway inflammation, which is normally observed after delivery of adenoviral particles. Taken together, these results indicate that NAL pretreatment followed by adenoviral vector-mediated gene delivery can be beneficial to CF patients by increasing the efficiency of gene transfer to the airways, and reducing the acute toxicity associated with the administration of adenoviral vectors.

  11. Genetic modification of adeno-associated viral vector type 2 capsid enhances gene transfer efficiency in polarized human airway epithelial cells.

    PubMed

    White, April F; Mazur, Marina; Sorscher, Eric J; Zinn, Kurt R; Ponnazhagan, Selvarangan

    2008-12-01

    Cystic fibrosis (CF) is a common genetic disease characterized by defects in the expression of the CF transmembrane conductance regulator (CFTR) gene. Gene therapy offers better hope for the treatment of CF. Adeno-associated viral (AAV) vectors are capable of stable expression with low immunogenicity. Despite their potential in CF gene therapy, gene transfer efficiency by AAV is limited because of pathophysiological barriers in these patients. Although a few AAV serotypes have shown better transduction compared with the AAV2-based vectors, gene transfer efficiency in human airway epithelium has still not reached therapeutic levels. To engineer better AAV vectors for enhanced gene delivery in human airway epithelium, we developed and characterized mutant AAV vectors by genetic capsid modification, modeling the well-characterized AAV2 serotype. We genetically incorporated putative high-affinity peptide ligands to human airway epithelium on the GH loop region of AAV2 capsid protein. Six independent mutant AAV were constructed, containing peptide ligands previously reported to bind with high affinity for known and unknown receptors on human airway epithelial cells. The vectors were tested on nonairway cells and nonpolarized and polarized human airway epithelial cells for enhanced infectivity. One of the mutant vectors, with the peptide sequence THALWHT, not only showed the highest transduction in undifferentiated human airway epithelial cells but also indicated significant transduction in polarized cells. Interestingly, this modified vector was also able to infect cells independently of the heparan sulfate proteoglycan receptor. Incorporation of this ligand on other AAV serotypes, which have shown improved gene transfer efficiency in the human airway epithelium, may enhance the application of AAV vectors in CF gene therapy.

  12. Endotoxin responsiveness of human airway epithelia is limited by low expression of MD-2.

    PubMed

    Jia, Hong Peng; Kline, Joel N; Penisten, Andrea; Apicella, Michael A; Gioannini, Theresa L; Weiss, Jerrold; McCray, Paul B

    2004-08-01

    The expression of inducible antimicrobial peptides, such as human beta-defensin-2 (HBD-2) by epithelia, comprises a component of innate pulmonary defenses. We hypothesized that HBD-2 induction in airway epithelia is linked to pattern recognition receptors such as the Toll-like receptors (TLRs). We found that primary cultures of well-differentiated human airway epithelia express the mRNA for TLR-4, but little or no MD-2 mRNA, and display little HBD-2 expression in response to treatment with purified endotoxin +/- LPS binding protein (LBP) and soluble CD14. Expression of endogenous MD-2 by transduction of airway epithelial cells with an adenoviral vector encoding MD-2 or extracellular addition of recombinant MD-2 both increased the responses of airway epithelia to endotoxin + LBP and sCD14 by >100-fold, as measured by NF-kappaB-luciferase activity and HBD-2 mRNA expression. MD-2 mRNA could be induced in airway epithelia by exposure of these cells to specific bacterial or host products (e.g., killed Haemophilus influenzae, the P6 outer membrane protein from H. influenzae, or TNF-alpha + IFN-gamma). These findings suggest that MD-2, either coexpressed with TLR-4 or secreted when produced in excess of TLR-4 from neighboring cells, is required for airway epithelia to respond sensitively to endotoxin. The regulation of MD-2 expression in airway epithelia and pulmonary macrophages may serve as a means to modify endotoxin responsiveness in the airway.

  13. Human airway smooth muscle cells secrete amphiregulin via bradykinin/COX-2/PGE2, inducing COX-2, CXCL8, and VEGF expression in airway epithelial cells

    PubMed Central

    Knox, Alan J.

    2015-01-01

    Human airway smooth muscle cells (HASMC) contribute to asthma pathophysiology through an increased smooth muscle mass and elevated cytokine/chemokine output. Little is known about how HASMC and the airway epithelium interact to regulate chronic airway inflammation and remodeling. Amphiregulin is a member of the family of epidermal growth factor receptor (EGFR) agonists with cell growth and proinflammatory roles and increased expression in the lungs of asthma patients. Here we show that bradykinin (BK) stimulation of HASMC increases amphiregulin secretion in a mechanism dependent on BK-induced COX-2 expression, increased PGE2 output, and the stimulation of HASMC EP2 and EP4 receptors. Conditioned medium from BK treated HASMC induced CXCL8, VEGF, and COX-2 mRNA and protein accumulation in airway epithelial cells, which were blocked by anti-amphiregulin antibodies and amphiregulin siRNA, suggesting a paracrine effect of HASMC-derived amphiregulin on airway epithelial cells. Consistent with this, recombinant amphiregulin induced CXCL8, VEGF, and COX-2 in airway epithelial cells. Finally, we found that conditioned media from amphiregulin-stimulated airway epithelial cells induced amphiregulin expression in HASMC and that this was dependent on airway epithelial cell COX-2 activity. Our study provides evidence of a dynamic axis of interaction between HASMC and epithelial cells that amplifies CXCL8, VEGF, COX-2, and amphiregulin production. PMID:26047642

  14. Histone deacetylase inhibitors up-regulate LL-37 expression independent of toll-like receptor mediated signalling in airway epithelial cells.

    PubMed

    Liu, Quan; Liu, Juan; Roschmann, Kristina Irene Lisolette; van Egmond, Danielle; Golebski, Korneliusz; Fokkens, Wytske Johanna; Wang, Dehui; van Drunen, Cornelis Maria

    2013-04-11

    HDAC inhibitors have been proposed as anticancer agents. However, their roles in innate genes expression remain not well known. Cathelicidin LL-37 is one of the few human bactericidal peptides, but the regulation of histone acetylation on LL-37 expression in airway epithelium remains largely unknown. Therefore, we investigated the effects of two non-selective HDACi, trichostatin A (TSA) and sodium butyrate (SB), on the expression of the cathelicidin LL-37 in human airway epithelial cells. LL37 in human NCI-H292 airway epithelial cells and the primary cultures of normal nasal epithelial cells(PNEC) in response to HDAC inhibitors with or without poly (I:C) stimulation was assessed using real-time PCR and western blot. In parallel, IL-6 expression was evaluated by ELISA. Our results showed that HDAC inhibitors up-regulated LL-37 gene expression independent of poly (I:C) stimulation in PNEC as well as in NCI-H292 cells. HDAC inhibitors increased LL37 protein expression in NCI-H292 cells but not in PNEC. In addition, HDAC inhibitors significantly inhibited poly (I:C)-induced IL-6 production in both of the epithelial cells. In conclusion, HDAC inhibitors directly up-regulated LL-37 gene expression in human airway epithelial cells.

  15. Histone deacetylase inhibitors up-regulate LL-37 expression independent of toll-like receptor mediated signalling in airway epithelial cells

    PubMed Central

    2013-01-01

    HDAC inhibitors have been proposed as anticancer agents. However, their roles in innate genes expression remain not well known. Cathelicidin LL-37 is one of the few human bactericidal peptides, but the regulation of histone acetylation on LL-37 expression in airway epithelium remains largely unknown. Therefore, we investigated the effects of two non-selective HDACi, trichostatin A (TSA) and sodium butyrate (SB), on the expression of the cathelicidin LL-37 in human airway epithelial cells. LL37 in human NCI-H292 airway epithelial cells and the primary cultures of normal nasal epithelial cells(PNEC) in response to HDAC inhibitors with or without poly (I:C) stimulation was assessed using real-time PCR and western blot. In parallel, IL-6 expression was evaluated by ELISA. Our results showed that HDAC inhibitors up-regulated LL-37 gene expression independent of poly (I:C) stimulation in PNEC as well as in NCI-H292 cells. HDAC inhibitors increased LL37 protein expression in NCI-H292 cells but not in PNEC. In addition, HDAC inhibitors significantly inhibited poly (I:C)-induced IL-6 production in both of the epithelial cells. In conclusion, HDAC inhibitors directly up-regulated LL-37 gene expression in human airway epithelial cells. PMID:23577829

  16. Interleukin 12 inhibits antigen-induced airway hyperresponsiveness, inflammation, and Th2 cytokine expression in mice

    PubMed Central

    1995-01-01

    Allergic asthma is characterized by airway hyperresponsiveness and pulmonary eosinophilia, and may be mediated by T helper (Th) lymphocytes expressing a Th2 cytokine pattern. Interleukin (IL) 12 suppresses the expression of Th2 cytokines and their associated responses, including eosinophilia, serum immunoglobulin E, and mucosal mastocytosis. We have previously shown in a murine model that antigen- induced increases in airway hyperresponsiveness and pulmonary eosinophilia are CD4+ T cell dependent. We used this model to determine the ability of IL-12 to prevent antigen-induced increases in airway hyperresponsiveness, bronchoalveolar lavage (BAL) eosinophils, and lung Th2 cytokine expression. Sensitized A/J mice developed airway hyperresponsiveness and increased numbers of BAL eosinophils and other inflammatory cells after single or repeated intratracheal challenges with sheep red blood cell antigen. Pulmonary mRNA and protein levels of the Th2 cytokines IL-4 and IL-5 were increased after antigen challenge. Administration of IL-12 (1 microgram/d x 5 d) at the time of a single antigen challenge abolished the airway hyperresponsiveness and pulmonary eosinophilia and promoted an increase in interferon (IFN) gamma and decreases in IL-4 and IL-5 expression. The effects of IL-12 were partially dependent on IFN-gamma, because concurrent treatment with IL-12 and anti-IFN-gamma monoclonal antibody partially reversed the inhibition of airway hyperresponsiveness and eosinophilia by IL-12. Treatment of mice with IL-12 at the time of a second antigen challenge also prevented airway hyperresponsiveness and significantly reduced numbers of BAL inflammatory cells, reflecting the ability of IL-12 to inhibit responses associated with ongoing antigen-induced pulmonary inflammation. These data show that antigen-induced airway hyperresponsiveness and inflammation can be blocked by IL-12, which suppresses Th2 cytokine expression. Local administration of IL-12 may provide a novel

  17. N-acetylcysteine enhances cystic fibrosis sputum penetration and airway gene transfer by highly compacted DNA nanoparticles.

    PubMed

    Suk, Jung Soo; Boylan, Nicholas J; Trehan, Kanika; Tang, Benjamin C; Schneider, Craig S; Lin, Jung-Ming G; Boyle, Michael P; Zeitlin, Pamela L; Lai, Samuel K; Cooper, Mark J; Hanes, Justin

    2011-11-01

    For effective airway gene therapy of cystic fibrosis (CF), inhaled gene carriers must first penetrate the hyperviscoelastic sputum covering the epithelium. Whether clinically studied gene carriers can penetrate CF sputum remains unknown. Here, we measured the diffusion of a clinically tested nonviral gene carrier, composed of poly-l-lysine conjugated with a 10 kDa polyethylene glycol segment (CK(30)PEG(10k)). We found that CK(30)PEG(10k)/DNA nanoparticles were trapped in CF sputum. To improve gene carrier diffusion across sputum, we tested adjuvant regimens consisting of N-acetylcysteine (NAC), recombinant human DNase (rhDNase) or NAC together with rhDNase. While rhDNase alone did not enhance gene carrier diffusion, NAC and NAC + rhDNase increased average effective diffusivities by 6-fold and 13-fold, respectively, leading to markedly greater fractions of gene carriers that may penetrate sputum layers. We further tested the adjuvant effects of NAC in the airways of mice with Pseudomonas aeruginosa lipopolysaccharide (LPS)-induced mucus hypersecretion. Intranasal dosing of NAC prior to CK(30)PEG(10k)/DNA nanoparticles enhanced gene expression by up to ~12-fold compared to saline control, reaching levels observed in the lungs of mice without LPS challenge. Our findings suggest that a promising synthetic nanoparticle gene carrier may transfer genes substantially more effectively to lungs of CF patients if administered following adjuvant mucolytic therapy with NAC or NAC + rhDNase.

  18. N-acetylcysteine Enhances Cystic Fibrosis Sputum Penetration and Airway Gene Transfer by Highly Compacted DNA Nanoparticles.

    PubMed

    Suk, Jung Soo; Boylan, Nicholas J; Trehan, Kanika; Tang, Benjamin C; Schneider, Craig S; Lin, Jung-Ming G; Boyle, Michael P; Zeitlin, Pamela L; Lai, Samuel K; Cooper, Mark J; Hanes, Justin

    2011-11-01

    For effective airway gene therapy of cystic fibrosis (CF), inhaled gene carriers must first penetrate the hyperviscoelastic sputum covering the epithelium. Whether clinically studied gene carriers can penetrate CF sputum remains unknown. Here, we measured the diffusion of a clinically tested nonviral gene carrier, composed of poly-l-lysine conjugated with a 10 kDa polyethylene glycol segment (CK30PEG10k). We found that CK30PEG10k/DNA nanoparticles were trapped in CF sputum. To improve gene carrier diffusion across sputum, we tested adjuvant regimens consisting of N-acetylcysteine (NAC), recombinant human DNase (rhDNase) or NAC together with rhDNase. While rhDNase alone did not enhance gene carrier diffusion, NAC and NAC + rhDNase increased average effective diffusivities by 6-fold and 13-fold, respectively, leading to markedly greater fractions of gene carriers that may penetrate sputum layers. We further tested the adjuvant effects of NAC in the airways of mice with Pseudomonas aeruginosa lipopolysaccharide (LPS)-induced mucus hypersecretion. Intranasal dosing of NAC prior to CK30PEG10k/DNA nanoparticles enhanced gene expression by up to ~12-fold compared to saline control, reaching levels observed in the lungs of mice without LPS challenge. Our findings suggest that a promising synthetic nanoparticle gene carrier may transfer genes substantially more effectively to lungs of CF patients if administered following adjuvant mucolytic therapy with NAC or NAC + rhDNase.

  19. Cystic fibrosis gene modifier SLC26A9 modulates airway response to CFTR-directed therapeutics.

    PubMed

    Strug, Lisa J; Gonska, Tanja; He, Gengming; Keenan, Katherine; Ip, Wan; Boëlle, Pierre-Yves; Lin, Fan; Panjwani, Naim; Gong, Jiafen; Li, Weili; Soave, David; Xiao, Bowei; Tullis, Elizabeth; Rabin, Harvey; Parkins, Michael D; Price, April; Zuberbuhler, Peter C; Corvol, Harriet; Ratjen, Felix; Sun, Lei; Bear, Christine E; Rommens, Johanna M

    2016-08-29

    Cystic fibrosis is realizing the promise of personalized medicine. Recent advances in drug development that target the causal CFTR directly result in lung function improvement, but variability in response is demanding better prediction of outcomes to improve management decisions. The genetic modifier SLC26A9 contributes to disease severity in the CF pancreas and intestine at birth and here we assess its relationship with disease severity and therapeutic response in the airways. SLC26A9 association with lung disease was assessed in individuals from the Canadian and French CF Gene Modifier consortia with CFTR-gating mutations and in those homozygous for the common Phe508del mutation. Variability in response to a CFTR-directed therapy attributed to SLC26A9 genotype was assessed in Canadian patients with gating mutations. A primary airway model system determined if SLC26A9 shows modification of Phe508del CFTR function upon treatment with a CFTR corrector.In those with gating mutations that retain cell surface-localized CFTR we show that SLC26A9 modifies lung function while this is not the case in individuals homozygous for Phe508del where cell surface expression is lacking. Treatment response to ivacaftor, which aims to improve CFTR-channel opening probability in patients with gating mutations, shows substantial variability in response, 28% of which can be explained by rs7512462 in SLC26A9 (P = 0.0006). When homozygous Phe508del primary bronchial cells are treated to restore surface CFTR, SLC26A9 likewise modifies treatment response (P = 0.02). Our findings indicate that SLC26A9 airway modification requires CFTR at the cell surface, and that a common variant in SLC26A9 may predict response to CFTR-directed therapeutics.

  20. Expression of beta-defensin-1 in chimpanzee (Pan troglodytes) airways.

    PubMed

    Duits, L A; Langermans, J A; Paltansing, S; van der Straaten, T; Vervenne, R A; Frost, P A; Hiemstra, P S; Thomas, A W; Nibbering, P H

    2000-10-01

    In human airways, beta-defensins function in the elimination of various pathogens. They have been identified in a wide range of species. Here we report the identification and expression of chimpanzee beta-defensin-1 (cBD1), which is a homolog of human beta-defensin-1, in chimpanzee airways and skin. The cBD1 cDNA sequence differs by only one synonymous nucleotide substitution compared to the human cDNA sequence. In situ hybridization revealed that in lung tissue beside alveolar macrophages also airway epithelial cells, endothelial cells and type II pneumocytes express cBD1 mRNA. In skin, cBD1 mRNA was expressed in keratinocytes and endothelial cells. Together, these results show similarity in structure and expression pattern and perhaps in function.

  1. Klotho expression is reduced in COPD airway epithelial cells: effects on inflammation and oxidant injury.

    PubMed

    Gao, Wei; Yuan, Cheng; Zhang, Jingying; Li, Lingling; Yu, Like; Wiegman, Coen H; Barnes, Peter J; Adcock, Ian M; Huang, Mao; Yao, Xin

    2015-12-01

    COPD (chronic obstructive pulmonary disease) is associated with sustained inflammation, excessive injury, and accelerated lung aging. Human Klotho (KL) is an anti-aging protein that protects cells against inflammation and damage. In the present study, we quantified KL expression in the lungs of COPD patients and in an ozone-induced mouse model of COPD, and investigated the mechanisms that control KL expression and function in the airways. KL distribution and levels in human and mouse airways were measured by immunohistochemistry and Western blotting. The effect of CSE (cigarette smoke extract) on KL expression was detected in human bronchial epithelial cells. Moreover, the effect of KL on CSE-mediated inflammation and hydrogen peroxide-induced cellular injury/apoptosis was determined using siRNAs. KL expression was decreased in the lungs of smokers and further reduced in patients with COPD. Similarly, 6 weeks of exposure to ozone decreased KL levels in airway epithelial cells. CSE and TNFα (tumour necrosis factor α) decreased KL expression and release from airway epithelial cells, which was associated with enhanced pro-inflammatory cytokine expression. Moreover, KL depletion increased cell sensitivity to cigarette smoke-induced inflammation and oxidative stress-induced cell damage. These effects involved the NF-κB (nuclear factor κB), MAPK (mitogen-activated protein kinase) and Nrf2 (nuclear factor erythroid 2-related factor 2) pathways. Reduced KL expression in COPD airway epithelial cells was associated with increased oxidative stress, inflammation and apoptosis. These data provide new insights into the mechanisms associated with the accelerated lung aging in COPD development.

  2. Klotho expression is reduced in COPD airway epithelial cells: effects on inflammation and oxidant injury

    PubMed Central

    Gao, Wei; Yuan, Cheng; Zhang, Jingying; Li, Lingling; Yu, Like; Wiegman, Coen H.; Barnes, Peter J.; Adcock, Ian M.; Huang, Mao

    2015-01-01

    COPD (chronic obstructive pulmonary disease) is associated with sustained inflammation, excessive injury, and accelerated lung aging. Human Klotho (KL) is an anti-aging protein that protects cells against inflammation and damage. In the present study, we quantified KL expression in the lungs of COPD patients and in an ozone-induced mouse model of COPD, and investigated the mechanisms that control KL expression and function in the airways. KL distribution and levels in human and mouse airways were measured by immunohistochemistry and Western blotting. The effect of CSE (cigarette smoke extract) on KL expression was detected in human bronchial epithelial cells. Moreover, the effect of KL on CSE-mediated inflammation and hydrogen peroxide-induced cellular injury/apoptosis was determined using siRNAs. KL expression was decreased in the lungs of smokers and further reduced in patients with COPD. Similarly, 6 weeks of exposure to ozone decreased KL levels in airway epithelial cells. CSE and TNFα (tumour necrosis factor α) decreased KL expression and release from airway epithelial cells, which was associated with enhanced pro-inflammatory cytokine expression. Moreover, KL depletion increased cell sensitivity to cigarette smoke-induced inflammation and oxidative stress-induced cell damage. These effects involved the NF-κB (nuclear factor κB), MAPK (mitogen-activated protein kinase) and Nrf2 (nuclear factor erythroid 2-related factor 2) pathways. Reduced KL expression in COPD airway epithelial cells was associated with increased oxidative stress, inflammation and apoptosis. These data provide new insights into the mechanisms associated with the accelerated lung aging in COPD development. PMID:26201096

  3. Cyclooxygenase-2 and microRNA-155 expression are elevated in asthmatic airway smooth muscle cells.

    PubMed

    Comer, Brian S; Camoretti-Mercado, Blanca; Kogut, Paul C; Halayko, Andrew J; Solway, Julian; Gerthoffer, William T

    2015-04-01

    Cyclooxygenase-2 (COX-2) expression and PGE2 secretion from human airway smooth muscle cells (hASMCs) may contribute to β2-adrenoceptor hyporesponsiveness, a clinical feature observed in some patients with asthma. hASMCs from patients with asthma exhibit elevated expression of cytokine-responsive genes, and in some instances this is attributable to an altered histone code and/or microRNA expression. We hypothesized that COX-2 expression and PGE2 secretion might be elevated in asthmatic hASMCs in response to proinflammatory signals in part due to altered histone acetylation and/or microRNA expression. hASMCs obtained from nonasthmatic and asthmatic human subjects were treated with cytomix (IL-1β, TNF-α, and IFN-γ). A greater elevation of COX-2 mRNA, COX-2 protein, and PGE2 secretion was observed in the asthmatic cells. We investigated histone H3/H4-acetylation, transcription factor binding, mRNA stability, p38 mitogen-activated protein kinase signaling, and microRNA (miR)-155 expression as potential mechanisms responsible for the differential elevation of COX-2 expression. We found that histone H3/H4-acetylation and transcription factor binding to the COX-2 promoter were similar in both groups, and histone H3/H4-acetylation did not increase after cytomix treatment. Cytomix treatment elevated NF-κB and RNA polymerase II binding to similar levels in both groups. COX-2 mRNA stability was increased in asthmatic cells. MiR-155 expression was higher in cytomix-treated asthmatic cells, and we show it enhances COX-2 expression and PGE2 secretion in asthmatic and nonasthmatic hASMCs. Thus, miR-155 expression positively correlates with COX-2 expression in the asthmatic hASMCs and may contribute to the elevated expression observed in these cells. These findings may explain, at least in part, β2-adrenoceptor hyporesponsiveness in patients with asthma.

  4. MD2 expression is reduced in large airways of smokers and COPD smokers.

    PubMed

    Pace, Elisabetta; Ferraro, Maria; Chiappara, Giuseppina; Vitulo, Patrizio; Pipitone, Loredana; Di Vincenzo, Serena; Gjomarkaj, Mark

    2015-09-01

    Toll-like receptor 4 (TLR4) signaling requires a number of accessory proteins to initiate a signal. MD-2 is one of the accessory proteins with a relevant role in lipopolysaccharide responses. Although cigarette smoke increases TLR4 expression, TLR4 signaling is altered in smokers and in smokers COPD patients. The main aims of this study were to explore whether MD2 is altered in large and small airways of COPD and of smokers without COPD. The expression of MD2 ex vivo was assessed by immunohistochemistry in surgical specimens from current smokers COPD (s-COPD; n = 14), smokers without COPD (S; n = 7), and from non-smoker non-COPD subjects (C; n = 11. The in vitro effects of cigarette smoke extracts on the MD2 expression in a human bronchial epithelial cell line (16-HBE) were also assessed by flow cytometry. MD2 is reduced in the epithelium and in the submucosa in large airways but not in the epithelium and in the submucosa in small airways of smokers and of s-COPD. The expression of MD2 in the submucosa of the large airways is significantly higher in comparison to the submucosa of the small airways in all the studied groups. In vitro, cigarette smoke is able to increase TLR4 but it reduces MD2 in a dose-dependent manner in bronchial epithelial cells. Cigarette smoke may alter innate immune responses reducing the expression of the MD2, a molecule with an important role in TLR4 signaling.

  5. Gene Transfer by Guanidinium-Cholesterol Cationic Lipids into Airway Epithelial Cells in vitro and in vivo

    NASA Astrophysics Data System (ADS)

    Oudrhiri, Noufissa; Vigneron, Jean-Pierre; Peuchmaur, Michel; Leclerc, Tony; Lehn, Jean-Marie; Lehn, Pierre

    1997-03-01

    Synthetic vectors represent an attractive alternative approach to viral vectors for gene transfer, in particular into airway epithelial cells for lung-directed gene therapy for cystic fibrosis. Having recently found that guanidinium-cholesterol cationic lipids are efficient reagents for gene transfer into mammalian cell lines in vitro, we have investigated their use for gene delivery into primary airway epithelial cells in vitro and in vivo. The results obtained indicate that the lipid bis (guanidinium)-tren-cholesterol (BGTC) can be used to transfer a reporter gene into primary human airway epithelial cells in culture. Furthermore, liposomes composed of BGTC and dioleoyl phosphatidylethanolamine (DOPE) are efficient for gene delivery to the mouse airway epithelium in vivo. Transfected cells were detected both in the surface epithelium and in submucosal glands. In addition, the transfection efficiency of BGTC/DOPE liposomes in vivo was quantitatively assessed by using the luciferase reporter gene system.

  6. Matrix metalloproteinase expression and activity in human airway smooth muscle cells

    PubMed Central

    Elshaw, Shona R; Henderson, Neil; Knox, Alan J; Watson, Susan A; Buttle, David J; Johnson, Simon R

    2004-01-01

    Airway remodelling is a feature of chronic asthma comprising smooth muscle hypertrophy and deposition of extracellular matrix (ECM) proteins. Matrix metalloproteinases (MMPs) breakdown ECM, are involved in tissue remodelling and have been implicated in airway remodelling. Although mesenchymal cells are an important source of MMPs, little data are available on airway smooth muscle (ASM) derived MMPs. We therefore investigated MMP and tissue inhibitor of metalloproteinase (TIMP) production and activity in human ASM cells.MMPs and TIMPs were examined using quantitative real-time RT–PCR, Western blotting, zymography and a quench fluorescence (QF) assay of total MMP activity.The most abundant MMPs were pro-MMP-2, pro- MMP-3, active MMP-3 and MT1-MMP. TIMP-1 and TIMP-2 expression was low in cell lysates but high in conditioned medium. High TIMP secretion was confirmed by the ability of ASM-conditioned medium to inhibit recombinant MMP-2 in a QF assay. Thrombin increased MMP activity by activation of pro-MMP-2 independent of the conventional smooth muscle thrombin receptors PAR 1 and 4.In conclusion, ASM cells express pro-MMP-2, pro and active MMP-3, MMP-9 and MT1-MMP. Unstimulated cells secrete excess TIMP 1 and 2, preventing proteolytic activity. MMP-2 can be activated by thrombin which may contribute to airway remodelling. PMID:15265805

  7. Persistence of Smoking-Induced Dysregulation of MiRNA Expression in the Small Airway Epithelium Despite Smoking Cessation

    PubMed Central

    Strulovici-Barel, Yael; Salit, Jacqueline; Staudt, Michelle R.; Ahmed, Joumana; Tilley, Ann E.; Yee-Levin, Jenny; Hollmann, Charleen; Harvey, Ben-Gary; Kaner, Robert J.; Mezey, Jason G.; Sridhar, Sriram; Pillai, Sreekumar G.; Hilton, Holly; Wolff, Gerhard; Bitter, Hans; Visvanathan, Sudha; Fine, Jay S.; Stevenson, Christopher S.; Crystal, Ronald G.

    2015-01-01

    Even after quitting smoking, the risk of the development of chronic obstructive pulmonary disease (COPD) and lung cancer remains significantly higher compared to healthy nonsmokers. Based on the knowledge that COPD and most lung cancers start in the small airway epithelium (SAE), we hypothesized that smoking modulates miRNA expression in the SAE linked to the pathogenesis of smoking-induced airway disease, and that some of these changes persist after smoking cessation. SAE was collected from 10th to 12th order bronchi using fiberoptic bronchoscopy. Affymetrix miRNA 2.0 arrays were used to assess miRNA expression in the SAE from 9 healthy nonsmokers and 10 healthy smokers, before and after they quit smoking for 3 months. Smoking status was determined by urine nicotine and cotinine measurement. There were significant differences in the expression of 34 miRNAs between healthy smokers and healthy nonsmokers (p<0.01, fold-change >1.5), with functions associated with lung development, airway epithelium differentiation, inflammation and cancer. After quitting smoking for 3 months, 12 out of the 34 miRNAs did not return to normal levels, with Wnt/β-catenin signaling pathway being the top identified enriched pathway of the target genes of the persistent dysregulated miRNAs. In the context that many of these persistent smoking-dependent miRNAs are associated with differentiation, inflammatory diseases or lung cancer, it is likely that persistent smoking-related changes in SAE miRNAs play a role in the subsequent development of these disorders. PMID:25886353

  8. Down-regulation of 8-oxoguanine DNA glycosylase 1 expression in the airway epithelium ameliorates allergic lung inflammation.

    PubMed

    Bacsi, Attila; Aguilera-Aguirre, Leopoldo; Szczesny, Bartosz; Radak, Zsolt; Hazra, Tapas K; Sur, Sanjiv; Ba, Xueqing; Boldogh, Istvan

    2013-01-01

    Allergic airway inflammation is characterized by increased expression of pro-inflammatory mediators, inflammatory cell infiltration, mucus hypersecretion, and airway hyperresponsiveness, in parallel with oxidative DNA base and strand damage, whose etiological role is not understood. Our goal was to establish the role of 8-oxoguanine (8-oxoG), a common oxidatively damaged base, and its repair by 8-oxoguanine DNA glycosylase 1 (Ogg1) in allergic airway inflammatory processes. Airway inflammation was induced by intranasally administered ragweed (Ambrosia artemisiifolia) pollen grain extract (RWPE) in sensitized BALB/c mice. We utilized siRNA technology to deplete Ogg1 from airway epithelium; 8-oxoG and DNA strand break levels were quantified by Comet assays. Inflammatory cell infiltration and epithelial methaplasia were determined histologically, mucus and cytokines levels biochemically and enhanced pause was used as the main index of airway hyperresponsiveness. Decreased Ogg1 expression and thereby 8-oxoG repair in the airway epithelium conveyed a lower inflammatory response after RWPE challenge of sensitized mice, as determined by expression of Th2 cytokines, eosinophilia, epithelial methaplasia, and airway hyperresponsiveness. In contrast, 8-oxoG repair in Ogg1-proficient airway epithelium was coupled to an increase in DNA single-strand break (SSB) levels and exacerbation of allergen challenge-dependent inflammation. Decreased expression of the Nei-like glycosylases Neil1 and Neil2 that preferentially excise ring-opened purines and 5-hydroxyuracil, respectively, did not alter the above parameters of allergic immune responses to RWPE. These results show that DNA SSBs formed during Ogg1-mediated repair of 8-oxoG augment antigen-driven allergic immune responses. A transient modulation of OGG1 expression/activity in airway epithelial cells could have clinical benefits.

  9. Endosomal processing limits gene transfer to polarized airway epithelia by adeno-associated virus

    PubMed Central

    Duan, Dongsheng; Yue, Yongping; Yan, Ziying; Yang, Jusan; Engelhardt, John F.

    2000-01-01

    The restriction of viral receptors and coreceptors to the basolateral surface of airway epithelial cells has been blamed for the inefficient transfer of viral vectors to the apical surface of this tissue. We now report, however, that differentiated human airway epithelia internalize rAAV type-2 virus efficiently from their apical surfaces, despite the absence of known adeno-associated virus–2 (AAV-2) receptors or coreceptors at these sites. The dramatically lower transduction efficiency of rAAV infection from the apical surface of airway cells appears to result instead from differences in endosomal processing and nuclear trafficking of apically or basolaterally internalized virions. AAV capsid proteins are ubiquitinated after endocytosis, and gene transfer can be significantly enhanced by proteasome or ubiquitin ligase inhibitors. Tripeptide proteasome inhibitors increased persistent rAAV gene delivery from the apical surface >200-fold, to a level nearly equivalent to that achieved with basolateral infection. In vivo application of proteasome inhibitor in mouse lung augmented rAAV gene transfer from undetectable levels to a mean of 10.4 ± 1.6% of the epithelial cells in large bronchioles. Proteasome inhibitors also increased rAAV-2–mediated gene transfer to the liver tenfold, but they did not affect transduction of skeletal or cardiac muscle. These findings suggest that tissue-specific ubiquitination of viral capsid proteins interferes with rAAV-2 transduction and provides new approaches to circumvent this barrier for gene therapy of diseases such as cystic fibrosis. PMID:10841516

  10. Human-specific cystic fibrosis transmembrane conductance regulator antibodies detect in vivo gene transfer to ovine airways.

    PubMed

    Davidson, Heather; McLachlan, Gerry; Wilson, Abigail; Boyd, A Christopher; Doherty, Ann; MacGregor, Gordon; Davies, Lee; Painter, Hazel A; Coles, Rebecca; Hyde, Stephen C; Gill, Deborah R; Amaral, Margarida D; Collie, David D S; Porteous, David J; Penque, Deborah

    2006-07-01

    A panel of 11 human cystic fibrosis transmembrane conductance regulator (hCFTR) antibodies were tested in ovine nasal, tracheal, and bronchial epithelial brushings. Two of these, G449 (polyclonal) and MATG1104 (monoclonal), recognized hCFTR but did not cross react with endogenous sheep CFTR. This specificity allows immunologic detection of hCFTR expressed in gene transfer studies in sheep against the background of endogenous ovine CFTR, thus enhancing the value of the sheep as a model animal in which to study CFTR gene transfer. Studies on mixed populations of human and sheep nasal epithelial cells showed that detection of hCFTR by these two antibodies was possible even at the lowest proportion of human cells (1:100). The hCFTR gene was delivered in vivo by local instillation using polyethylenimine-mediated gene transfer to the ventral surface of the ovine trachea and hCFTR mRNA and protein levels scored in a blinded fashion. Despite abundant hCFTR mRNA expression, the number of cells expressing hCFTR protein detectable by G449 was low (approximately 0.006-0.05%). Immunohistochemistry for hCFTR in animals treated by whole-lung aerosol demonstrated positive cells in sections of tracheal epithelium and in distal conducting airways. The strategic use of hCFTR-specific antibodies supports the utility of the normal sheep as a model for hCFTR gene transfer studies.

  11. Semaphorin 7A is expressed on airway eosinophils and upregulated by IL-5 family cytokines.

    PubMed

    Esnault, Stephane; Kelly, Elizabeth A; Johansson, Mats W; Liu, Lin Ying; Han, Shih-Tsung; Akhtar, Moneeb; Sandbo, Nathan; Mosher, Deane F; Denlinger, Loren C; Mathur, Sameer K; Malter, James S; Jarjour, Nizar N

    2014-01-01

    Semaphorin 7A (sema7a) plays a major role in TGF-β1-induced lung fibrosis. Based on the accumulating evidence that eosinophils contribute to fibrosis/remodeling in the airway, we hypothesized that airway eosinophils may be a significant source of sema7a. In vivo, sema7a was expressed on the surface of circulating eosinophils and upregulated on bronchoalveolar lavage eosinophils obtained after segmental bronchoprovocation with allergen. Based on mRNA levels in unfractionated and isolated bronchoalveolar cells, eosinophils are the predominant source of sema7a. In vitro, among the members of the IL-5-family cytokines, sema7a protein on the surface of blood eosinophils was increased more by IL-3 than by GM-CSF or IL-5. Cytokine-induced expression of cell surface sema7a required translation of newly synthesized protein. Finally, a recombinant sema7a induced alpha-smooth muscle actin production in human bronchial fibroblasts. semaphorin 7A is a potentially important modulator of eosinophil profibrotic functions in the airway remodeling of patients with chronic asthma.

  12. Vitronectin Expression in the Airways of Subjects with Asthma and Chronic Obstructive Pulmonary Disease

    PubMed Central

    Salazar-Peláez, Lina M.; Abraham, Thomas; Herrera, Ana M.; Correa, Mario A.; Ortega, Jorge E.; Paré, Peter D.; Seow, Chun Y.

    2015-01-01

    Vitronectin, a multifunctional glycoprotein, is involved in coagulation, inhibition of the formation of the membrane attack complex (MAC), cell adhesion and migration, wound healing, and tissue remodeling. The primary cellular source of vitronectin is hepatocytes; it is not known whether resident cells of airways produce vitronectin, even though the glycoprotein has been found in exhaled breath condensate and bronchoalveolar lavage from healthy subjects and patients with interstitial lung disease. It is also not known whether vitronectin expression is altered in subjects with asthma and COPD. In this study, bronchial tissue from 7 asthmatic, 10 COPD and 14 control subjects was obtained at autopsy and analyzed by immunohistochemistry to determine the percent area of submucosal glands occupied by vitronectin. In a separate set of experiments, quantitative colocalization analysis was performed on tracheobronchial tissue sections obtained from donor lungs (6 asthmatics, 4 COPD and 7 controls). Vitronectin RNA and protein expressions in bronchial surface epithelium were examined in 12 subjects who undertook diagnostic bronchoscopy. Vitronectin was found in the tracheobronchial epithelium from asthmatic, COPD, and control subjects, although its expression was significantly lower in the asthmatic group. Colocalization analysis of 3D confocal images indicates that vitronectin is expressed in the glandular serous epithelial cells and in respiratory surface epithelial cells other than goblet cells. Expression of the 65-kDa vitronectin isoform was lower in bronchial surface epithelium from the diseased subjects. The cause for the decreased vitronectin expression in asthma is not clear, however, the reduced concentration of vitronectin in the epithelial/submucosal layer of airways may be linked to airway remodeling. PMID:25768308

  13. Asthmatic airway epithelial cells differentially regulate fibroblast expression of extracellular matrix components

    PubMed Central

    Reeves, Stephen R.; Kolstad, Tessa; Lien, Tin-Yu; Elliott, Molly; Ziegler, Steven F.; Wight, Thomas N.; Debley, Jason S.

    2014-01-01

    Background Airway remodeling may explain lung function decline among asthmatic children. Extracellular matrix (ECM) deposition by human lung fibroblasts (HLFs) is implicated in airway remodeling. Airway epithelial cell (AEC) signaling may regulate HLF ECM expression. Objectives Determine whether AECs from asthmatic children differentially regulate HLF expression of ECM constituents. Methods Primary AECs were obtained from well-characterized atopic-asthmatic (N=10) and healthy children (N=10) intubated under anesthesia for an elective surgical procedure. AECs were differentiated at an air-liquid interface (ALI) for 3 weeks, then co-cultured with HLFs from a healthy child for 96 hours. Collagen I (COL1A1), collagen III (COL3A1), hyaluronan synthase 2 (HAS2), and fibronectin (FNDC) expression by HLFs and prostaglandin E2 synthase (PGE2S) expression by AECs was assessed by RT-PCR. TGFb1&2 concentrations in media were measured by ELISA. Results COL1A1 and COL3A1 expression by HLFs co-cultured with asthmatic AECs was greater than HLFs co-cultured with healthy AECs (2.2 fold, p<0.02; 10.8 fold, p<0.02). HAS2 expression by HLFs co-cultured with asthmatic AECs was 2.5-fold higher than by HLFs co-cultured with healthy AECs (p<0.002). FNDC expression by HLFs co-cultured with asthmatic AECs was significantly greater than by HLFs alone. TGFb2 activity was elevated in asthmatic AEC-HLF co-cultures (p<0.05) while PGES2 was down regulated in AEC-HLF co-cultures (2.2 fold, p<0.006). Conclusions HLFs co-cultured with asthmatic AECs showed differential expression of ECM constituents COL1A1 & COL3A1, and HAS2 compared to HLFs co-cultured with healthy AECs. These findings support a role for altered ECM production in asthmatic airway remodeling, possibly regulated by unbalanced AEC signaling. PMID:24875618

  14. Patterns of airway inflammation and MMP-12 expression in smokers and ex-smokers with COPD

    PubMed Central

    Babusyte, Agne; Stravinskaite, Kristina; Jeroch, Jolanta; Lötvall, Jan; Sakalauskas, Raimundas; Sitkauskiene, Brigita

    2007-01-01

    Background Smoking activates and recruits inflammatory cells and proteases to the airways. Matrix metalloproteinase (MMP)-12 may be a key mediator in smoke induced emphysema. However, the influence of smoking and its cessation on airway inflammation and MMP-12 expression during COPD is still unknown. We aimed to analyse airway inflammatory cell patterns in induced sputum (IS) and bronchoalveolar lavage (BAL) from COPD patients who are active smokers and who have ceased smoking >2 years ago. Methods 39 COPD outpatients – smokers (n = 22) and ex-smokers (n = 17) were studied. 8 'healthy' smokers and 11 healthy never-smokers were tested as the control groups. IS and BAL samples were obtained for differential and MMP-12+-macrophages count analysis. Results The number of IS neutrophils was higher in both COPD groups compared to both controls. The amount of BAL neutrophils was higher in COPD smokers compared to healthy never-smokers. The number of BAL MMP-12+-macrophages was higher in COPD smokers (1.6 ± 0.3 × 106/ml) compared to COPD ex-smokers, 'healthy' smokers and healthy never-smokers (0.9 ± 0.4, 0.4 ± 0.2, 0.2 ± 0.1 × 106/ml respectively, p < 0.05). Conclusion The lower amount of BAL neutrophils in COPD ex-smokers, compared to COPD smokers, suggests positive alterations in alveolar compartment after smoking cessation. Smoking and disease itself may stimulate MMP-12 expression in airway compartments (IS and BAL) from COPD patients. PMID:18001475

  15. Nasal administration of interleukin-33 induces airways angiogenesis and expression of multiple angiogenic factors in a murine asthma surrogate.

    PubMed

    Shan, Shan; Li, Yan; Wang, Jingjing; Lv, Zhe; Yi, Dawei; Huang, Qiong; Corrigan, Chris J; Wang, Wei; Quangeng, Zhang; Ying, Sun

    2016-05-01

    The T-helper cell type 2-promoting cytokine interleukin-33 (IL-33) has been implicated in asthma pathogenesis. Angiogenesis is a feature of airways remodelling in asthma. We hypothesized that IL-33 induces airways angiogenesis and expression of angiogenic factors in an established murine surrogate of asthma. In the present study, BALB/c mice were subjected to serial intranasal challenge with IL-33 alone for up to 70 days. In parallel, ovalbumin (OVA) -sensitized mice were subjected to serial intranasal challenge with OVA or normal saline to serve as positive and negative controls, respectively. Immunohistochemical analysis of expression of von Willebrand factor and erythroblast transformation-specific-related gene, both blood vessel markers, and angiogenic factors angiogenin, insulin-like growth factor-1, endothelin-1, epidermal growth factor and amphiregulin was performed in lung sections ex vivo. An established in-house assay was used to test whether IL-33 was able to induce microvessel formation by human vascular endothelial cells. Results showed that serial intranasal challenge of mice with IL-33 or OVA resulted in proliferation of peribronchial von Willebrand factor-positive blood vessels to a degree closely related to the total expression of the angiogenic factors amphiregulin, angiogenin, endothelin-1, epidermal growth factor and insulin-like growth factor-1. IL-33 also induced microvessel formation by human endothelial cells in a concentration-dependent fashion in vitro. Our data are consistent with the hypothesis that IL-33 has the capacity to induce angiogenesis at least partly by increasing local expression of multiple angiogenic factors in an allergen-independent murine asthma surrogate, and consequently that IL-33 or its receptor is a potential novel molecular target for asthma therapy.

  16. Use of Phage Display To Identify Potential Pseudomonas aeruginosa Gene Products Relevant to Early Cystic Fibrosis Airway Infections

    PubMed Central

    Beckmann, Christiane; Brittnacher, Mitchell; Ernst, Robert; Mayer-Hamblett, Nicole; Miller, Samuel I.; Burns, Jane L.

    2005-01-01

    Pseudomonas aeruginosa airway infections are a major cause of morbidity and mortality in patients with cystic fibrosis. Treatment of established infections is difficult, even with microbiologically active agents. Thus, prevention of infection is an important goal of management. Isolates from cystic fibrosis patients appear to originate from the environment but adapt to the milieu of the airway of the cystic fibrosis patient and evolve toward a common phenotype. Identification of the antigens expressed early in infection may lead to novel targets for vaccine development. Immunogenic peptides were identified in a J404 random nonapeptide phage display library with serum from cystic fibrosis patients obtained within the first year of P. aeruginosa infection. One hundred sixty-five reactive clones were verified by plaque lift assays, and their inserts were sequenced. The sequenced nonapeptides were compared with the published sequence of strain PAO1, identifying homologies to 76 genes encoding outer membrane and secreted proteins. The majority of these were proteins involved in small-molecule transport, membrane structural proteins, and secreted factors. An in silico analysis was performed that suggested that the occurrence of multiple matches to predominantly outer membrane and secreted proteins was not attributable to random chance. Finally, gene expression array data from early isolates of P. aeruginosa from cystic fibrosis patients was compared with the results from phage display analysis. Eleven outer membrane and secreted proteins were common between the two data sets. These included genes involved in iron acquisition, antibiotic efflux, fimbrial biogenesis, and pyocin synthesis. These results demonstrate the feasibility and validity of this novel approach and suggest potential targets for future development. PMID:15618183

  17. Effect of stress on eotaxin and expression of adhesion molecules in a murine model of allergic airway inflammation.

    PubMed

    Joachim, Ricarda A; Sagach, Viktoriya; Quarcoo, David; Dinh, Q Thai; Arck, Petra C; Klapp, Burghard F

    2007-01-01

    Recently we have shown that sound stress enhances allergic airway inflammation in a combined murine model. In the current study we investigated mediating factors and early kinetics of stress exacerbated allergic airway inflammation. Stress significantly increased allergen induced airway inflammation as identified by leukocyte numbers in BAL fluids. Eotaxin levels from stressed mice were significantly higher 24 h after stress. No differences were found for vascular or cellular adhesion molecule expression or cytokine levels. Our data indicate that the effect of stress on allergic airway inflammation might be mediated by the chemoattractant eotaxin, while Th2 cytokines and expression of adhesion molecules seem not to be differently regulated in stressed and non-stressed mice.

  18. Novel expression of a functional glycine receptor chloride channel that attenuates contraction in airway smooth muscle

    PubMed Central

    Yim, Peter D.; Gallos, George; Xu, Dingbang; Zhang, Yi; Emala, Charles W.

    2011-01-01

    Airway smooth muscle (ASM) contraction is an important component of the pathophysiology of asthma. Taurine, an agonist of glycine receptor chloride (GlyR Cl−) channels, was found to relax contracted ASM, which led us to question whether functional GlyR Cl− channels are expressed in ASM. Messenger RNA for β (GLRB), α1 (GLRA1), α2 (GLRA2), and α4 (GLRA4) subunits were found in human (Homo sapiens) and guinea pig (Cavia porcellus) tracheal smooth muscle. Immunoblotting confirmed the protein expression of GLRA1 and GLRB subunits in ASM. Electrical activity of cultured human ASM cells was assessed using a fluorescent potentiometric dye and electrophysiological recordings. Glycine increased current and significantly increased fluorescence in a dose-dependent manner. The GlyR Cl− channel antagonist strychnine significantly blocked the effects of glycine on potentiometric fluorescence in ASM cells. Guinea pig airway ring relaxation of ACh-induced contractions by isoproterenol was significantly left-shifted in the presence of glycine. This effect of glycine was blocked by pretreatment with the GlyR Cl− channel antagonist strychnine. Glycine treatment during tachykinin- and acetylcholine-induced contractions significantly decreased the maintenance of muscle force compared to control. GlyR Cl− channels are expressed on ASM and regulate smooth muscle force and offer a novel target for therapeutic relaxation of ASM.—Yim, P. D., Gallos, G., Xu, D., Zhang, Y., Emala, C. W. Novel expression of a functional glycine receptor chloride channel that attenuates contraction in airway smooth muscle. PMID:21282206

  19. Method of controlling gene expression

    DOEpatents

    Peters, Norman K.; Frost, John W.; Long, Sharon R.

    1991-12-03

    A method of controlling expression of a DNA segment under the control of a nod gene promoter which comprises administering to a host containing a nod gene promoter an amount sufficient to control expression of the DNA segment of a compound of the formula: ##STR1## in which each R is independently H or OH, is described.

  20. The flow of gene expression.

    PubMed

    Misteli, Tom

    2004-03-01

    Gene expression is a highly interconnected multistep process. A recent meeting in Iguazu Falls, Argentina, highlighted the need to uncover both the molecular details of each single step as well as the mechanisms of coordination among processes in order to fully understand the expression of genes.

  1. Discovering modulators of gene expression

    PubMed Central

    Babur, Özgün; Demir, Emek; Gönen, Mithat; Sander, Chris; Dogrusoz, Ugur

    2010-01-01

    Proteins that modulate the activity of transcription factors, often called modulators, play a critical role in creating tissue- and context-specific gene expression responses to the signals cells receive. GEM (Gene Expression Modulation) is a probabilistic framework that predicts modulators, their affected targets and mode of action by combining gene expression profiles, protein–protein interactions and transcription factor–target relationships. Using GEM, we correctly predicted a significant number of androgen receptor modulators and observed that most modulators can both act as co-activators and co-repressors for different target genes. PMID:20466809

  2. The Expression of NOX4 in Smooth Muscles of Small Airway Correlates with the Disease Severity of COPD

    PubMed Central

    2016-01-01

    Airway smooth muscle (ASM) remodeling is a hallmark in chronic obstructive pulmonary disease (COPD), and nicotinamide-adenine dinucleotide phosphate (NADPH) oxidases (NOXs) produced reactive oxygen species (ROS) play a crucial role in COPD pathogenesis. In the present study, the expression of NOX4 and its correlation with the ASM hypertrophy/hyperplasia, clinical pulmonary functions, and the expression of transforming growth factor β (TGF-β) in the ASM of COPD small airways were investigated by semiquantitative morphological and/or immunohistochemistry staining methods. The results showed that an elevated expression of NOX4 and TGF-β, along with an increased volume of ASM mass, was found in the ASM of small airways in COPD patients. The abundance of NOX4 protein in the ASM was increased with disease severity and inversely correlated with the pulmonary functions in COPD patients. In addition, the expression of NOX4 and ASM marker α-SMA was colocalized, and the increased NOX4 expression was found to accompany an upregulated expression of TGF-β in the ASM of small airways of COPD lung. These results indicate that NOX4 may be a key regulator in ASM remodeling of small airway, in part through a mechanism interacting with TGF-β signaling in the pathogenesis of COPD, which warrants further investigation. PMID:27656649

  3. [Prospects of gene therapy in mucoviscidosis using viral infection of the airway epithelium].

    PubMed

    Bayle, J Y; Boucher, R C

    1994-01-01

    Mucoviscidosis is the most common severe inherited autosomal recessive disease. Since the gene has been recognised (cystic fibrosis transmembrane conductance regulator gene) (CFTR) the technique of genetic transfer has been applied to the airway epithelium. The prospect for gene therapy to treat the consequences of bronchopulmonary mucoviscidosis is now evident. The in vitro introduction of the normal CFTR human gene in epithelial cells has been obtained using recombinant retrovirus, adenovirus and parvovirus rendered defective for replication. The abnormal bioelectric phenotype of the cells from patients with mucoviscidosis has been corrected. Of these, only adenovirus and parvovirus have been capable of assuring effective genetic transfer by direct introduction into the airways. This data has been considered sufficient to justify starting clinical trials in man with adenovirus; the preliminary results confirm the possibility of correcting the chloride transport. Nevertheless the observation of an immune response and secondary inflammation raises ethical questions relative to the safety of such trials. This observation justifies research into an alternative non-viral technique such as employing liposomes. The authors have made a review of the data which may be established as a basis for genetic therapy for mucoviscidosis.

  4. IL-10 is necessary for the expression of airway hyperresponsiveness but not pulmonary inflammation after allergic sensitization

    NASA Astrophysics Data System (ADS)

    Mäkelä, M. J.; Kanehiro, A.; Borish, L.; Dakhama, A.; Loader, J.; Joetham, A.; Xing, Z.; Jordana, M.; Larsen, G. L.; Gelfand, E. W.

    2000-05-01

    Cytokines play an important role in modulating inflammatory responses and, as a result, airway tone. IL-10 is a regulatory cytokine that has been suggested for treatment of asthma because of its immunosuppressive and anti-inflammatory properties. In contrast to these suggestions, we demonstrate in a model of allergic sensitization that mice deficient in IL-10 (IL-10/) develop a pulmonary inflammatory response but fail to exhibit airway hyperresponsiveness in both in vitro and in vivo assessments of lung function. Reconstitution of these deficient mice with the IL-10 gene fully restores development of airway hyperresponsiveness comparable to control mice. These results identify an important role of IL-10, downstream of the inflammatory cascade, in regulating the tone of the airways after allergic sensitization and challenge.

  5. Human Lacrimal Gland Gene Expression

    PubMed Central

    Aakalu, Vinay Kumar; Parameswaran, Sowmya; Maienschein-Cline, Mark; Bahroos, Neil; Shah, Dhara; Ali, Marwan; Krishnakumar, Subramanian

    2017-01-01

    Background The study of human lacrimal gland biology and development is limited. Lacrimal gland tissue is damaged or poorly functional in a number of disease states including dry eye disease. Development of cell based therapies for lacrimal gland diseases requires a better understanding of the gene expression and signaling pathways in lacrimal gland. Differential gene expression analysis between lacrimal gland and other embryologically similar tissues may be helpful in furthering our understanding of lacrimal gland development. Methods We performed global gene expression analysis of human lacrimal gland tissue using Affymetrix ® gene expression arrays. Primary data from our laboratory was compared with datasets available in the NLM GEO database for other surface ectodermal tissues including salivary gland, skin, conjunctiva and corneal epithelium. Results The analysis revealed statistically significant difference in the gene expression of lacrimal gland tissue compared to other ectodermal tissues. The lacrimal gland specific, cell surface secretory protein encoding genes and critical signaling pathways which distinguish lacrimal gland from other ectodermal tissues are described. Conclusions Differential gene expression in human lacrimal gland compared with other ectodermal tissue types revealed interesting patterns which may serve as the basis for future studies in directed differentiation among other areas. PMID:28081151

  6. Monoallelic Gene Expression in Mammals.

    PubMed

    Chess, Andrew

    2016-11-23

    Monoallelic expression not due to cis-regulatory sequence polymorphism poses an intriguing problem in epigenetics because it requires the unequal treatment of two segments of DNA that are present in the same nucleus and that can indeed have absolutely identical sequences. Here, I focus on a few recent developments in the field of monoallelic expression that are of particular interest and raise interesting questions for future work. One development is regarding analyses of imprinted genes, in which recent work suggests the possibility that intriguing networks of imprinted genes exist and are important for genetic and physiological studies. Another issue that has been raised in recent years by a number of publications is the question of how skewed allelic expression should be for it to be designated as monoallelic expression and, further, what methods are appropriate or inappropriate for analyzing genomic data to examine allele-specific expression. Perhaps the most exciting recent development in mammalian monoallelic expression is a clever and carefully executed analysis of genetic diversity of autosomal genes subject to random monoallelic expression (RMAE), which provides compelling evidence for distinct evolutionary forces acting on random monoallelically expressed genes.

  7. Tuning noise in gene expression.

    PubMed

    Tyagi, Sanjay

    2015-05-05

    The relative contribution of promoter architecture and the associated chromatin environment in regulating gene expression noise has remained elusive. In their recent work, Arkin, Schaffer and colleagues (Dey et al, 2015) show that mean expression and noise for a given promoter at different genomic loci are uncorrelated and influenced by the local chromatin environment.

  8. Smooth muscle actin and myosin expression in cultured airway smooth muscle cells.

    PubMed

    Wong, J Z; Woodcock-Mitchell, J; Mitchell, J; Rippetoe, P; White, S; Absher, M; Baldor, L; Evans, J; McHugh, K M; Low, R B

    1998-05-01

    In this study, the expression of smooth muscle actin and myosin was examined in cultures of rat tracheal smooth muscle cells. Protein and mRNA analyses demonstrated that these cells express alpha- and gamma-smooth muscle actin and smooth muscle myosin and nonmuscle myosin-B heavy chains. The expression of the smooth muscle specific actin and myosin isoforms was regulated in the same direction when growth conditions were changed. Thus, at confluency in 1 or 10% serum-containing medium as well as for low-density cells (50-60% confluent) deprived of serum, the expression of the smooth muscle forms of actin and myosin was relatively high. Conversely, in rapidly proliferating cultures at low density in 10% serum, smooth muscle contractile protein expression was low. The expression of nonmuscle myosin-B mRNA and protein was more stable and was upregulated only to a small degree in growing cells. Our results provide new insight into the molecular basis of differentiation and contractile function in airway smooth muscle cells.

  9. NEU1 sialidase expressed in human airway epithelia regulates epidermal growth factor receptor (EGFR) and MUC1 protein signaling.

    PubMed

    Lillehoj, Erik P; Hyun, Sang Won; Feng, Chiguang; Zhang, Lei; Liu, Anguo; Guang, Wei; Nguyen, Chinh; Luzina, Irina G; Atamas, Sergei P; Passaniti, Antonino; Twaddell, William S; Puché, Adam C; Wang, Lai-Xi; Cross, Alan S; Goldblum, Simeon E

    2012-03-09

    Epithelial cells (ECs) lining the airways provide a protective barrier between the external environment and the internal host milieu. These same airway epithelia express receptors that respond to danger signals and initiate repair programs. Because the sialylation state of a receptor can influence its function and is dictated in part by sialidase activity, we asked whether airway epithelia express catalytically active sialidase(s). Human primary small airway and A549 ECs expressed NEU1 sialidase at the mRNA and protein levels, and NEU1 accounted for >70% of EC sialidase activity. Blotting with Maackia amurensis and peanut agglutinin lectins established epidermal growth factor receptor (EGFR) and MUC1 as in vivo substrates for NEU1. NEU1 associated with EGFR and MUC1, and NEU1-EGFR association was regulated by EGF stimulation. NEU1 overexpression diminished EGF-stimulated EGFR Tyr-1068 autophosphorylation by up to 44% but enhanced MUC1-dependent Pseudomonas aeruginosa adhesion by 1.6-1.7-fold and flagellin-stimulated ERK1/2 activation by 1.7-1.9-fold. In contrast, NEU1 depletion increased EGFR activation (1.5-fold) and diminished MUC1-mediated bacterial adhesion (38-56%) and signaling (73%). These data indicate for the first time that human airway epithelia express catalytically active NEU1 sialidase that regulates EGFR- and MUC1-dependent signaling and bacterial adhesion. NEU1 catalytic activity may offer an additional level of regulation over the airway epithelial response to ligands, pathogens, and injurious stimuli.

  10. Cigarette smoke alters IL-33 expression and release in airway epithelial cells.

    PubMed

    Pace, Elisabetta; Di Sano, Caterina; Sciarrino, Serafina; Scafidi, Valeria; Ferraro, Maria; Chiappara, Giuseppina; Siena, Liboria; Gangemi, Sebastiano; Vitulo, Patrizio; Giarratano, Antonino; Gjomarkaj, Mark

    2014-09-01

    Airway epithelium is a regulator of innate immune responses to a variety of insults including cigarette smoke. Cigarette smoke alters the expression and the activation of Toll Like Receptor 4 (TLR4), an innate immunity receptor. IL-33, an alarmin, increases innate immunity Th2 responses. The aims of this study were to explore whether mini-bronchoalveolar lavage (mini-BAL) or sera from smokers have altered concentrations of IL-33 and whether cigarette smoke extracts (CSE) alter both intracellular expression (mRNA and protein) and release of IL-33 in bronchial epithelial cells. The role of TLR4 in the expression of IL-33 was also explored. Mini-BALs, but not sera, from smokers show reduced concentrations of IL-33. The expression of IL-33 was increased also in bronchial epithelium from smokers. 20% CSE reduced IL-33 release but increased the mRNA for IL-33 by real time PCR and the intracellular expression of IL-33 in bronchial epithelial cells as confirmed by flow cytometry, immunocytochemistry and western blot analysis. The effect of CSE on IL-33 expression was also observed in primary bronchial epithelial cells. IL-33 expression was mainly concentrated within the cytoplasm of the cells. LPS, an agonist of TLR4, reduced IL-33 expression, and an inhibitor of TLR4 increased the intracellular expression of IL-33. In conclusion, the release of IL-33 is tightly controlled and, in smokers, an altered activation of TLR4 may lead to an increased intracellular expression of IL-33 with a limited IL-33 release.

  11. Downregulation of Orai1 expression in the airway alleviates murine allergic rhinitis

    PubMed Central

    Wang, Yi; Lin, Lin

    2012-01-01

    Orai1 is the key subunit of the Ca2+-release-activated Ca2+ channel. Our previous report has demonstrated that Orai1 expression in the airway was upregulated in the ovalbumin (OVA)-induced allergic rhinitis (AR) mouse models. To observe whether inhibition of Orai1 expression in the airway could suppress symptoms in a murine model of AR and to assess the impacts of this inhibition on the responses of local and systemic immunocytes, we administered recombinant lentivirus vectors that encoded shRNA against ORAI1 (lenti-ORAI1) into the nostrils of OVA-sensitized mice before the challenges, and analyzed its effect on allergic responses, as compared with the unsensitized mice and untreated AR mice. Administration of lenti-ORAI1 into the nasal cavity successfully infected cells in the epithelial layer of the nasal mucosa, and significantly decreased the frequencies of sneezing and nasal rubbing of the mice. Protein levels of leukotriene C4, OVA-specific IgE, and IL-4 in the nasal lavage fluid and serum and eosinophil cation protein in the serum were also significantly reduced by lenti-ORAI1, as were the mRNA levels of these factors in the nasal mucosa and spleen. These data suggested that administration of lenti-ORAI1 into the nasal cavity effectively decreased Orai1 expression in the nasal mucosa, alleviated AR symptoms, and partially inhibited the hyperresponsiveness of the local and systemic immune cells including T cells, B cells, mast cells and eosinophils that are involved in the pathogenesis of AR. PMID:22170034

  12. Elevated expression of placental growth factor is associated with airway-wall vascular remodelling and thickening in smokers with asthma

    PubMed Central

    Wu, Dong; Lai, Tianwen; Yuan, Yalian; Chen, Min; Xia, Jun; Li, Wen; Pan, Guihai; Yuan, Binfan; Lv, Quanchao; Li, Yanyu; Li, Dongmin; Wu, Bin

    2017-01-01

    The increased expression of placental growth factor (PlGF) in chronic obstructive pulmonary disease and allergy-related asthma suggests its role in the pathogenesis of these diseases. In asthmatic smokers, airway remodelling is accompanied by an accelerated decline in lung function. However, whether PlGF contributes to the persistent airflow obstruction and vascular remodelling typically seen in asthmatic smokers is unknown. In this study we measured lung function, airway-wall thickening, and PlGF levels in serum and induced sputum in 74 asthmatic and 42 healthy smokers and never-smokers. Using human lung microvascular endothelial cells (HLMECs), we evaluated the in vitro effects of PlGF on each step of vascular remodelling, including proliferation, migration, stress-fibre expression, and tubule formation. Our data showed significantly higher serum and sputum PlGF levels in asthma patients, especially asthmatic smokers, than in healthy controls. Serum and sputum PlGF levels correlated negatively with post-bronchodilator forced expiratory volume in 1 s (FEV1) and the FEV1/forced vital capacity, but positively with airway-wall thickening. Stimulation of HLMECs with rhPlGF promoted all of the steps of airway-microvascular remodelling. These findings provide insights into the influence of cigarette smoking on the structural changes in the airways of asthmatics and the important pathogenic role played by PlGF. PMID:28220848

  13. Persistent gene expression in mouse nasal epithelia following feline immunodeficiency virus-based vector gene transfer.

    PubMed

    Sinn, Patrick L; Burnight, Erin R; Hickey, Melissa A; Blissard, Gary W; McCray, Paul B

    2005-10-01

    Gene transfer development for treatment or prevention of cystic fibrosis lung disease has been limited by the inability of vectors to efficiently and persistently transduce airway epithelia. Influenza A is an enveloped virus with natural lung tropism; however, pseudotyping feline immunodeficiency virus (FIV)-based lentiviral vector with the hemagglutinin envelope protein proved unsuccessful. Conversely, pseudotyping FIV with the envelope protein from influenza D (Thogoto virus GP75) resulted in titers of 10(6) transducing units (TU)/ml and conferred apical entry into well-differentiated human airway epithelial cells. Baculovirus GP64 envelope glycoproteins share sequence identity with influenza D GP75 envelope glycoproteins. Pseudotyping FIV with GP64 from three species of baculovirus resulted in titers of 10(7) to 10(9) TU/ml. Of note, GP64 from Autographa californica multicapsid nucleopolyhedrovirus resulted in high-titer FIV preparations (approximately 10(9) TU/ml) and conferred apical entry into polarized primary cultures of human airway epithelia. Using a luciferase reporter gene and bioluminescence imaging, we observed persistent gene expression from in vivo gene transfer in the mouse nose with A. californica GP64-pseudotyped FIV (AcGP64-FIV). Longitudinal bioluminescence analysis documented persistent expression in nasal epithelia for approximately 1 year without significant decline. According to histological analysis using a LacZ reporter gene, olfactory and respiratory epithelial cells were transduced. In addition, methylcellulose-formulated AcGP64-FIV transduced mouse nasal epithelia with much greater efficiency than similarly formulated vesicular stomatitis virus glycoprotein-pseudotyped FIV. These data suggest that AcGP64-FIV efficiently transduces and persistently expresses a transgene in nasal epithelia in the absence of agents that disrupt the cellular tight junction integrity.

  14. EFFECT OF INHALED ENDOTOXIN ON AIRWAY AND CIRCULATING INFLAMMATORY CELL PHAGOCYTOSIS AND CD11B EXPRESSION IN ATOPIC ASTHMATIC SUBJECTS

    EPA Science Inventory

    Effect of inhaled endotoxin on airway and circulating inflammatory cell phagocytosis and CD11b expression in atopic asthmatic subjects

    Neil E. Alexis, PhD, Marlowe W. Eldridge, MD, David B. Peden, MD, MS

    Chapel Hill and Research Triangle Park, NC

    Backgrou...

  15. Differential Gene Expression in Glaucoma

    PubMed Central

    Jakobs, Tatjana C.

    2014-01-01

    In glaucoma, regardless of its etiology, retinal ganglion cells degenerate and eventually die. Although age and elevated intraocular pressure (IOP) are the main risk factors, there are still many mysteries in the pathogenesis of glaucoma. The advent of genome-wide microarray expression screening together with the availability of animal models of the disease has allowed analysis of differential gene expression in all parts of the eye in glaucoma. This review will outline the findings of recent genome-wide expression studies and discuss their commonalities and differences. A common finding was the differential regulation of genes involved in inflammation and immunity, including the complement system and the cytokines transforming growth factor β (TGFβ) and tumor necrosis factor α (TNFα). Other genes of interest have roles in the extracellular matrix, cell–matrix interactions and adhesion, the cell cycle, and the endothelin system. PMID:24985133

  16. Differential gene expression in glaucoma.

    PubMed

    Jakobs, Tatjana C

    2014-07-01

    In glaucoma, regardless of its etiology, retinal ganglion cells degenerate and eventually die. Although age and elevated intraocular pressure (IOP) are the main risk factors, there are still many mysteries in the pathogenesis of glaucoma. The advent of genome-wide microarray expression screening together with the availability of animal models of the disease has allowed analysis of differential gene expression in all parts of the eye in glaucoma. This review will outline the findings of recent genome-wide expression studies and discuss their commonalities and differences. A common finding was the differential regulation of genes involved in inflammation and immunity, including the complement system and the cytokines transforming growth factor β (TGFβ) and tumor necrosis factor α (TNFα). Other genes of interest have roles in the extracellular matrix, cell-matrix interactions and adhesion, the cell cycle, and the endothelin system.

  17. Cadmium regulates the expression of the CFTR chloride channel in human airway epithelial cells.

    PubMed

    Rennolds, Jessica; Butler, Susie; Maloney, Kevin; Boyaka, Prosper N; Davis, Ian C; Knoell, Daren L; Parinandi, Narasimham L; Cormet-Boyaka, Estelle

    2010-07-01

    Cadmium is a toxic heavy metal ranked seventh on the Priority List of Hazardous Substances. As a byproduct of smelters, cadmium is a prevalent environmental contaminant. It is also a major component of cigarette smoke, and its inhalation is associated with decreased pulmonary function, lung cancer, and chronic obstructive pulmonary disease. Ion channels, including the cystic fibrosis transmembrane conductance regulator (CFTR), play a central role in maintaining fluid homeostasis and lung functions. CFTR is mostly expressed in epithelial cells, and little is known about the effect of cadmium exposure on lung epithelial cell function. We show that exposure to cadmium decreases the expression of the CFTR protein and subsequent chloride transport in human airway epithelial cells in vitro. Impairment of CFTR protein expression was also observed in vivo in the lung of mice after intranasal instillation of cadmium. We established that the inhibitory effect of cadmium was not a nonspecific effect of heavy metals, as nickel had no effect on CFTR protein levels. Finally, we show that selected antioxidants, including alpha-tocopherol (vitamin E), but not N-acetylcysteine, can prevent the cadmium-induced suppression of CFTR. In summary, we have identified cadmium as a regulator of the CFTR chloride channel present in lung epithelial cells. Future strategies to prevent the deleterious effect of cadmium on epithelial cells and lung functions may benefit from the finding that alpha-tocopherol protects CFTR expression and function.

  18. Transgenic Arabidopsis Gene Expression System

    NASA Technical Reports Server (NTRS)

    Ferl, Robert; Paul, Anna-Lisa

    2009-01-01

    The Transgenic Arabidopsis Gene Expression System (TAGES) investigation is one in a pair of investigations that use the Advanced Biological Research System (ABRS) facility. TAGES uses Arabidopsis thaliana, thale cress, with sensor promoter-reporter gene constructs that render the plants as biomonitors (an organism used to determine the quality of the surrounding environment) of their environment using real-time nondestructive Green Fluorescent Protein (GFP) imagery and traditional postflight analyses.

  19. Zipf's Law in Gene Expression

    NASA Astrophysics Data System (ADS)

    Furusawa, Chikara; Kaneko, Kunihiko

    2003-02-01

    Using data from gene expression databases on various organisms and tissues, including yeast, nematodes, human normal and cancer tissues, and embryonic stem cells, we found that the abundances of expressed genes exhibit a power-law distribution with an exponent close to -1; i.e., they obey Zipf’s law. Furthermore, by simulations of a simple model with an intracellular reaction network, we found that Zipf’s law of chemical abundance is a universal feature of cells where such a network optimizes the efficiency and faithfulness of self-reproduction. These findings provide novel insights into the nature of the organization of reaction dynamics in living cells.

  20. Neighboring Genes Show Correlated Evolution in Gene Expression.

    PubMed

    Ghanbarian, Avazeh T; Hurst, Laurence D

    2015-07-01

    When considering the evolution of a gene's expression profile, we commonly assume that this is unaffected by its genomic neighborhood. This is, however, in contrast to what we know about the lack of autonomy between neighboring genes in gene expression profiles in extant taxa. Indeed, in all eukaryotic genomes genes of similar expression-profile tend to cluster, reflecting chromatin level dynamics. Does it follow that if a gene increases expression in a particular lineage then the genomic neighbors will also increase in their expression or is gene expression evolution autonomous? To address this here we consider evolution of human gene expression since the human-chimp common ancestor, allowing for both variation in estimation of current expression level and error in Bayesian estimation of the ancestral state. We find that in all tissues and both sexes, the change in gene expression of a focal gene on average predicts the change in gene expression of neighbors. The effect is highly pronounced in the immediate vicinity (<100 kb) but extends much further. Sex-specific expression change is also genomically clustered. As genes increasing their expression in humans tend to avoid nuclear lamina domains and be enriched for the gene activator 5-hydroxymethylcytosine, we conclude that, most probably owing to chromatin level control of gene expression, a change in gene expression of one gene likely affects the expression evolution of neighbors, what we term expression piggybacking, an analog of hitchhiking.

  1. Expression of a TGF-{beta} regulated cyclin-dependent kinase inhibitor in normal and immortalized airway epithelial cells

    SciTech Connect

    Tierney, L.A.; Bloomfield, C.; Johnson, N.F.

    1995-12-01

    Tumors arising from epithelial cells, including lung cancers are frequently resistant to factors that regulate growth and differentiation in normal in normal cells. Once such factor is transforming growth factor-{Beta} (TGF-{Beta}). Escape from the growth-inhibitory effects of TGF-{Beta} is thought to be a key step in the transformation of airway epithelial cells. most lung cancer cell lines require serum for growth. In contrast, normal human bronchial epithelial (NHBE) cells are exquisitely sensitive to growth-inhibitory and differentiating effects of TGF-{Beta}. The recent identification of a novel cyclin-dependent kinase inhibitor, p15{sup INK4B}, which is regulated by TGF-{Beta}, suggests a mechanism by which TGF-{Beta} mediates growth arrest in NHBE cells. The purpose of this study was two-fold: (1) to determine if p15{sup INK4B} is induced by TGF-{Beta} in NHBE cells or immortalized bronchial epithelial (R.1) cells and if that induction corresponds to a G1/S cell-cycle arrest; (2) to determine the temporal relationship between p15{sup INK4B} induction, cell-cycle arrest, and the phosphorylation state of the pRB because it is thought that p15{sup INK4B} acts indirectly by preventing phosphorylation of the RB gene product. In this study, expression of p15{sup INK4B} was examined in NHBE cells and R.1 cells at different time intervals following TGF-{Beta} treatment. The expression of this kinase inhibitor and its relationship to the cell and the pRb phosphorylation state were examined in cells that were both sensitive (NHBE) and resistant (R.1) to the effects of TGF-{Beta}. These results suggest that the cyclin-dependent kinase inhibitor, p15{sup INK4B}, is involved in airway epithelial cell differentiation and that loss or reduction of expression plays a role in the resistance of transformed or neoplastic cells to the growth-inhibitory effects of TGF-{Beta}.

  2. Expression of surface platelet receptors (CD62P and CD41/61) in horses with recurrent airway obstruction (RAO).

    PubMed

    Iwaszko-Simonik, Alicja; Niedzwiedz, Artur; Graczyk, Stanislaw; Slowikowska, Malwina; Pliszczak-Krol, Aleksandra

    2015-03-15

    Recurrent airway obstruction (RAO) is an allergic disease of horses similar to human asthma, which is characterized by airway inflammation and activation of neutrophils, lymphocytes and platelets. Platelet activation and an increase in circulating platelet-leukocyte aggregates may lead to airway remodeling. The aim of this study was to investigate platelet status in RAO-affected horses based on the platelet morphology and platelet surface expression of CD41/61 and CD62P. Ten RAO-affected horses and ten healthy horses were included in this study. Blood samples were obtained to determine the platelet count (PLT), mean platelet volume (MPV) and platelet large cell ratio (P-LCR). Expression of CD62P and CD41/61 was detected by flow cytometry on activated platelets. The median PLT was significantly reduced in horses with RAO compared to the controls. The MPV and the P-LCR values were significantly higher in RAO horses than controls. Expression of CD41/61 on platelets was increased in RAO horses, while CD62P expression was reduced. This study demonstrated the morphological changes in platelets and expression of platelet surface receptors. Despite the decrease of CD62P expression, the observed increased surface expression of CD41/61 on platelets in horses with RAO may contribute to the formation of platelet aggregates in their respiratory system.

  3. Transcriptional Regionalization of the Fruit Fly’s Airway Epithelium

    PubMed Central

    Faisal, Muhammad N.; Hoffmann, Julia; El-Kholy, Samar; Kallsen, Kimberley; Wagner, Christina; Bruchhaus, Iris; Fink, Christine; Roeder, Thomas

    2014-01-01

    Although airway epithelia are primarily devoted to gas exchange, they have to fulfil a number of different tasks including organ maintenance and the epithelial immune response to fight airborne pathogens. These different tasks are at least partially accomplished by specialized cell types in the epithelium. In addition, a proximal to distal gradient mirroring the transition from airflow conduction to real gas exchange, is also operative. We analysed the airway system of larval Drosophila melanogaster with respect to region-specific expression in the proximal to distal axis. The larval airway system is made of epithelial cells only. We found differential expression between major trunks of the airways and more distal ones comprising primary, secondary and terminal ones. A more detailed analysis was performed using DNA-microarray analysis to identify cohorts of genes that are either predominantly expressed in the dorsal trunks or in the primary/secondary/terminal branches of the airways. Among these differentially expressed genes are especially those involved in signal transduction. Wnt-signalling associated genes for example are predominantly found in secondary/terminal airways. In addition, some G-protein coupled receptors are differentially expressed between both regions of the airways, exemplified by those activated by octopamine or tyramine, the invertebrate counterparts of epinephrine and norepinephrine. Whereas the OAMB is predominantly found in terminal airway regions, the oct3βR has higher expression levels in dorsal trunks. In addition, we observed a significant association of both, genes predominantly expressed in dorsal trunks or in primary to terminal branches branches with those regulated by hypoxia. Taken together, this observed differential expression is indicative for a proximal to distal transcriptional regionalization presumably reflecting functional differences in these parts of the fly’s airway system. PMID:25020150

  4. Toll-like Receptors, Triggering Receptor Expressed on Myeloid Cells Family Members and Receptor for Advanced Glycation End-products in Allergic Airway Inflammation

    PubMed Central

    Hall, Sannette C.; Agrawal, Devendra K.

    2016-01-01

    Asthma is a chronic disorder of the airways characterized by cellular infiltration, airway hyper-responsive and airway inflammation. Innate immune cells are the first line of defense against endogenous and exogenous signals in the airways and as such possess a diverse array of pattern recognition receptors. Toll-like receptors are crucial sentinels which when activated, can either promote or ameliorate the inflammatory response in predisposed individuals. The recently discovered triggering receptor expressed on myeloid cells family members are emerging mediators of inflammation. These receptors are believed to modulate inflammatory responses by collaborating with classic PRRs. Endogenous signals like HMGB-1, signaling through the receptor for advanced glycation end products, also promotes inflammation, however, its contribution to inflammation in the airways is not well known. Here, we discuss the role of each receptor in airway inflammation and highlight potential synergistic mechanisms, which contribute to disease pathogenesis in allergic asthma. PMID:26678062

  5. Airway acidification initiates host defense abnormalities in cystic fibrosis mice

    PubMed Central

    Shah, Viral S.; Meyerholz, David K.; Tang, Xiao Xiao; Reznikov, Leah; Alaiwa, Mahmoud Abou; Ernst, Sarah E.; Karp, Philip H.; Wohlford-Lenane, Christine L.; Heilmann, Kristopher P.; Leidinger, Mariah R.; Allen, Patrick D.; Zabner, Joseph; McCray, Paul B.; Ostedgaard, Lynda S.; Stoltz, David A.; Randak, Christoph O.; Welsh, Michael J.

    2016-01-01

    Cystic fibrosis (CF) is caused by mutations in the gene that encodes the cystic fibrosis transmembrane conductance regulator (CFTR) anion channel. In humans and pigs, the loss of CFTR impairs respiratory host defenses, causing airway infection. But CF mice are spared. We found that in all three species, CFTR secreted bicarbonate into airway surface liquid. In humans and pigs lacking CFTR, unchecked H+ secretion by the nongastric H+/K+ adenosine triphosphatase (ATP12A) acidified airway surface liquid, which impaired airway host defenses. In contrast, mouse airways expressed little ATP12A and secreted minimal H+; consequently, airway surface liquid in CF and non-CF mice had similar pH. Inhibiting ATP12A reversed host defense abnormalities in human and pig airways. Conversely, expressing ATP12A in CF mouse airways acidified airway surface liquid, impaired defenses, and increased airway bacteria. These findings help explain why CF mice are protected from infection and nominate ATP12A as a potential therapeutic target for CF. PMID:26823428

  6. Regulation of ABO gene expression.

    PubMed

    Kominato, Yoshihiko; Hata, Yukiko; Matsui, Kazuhiro; Takizawa, Hisao

    2005-07-01

    The ABO blood group system is important in blood transfusions and in identifying individuals during criminal investigations. Two carbohydrate antigens, the A and B antigens, and their antibodies constitute this system. Although biochemical and molecular genetic studies have demonstrated the molecular basis of the histo-blood group ABO system, some aspects remain to be elucidated. To explain the molecular basis of how the ABO genes are controlled in cell type-specific expression, during normal cell differentiation, and in cancer cells with invasive and metastatic potential that lack A/B antigens, it is essential to understand the regulatory mechanism of ABO gene transcription. We review the transcriptional regulation of the ABO gene, including positive and negative elements in the upstream region of the gene, and draw some inferences that help to explain the phenomena described above.

  7. Gene expression profile of pulpitis

    PubMed Central

    Galicia, Johnah C.; Henson, Brett R.; Parker, Joel S.; Khan, Asma A.

    2016-01-01

    The cost, prevalence and pain associated with endodontic disease necessitate an understanding of the fundamental molecular aspects of its pathogenesis. This study was aimed to identify the genetic contributors to pulpal pain and inflammation. Inflamed pulps were collected from patients diagnosed with irreversible pulpitis (n=20). Normal pulps from teeth extracted for various reasons served as controls (n=20). Pain level was assessed using a visual analog scale (VAS). Genome-wide microarray analysis was performed using Affymetrix GeneTitan Multichannel Instrument. The difference in gene expression levels were determined by the Significance Analysis of Microarray program using a false discovery rate (q-value) of 5%. Genes involved in immune response, cytokine-cytokine receptor interaction and signaling, integrin cell surface interactions, and others were expressed at relatively higher levels in the in the pulpitis group. Moreover, several genes known to modulate pain and inflammation showed differential expression in asymptomatic and mild pain patients (≥30mm on VAS) compared to those with moderate to severe pain. This exploratory study provides a molecular basis for the clinical diagnosis of pulpitis. With an enhanced understanding of pulpal inflammation, future studies on treatment and management of pulpitis and on pain associated with it can have a biological reference to bridge treatment strategies with pulpal biology. PMID:27052691

  8. Gene expression profile of pulpitis.

    PubMed

    Galicia, J C; Henson, B R; Parker, J S; Khan, A A

    2016-06-01

    The cost, prevalence and pain associated with endodontic disease necessitate an understanding of the fundamental molecular aspects of its pathogenesis. This study was aimed to identify the genetic contributors to pulpal pain and inflammation. Inflamed pulps were collected from patients diagnosed with irreversible pulpitis (n=20). Normal pulps from teeth extracted for various reasons served as controls (n=20). Pain level was assessed using a visual analog scale (VAS). Genome-wide microarray analysis was performed using Affymetrix GeneTitan Multichannel Instrument. The difference in gene expression levels were determined by the significance analysis of microarray program using a false discovery rate (q-value) of 5%. Genes involved in immune response, cytokine-cytokine receptor interaction and signaling, integrin cell surface interactions, and others were expressed at relatively higher levels in the pulpitis group. Moreover, several genes known to modulate pain and inflammation showed differential expression in asymptomatic and mild pain patients (⩾30 mm on VAS) compared with those with moderate to severe pain. This exploratory study provides a molecular basis for the clinical diagnosis of pulpitis. With an enhanced understanding of pulpal inflammation, future studies on treatment and management of pulpitis and on pain associated with it can have a biological reference to bridge treatment strategies with pulpal biology.

  9. Early life ozone exposure results in dysregulated innate immune function and altered microRNA expression in airway epithelium.

    PubMed

    Clay, Candice C; Maniar-Hew, Kinjal; Gerriets, Joan E; Wang, Theodore T; Postlethwait, Edward M; Evans, Michael J; Fontaine, Justin H; Miller, Lisa A

    2014-01-01

    Exposure to ozone has been associated with increased incidence of respiratory morbidity in humans; however the mechanism(s) behind the enhancement of susceptibility are unclear. We have previously reported that exposure to episodic ozone during postnatal development results in an attenuated peripheral blood cytokine response to lipopolysaccharide (LPS) that persists with maturity. As the lung is closely interfaced with the external environment, we hypothesized that the conducting airway epithelium of neonates may also be a target of immunomodulation by ozone. To test this hypothesis, we evaluated primary airway epithelial cell cultures derived from juvenile rhesus macaque monkeys with a prior history of episodic postnatal ozone exposure. Innate immune function was measured by expression of the proinflammatory cytokines IL-6 and IL-8 in primary cultures established following in vivo LPS challenge or, in response to in vitro LPS treatment. Postnatal ozone exposure resulted in significantly attenuated IL-6 mRNA and protein expression in primary cultures from juvenile animals; IL-8 mRNA was also significantly reduced. The effect of antecedent ozone exposure was modulated by in vivo LPS challenge, as primary cultures exhibited enhanced cytokine expression upon secondary in vitro LPS treatment. Assessment of potential IL-6-targeting microRNAs miR-149, miR-202, and miR-410 showed differential expression in primary cultures based upon animal exposure history. Functional assays revealed that miR-149 is capable of binding to the IL-6 3' UTR and decreasing IL-6 protein synthesis in airway epithelial cell lines. Cumulatively, our findings suggest that episodic ozone during early life contributes to the molecular programming of airway epithelium, such that memory from prior exposures is retained in the form of a dysregulated IL-6 and IL-8 response to LPS; differentially expressed microRNAs such as miR-149 may play a role in the persistent modulation of the epithelial innate

  10. Reduction of DNA mismatch repair protein expression in airway epithelial cells of premenopausal women chronically exposed to biomass smoke.

    PubMed

    Mukherjee, Bidisha; Dutta, Anindita; Chowdhury, Saswati; Roychoudhury, Sanghita; Ray, Manas Ranjan

    2014-02-01

    Biomass burning is a major source of indoor air pollution in rural India. This study examined whether chronic inhalation of biomass smoke causes change in the DNA mismatch repair (MMR) pathway in the airway cells. For this, airway cells exfoliated in sputum were collected from 72 premenopausal nonsmoking rural women (median age 34 years) who cooked with biomass (wood, dung, crop residues) and 68 control women who cooked with cleaner fuel liquefied petroleum gas (LPG) for the past 5 years or more. The levels of particulate matters with diameters less than 10 and 2.5 μm (PM10 and PM2.5) in indoor air were measured by real-time aerosol monitor. Benzene exposure was monitored by measuring trans,trans-muconic acid (t,t-MA) in urine by high-performance liquid chromatography with ultraviolet detector. Generation of reactive oxygen species (ROS) and level of superoxide dismutase (SOD) in airway cells were measured by flow cytometry and spectrophotometry, respectively. Immunocytochemical assay revealed lower percentage of airway epithelial cells expressing MMR proteins mutL homolog 1 (MLH1) and mutS homolog 2 (MSH2) in biomass-using women compared to LPG-using controls. Women who cooked with biomass had 6.7 times higher level of urinary t,t-MA, twofold increase in ROS generation, and 31 % depletion of SOD. Indoor air of biomass-using households had three times more particulate matters than that of controls. ROS, urinary t,t-MA, and particulate pollution in biomass-using kitchen had negative correlation, while SOD showed positive correlation with MSH2 and MLH1 expression. It appears that chronic exposure to biomass smoke reduces MMR response in airway epithelial cells, and oxidative stress plays an important role in the process.

  11. Gene expression throughout a vertebrate's embryogenesis

    PubMed Central

    2011-01-01

    Background Describing the patterns of gene expression during embryonic development has broadened our understanding of the processes and patterns that define morphogenesis. Yet gene expression patterns have not been described throughout vertebrate embryogenesis. This study presents statistical analyses of gene expression during all 40 developmental stages in the teleost Fundulus heteroclitus using four biological replicates per stage. Results Patterns of gene expression for 7,000 genes appear to be important as they recapitulate developmental timing. Among the 45% of genes with significant expression differences between pairs of temporally adjacent stages, significant differences in gene expression vary from as few as five to more than 660. Five adjacent stages have disproportionately more significant changes in gene expression (> 200 genes) relative to other stages: four to eight and eight to sixteen cell stages, onset of circulation, pre and post-hatch, and during complete yolk absorption. The fewest differences among adjacent stages occur during gastrulation. Yet, at stage 16, (pre-mid-gastrulation) the largest number of genes has peak expression. This stage has an over representation of genes in oxidative respiration and protein expression (ribosomes, translational genes and proteases). Unexpectedly, among all ribosomal genes, both strong positive and negative correlations occur. Similar correlated patterns of expression occur among all significant genes. Conclusions These data provide statistical support for the temporal dynamics of developmental gene expression during all stages of vertebrate development. PMID:21356103

  12. Does FACS perturb gene expression?

    PubMed

    Richardson, Graham M; Lannigan, Joanne; Macara, Ian G

    2015-02-01

    Fluorescence activated cell sorting is the technique most commonly used to separate primary mammary epithelial sub-populations. Many studies incorporate this technique before analyzing gene expression within specific cellular lineages. However, to our knowledge, no one has examined the effects of fluorescence activated cell sorting (FACS) separation on short-term transcriptional profiles. In this study, we isolated a heterogeneous mixture of cells from the mouse mammary gland. To determine the effects of the isolation and separation process on gene expression, we harvested RNA from the cells before enzymatic digestion, following enzymatic digestion, and following a mock FACS sort where the entire cohort of cells was retained. A strict protocol was followed to minimize disruption to the cells, and to ensure that no subpopulations were enriched or lost. Microarray analysis demonstrated that FACS causes minimal disruptions to gene expression patterns, but prior steps in the mammary cell isolation process are followed by upregulation of 18 miRNA's and rapid decreases in their predicted target transcripts. © 2015 International Society for Advancement of Cytometry.

  13. The Gene Expression Omnibus Database.

    PubMed

    Clough, Emily; Barrett, Tanya

    2016-01-01

    The Gene Expression Omnibus (GEO) database is an international public repository that archives and freely distributes high-throughput gene expression and other functional genomics data sets. Created in 2000 as a worldwide resource for gene expression studies, GEO has evolved with rapidly changing technologies and now accepts high-throughput data for many other data applications, including those that examine genome methylation, chromatin structure, and genome-protein interactions. GEO supports community-derived reporting standards that specify provision of several critical study elements including raw data, processed data, and descriptive metadata. The database not only provides access to data for tens of thousands of studies, but also offers various Web-based tools and strategies that enable users to locate data relevant to their specific interests, as well as to visualize and analyze the data. This chapter includes detailed descriptions of methods to query and download GEO data and use the analysis and visualization tools. The GEO homepage is at http://www.ncbi.nlm.nih.gov/geo/.

  14. Hyperplasia of smooth muscle in mild to moderate asthma without changes in cell size or gene expression.

    PubMed

    Woodruff, Prescott G; Dolganov, Gregory M; Ferrando, Ronald E; Donnelly, Samantha; Hays, Steven R; Solberg, Owen D; Carter, Roderick; Wong, Hofer H; Cadbury, Peggy S; Fahy, John V

    2004-05-01

    Bronchial hyperresponsiveness in mild to moderate asthma may result from airway smooth muscle cell proliferation or acquisition of a hypercontractile phenotype. Because these cells have not been well characterized in mild to moderate asthma, we examined the morphometric and gene expression characteristics of smooth muscle cells in this subgroup of patients with asthma. Using bronchial biopsies from 14 subjects with mild to moderate asthma and 15 control subjects, we quantified smooth muscle cell morphology by stereology and the expression of a panel of genes related to a hypercontractile phenotype of airway smooth muscle, using laser microdissection and two-step real-time polymerase chain reaction. We found that airway smooth muscle cell size was similar in both groups, but cell number was nearly twofold higher in subjects with asthma (p = 0.03), and the amount of smooth muscle in the submucosa was increased 50-83% (p < 0.005). Gene expression profiling in smooth muscle cells showed no difference in the expression of genes encoding phenotypic markers in cells from healthy subjects and subjects with asthma (all p > 0.1). We conclude that airway smooth muscle proliferation is a pathologic characteristic of subjects with mild to moderate asthma. However, smooth muscle cells in mild to moderate asthma do not show hypertrophy or gene expression changes of a hypercontractile phenotype observed in vitro.

  15. Classification of genes based on gene expression analysis

    SciTech Connect

    Angelova, M. Myers, C. Faith, J.

    2008-05-15

    Systems biology and bioinformatics are now major fields for productive research. DNA microarrays and other array technologies and genome sequencing have advanced to the point that it is now possible to monitor gene expression on a genomic scale. Gene expression analysis is discussed and some important clustering techniques are considered. The patterns identified in the data suggest similarities in the gene behavior, which provides useful information for the gene functionalities. We discuss measures for investigating the homogeneity of gene expression data in order to optimize the clustering process. We contribute to the knowledge of functional roles and regulation of E. coli genes by proposing a classification of these genes based on consistently correlated genes in expression data and similarities of gene expression patterns. A new visualization tool for targeted projection pursuit and dimensionality reduction of gene expression data is demonstrated.

  16. MicroRNA in United Airway Diseases

    PubMed Central

    Liu, Zheng; Zhang, Xin-Hao; Callejas-Díaz, Borja; Mullol, Joaquim

    2016-01-01

    The concept of united airway diseases (UAD) has received increasing attention in recent years. Sustained and increased inflammation is a common feature of UAD, which is inevitably accompanied with marked gene modification and tight gene regulation. However, gene regulation in the common inflammatory processes in UAD remains unclear. MicroRNA (miRNA), a novel regulator of gene expression, has been considered to be involved in many inflammatory diseases. Although there are an increasing number of studies of miRNAs in inflammatory upper and lower airway diseases, few miRNAs have been identified that directly link the upper and lower airways. In this article, therefore, we reviewed the relevant studies available in order to improve the understanding of the roles of miRNAs in the interaction and pathogenesis of UAD. PMID:27187364

  17. Expression and regulation of immune-modulatory enzyme indoleamine 2,3-dioxygenase (IDO) by human airway epithelial cells and its effect on T cell activation

    PubMed Central

    Sewell, Herb F.; Knox, Alan; Ghaemmaghami, Amir M.

    2016-01-01

    Indoleamine 2,3-dioxygenase (IDO) catalyzes the degradation of tryptophan, which plays a critical role in immune suppression through regulating the production of a series of metabolites that are generally referred to as kynurenines. It has become increasingly clear that epithelial cells (ECs) play an active role in maintaining lung homeostasis by modulating the function of immune cells via producing cytokines, chemokines, and anti-microbial mediators. In this study we assessed the regulation of IDO activity and expression in human primary ECs and EC lines under steady state conditions and in response to bacterial and allergenic stimuli. We also investigated the potential immune modulatory functions of IDO expression in human airway ECs. Our data clearly show that airway ECs produce IDO, which is down-regulated in response to allergens and TLR ligands while up-regulated in response to IFN-γ. Using gene silencing, we further demonstrate that IDO plays a key role in the EC-mediated suppression of antigen-specific and polyclonal proliferation of T cells. Interestingly, our data also show that ECs lose their inhibitory effect on T cell activation in response to different TLR agonists mimicking bacterial or viral infections. In conclusion, our work provides an understanding of how IDO is regulated in ECs as well as demonstrates that “resting” ECs can suppress T cell activation in an IDO dependent manner. These data provide new insight into how ECs, through the production of IDO, can influence downstream innate and adaptive responses as part of their function in maintaining immune homeostasis in the airways. PMID:27613847

  18. Functional expression of the TMEM16 family of calcium-activated chloride channels in airway smooth muscle

    PubMed Central

    Remy, Kenneth E.; Danielsson, Jennifer; Funayama, Hiromi; Fu, Xiao Wen; Chang, Herng-Yu Sucie; Yim, Peter; Xu, Dingbang; Emala, Charles W.

    2013-01-01

    Airway smooth muscle hyperresponsiveness is a key component in the pathophysiology of asthma. Although calcium-activated chloride channel (CaCC) flux has been described in many cell types, including human airway smooth muscle (HASM), the true molecular identity of the channels responsible for this chloride conductance remains controversial. Recently, a new family of proteins thought to represent the true CaCCs was identified as the TMEM16 family. This led us to question whether members of this family are functionally expressed in native and cultured HASM. We further questioned whether expression of these channels contributes to the contractile function of HASM. We identified the mRNA expression of eight members of the TMEM16 family in HASM cells and show immunohistochemical evidence of TMEM16A in both cultured and native HASM. Functionally, we demonstrate that the classic chloride channel inhibitor, 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB), inhibited halide flux in cultured HASM cells. Moreover, HASM cells displayed classical electrophysiological properties of CaCCs during whole cell electrophysiological recordings, which were blocked by using an antibody selective for TMEM16A. Furthermore, two distinct TMEM16A antagonists (tannic acid and benzbromarone) impaired a substance P-induced contraction in isolated guinea pig tracheal rings. These findings demonstrate that multiple members of this recently described family of CaCCs are expressed in HASM cells, they display classic electrophysiological properties of CaCCs, and they modulate contractile tone in airway smooth muscle. The TMEM16 family may provide a novel therapeutic target for limiting airway constriction in asthma. PMID:23997176

  19. Adenovirus-mediated Foxp3 expression in lung epithelial cells reduces airway inflammation in ovalbumin and cockroach-induced asthma model

    PubMed Central

    Park, Soojin; Chung, Hwan-Suck; Shin, Dasom; Jung, Kyung-Hwa; Lee, Hyunil; Moon, Junghee; Bae, Hyunsu

    2016-01-01

    Foxp3 is a master regulator of CD4+CD25+ regulatory T-cell (Treg) function and is also a suppressor of SKP2 and HER2/ErbB2. There are an increasing number of reports describing the functions of Foxp3 in cell types other than Tregs. In this context, we evaluated the functions of Foxp3 in ovalbumin- and cockroach-induced asthma models. Foxp3-EGFP-expressing adenovirus or EGFP control adenovirus was administered intratracheally (i.t.), followed by challenge with ovalbumin (OVA) or cockroach extract to induce asthma. Th2 cytokine and immune cell profiles of bronchoalveolar lavage fluid (BALF), as well as serum IgE levels, were analyzed. Histological analyses were also conducted to demonstrate the effects of Foxp3 expression on airway remodeling, goblet cell hyperplasia and inflammatory responses in the lung. Adenoviral Foxp3 was expressed only in lung epithelial cells, and not in CD4+ or CD8+ cells. BALF from Foxp3 gene-delivered mice showed significantly reduced numbers of total immune cells, eosinophils, neutrophils, macrophages and lymphocytes in response to cockroach allergen or OVA. In addition, Foxp3 expression in the lung reduced the levels of Th2 cytokines and IgE in BALF and serum, respectively. Moreover, histopathological analysis also showed that Foxp3 expression substantially inhibited eosinophil infiltration into the airways, goblet cell hyperplasia and smooth muscle cell hypertrophy. Furthermore, when Tregs were depleted by diphtheria toxin in Foxp3DTR mice, the anti-asthmatic functions of Foxp3 were not altered in OVA-challenged asthma models. In this study, our results suggest that Foxp3 expression in lung epithelial cells, and not in Tregs, inhibited OVA- and cockroach extract-induced asthma. PMID:27633092

  20. Adenovirus-mediated Foxp3 expression in lung epithelial cells reduces airway inflammation in ovalbumin and cockroach-induced asthma model.

    PubMed

    Park, Soojin; Chung, Hwan-Suck; Shin, Dasom; Jung, Kyung-Hwa; Lee, Hyunil; Moon, Junghee; Bae, Hyunsu

    2016-09-16

    Foxp3 is a master regulator of CD4(+)CD25(+) regulatory T-cell (Treg) function and is also a suppressor of SKP2 and HER2/ErbB2. There are an increasing number of reports describing the functions of Foxp3 in cell types other than Tregs. In this context, we evaluated the functions of Foxp3 in ovalbumin- and cockroach-induced asthma models. Foxp3-EGFP-expressing adenovirus or EGFP control adenovirus was administered intratracheally (i.t.), followed by challenge with ovalbumin (OVA) or cockroach extract to induce asthma. Th2 cytokine and immune cell profiles of bronchoalveolar lavage fluid (BALF), as well as serum IgE levels, were analyzed. Histological analyses were also conducted to demonstrate the effects of Foxp3 expression on airway remodeling, goblet cell hyperplasia and inflammatory responses in the lung. Adenoviral Foxp3 was expressed only in lung epithelial cells, and not in CD4(+) or CD8(+) cells. BALF from Foxp3 gene-delivered mice showed significantly reduced numbers of total immune cells, eosinophils, neutrophils, macrophages and lymphocytes in response to cockroach allergen or OVA. In addition, Foxp3 expression in the lung reduced the levels of Th2 cytokines and IgE in BALF and serum, respectively. Moreover, histopathological analysis also showed that Foxp3 expression substantially inhibited eosinophil infiltration into the airways, goblet cell hyperplasia and smooth muscle cell hypertrophy. Furthermore, when Tregs were depleted by diphtheria toxin in Foxp3(DTR) mice, the anti-asthmatic functions of Foxp3 were not altered in OVA-challenged asthma models. In this study, our results suggest that Foxp3 expression in lung epithelial cells, and not in Tregs, inhibited OVA- and cockroach extract-induced asthma.

  1. Harnessing gene expression networks to prioritize candidate epileptic encephalopathy genes.

    PubMed

    Oliver, Karen L; Lukic, Vesna; Thorne, Natalie P; Berkovic, Samuel F; Scheffer, Ingrid E; Bahlo, Melanie

    2014-01-01

    We apply a novel gene expression network analysis to a cohort of 182 recently reported candidate Epileptic Encephalopathy genes to identify those most likely to be true Epileptic Encephalopathy genes. These candidate genes were identified as having single variants of likely pathogenic significance discovered in a large-scale massively parallel sequencing study. Candidate Epileptic Encephalopathy genes were prioritized according to their co-expression with 29 known Epileptic Encephalopathy genes. We utilized developing brain and adult brain gene expression data from the Allen Human Brain Atlas (AHBA) and compared this to data from Celsius: a large, heterogeneous gene expression data warehouse. We show replicable prioritization results using these three independent gene expression resources, two of which are brain-specific, with small sample size, and the third derived from a heterogeneous collection of tissues with large sample size. Of the nineteen genes that we predicted with the highest likelihood to be true Epileptic Encephalopathy genes, two (GNAO1 and GRIN2B) have recently been independently reported and confirmed. We compare our results to those produced by an established in silico prioritization approach called Endeavour, and finally present gene expression networks for the known and candidate Epileptic Encephalopathy genes. This highlights sub-networks of gene expression, particularly in the network derived from the adult AHBA gene expression dataset. These networks give clues to the likely biological interactions between Epileptic Encephalopathy genes, potentially highlighting underlying mechanisms and avenues for therapeutic targets.

  2. The TAK1→IKKβ→TPL2→MKK1/MKK2 Signaling Cascade Regulates IL-33 Expression in Cystic Fibrosis Airway Epithelial Cells Following Infection by Pseudomonas aeruginosa

    PubMed Central

    Farias, Raquel; Rousseau, Simon

    2016-01-01

    In cystic fibrosis (CF), chronic respiratory infections result in an exaggerated and uncontrolled inflammatory response that ultimately lead to a decrease in pulmonary function. We have previously described the presence of the alarmin IL-33 in lung explants from CF patients. The signals regulating IL-33 expression in the airway epithelium following a gram-negative bacterial infection are currently unknown. Our objective was to characterize the pathways in CF airway epithelial cells (AECs) leading to an increase in IL-33 expression. We found that, in CF AECs expressing a deletion of a phenylalanine at position 508 of the gene coding for Cystic Fibrosis Transmembrane Conductance Regulator (CFTRdelF508), exposure to live Pseudomonas aeruginosa upregulates IL-33 via the TLR2 and TLR5 signaling pathways. This up-regulation can be partially or fully reverted by pre-incubating CFTRdelF508 AECs with a CFTR corrector (VX-809) and/or a CFTR potentiator (VX-770). Similarly, incubation with the CFTR corrector and/or the CFTR potentiator also decreased IL-8 expression in response to infection. Moreover, using different protein kinase inhibitors that target elements downstream of TLR signaling, we show that the TAK1→IKKβ→TPL2→MKK1/MKK2 pathway regulates IL-33 expression following an infection with P. aeruginosa. Our findings represent the first characterization of the signals regulating IL-33 expression in CF airway epithelial cells in response to a bacterial infection. PMID:26793709

  3. Pulmonary Gene Expression Profiling of Inhaled Ricin

    DTIC Science & Technology

    2007-11-02

    in which 34 genes had statistically significant changes in gene expression. Transcripts identified by the assay included those that facilitate...gene expression. Transcripts identified by the assay included those that facilitate tissue healing (early growth response gene (egr)-1), regulate...impingement to determine aerosol concentration. Ricin concentrations from impinger samples were measured by protein assay (Pierce, MicroBCA, Rockford

  4. Phosphoinositide 3-kinase δ gene mutation predisposes to respiratory infection and airway damage

    PubMed Central

    Angulo, Ivan; Vadas, Oscar; Garçon, Fabien; Banham-Hall, Edward; Plagnol, Vincent; Leahy, Timothy R.; Baxendale, Helen; Coulter, Tanya; Curtis, James; Wu, Changxin; Blake-Palmer, Katherine; Perisic, Olga; Smyth, Deborah; Maes, Mailis; Fiddler, Christine; Juss, Jatinder; Cilliers, Deirdre; Markelj, Gašper; Chandra, Anita; Farmer, George; Kielkowska, Anna; Clark, Jonathan; Kracker, Sven; Debré, Marianne; Picard, Capucine; Pellier, Isabelle; Jabado, Nada; Morris, James A.; Barcenas-Morales, Gabriela; Fischer, Alain; Stephens, Len; Hawkins, Phillip; Barrett, Jeffrey C.; Abinun, Mario; Clatworthy, Menna; Durandy, Anne; Doffinger, Rainer; Chilvers, Edwin; Cant, Andrew J.; Kumararatne, Dinakantha; Okkenhaug, Klaus; Williams, Roger L.; Condliffe, Alison; Nejentsev, Sergey

    2014-01-01

    Genetic mutations cause primary immunodeficiencies (PIDs), which predispose to infections. Here we describe Activated PI3K-δ Syndrome (APDS), a PID associated with a dominant gain-of-function mutation E1021K in the p110δ protein, the catalytic subunit of phosphoinositide 3-kinase δ (PI3Kδ), encoded by the PIK3CD gene. We found E1021K in 17 patients from seven unrelated families, but not among 3,346 healthy subjects. APDS was characterized by recurrent respiratory infections, progressive airway damage, lymphopenia, increased circulating transitional B cells, increased IgM and reduced IgG2 levels in serum and impaired vaccine responses. The E1021K mutation enhanced membrane association and kinase activity of p110δ. Patient-derived lymphocytes had increased levels of phosphatidylinositol 3,4,5-trisphosphate and phosphorylated AKT protein and were prone to activation-induced cell death. Selective p110δ inhibitors IC87114 and GS-1101 reduced the activity of the mutant enzyme in vitro, suggesting a therapeutic approach for patients with APDS. PMID:24136356

  5. Glycerophosphorylcholine regulates Haemophilus influenzae glpQ gene expression.

    PubMed

    Alrousan, Enas; Fan, Xin

    2015-05-01

    An important virulence strategy adopted by Haemophilus influenzae to establish a niche on the mucosal surface of the host is the phosphorylcholine (ChoP) decoration of its lipopolysaccharides, which promotes adherence to the host cells. Haemophilus influenzae is able to use glycerophosphorylcholine (GPC) from host for ChoP synthesis. Utilization of GPC requires glpQ, which encodes a glycerophosphodiester phosphodiesterase enzyme. In this study, we investigate the transcriptional regulation of glpQ gene using real-time PCR and transcriptional fusion of H. influenzae glpQ promoter to the Escherichia coli lacZ reporter gene. The glpQ promoter activities were examined under environmental conditions including changes in temperature, oxygen, high salt and minimal growth medium. Our data showed that under room temperature and anaerobic conditions, the glpQ gene expression levels were significantly higher than under other growth conditions. In addition, the glpQ gene expression levels were upregulated in the presence of GPC. These results suggest that H. influenzae may upregulate glpQ expression in response to different environments it encounters during infection, from the airway surfaces (room temperature) to deep tissues (anaerobic). Upregulation of glpQ by GPC may allow efficient use of abundant GPC from mammalian cells by H. influenzae as a source of nutrient and for ChoP decoration of lipopolysaccharide that facilitates bacterial adhesion to host cells and growth during infection.

  6. Does inbreeding affect gene expression in birds?

    PubMed

    Hansson, Bengt; Naurin, Sara; Hasselquist, Dennis

    2014-09-01

    Inbreeding increases homozygosity, exposes genome-wide recessive deleterious alleles and often reduces fitness. The physiological and reproductive consequences of inbreeding may be manifested already during gene regulation, but the degree to which inbreeding influences gene expression is unknown in most organisms, including in birds. To evaluate the pattern of inbreeding-affected gene expression over the genome and in relation to sex, we performed a transcriptome-wide gene expression (10 695 genes) study of brain tissue of 10-day-old inbred and outbred, male and female zebra finches. We found significantly lower gene expression in females compared with males at Z-linked genes, confirming that dosage compensation is incomplete in female birds. However, inbreeding did not affect gene expression at autosomal or sex-linked genes, neither in males nor in females. Analyses of single genes again found a clear sex-biased expression at Z-linked genes, whereas only a single gene was significantly affected by inbreeding. The weak effect of inbreeding on gene expression in zebra finches contrasts to the situation, for example, in Drosophila where inbreeding has been found to influence gene expression more generally and at stress-related genes in particular.

  7. [Neuronal plasticity and gene expression].

    PubMed

    Sokolova, O O; Shtark, M B; Lisachev, P D

    2010-01-01

    Neuronal plasticity--a fundamental feature of brain--provides adequate interactions with dynamic environment. One of the most deeply investigated forms of the neuronal plasticity is a long-term potentiation (LTP)--a phenomenon underlying learning and memory. Signal paths activated during LTP converge into the nuclear of the neuron, giving rise to launch of the molecular-genetic programs, which mediate structural and functional remodeling of synapses. In the review data concerning involvement of multilevel gene expression into plastic change under neuronal activation are summarized.

  8. Corticosteroid-induced gene expression in allergen-challenged asthmatic subjects taking inhaled budesonide

    PubMed Central

    Kelly, MM; King, EM; Rider, CF; Gwozd, C; Holden, NS; Eddleston, J; Zuraw, B; Leigh, R; O'Byrne, PM; Newton, R

    2012-01-01

    BACKGROUND AND PURPOSE Inhaled corticosteroids (ICS) are the cornerstone of asthma pharmacotherapy and, acting via the glucocorticoid receptor (GR), reduce inflammatory gene expression. While this is often attributed to a direct inhibitory effect of the GR on inflammatory gene transcription, corticosteroids also induce the expression of anti-inflammatory genes in vitro. As there are no data to support this effect in asthmatic subjects taking ICS, we have assessed whether ICS induce anti-inflammatory gene expression in subjects with atopic asthma. EXPERIMENTAL APPROACH Bronchial biopsies from allergen-challenged atopic asthmatic subjects taking inhaled budesonide or placebo were subjected to gene expression analysis using real-time reverse transcriptase-PCR for the corticosteroid-inducible genes (official gene symbols with aliases in parentheses): TSC22D3 [glucocorticoid-induced leucine zipper (GILZ)], dual-specificity phosphatase-1 (MAPK phosphatase-1), both anti-inflammatory effectors, and FKBP5 [FK506-binding protein 51 (FKBP51)], a regulator of GR function. Cultured pulmonary epithelial and smooth muscle cells were also treated with corticosteroids before gene expression analysis. KEY RESULTS Compared with placebo, GILZ and FKBP51 mRNA expression was significantly elevated in budesonide-treated subjects. Budesonide also increased GILZ expression in human epithelial and smooth muscle cells in culture. Immunostaining of bronchial biopsies revealed GILZ expression in the airways epithelium and smooth muscle of asthmatic subjects. CONCLUSIONS AND IMPLICATIONS Expression of the corticosteroid-induced genes, GILZ and FKBP51, is up-regulated in the airways of allergen-challenged asthmatic subjects taking inhaled budesonide. Consequently, the biological effects of corticosteroid-induced genes should be considered when assessing the actions of ICS. Treatment modalities that increase or decrease GR-dependent transcription may correspondingly affect corticosteroid efficacy

  9. MicroRNA-146a and microRNA-146b expression and anti-inflammatory function in human airway smooth muscle.

    PubMed

    Comer, Brian S; Camoretti-Mercado, Blanca; Kogut, Paul C; Halayko, Andrew J; Solway, Julian; Gerthoffer, William T

    2014-11-01

    MicroRNA (miR)-146a and miR-146b are negative regulators of inflammatory gene expression in lung fibroblasts, epithelial cells, monocytes, and endothelial cells. The abundance of cyclooxygenase-2 (COX-2) and IL-1β is negatively regulated by the miR-146 family, suggesting miR-146a and/or miR-146b might modulate inflammatory mediator expression in airway smooth muscle thereby contributing to pathogenesis of asthma. To test this idea we compared miR-146a and miR-146b expression in human airway smooth muscle cells (hASMCs) from nonasthmatic and asthmatic subjects treated with cytomix (IL-1β, TNF-α, and IFNγ) and examined the miRNAs' effects on COX-2 and IL-1β expression. We found that cytomix treatment elevated miR-146a and miR-146b abundance. Induction with cytomix was greater than induction with individual cytokines, and asthmatic cells exhibited higher levels of miR-146a expression following cytomix treatment than nonasthmatic cells. Transfection of miR-146a or miR-146b mimics reduced COX-2 and IL-1β expression. A miR-146a inhibitor increased COX-2 and IL-1β expression, but a miR-146b inhibitor was ineffective. Repression of COX-2 and IL-1β expression by miR-146a correlated with reduced abundance of the RNA-binding protein human antigen R. These results demonstrate that miR-146a and miR-146b expression is inducible in hASMCs by proinflammatory cytokines and that miR-146a expression is greater in asthmatic cells. Both miR-146a and miR-146b can negatively regulate COX-2 and IL-1β expression at pharmacological levels, but loss-of-function studies showed that only miR-146a is an endogenous negative regulator in hASMCs. The results suggest miR-146 mimics may be an attractive candidate for further preclinical studies as an anti-inflammatory treatment of asthma.

  10. Interleukin-13 interferes with CFTR and AQP5 expression and localization during human airway epithelial cell differentiation

    SciTech Connect

    Skowron-zwarg, Marie; Boland, Sonja; Caruso, Nathalie; Coraux, Christelle; Marano, Francelyne; Tournier, Frederic . E-mail: f-tournier@paris7.jussieu.fr

    2007-07-15

    Interleukin-13 (IL-13) is a central regulator of Th2-dominated respiratory disorders such as asthma. Lesions of the airway epithelial barrier frequently observed in chronic respiratory inflammatory diseases are repaired through proliferation, migration and differentiation of epithelial cells. Our work is focused on the effects of IL-13 in human cellular models of airway epithelial cell regeneration. We have previously shown that IL-13 altered epithelial cell polarity during mucociliary differentiation of human nasal epithelial cells. In particular, the cytokine inhibited ezrin expression and interfered with its apical localization during epithelial cell differentiation in vitro. Here we show that CFTR expression is enhanced in the presence of the cytokine, that two additional CFTR protein isoforms are expressed in IL-13-treated cells and that part of the protein is retained within the endoplasmic reticulum. We further show that aquaporin 5 expression, a water channel localized within the apical membrane of epithelial cells, is completely abolished in the presence of the cytokine. These results show that IL-13 interferes with ion and water channel expression and localization during epithelial regeneration and may thereby influence mucus composition and hydration.

  11. Mechanoregulation of gene expression in fibroblasts

    PubMed Central

    Wang, James H.-C.; Thampatty, Bhavani P.; Lin, Jeen-Shang; Im, Hee-Jeong

    2010-01-01

    Mechanical loads placed on connective tissues alter gene expression in fibroblasts through mechanotransduction mechanisms by which cells convert mechanical signals into cellular biological events, such as gene expression of extracellular matrix components (e.g., collagen). This mechanical regulation of ECM gene expression affords maintenance of connective tissue homeostasis. However, mechanical loads can also interfere with homeostatic cellular gene expression and consequently cause the pathogenesis of connective tissue diseases such as tendinopathy and osteoarthritis. Therefore, the regulation of gene expression by mechanical loads is closely related to connective tissue physiology and pathology. This article reviews the effects of various mechanical loading conditions on gene regulation in fibroblasts and discusses several mechanotransduction mechanisms. Future research directions in mechanoregulation of gene expression are also suggested. PMID:17331678

  12. Comparison of Chromosome 4 gene expression profile between lung telocytes and other local cell types.

    PubMed

    Song, Dongli; Cretoiu, Dragos; Zheng, Minghuan; Qian, Mengjia; Zhang, Miaomiao; Cretoiu, Sanda M; Chen, Luonan; Fang, Hao; Popescu, Laurentiu M; Wang, Xiangdong

    2016-01-01

    Telocytes (TCs) are new cellular entities of mesenchymal origin described almost ubiquitously in human and mammalian organs (www.telocytes.com). Different subtypes of TCs were described, all forming networks in the interstitial space by homo- and heterocellular junctions. Previous studies analysed the gene expression profiles of chromosomes 1, 2, 3, 17 and 18 of murine pulmonary TCs. In this study, we analysed by bioinformatics tools the gene expression profiles of chromosome 4 for murine pulmonary TCs and compared it with mesenchymal stem cells (MSCs), fibroblasts (Fbs), alveolar type II cells (ATII), airway basal cells, proximal airway cells, CD8(+) T cells from bronchial lymph nodes (T-BL) and CD8(+) T cells from lungs (T-L). Key functional genes were identified with the aid of the reference library of the National Center for Biotechnology Information Gene Expression Omnibus database. Seventeen genes were up-regulated and 56 genes were down-regulated in chromosome 4 of TCs compared with other cells. Four genes (Akap2, Gpr153, Sdc3 and Tbc1d2) were up-regulated between one and fourfold and one gene, Svep1, was overexpressed over fourfold. The main functional networks were identified and analysed, pointing out to a TCs involvement in cellular signalling, regulation of tissue inflammation and cell expansion and movement.

  13. Differential Gene Expression in Human Cerebrovascular Malformations

    PubMed Central

    Shenkar, Robert; Elliott, J. Paul; Diener, Katrina; Gault, Judith; Hu, Ling-Jia; Cohrs, Randall J.; Phang, Tzulip; Hunter, Lawrence; Breeze, Robert E.; Awad, Issam A.

    2009-01-01

    OBJECTIVE We sought to identify genes with differential expression in cerebral cavernous malformations (CCMs), arteriovenous malformations (AVMs), and control superficial temporal arteries (STAs) and to confirm differential expression of genes previously implicated in the pathobiology of these lesions. METHODS Total ribonucleic acid was isolated from four CCM, four AVM, and three STA surgical specimens and used to quantify lesion-specific messenger ribonucleic acid expression levels on human gene arrays. Data were analyzed with the use of two separate methodologies: gene discovery and confirmation analysis. RESULTS The gene discovery method identified 42 genes that were significantly up-regulated and 36 genes that were significantly down-regulated in CCMs as compared with AVMs and STAs (P = 0.006). Similarly, 48 genes were significantly up-regulated and 59 genes were significantly down-regulated in AVMs as compared with CCMs and STAs (P = 0.006). The confirmation analysis showed significant differential expression (P < 0.05) in 11 of 15 genes (angiogenesis factors, receptors, and structural proteins) that previously had been reported to be expressed differentially in CCMs and AVMs in immunohistochemical analysis. CONCLUSION We identify numerous genes that are differentially expressed in CCMs and AVMs and correlate expression with the immunohistochemistry of genes implicated in cerebrovascular malformations. In future efforts, we will aim to confirm candidate genes specifically related to the pathobiology of cerebrovascular malformations and determine their biological systems and mechanistic relevance. PMID:12535382

  14. Norovirus gene expression and replication.

    PubMed

    Thorne, Lucy G; Goodfellow, Ian G

    2014-02-01

    Noroviruses are small, positive-sense RNA viruses within the family Caliciviridae, and are now accepted widely as a major cause of acute gastroenteritis in both developed and developing countries. Despite their impact, our understanding of the life cycle of noroviruses has lagged behind that of other RNA viruses due to the inability to culture human noroviruses (HuNVs). Our knowledge of norovirus biology has improved significantly over the past decade as a result of numerous technological advances. The use of a HuNV replicon, improved biochemical and cell-based assays, combined with the discovery of a murine norovirus capable of replication in cell culture, has improved greatly our understanding of the molecular mechanisms of norovirus genome translation and replication, as well as the interaction with host cell processes. In this review, the current state of knowledge of the intracellular life of noroviruses is discussed with particular emphasis on the mechanisms of viral gene expression and viral genome replication.

  15. Familial aggregation analysis of gene expressions

    PubMed Central

    Rao, Shao-Qi; Xu, Liang-De; Zhang, Guang-Mei; Li, Xia; Li, Lin; Shen, Gong-Qing; Jiang, Yang; Yang, Yue-Ying; Gong, Bin-Sheng; Jiang, Wei; Zhang, Fan; Xiao, Yun; Wang, Qing K

    2007-01-01

    Traditional studies of familial aggregation are aimed at defining the genetic (and non-genetic) causes of a disease from physiological or clinical traits. However, there has been little attempt to use genome-wide gene expressions, the direct phenotypic measures of genes, as the traits to investigate several extended issues regarding the distributions of familially aggregated genes on chromosomes or in functions. In this study we conducted a genome-wide familial aggregation analysis by using the in vitro cell gene expressions of 3300 human autosome genes (Problem 1 data provided to Genetic Analysis Workshop 15) in order to answer three basic genetics questions. First, we investigated how gene expressions aggregate among different types (degrees) of relative pairs. Second, we conducted a bioinformatics analysis of highly familially aggregated genes to see how they are distributed on chromosomes. Third, we performed a gene ontology enrichment test of familially aggregated genes to find evidence to support their functional consensus. The results indicated that 1) gene expressions did aggregate in families, especially between sibs. Of 3300 human genes analyzed, there were a total of 1105 genes with one or more significant (empirical p < 0.05) familial correlation; 2) there were several genomic hot spots where highly familially aggregated genes (e.g., the chromosome 6 HLA genes cluster) were clustered; 3) as we expected, gene ontology enrichment tests revealed that the 1105 genes were aggregating not only in families but also in functional categories. PMID:18466548

  16. Methods for monitoring multiple gene expression

    SciTech Connect

    Berka, Randy; Bachkirova, Elena; Rey, Michael

    2012-05-01

    The present invention relates to methods for monitoring differential expression of a plurality of genes in a first filamentous fungal cell relative to expression of the same genes in one or more second filamentous fungal cells using microarrays containing Trichoderma reesei ESTs or SSH clones, or a combination thereof. The present invention also relates to computer readable media and substrates containing such array features for monitoring expression of a plurality of genes in filamentous fungal cells.

  17. Methods for monitoring multiple gene expression

    SciTech Connect

    Berka, Randy; Bachkirova, Elena; Rey, Michael

    2013-10-01

    The present invention relates to methods for monitoring differential expression of a plurality of genes in a first filamentous fungal cell relative to expression of the same genes in one or more second filamentous fungal cells using microarrays containing Trichoderma reesei ESTs or SSH clones, or a combination thereof. The present invention also relates to computer readable media and substrates containing such array features for monitoring expression of a plurality of genes in filamentous fungal cells.

  18. Methods for monitoring multiple gene expression

    DOEpatents

    Berka, Randy; Bachkirova, Elena; Rey, Michael

    2008-06-01

    The present invention relates to methods for monitoring differential expression of a plurality of genes in a first filamentous fungal cell relative to expression of the same genes in one or more second filamentous fungal cells using microarrays containing Trichoderma reesei ESTs or SSH clones, or a combination thereof. The present invention also relates to computer readable media and substrates containing such array features for monitoring expression of a plurality of genes in filamentous fungal cells.

  19. A novel small compound SH-2251 suppresses Th2 cell-dependent airway inflammation through selective modulation of chromatin status at the Il5 gene locus.

    PubMed

    Suzuki, Junpei; Kuwahara, Makoto; Tofukuji, Soichi; Imamura, Masashi; Kato, Fuminori; Nakayama, Toshinori; Ohara, Osamu; Yamashita, Masakatsu

    2013-01-01

    IL-5 is a key cytokine that plays an important role in the development of pathological conditions in allergic inflammation. Identifying strategies to inhibit IL-5 production is important in order to establish new therapies for treating allergic inflammation. We found that SH-2251, a novel thioamide-related small compound, selectively inhibits the differentiation of IL-5-producing Th2 cells. SH-2251 inhibited the induction of active histone marks at the Il5 gene locus during Th2 cell differentiation. The recruitment of RNA polymerase II, and following expression of the Th2 cell-specific intergenic transcripts around the Il5 gene locus was also inhibited. Furthermore, Th2 cell-dependent airway inflammation in mice was suppressed by the oral administration of SH-2251. Gfi1, a transcriptional repressor, was identified as a downstream target molecule of SH-2251 using a DNA microarray analysis. The Gfi1 expression dramatically decreased in SH-2251-treated Th2 cells, and the SH-2251-mediated inhibition of IL-5-producing Th2 cell differentiation was restored by transduction of Gfi1. Therefore, our study unearthed SH-2251 as a novel therapeutic candidate for allergic inflammation that selectively inhibits active histone marks at the Il5 gene locus.

  20. Estimation and Testing of Gene Expression Heterosis

    PubMed Central

    Liu, Peng; Nettleton, Dan

    2014-01-01

    Heterosis, also known as the hybrid vigor, occurs when the mean phenotype of hybrid off-spring is superior to that of its two inbred parents. The heterosis phenomenon is extensively utilized in agriculture though the molecular basis is still unknown. In an effort to understand phenotypic heterosis at the molecular level, researchers have begun to compare expression levels of thousands of genes between parental inbred lines and their hybrid offspring to search for evidence of gene expression heterosis. Standard statistical approaches for separately analyzing expression data for each gene can produce biased and highly variable estimates and unreliable tests of heterosis. To address these shortcomings, we develop a hierarchical model to borrow information across genes. Using our modeling framework, we derive empirical Bayes estimators and an inference strategy to identify gene expression heterosis. Simulation results show that our proposed method outperforms the more traditional strategy used to detect gene expression heterosis. This article has supplementary material online. PMID:25435758

  1. Estimation and Testing of Gene Expression Heterosis.

    PubMed

    Ji, Tieming; Liu, Peng; Nettleton, Dan

    2014-09-01

    Heterosis, also known as the hybrid vigor, occurs when the mean phenotype of hybrid off-spring is superior to that of its two inbred parents. The heterosis phenomenon is extensively utilized in agriculture though the molecular basis is still unknown. In an effort to understand phenotypic heterosis at the molecular level, researchers have begun to compare expression levels of thousands of genes between parental inbred lines and their hybrid offspring to search for evidence of gene expression heterosis. Standard statistical approaches for separately analyzing expression data for each gene can produce biased and highly variable estimates and unreliable tests of heterosis. To address these shortcomings, we develop a hierarchical model to borrow information across genes. Using our modeling framework, we derive empirical Bayes estimators and an inference strategy to identify gene expression heterosis. Simulation results show that our proposed method outperforms the more traditional strategy used to detect gene expression heterosis. This article has supplementary material online.

  2. Relationships among CFTR expression, HCO3- secretion, and host defense may inform gene- and cell-based cystic fibrosis therapies.

    PubMed

    Shah, Viral S; Ernst, Sarah; Tang, Xiao Xiao; Karp, Philip H; Parker, Connor P; Ostedgaard, Lynda S; Welsh, Michael J

    2016-05-10

    Cystic fibrosis (CF) is caused by mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR) anion channel. Airway disease is the major source of morbidity and mortality. Successful implementation of gene- and cell-based therapies for CF airway disease requires knowledge of relationships among percentages of targeted cells, levels of CFTR expression, correction of electrolyte transport, and rescue of host defense defects. Previous studies suggested that, when ∼10-50% of airway epithelial cells expressed CFTR, they generated nearly wild-type levels of Cl(-) secretion; overexpressing CFTR offered no advantage compared with endogenous expression levels. However, recent discoveries focused attention on CFTR-mediated HCO3 (-) secretion and airway surface liquid (ASL) pH as critical for host defense and CF pathogenesis. Therefore, we generated porcine airway epithelia with varying ratios of CF and wild-type cells. Epithelia with a 50:50 mix secreted HCO3 (-) at half the rate of wild-type epithelia. Likewise, heterozygous epithelia (CFTR(+/-) or CFTR(+/∆F508)) expressed CFTR and secreted HCO3 (-) at ∼50% of wild-type values. ASL pH, antimicrobial activity, and viscosity showed similar relationships to the amount of CFTR. Overexpressing CFTR increased HCO3 (-) secretion to rates greater than wild type, but ASL pH did not exceed wild-type values. Thus, in contrast to Cl(-) secretion, the amount of CFTR is rate-limiting for HCO3 (-) secretion and for correcting host defense abnormalities. In addition, overexpressing CFTR might produce a greater benefit than expressing CFTR at wild-type levels when targeting small fractions of cells. These findings may also explain the risk of airway disease in CF carriers.

  3. Fetal Exposure of Rhesus Macaques to Bisphenol A Alters Cellular Development of the Conducting Airway by Changing Epithelial Secretory Product Expression

    PubMed Central

    Murphy, Shannon R.; Boetticher, Miriam V.; VandeVoort, Catherine A.

    2013-01-01

    Background: Bisphenol A (BPA) exposure early in life results in organizational changes in reproductive organs, but the effect of BPA on conducting airway cellular maturation has not been studied. Late gestation is characterized by active differentiation of secretory cells in the lung epithelium. Objective: We evaluated the hypothesis that BPA exposure disrupts epithelial secretory cell development in the fetal conducting airway of the rhesus macaque. Methods: We exposed animals to BPA during either the second (early term) or the third (late term) trimester. There were four treatment groups: a) sham control early term, b) sham control late term, c) BPA early term (BPA-early), and d) BPA late term (BPA-late). Because cellular maturation occurs nonuniformly in the lung, we defined mRNA and protein expression by airway level using microdissection. Results: BPA exposure of the dam during late term significantly accelerated secretory cell maturation in the proximal airways of the fetus; both Clara cell secretory protein (CCSP) and MUC5AC/5B mRNA and protein expression increased. Conclusions: BPA exposure during late gestation accelerates secretory cell maturation in the proximal conducting airways. We identified a critical window of fetal susceptibility for BPA effects on lung epithelial cell maturation in the third trimester. This is of environmental health importance because increases in airway mucins are hallmarks of a number of childhood lung diseases that may be affected by BPA exposure. PMID:23757601

  4. Cardiac gene expression data and in silico analysis provide novel insights into human and mouse taste receptor gene regulation.

    PubMed

    Foster, Simon R; Porrello, Enzo R; Stefani, Maurizio; Smith, Nicola J; Molenaar, Peter; dos Remedios, Cristobal G; Thomas, Walter G; Ramialison, Mirana

    2015-10-01

    G protein-coupled receptors are the principal mediators of the sweet, umami, bitter, and fat taste qualities in mammals. Intriguingly, the taste receptors are also expressed outside of the oral cavity, including in the gut, airways, brain, and heart, where they have additional functions and contribute to disease. However, there is little known about the mechanisms governing the transcriptional regulation of taste receptor genes. Following our recent delineation of taste receptors in the heart, we investigated the genomic loci encoding for taste receptors to gain insight into the regulatory mechanisms that drive their expression in the heart. Gene expression analyses of healthy and diseased human and mouse hearts showed coordinated expression for a subset of chromosomally clustered taste receptors. This chromosomal clustering mirrored the cardiac expression profile, suggesting that a common gene regulatory block may control the taste receptor locus. We identified unique domains with strong regulatory potential in the vicinity of taste receptor genes. We also performed de novo motif enrichment in the proximal promoter regions and found several overrepresented DNA motifs in cardiac taste receptor gene promoters corresponding to ubiquitous and cardiac-specific transcription factor binding sites. Thus, combining cardiac gene expression data with bioinformatic analyses, this study has provided insights into the noncoding regulatory landscape for taste GPCRs. These findings also have broader relevance for the study of taste GPCRs outside of the classical gustatory system, where understanding the mechanisms controlling the expression of these receptors may have implications for future therapeutic development.

  5. Regulation of mucous differentiation and mucin gene expression in the tracheobronchial epithelium.

    PubMed

    Gray, T; Koo, J S; Nettesheim, P

    2001-03-07

    The goal of our studies is to elucidate mechanisms that control and modulate mucous differentiation and mucin gene expression in the conducting airways. We used cultures of normal human tracheobronchial epithelial (NHTBE) cells that were shown to secrete two major airway mucins, namely MUC5AC and MUC5B as well as several other secretory products. Mucous differentiation and expression of MUC2, MUC5AC, MUC5B and MUC7, but not MUCi, MUC4, and MUC8 mucin genes, were shown to be retinoic acid- (RA) or retinol-dependent. We found that RA control of mucin genes was mediated by the retinoid acid receptors RAR alpha and, to a lesser extent, by RAR gamma. Our studies also showed that other important bioregulators such as thyroid hormone (T3) and epidermal growth factor (EGF) modulate basal expression of mucin genes, interacting with RA in a concentration-dependent manner. T3, which binds to thyroid receptors (TRs) belonging to the same superfamily of steroid hormone nuclear receptors as the RARs, inhibits mucin gene expression, particularly MUC5AC. One possible mechanism of this T3 effect is downregulation of RAR proteins, which are critical for mucin gene expression. However, we also found that T3 inhibits MUC5AC transcription.EGF, which had previously been shown to stimulate mucin expression and mucin secretion in cultured rat tracheal epithelial (RTE) cells, inhibited mucin secretion in human bronchial epithelial cell cultures. This effect was EGF concentration- and time-dependent and was progressively abolished by increasing the RA concentration. Subsequent studies suggested that the inhibitory effects of high concentrations of EGF may result from selective reduction of MUC5AC expression. These studies thus point to potentially important species differences in the mechanisms regulating mucous production, and they also confirm previous findings indicating differential regulation of MUC5AC and MUC5B gene expression.

  6. Cox4i2, Ifit2, and Prdm11 Mutant Mice: Effective Selection of Genes Predisposing to an Altered Airway Inflammatory Response from a Large Compendium of Mutant Mouse Lines.

    PubMed

    Horsch, Marion; Aguilar-Pimentel, Juan Antonio; Bönisch, Clemens; Côme, Christophe; Kolster-Fog, Cathrine; Jensen, Klaus T; Lund, Anders H; Lee, Icksoo; Grossman, Lawrence I; Sinkler, Christopher; Hüttemann, Maik; Bohn, Erwin; Fuchs, Helmut; Ollert, Markus; Gailus-Durner, Valérie; de Angelis, Martin Hrabĕ; Beckers, Johannes

    2015-01-01

    We established a selection strategy to identify new models for an altered airway inflammatory response from a large compendium of mutant mouse lines that were systemically phenotyped in the German Mouse Clinic (GMC). As selection criteria we included published gene functional data, as well as immunological and transcriptome data from GMC phenotyping screens under standard conditions. Applying these criteria we identified a few from several hundred mutant mouse lines and further characterized the Cox4i2tm1Hutt, Ifit2tm1.1Ebsb, and Prdm11tm1.1ahl lines following ovalbumin (OVA) sensitization and repeated OVA airway challenge. Challenged Prdm11tm1.1ahl mice exhibited changes in B cell counts, CD4+ T cell counts, and in the number of neutrophils in bronchoalveolar lavages, whereas challenged Ifit2tm1.1Ebsb mice displayed alterations in plasma IgE, IgG1, IgG3, and IgM levels compared to the challenged wild type littermates. In contrast, challenged Cox4i2tm1Hutt mutant mice did not show alterations in the humoral or cellular immune response compared to challenged wild type mice. Transcriptome analyses from lungs of the challenged mutant mouse lines showed extensive changes in gene expression in Prdm11tm1.1ahl mice. Functional annotations of regulated genes of all three mutant mouse lines were primarily related to inflammation and airway smooth muscle (ASM) remodeling. We were thus able to define an effective selection strategy to identify new candidate genes for the predisposition to an altered airway inflammatory response under OVA challenge conditions. Similar selection strategies may be used for the analysis of additional genotype-envirotype interactions for other diseases.

  7. Gene Expression Patterns in Ovarian Carcinomas

    PubMed Central

    Schaner, Marci E.; Ross, Douglas T.; Ciaravino, Giuseppe; Sørlie, Therese; Troyanskaya, Olga; Diehn, Maximilian; Wang, Yan C.; Duran, George E.; Sikic, Thomas L.; Caldeira, Sandra; Skomedal, Hanne; Tu, I-Ping; Hernandez-Boussard, Tina; Johnson, Steven W.; O'Dwyer, Peter J.; Fero, Michael J.; Kristensen, Gunnar B.; Børresen-Dale, Anne-Lise; Hastie, Trevor; Tibshirani, Robert; van de Rijn, Matt; Teng, Nelson N.; Longacre, Teri A.; Botstein, David; Brown, Patrick O.; Sikic, Branimir I.

    2003-01-01

    We used DNA microarrays to characterize the global gene expression patterns in surface epithelial cancers of the ovary. We identified groups of genes that distinguished the clear cell subtype from other ovarian carcinomas, grade I and II from grade III serous papillary carcinomas, and ovarian from breast carcinomas. Six clear cell carcinomas were distinguished from 36 other ovarian carcinomas (predominantly serous papillary) based on their gene expression patterns. The differences may yield insights into the worse prognosis and therapeutic resistance associated with clear cell carcinomas. A comparison of the gene expression patterns in the ovarian cancers to published data of gene expression in breast cancers revealed a large number of differentially expressed genes. We identified a group of 62 genes that correctly classified all 125 breast and ovarian cancer specimens. Among the best discriminators more highly expressed in the ovarian carcinomas were PAX8 (paired box gene 8), mesothelin, and ephrin-B1 (EFNB1). Although estrogen receptor was expressed in both the ovarian and breast cancers, genes that are coregulated with the estrogen receptor in breast cancers, including GATA-3, LIV-1, and X-box binding protein 1, did not show a similar pattern of coexpression in the ovarian cancers. PMID:12960427

  8. Arabidopsis gene expression patterns during spaceflight

    NASA Astrophysics Data System (ADS)

    Paul, A.-L.; Ferl, R. J.

    The exposure of Arabidopsis thaliana (Arabidopsis) plants to spaceflight environments resulted in the differential expression of hundreds of genes. A 5 day mission on orbiter Columbia in 1999 (STS-93) carried transgenic Arabidopsis plants engineered with a transgene composed of the alcohol dehydrogenase (Adh) gene promoter linked to the β -Glucuronidase (GUS) reporter gene. The plants were used to evaluate the effects of spaceflight on two fronts. First, expression patterns visualized with the Adh/GUS transgene were used to address specifically the possibility that spaceflight induces a hypoxic stress response, and to assess whether any spaceflight response was similar to control terrestrial hypoxia-induced gene expression patterns. (Paul et al., Plant Physiol. 2001, 126:613). Second, genome-wide patterns of native gene expression were evaluated utilizing the Affymetrix ATH1 GeneChip? array of 8,000 Arabidopsis genes. As a control for the veracity of the array analyses, a selection of genes identified with the arrays was further characterized with quantitative Real-Time RT PCR (ABI - TaqmanTM). Comparison of the patterns of expression for arrays of hybridized with RNA isolated from plants exposed to spaceflight compared to the control arrays revealed hundreds of genes that were differentially expressed in response to spaceflight, yet most genes that are hallmarks of hypoxic stress were unaffected. These results will be discussed in light of current models for plant responses to the spaceflight environment, and with regard to potential future flight opportunities.

  9. Overexpression of farnesoid X receptor in small airways contributes to epithelial to mesenchymal transition and COX-2 expression in chronic obstructive pulmonary disease

    PubMed Central

    Chen, Bi; You, Wen-Jie; Xue, Shan; Qin, Hui; Zhao, Xu-Ji; Zhang, Miao; Liu, Xue-Qing; Zhu, Shu-Yang

    2016-01-01

    Background Epithelial-mesenchymal transition (EMT) and cyclooxygenase-2 (COX-2) contribute to airway remodelling and inflammation in chronic obstructive pulmonary disease (COPD). Recent data suggest that the farnesoid X receptor (FXR), a nuclear receptor traditionally considered as bile acid-activated receptor, is also expressed in non-classical bile acids target tissues with novel functions beyond regulating bile acid homeostasis. This study aimed to investigate the potential role of FXR in the development of COPD, as well as factors that affect FXR expression. Methods Expression of FXR, EMT biomarkers and COX-2 was examined by immunohistochemistry in lung tissues from non-smokers, smokers, and smokers with COPD. The role of FXR in TGF-β1-induced EMT and COX-2 expression in human bronchial epithelial (HBE) cells was evaluated in vitro. Factors regulating FXR expression were assessed in cultured HBE cells and a cigarette smoke-induced rat model of COPD. Results Expression of FXR, EMT markers and COX-2 was significantly elevated in small airway epithelium of COPD patients compared with controls. The staining scores of FXR in small airway epithelium were negatively related with FEV1% of predicted of smokers without and with COPD. FXR agonist GW4064 remarkably enhanced and FXR antagonist Z-Guggulsterone significantly inhibited EMT changes in TGF-β1-treated HBE cells. Both chenodeoxycholic acid (CDCA) and GW4064 increased COX-2 expression in HBE cells, whereas Z-Guggulsterone dramatically restrained CDCA-induced COX-2 expression. Finally, FXR expression is induced by IL-4 and IL-13 in HBE cells, as well as by cigarette smoke exposure in a rat model of COPD. Conclusions Overexpression of FXR in small airway may contribute to airway remodelling and inflammation in COPD by regulating EMT and COX-2 expression. PMID:28066584

  10. Stratified gene expression analysis identifies major amyotrophic lateral sclerosis genes.

    PubMed

    Jones, Ashley R; Troakes, Claire; King, Andrew; Sahni, Vibhu; De Jong, Simone; Bossers, Koen; Papouli, Efterpi; Mirza, Muddassar; Al-Sarraj, Safa; Shaw, Christopher E; Shaw, Pamela J; Kirby, Janine; Veldink, Jan H; Macklis, Jeffrey D; Powell, John F; Al-Chalabi, Ammar

    2015-05-01

    Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease of motor neurons resulting in progressive paralysis. Gene expression studies of ALS only rarely identify the same gene pathways as gene association studies. We hypothesized that analyzing tissues by matching on degree of disease severity would identify different patterns of gene expression from a traditional case-control comparison. We analyzed gene expression changes in 4 postmortem central nervous system regions, stratified by severity of motor neuron loss. An overall comparison of cases (n = 6) and controls (n = 3) identified known ALS gene, SOX5, as showing differential expression (log2 fold change = 0.09, p = 5.5 × 10(-5)). Analyses stratified by disease severity identified expression changes in C9orf72 (p = 2.77 × 10(-3)), MATR3 (p = 3.46 × 10(-3)), and VEGFA (p = 8.21 × 10(-4)), all implicated in ALS through genetic studies, and changes in other genes in pathways involving RNA processing and immune response. These findings suggest that analysis of gene expression stratified by disease severity can identify major ALS genes and may be more efficient than traditional case-control comparison.

  11. Syk Regulates Neutrophilic Airway Hyper-Responsiveness in a Chronic Mouse Model of Allergic Airways Inflammation

    PubMed Central

    Juvet, Stephen; Scott, Jeremy A.; Chow, Chung-Wai

    2017-01-01

    Background Asthma is a chronic inflammatory disease characterized by airways hyper-responsiveness (AHR), reversible airway obstruction, and airway inflammation and remodeling. We previously showed that Syk modulates methacholine-induced airways contractility in naïve mice and in mice with allergic airways inflammation. We hypothesize that Syk plays a role in the pathogenesis of AHR; this was evaluated in a chronic 8-week mouse model of house dust mite (HDM)-induced allergic airways inflammation. Methods We used the Sykflox/flox//rosa26CreERT2 conditional Syk knock-out mice to assess the role of Syk prior to HDM exposure, and treated HDM-sensitized mice with the Syk inhibitor, GSK143, to evaluate its role in established allergic airways inflammation. Respiratory mechanics and methacholine (MCh)-responsiveness were assessed using the flexiVent® system. Lungs underwent bronchoalveolar lavage to isolate inflammatory cells or were frozen for determination of gene expression in tissues. Results MCh-induced AHR was observed following HDM sensitization in the Syk-intact (Sykflox/flox) and vehicle-treated BALB/c mice. MCh responsiveness was reduced to control levels in HDM-sensitized Sykdel/del mice and in BALB/c and Sykflox/flox mice treated with GSK143. Both Sykdel/del and GSK143-treated mice mounted appropriate immune responses to HDM, with HDM-specific IgE levels that were comparable to Sykflox/flox and vehicle-treated BALB/c mice. HDM-induced increases in bronchoalveolar lavage cell counts were attenuated in both Sykdel/del and GSK143-treated mice, due primarily to decreased neutrophil recruitment. Gene expression analysis of lung tissues revealed that HDM-induced expression of IL-17 and CXCL-1 was significantly attenuated in both Sykdel/del and GSK143-treated mice. Conclusion Syk inhibitors may play a role in the management of neutrophilic asthma. PMID:28107345

  12. Gene Expression Noise, Fitness Landscapes, and Evolution

    NASA Astrophysics Data System (ADS)

    Charlebois, Daniel

    The stochastic (or noisy) process of gene expression can have fitness consequences for living organisms. For example, gene expression noise facilitates the development of drug resistance by increasing the time scale at which beneficial phenotypic states can be maintained. The present work investigates the relationship between gene expression noise and the fitness landscape. By incorporating the costs and benefits of gene expression, we track how the fluctuation magnitude and timescale of expression noise evolve in simulations of cell populations under stress. We find that properties of expression noise evolve to maximize fitness on the fitness landscape, and that low levels of expression noise emerge when the fitness benefits of gene expression exceed the fitness costs (and that high levels of noise emerge when the costs of expression exceed the benefits). The findings from our theoretical/computational work offer new hypotheses on the development of drug resistance, some of which are now being investigated in evolution experiments in our laboratory using well-characterized synthetic gene regulatory networks in budding yeast. Nserc Postdoctoral Fellowship (Grant No. PDF-453977-2014).

  13. Gene expression in the etiology of schizophrenia.

    PubMed

    Bray, Nicholas J

    2008-05-01

    Gene expression represents a fundamental interface between genes and environment in the development and ongoing plasticity of the human brain. Individual differences in gene expression are likely to underpin much of human diversity, including psychiatric illness. In the past decade, the development of microarray and proteomic technology has enabled global description of gene expression in schizophrenia. However, it is difficult on the basis of gene expression assays alone to distinguish between those changes that constitute primary etiology and those that reflect secondary pathology, compensatory mechanisms, or confounding influences. In this respect, tests of genetic association with schizophrenia will be instructive because changes in gene expression that result from gene variants that are associated with the disorder are likely to be of primary etiological significance. However, regulatory polymorphism is extremely difficult to recognize on the basis of sequence interrogation alone. Functional assays at the messenger RNA and/or protein level will be essential in elucidating the molecular mechanisms underlying genetic association with schizophrenia and are likely to become increasingly important in the identification of regulatory variants with which to test for association with the disorder and related traits. Once established, etiologically relevant changes in gene expression can be recapitulated in model systems in order to elucidate the molecular and physiological pathways that may ultimately give rise to the condition.

  14. Noise minimisation in gene expression switches.

    PubMed

    Monteoliva, Diana; McCarthy, Christina B; Diambra, Luis

    2013-01-01

    Gene expression is subject to stochastic variation which leads to fluctuations in the rate of protein production. Recently, a study in yeast at a genomic scale showed that, in some cases, gene expression variability alters phenotypes while, in other cases, these remain unchanged despite fluctuations in the expression of other genes. These studies suggested that noise in gene expression is a physiologically relevant trait and, to prevent harmful stochastic variation in the expression levels of some genes, it can be subject to minimisation. However, the mechanisms for noise minimisation are still unclear. In the present work, we analysed how noise expression depends on the architecture of the cis-regulatory system, in particular on the number of regulatory binding sites. Using analytical calculations and stochastic simulations, we found that the fluctuation level in noise expression decreased with the number of regulatory sites when regulatory transcription factors interacted with only one other bound transcription factor. In contrast, we observed that there was an optimal number of binding sites when transcription factors interacted with many bound transcription factors. This finding suggested a new mechanism for preventing large fluctuations in the expression of genes which are sensitive to the concentration of regulators.

  15. Noise Minimisation in Gene Expression Switches

    PubMed Central

    Monteoliva, Diana; McCarthy, Christina B.; Diambra, Luis

    2013-01-01

    Gene expression is subject to stochastic variation which leads to fluctuations in the rate of protein production. Recently, a study in yeast at a genomic scale showed that, in some cases, gene expression variability alters phenotypes while, in other cases, these remain unchanged despite fluctuations in the expression of other genes. These studies suggested that noise in gene expression is a physiologically relevant trait and, to prevent harmful stochastic variation in the expression levels of some genes, it can be subject to minimisation. However, the mechanisms for noise minimisation are still unclear. In the present work, we analysed how noise expression depends on the architecture of the cis-regulatory system, in particular on the number of regulatory binding sites. Using analytical calculations and stochastic simulations, we found that the fluctuation level in noise expression decreased with the number of regulatory sites when regulatory transcription factors interacted with only one other bound transcription factor. In contrast, we observed that there was an optimal number of binding sites when transcription factors interacted with many bound transcription factors. This finding suggested a new mechanism for preventing large fluctuations in the expression of genes which are sensitive to the concentration of regulators. PMID:24376783

  16. Nucleosome repositioning underlies dynamic gene expression.

    PubMed

    Nocetti, Nicolas; Whitehouse, Iestyn

    2016-03-15

    Nucleosome repositioning at gene promoters is a fundamental aspect of the regulation of gene expression. However, the extent to which nucleosome repositioning is used within eukaryotic genomes is poorly understood. Here we report a comprehensive analysis of nucleosome positions as budding yeast transit through an ultradian cycle in which expression of >50% of all genes is highly synchronized. We present evidence of extensive nucleosome repositioning at thousands of gene promoters as genes are activated and repressed. During activation, nucleosomes are relocated to allow sites of general transcription factor binding and transcription initiation to become accessible. The extent of nucleosome shifting is closely related to the dynamic range of gene transcription and generally related to DNA sequence properties and use of the coactivators TFIID or SAGA. However, dynamic gene expression is not limited to SAGA-regulated promoters and is an inherent feature of most genes. While nucleosome repositioning occurs pervasively, we found that a class of genes required for growth experience acute nucleosome shifting as cells enter the cell cycle. Significantly, our data identify that the ATP-dependent chromatin-remodeling enzyme Snf2 plays a fundamental role in nucleosome repositioning and the expression of growth genes. We also reveal that nucleosome organization changes extensively in concert with phases of the cell cycle, with large, regularly spaced nucleosome arrays being established in mitosis. Collectively, our data and analysis provide a framework for understanding nucleosome dynamics in relation to fundamental DNA-dependent transactions.

  17. Protein tyrosine phosphatase SHP2 regulates TGF-β1 production in airway epithelia and asthmatic airway remodeling in mice

    PubMed Central

    Qin, X.-J.; Zhang, G.-S.; Zhang, X.; Qiu, Z.-W.; Wang, P.-L.; Li, Y.-W.; Li, W.; Xie, Q.-M.; Ke, Y.-H.; Lee, J. J.; Shen, H.-H.

    2014-01-01

    Background Transforming growth factor (TGF)-β1 produced in airway epithelia has been suggested as a contributor to the airway remodeling observed in asthma patients. The protein tyrosine phosphatase SHP2 is a demonstrable modulator of TGF-β1 production and thus a potential regulator of airway remodeling. Objectives To define the signal event by which SHP2 regulates asthmatic responses in airway epithelial cells by using a mouse model of experimental OVA-induced airway remodeling. Methods The airways of Shp2flox/flox mice were infected with recombinant adenovirus vectors expressing a Cre recombinase–green fluorescence protein (GFP) fusion protein as part of allergen provocation studies using mice sensitized with ovalbumin (OVA) and repeatedly challenged with OVA. Several endpoint pathologies were assessed, including airway hyper-responsiveness (AHR), lung inflammatory score, peribronchial collagen deposition, and α-smooth muscle actin (SMA) hyperplasia. In vitro studies using airway epithelial cells (BEAS-2B) were used to investigate the role of SHP2 in the regulation of pulmonary remodeling events, including the expression of collagen, α-SMA, and TGF-β1. Results Chronic OVA challenges in wild-type mice resulted in airway remodeling and lung dysfunction (e.g., increased inflammatory scores, collagen deposition (fibrosis), smooth muscle hyperplasia, and a significant increase in AHR). These endpoint pathology metrics were each significantly attenuated by conditional shp2 gene knockdown in airway epithelia. In vitro studies using BEAS-2B cells also demonstrated that the level of TGF-β1 production by these cells correlated with the extent of shp2 gene expression. Conclusions SHP2 activities in airway epithelial cells appear to modulate TGF-β1 production and, in turn, regulate allergic airway remodeling following allergen provocation. Clinical Implications Our findings identify SHP2 as a previously underappreciated contributor to the airway remodeling and lung

  18. Gene Expression Patterns in Human Liver Cancers

    PubMed Central

    Chen, Xin; Cheung, Siu Tim; So, Samuel; Fan, Sheung Tat; Barry, Christopher; Higgins, John; Lai, Kin-Man; Ji, Jiafu; Dudoit, Sandrine; Ng, Irene O.L.; van de Rijn, Matt; Botstein, David; Brown, Patrick O.

    2002-01-01

    Hepatocellular carcinoma (HCC) is a leading cause of death worldwide. Using cDNA microarrays to characterize patterns of gene expression in HCC, we found consistent differences between the expression patterns in HCC compared with those seen in nontumor liver tissues. The expression patterns in HCC were also readily distinguished from those associated with tumors metastatic to liver. The global gene expression patterns intrinsic to each tumor were sufficiently distinctive that multiple tumor nodules from the same patient could usually be recognized and distinguished from all the others in the large sample set on the basis of their gene expression patterns alone. The distinctive gene expression patterns are characteristic of the tumors and not the patient; the expression programs seen in clonally independent tumor nodules in the same patient were no more similar than those in tumors from different patients. Moreover, clonally related tumor masses that showed distinct expression profiles were also distinguished by genotypic differences. Some features of the gene expression patterns were associated with specific phenotypic and genotypic characteristics of the tumors, including growth rate, vascular invasion, and p53 overexpression. PMID:12058060

  19. Prostaglandin E2 induces expression of MAPK phosphatase 1 (MKP-1) in airway smooth muscle cells.

    PubMed

    Rumzhum, Nowshin N; Ammit, Alaina J

    2016-07-05

    Prostaglandin E2 (PGE2) is a prostanoid with diverse actions in health and disease. In chronic respiratory diseases driven by inflammation, PGE2 has both positive and negative effects. An enhanced understanding of the receptor-mediated cellular signalling pathways induced by PGE2 may help us separate the beneficial properties from unwanted actions of this important prostaglandin. PGE2 is known to exert anti-inflammatory and bronchoprotective actions in human airways. To date however, whether PGE2 increases production of the anti-inflammatory protein MAPK phosphatase 1 (MKP-1) was unknown. We address this herein and use primary cultures of human airway smooth muscle (ASM) cells to show that PGE2 increases MKP-1 mRNA and protein upregulation in a concentration-dependent manner. We explore the signalling pathways responsible and show that PGE2-induces CREB phosphorylation, not p38 MAPK activation, in ASM cells. Moreover, we utilize selective antagonists of EP2 (PF-04418948) and EP4 receptors (GW 627368X) to begin to identify EP-mediated functional outcomes in ASM cells in vitro. Taken together with earlier studies, our data suggest that PGE2 increases production of the anti-inflammatory protein MKP-1 via cAMP/CREB-mediated cellular signalling in ASM cells and demonstrates that EP2 may, in part, be involved.

  20. Digital gene expression signatures for maize development.

    PubMed

    Eveland, Andrea L; Satoh-Nagasawa, Namiko; Goldshmidt, Alexander; Meyer, Sandra; Beatty, Mary; Sakai, Hajime; Ware, Doreen; Jackson, David

    2010-11-01

    Genome-wide expression signatures detect specific perturbations in developmental programs and contribute to functional resolution of key regulatory networks. In maize (Zea mays) inflorescences, mutations in the RAMOSA (RA) genes affect the determinacy of axillary meristems and thus alter branching patterns, an important agronomic trait. In this work, we developed and tested a framework for analysis of tag-based, digital gene expression profiles using Illumina's high-throughput sequencing technology and the newly assembled B73 maize reference genome. We also used a mutation in the RA3 gene to identify putative expression signatures specific to stem cell fate in axillary meristem determinacy. The RA3 gene encodes a trehalose-6-phosphate phosphatase and may act at the interface between developmental and metabolic processes. Deep sequencing of digital gene expression libraries, representing three biological replicate ear samples from wild-type and ra3 plants, generated 27 million 20- to 21-nucleotide reads with frequencies spanning 4 orders of magnitude. Unique sequence tags were anchored to 3'-ends of individual transcripts by DpnII and NlaIII digests, which were multiplexed during sequencing. We mapped 86% of nonredundant signature tags to the maize genome, which associated with 37,117 gene models and unannotated regions of expression. In total, 66% of genes were detected by at least nine reads in immature maize ears. We used comparative genomics to leverage existing information from Arabidopsis (Arabidopsis thaliana) and rice (Oryza sativa) in functional analyses of differentially expressed maize genes. Results from this study provide a basis for the analysis of short-read expression data in maize and resolved specific expression signatures that will help define mechanisms of action for the RA3 gene.

  1. Gene expression homeostasis and chromosome architecture

    PubMed Central

    Seshasayee, Aswin Sai Narain

    2014-01-01

    In rapidly growing populations of bacterial cells, including those of the model organism Escherichia coli, genes essential for growth - such as those involved in protein synthesis - are expressed at high levels; this is in contrast to many horizontally-acquired genes, which are maintained at low transcriptional levels.1 This balance in gene expression states between 2 distinct classes of genes is established by a galaxy of transcriptional regulators, including the so-called nucleoid associated proteins (NAP) that contribute to shaping the chromosome.2 Besides these active players in gene regulation, it is not too far-fetched to anticipate that genome organization in terms of how genes are arranged on the chromosome,3 which is the result of long-drawn transactions among genome rearrangement processes and selection, and the manner in which it is structured inside the cell, plays a role in establishing this balance. A recent study from our group has contributed to the literature investigating the interplay between global transcriptional regulators and genome organization in establishing gene expression homeostasis.4 In particular, we address a triangle of functional interactions among genome organization, gene expression homeostasis and horizontal gene transfer. PMID:25997086

  2. Unmasking ultradian rhythms in gene expression

    PubMed Central

    van der Veen, Daan R.; Gerkema, Menno P.

    2017-01-01

    Biological oscillations with an ultradian time scale of 1 to several hours include cycles in behavioral arousal, episodic glucocorticoid release, and gene expression. Ultradian rhythms are thought to have an extrinsic origin because of a perceived absence of ultradian rhythmicity in vitro and a lack of known molecular ultradian oscillators. We designed a novel, non–spectral-analysis method of separating ultradian from circadian components and applied it to a published gene expression dataset with an ultradian sampling resolution. Ultradian rhythms in mouse hepatocytes in vivo have been published, and we validated our approach using this control by confirming 175 of 323 ultradian genes identified in a prior study and found 862 additional ultradian genes. For the first time, we now report ultradian expression of >900 genes in vitro. Sixty genes exhibited ultradian transcriptional rhythmicity, both in vivo and in vitro, including 5 genes involved in the cell cycle. Within these 60 genes, we identified significant enrichment of specific DNA motifs in the 1000 bp proximal promotor, some of which associate with known transcriptional factors. These findings are in strong support of instrinsically driven ultradian rhythms and expose potential molecular mechanisms and functions underlying ultradian rhythms that remain unknown.—Van der Veen, D. R., Gerkema, M. P. Unmasking ultradian rhythms in gene expression. PMID:27871062

  3. Expression of polarity genes in human cancer.

    PubMed

    Lin, Wan-Hsin; Asmann, Yan W; Anastasiadis, Panos Z

    2015-01-01

    Polarity protein complexes are crucial for epithelial apical-basal polarity and directed cell migration. Since alterations of these processes are common in cancer, polarity proteins have been proposed to function as tumor suppressors or oncogenic promoters. Here, we review the current understanding of polarity protein functions in epithelial homeostasis, as well as tumor formation and progression. As most previous studies focused on the function of single polarity proteins in simplified model systems, we used a genomics approach to systematically examine and identify the expression profiles of polarity genes in human cancer. The expression profiles of polarity genes were distinct in different human tissues and classified cancer types. Additionally, polarity expression profiles correlated with disease progression and aggressiveness, as well as with identified cancer types, where specific polarity genes were commonly altered. In the case of Scribble, gene expression analysis indicated its common amplification and upregulation in human cancer, suggesting a tumor promoting function.

  4. Regulation of Gene Expression in Protozoa Parasites

    PubMed Central

    Gomez, Consuelo; Esther Ramirez, M.; Calixto-Galvez, Mercedes; Medel, Olivia; Rodríguez, Mario A.

    2010-01-01

    Infections with protozoa parasites are associated with high burdens of morbidity and mortality across the developing world. Despite extensive efforts to control the transmission of these parasites, the spread of populations resistant to drugs and the lack of effective vaccines against them contribute to their persistence as major public health problems. Parasites should perform a strict control on the expression of genes involved in their pathogenicity, differentiation, immune evasion, or drug resistance, and the comprehension of the mechanisms implicated in that control could help to develop novel therapeutic strategies. However, until now these mechanisms are poorly understood in protozoa. Recent investigations into gene expression in protozoa parasites suggest that they possess many of the canonical machineries employed by higher eukaryotes for the control of gene expression at transcriptional, posttranscriptional, and epigenetic levels, but they also contain exclusive mechanisms. Here, we review the current understanding about the regulation of gene expression in Plasmodium sp., Trypanosomatids, Entamoeba histolytica and Trichomonas vaginalis. PMID:20204171

  5. Dynamic modeling of gene expression data

    NASA Technical Reports Server (NTRS)

    Holter, N. S.; Maritan, A.; Cieplak, M.; Fedoroff, N. V.; Banavar, J. R.

    2001-01-01

    We describe the time evolution of gene expression levels by using a time translational matrix to predict future expression levels of genes based on their expression levels at some initial time. We deduce the time translational matrix for previously published DNA microarray gene expression data sets by modeling them within a linear framework by using the characteristic modes obtained by singular value decomposition. The resulting time translation matrix provides a measure of the relationships among the modes and governs their time evolution. We show that a truncated matrix linking just a few modes is a good approximation of the full time translation matrix. This finding suggests that the number of essential connections among the genes is small.

  6. Mining Gene Expression Data of Multiple Sclerosis

    PubMed Central

    Zhu, Zhenli; Huang, Zhengliang; Li, Ke

    2014-01-01

    Objectives Microarray produces a large amount of gene expression data, containing various biological implications. The challenge is to detect a panel of discriminative genes associated with disease. This study proposed a robust classification model for gene selection using gene expression data, and performed an analysis to identify disease-related genes using multiple sclerosis as an example. Materials and methods Gene expression profiles based on the transcriptome of peripheral blood mononuclear cells from a total of 44 samples from 26 multiple sclerosis patients and 18 individuals with other neurological diseases (control) were analyzed. Feature selection algorithms including Support Vector Machine based on Recursive Feature Elimination, Receiver Operating Characteristic Curve, and Boruta algorithms were jointly performed to select candidate genes associating with multiple sclerosis. Multiple classification models categorized samples into two different groups based on the identified genes. Models’ performance was evaluated using cross-validation methods, and an optimal classifier for gene selection was determined. Results An overlapping feature set was identified consisting of 8 genes that were differentially expressed between the two phenotype groups. The genes were significantly associated with the pathways of apoptosis and cytokine-cytokine receptor interaction. TNFSF10 was significantly associated with multiple sclerosis. A Support Vector Machine model was established based on the featured genes and gave a practical accuracy of ∼86%. This binary classification model also outperformed the other models in terms of Sensitivity, Specificity and F1 score. Conclusions The combined analytical framework integrating feature ranking algorithms and Support Vector Machine model could be used for selecting genes for other diseases. PMID:24932510

  7. Intrapulmonary Versus Nasal Transduction of Murine Airways With GP64-pseudotyped Viral Vectors

    PubMed Central

    Oakland, Mayumi; Maury, Wendy; McCray, Paul B; Sinn, Patrick L

    2013-01-01

    Persistent viral vector-mediated transgene expression in the airways requires delivery to cells with progenitor capacity and avoidance of immune responses. Previously, we observed that GP64-pseudotyped feline immunodeficiency virus (FIV)-mediated gene transfer was more efficient in the nasal airways than the large airways of the murine lung. We hypothesized that in vivo gene transfer was limited by immunological and physiological barriers in the murine intrapulmonary airways. Here, we systematically investigate multiple potential barriers to lentiviral gene transfer in the airways of mice. We show that GP64-FIV vector transduced primary cultures of well-differentiated murine nasal epithelia with greater efficiency than primary cultures of murine tracheal epithelia. We further demonstrate that neutrophils, type I interferon (IFN) responses, as well as T and B lymphocytes are not the major factors limiting the transduction of murine conducting airways. In addition, we observed better transduction of GP64-pseudotyped vesicular stomatitis virus (VSV) in the nasal epithelia compared with the intrapulmonary airways in mice. VSVG glycoprotein pseudotyped VSV transduced intrapulmonary epithelia with similar efficiency as nasal epithelia. Our results suggest that the differential transduction efficiency of nasal versus intrapulmonary airways by FIV vector is not a result of immunological barriers or surface area, but rather differential expression of cellular factors specific for FIV vector transduction. PMID:23360952

  8. Amino acid regulation of gene expression.

    PubMed Central

    Fafournoux, P; Bruhat, A; Jousse, C

    2000-01-01

    The impact of nutrients on gene expression in mammals has become an important area of research. Nevertheless, the current understanding of the amino acid-dependent control of gene expression is limited. Because amino acids have multiple and important functions, their homoeostasis has to be finely maintained. However, amino-acidaemia can be affected by certain nutritional conditions or various forms of stress. It follows that mammals have to adjust several of their physiological functions involved in the adaptation to amino acid availability by regulating the expression of numerous genes. The aim of the present review is to examine the role of amino acids in regulating mammalian gene expression and protein turnover. It has been reported that some genes involved in the control of growth or amino acid metabolism are regulated by amino acid availability. For instance, limitation of several amino acids greatly increases the expression of the genes encoding insulin-like growth factor binding protein-1, CHOP (C/EBP homologous protein, where C/EBP is CCAAT/enhancer binding protein) and asparagine synthetase. Elevated mRNA levels result from both an increase in the rate of transcription and an increase in mRNA stability. Several observations suggest that the amino acid regulation of gene expression observed in mammalian cells and the general control process described in yeast share common features. Moreover, amino acid response elements have been characterized in the promoters of the CHOP and asparagine synthetase genes. Taken together, the results discussed in the present review demonstrate that amino acids, by themselves, can, in concert with hormones, play an important role in the control of gene expression. PMID:10998343

  9. Imputing gene expression to maximize platform compatibility.

    PubMed

    Zhou, Weizhuang; Han, Lichy; Altman, Russ B

    2017-02-15

    Microarray measurements of gene expression constitute a large fraction of publicly shared biological data, and are available in the Gene Expression Omnibus (GEO). Many studies use GEO data to shape hypotheses and improve statistical power. Within GEO, the Affymetrix HG-U133A and HG-U133 Plus 2.0 are the two most commonly used microarray platforms for human samples; the HG-U133 Plus 2.0 platform contains 54 220 probes and the HG-U133A array contains a proper subset (21 722 probes). When different platforms are involved, the subset of common genes is most easily compared. This approach results in the exclusion of substantial measured data and can limit downstream analysis. To predict the expression values for the genes unique to the HG-U133 Plus 2.0 platform, we constructed a series of gene expression inference models based on genes common to both platforms. Our model predicts gene expression values that are within the variability observed in controlled replicate studies and are highly correlated with measured data. Using six previously published studies, we also demonstrate the improved performance of the enlarged feature space generated by our model in downstream analysis.

  10. Optimal Reference Genes for Gene Expression Normalization in Trichomonas vaginalis.

    PubMed

    dos Santos, Odelta; de Vargas Rigo, Graziela; Frasson, Amanda Piccoli; Macedo, Alexandre José; Tasca, Tiana

    2015-01-01

    Trichomonas vaginalis is the etiologic agent of trichomonosis, the most common non-viral sexually transmitted disease worldwide. This infection is associated with several health consequences, including cervical and prostate cancers and HIV acquisition. Gene expression analysis has been facilitated because of available genome sequences and large-scale transcriptomes in T. vaginalis, particularly using quantitative real-time polymerase chain reaction (qRT-PCR), one of the most used methods for molecular studies. Reference genes for normalization are crucial to ensure the accuracy of this method. However, to the best of our knowledge, a systematic validation of reference genes has not been performed for T. vaginalis. In this study, the transcripts of nine candidate reference genes were quantified using qRT-PCR under different cultivation conditions, and the stability of these genes was compared using the geNorm and NormFinder algorithms. The most stable reference genes were α-tubulin, actin and DNATopII, and, conversely, the widely used T. vaginalis reference genes GAPDH and β-tubulin were less stable. The PFOR gene was used to validate the reliability of the use of these candidate reference genes. As expected, the PFOR gene was upregulated when the trophozoites were cultivated with ferrous ammonium sulfate when the DNATopII, α-tubulin and actin genes were used as normalizing gene. By contrast, the PFOR gene was downregulated when the GAPDH gene was used as an internal control, leading to misinterpretation of the data. These results provide an important starting point for reference gene selection and gene expression analysis with qRT-PCR studies of T. vaginalis.

  11. Perspectives: Gene Expression in Fisheries Management

    USGS Publications Warehouse

    Nielsen, Jennifer L.; Pavey, Scott A.

    2010-01-01

    Functional genes and gene expression have been connected to physiological traits linked to effective production and broodstock selection in aquaculture, selective implications of commercial fish harvest, and adaptive changes reflected in non-commercial fish populations subject to human disturbance and climate change. Gene mapping using single nucleotide polymorphisms (SNPs) to identify functional genes, gene expression (analogue microarrays and real-time PCR), and digital sequencing technologies looking at RNA transcripts present new concepts and opportunities in support of effective and sustainable fisheries. Genomic tools have been rapidly growing in aquaculture research addressing aspects of fish health, toxicology, and early development. Genomic technologies linking effects in functional genes involved in growth, maturation and life history development have been tied to selection resulting from harvest practices. Incorporating new and ever-increasing knowledge of fish genomes is opening a different perspective on local adaptation that will prove invaluable in wild fish conservation and management. Conservation of fish stocks is rapidly incorporating research on critical adaptive responses directed at the effects of human disturbance and climate change through gene expression studies. Genomic studies of fish populations can be generally grouped into three broad categories: 1) evolutionary genomics and biodiversity; 2) adaptive physiological responses to a changing environment; and 3) adaptive behavioral genomics and life history diversity. We review current genomic research in fisheries focusing on those that use microarrays to explore differences in gene expression among phenotypes and within or across populations, information that is critically important to the conservation of fish and their relationship to humans.

  12. Myofibroblast expression in airways and alveoli is affected by smoking and COPD

    PubMed Central

    2013-01-01

    Background Chronic obstructive pulmonary disease (COPD) is characterized by structural changes in alveoli and airways. Our aim was to analyse the numbers of alpha-smooth muscle actin (α-SMA) positive cells, as a marker of myofibroblasts, in different lung compartments in non-smokers and smokers with normal lung function or COPD. Methods α-SMA, tenascin-C (Tn-C) and EDA-fibronectin in alveolar level and airways were assayed by immunohistochemistry and quantified by image analysis. Immunohistochemical findings were correlated with clinical data. α-SMA protein was also analysed by Western blotting from fibroblastic cells cultured from peripheral lung of non-smokers, smokers without COPD and smokers with COPD. Results In many cases, the endings of the detached alveolar walls were widened, the structures of which were named as widened alveolar tips. Widened alveolar tips contained α-SMA positive cells, which were obviously myofibroblasts. There were less alveolar tips containing positive cells for α-SMA in alveoli and α-SMA positive cells in bronchioles in smokers and in COPD compared to non-smokers. The quantity of α-SMA positive cells was increased in bronchi in COPD. Tn-C was elevated in bronchi in COPD and smokers’ lung. The α-SMA protein level was 1.43-fold higher in stromal cells cultured from non-smokers than in those of smokers. Conclusions Myofibroblasts are localized variably in normal and diseased lung. This indicates that they have roles in both regeneration of lung and pathogenesis of COPD. The widened alveolar tips, these newly characterized histological structures, seemed to be the source of myofibroblasts at the alveolar level. PMID:23937155

  13. Airway epithelial cell wound repair mediated by alpha-dystroglycan.

    PubMed

    White, S R; Wojcik, K R; Gruenert, D; Sun, S; Dorscheid, D R

    2001-02-01

    Dystroglycans (DGs) bind laminin matrix proteins in skeletal and cardiac muscle and are expressed in other nonmuscle tissues. However, their expression in airway epithelial cells has not been demonstrated. We examined expression of DGs in the human airway epithelial cell line 1HAEo(-), and in human primary airway epithelial cells. Expression of the common gene for alpha- and beta-DG was demonstrated by reverse transcriptase/ polymerase chain reaction in 1HAEo(-) cells. Protein expression of beta-DG was demonstrated by both Western blot and flow cytometry in cultured cells. Localization of alpha-DG, using both a monoclonal antibody and the alpha-DG binding lectin wheat-germ agglutinin (WGA), was to the cell membrane and nucleus. We then examined the function of DGs in modulating wound repair over laminin matrix. Blocking alpha-DG binding to laminin in 1HAEo(-) monolayers using either glycosyaminoglycans or WGA attenuated cell migration and spreading after mechanical injury. alpha-DG was not expressed in epithelial cells at the wound edge immediately after wound creation, but localized to the cell membrane in these cells within 12 h of injury. These data demonstrate the presence of DGs in airway epithelium. alpha-DG is dynamically expressed and serves as a lectin to bind laminin during airway epithelial cell repair.

  14. CDK9-dependent transcriptional elongation in the innate interferon-stimulated gene response to respiratory syncytial virus infection in airway epithelial cells.

    PubMed

    Tian, Bing; Zhao, Yingxin; Kalita, Mridul; Edeh, Chukwudi B; Paessler, Slobodan; Casola, Antonella; Teng, Michael N; Garofalo, Roberto P; Brasier, Allan R

    2013-06-01

    Respiratory syncytial virus (RSV) is a negative-sense single-stranded RNA virus responsible for lower respiratory tract infections. During infection, the presence of double-stranded RNA (dsRNA) activates the interferon (IFN) regulatory factor 3 (IRF3) transcription factor, an event triggering expression of immediate early, IFN-stimulated genes (ISGs). We examine the role of transcriptional elongation in control of IRF3-dependent ISG expression. RSV infection induces ISG54, ISG56, and CIG5 gene expression in an IRF3-dependent manner demonstrated by IRF3 small interfering RNA (siRNA) silencing in both A549 epithelial cells and IRF3(-/-) MEFs. ISG expression was mediated by the recruitment of IRF3, CDK9, polymerase II (Pol II), and phospho-Ser(2) carboxy-terminal domain (CTD) Pol II to the IFN-stimulated response element (ISRE) binding sites of the IRF3-dependent ISG promoters in native chromatin. We find that RSV infection enhances the activated fraction of cyclin-dependent kinase 9 (CDK9) by promoting its association with bromodomain 4 (BRD4) and disrupting its association with the inhibitory 7SK small nuclear RNA. The requirement of CDK9 activity for ISG expression was shown by siRNA-mediated silencing of CDK9 and by a selective CDK9 inhibitor in A549 cells. In contrast, RSV-induced beta interferon (IFN-β) expression is not influenced by CDK9 inhibition. Using transcript-selective quantitative real-time reverse transcription-PCR (Q-RT-PCR) assays for the ISG54 gene, we observed that RSV induces transition from short to fully spliced mRNA transcripts and that this transition is blocked by CDK9 inhibition in both A549 and primary human small airway epithelial cells. These data indicate that transcription elongation plays a major role in RSV-induced ISG expression and is mediated by IRF3-dependent recruitment of activated CDK9. CDK9 activity may be a target for immunomodulation in RSV-induced lung disease.

  15. Airway epithelial IL-15 transforms monocytes into dendritic cells.

    PubMed

    Regamey, Nicolas; Obregon, Carolina; Ferrari-Lacraz, Sylvie; van Leer, Coretta; Chanson, Marc; Nicod, Laurent P; Geiser, Thomas

    2007-07-01

    IL-15 has recently been shown to induce the differentiation of functional dendritic cells (DCs) from human peripheral blood monocytes. Since DCs lay in close proximity to epithelial cells in the airway mucosa, we investigated whether airway epithelial cells release IL-15 in response to inflammatory stimuli and thereby induce differentiation and maturation of DCs. Alveolar (A549) and bronchial (BEAS-2B) epithelial cells produced IL-15 spontaneously and in a time- and dose-dependent manner after stimulation with IL-1beta, IFN-gamma, or TNF-alpha. Airway epithelial cell supernatants induced an increase of IL-15Ralpha gene expression in ex vivo monocytes, and stimulated DCs enhanced their IL-15Ralpha gene expression up to 300-fold. Airway epithelial cell-conditioned media induced the differentiation of ex vivo monocytes into partially mature DCs (HLA-DR+, DC-SIGN+, CD14+, CD80-, CD83+, CD86+, CCR3+, CCR6(+), CCR7-). Based on their phenotypic (CD123+, BDCA2+, BDCA4+, BDCA1(-), CD1a-) and functional properties (limited maturation upon stimulation with LPS and limited capacity to induce T cell proliferation), these DCs resembled plasmacytoid DCs. The effects of airway epithelial cell supernatants were largely blocked by a neutralizing monoclonal antibody to IL-15. Thus, our results demonstrate that airway epithelial cell-conditioned media have the capacity to differentiate monocytes into functional DCs, a process substantially mediated by epithelial-derived IL-15.

  16. Control of gene expression in trypanosomes.

    PubMed Central

    Vanhamme, L; Pays, E

    1995-01-01

    Trypanosomes are protozoan agents of major parasitic diseases such as Chagas' disease in South America and sleeping sickness of humans and nagana disease of cattle in Africa. They are transmitted to mammalian hosts by specific insect vectors. Their life cycle consists of a succession of differentiation and growth phases requiring regulated gene expression to adapt to the changing extracellular environment. Typical of such stage-specific expression is that of the major surface antigens of Trypanosoma brucei, procyclin in the procyclic (insect) form and the variant surface glycoprotein (VSG) in the bloodstream (mammalian) form. In trypanosomes, the regulation of gene expression is effected mainly at posttranscriptional levels, since primary transcription of most of the genes occurs in long polycistronic units and is constitutive. The transcripts are processed by transsplicing and polyadenylation under the influence of intergenic polypyrimidine tracts. These events show some developmental regulation. Untranslated sequences of the mRNAs seem to play a prominent role in the stage-specific control of individual gene expression, through a modulation of mRNA abundance. The VSG and procyclin transcription units exhibit particular features that are probably related to the need for a high level of expression. The promoters and RNA polymerase driving the expression of these units resemble those of the ribosomal genes. Their mutually exclusive expression is ensured by controls operating at several levels, including RNA elongation. Antigenic variation in the bloodstream is achieved through DNA rearrangements or alternative activation of the telomeric VSG gene expression sites. Recent discoveries, such as the existence of a novel nucleotide in telomeric DNA and the generation of point mutations in VSG genes, have shed new light on the mechanisms and consequences of antigenic variation. PMID:7603410

  17. Resource Sharing Controls Gene Expression Bursting.

    PubMed

    Caveney, Patrick M; Norred, S Elizabeth; Chin, Charles W; Boreyko, Jonathan B; Razooky, Brandon S; Retterer, Scott T; Collier, C Patrick; Simpson, Michael L

    2017-02-17

    Episodic gene expression, with periods of high expression separated by periods of no expression, is a pervasive biological phenomenon. This bursty pattern of expression draws from a finite reservoir of expression machinery in a highly time variant way, i.e., requiring no resources most of the time but drawing heavily on them during short intense bursts, that intimately links expression bursting and resource sharing. Yet, most recent investigations have focused on specific molecular mechanisms intrinsic to the bursty behavior of individual genes, while little is known about the interplay between resource sharing and global expression bursting behavior. Here, we confine Escherichia coli cell extract in both cell-sized microfluidic chambers and lipid-based vesicles to explore how resource sharing influences expression bursting. Interestingly, expression burst size, but not burst frequency, is highly sensitive to the size of the shared transcription and translation resource pools. The intriguing implication of these results is that expression bursts are more readily amplified than initiated, suggesting that burst formation occurs through positive feedback or cooperativity. When extrapolated to prokaryotic cells, these results suggest that large translational bursts may be correlated with large transcriptional bursts. This correlation is supported by recently reported transcription and translation bursting studies in E. coli. The results reported here demonstrate a strong intimate link between global expression burst patterns and resource sharing, and they suggest that bursting plays an important role in optimizing the use of limited, shared expression resources.

  18. Application of multidisciplinary analysis to gene expression.

    SciTech Connect

    Wang, Xuefel; Kang, Huining; Fields, Chris; Cowie, Jim R.; Davidson, George S.; Haaland, David Michael; Sibirtsev, Valeriy; Mosquera-Caro, Monica P.; Xu, Yuexian; Martin, Shawn Bryan; Helman, Paul; Andries, Erik; Ar, Kerem; Potter, Jeffrey; Willman, Cheryl L.; Murphy, Maurice H.

    2004-01-01

    Molecular analysis of cancer, at the genomic level, could lead to individualized patient diagnostics and treatments. The developments to follow will signal a significant paradigm shift in the clinical management of human cancer. Despite our initial hopes, however, it seems that simple analysis of microarray data cannot elucidate clinically significant gene functions and mechanisms. Extracting biological information from microarray data requires a complicated path involving multidisciplinary teams of biomedical researchers, computer scientists, mathematicians, statisticians, and computational linguists. The integration of the diverse outputs of each team is the limiting factor in the progress to discover candidate genes and pathways associated with the molecular biology of cancer. Specifically, one must deal with sets of significant genes identified by each method and extract whatever useful information may be found by comparing these different gene lists. Here we present our experience with such comparisons, and share methods developed in the analysis of an infant leukemia cohort studied on Affymetrix HG-U95A arrays. In particular, spatial gene clustering, hyper-dimensional projections, and computational linguistics were used to compare different gene lists. In spatial gene clustering, different gene lists are grouped together and visualized on a three-dimensional expression map, where genes with similar expressions are co-located. In another approach, projections from gene expression space onto a sphere clarify how groups of genes can jointly have more predictive power than groups of individually selected genes. Finally, online literature is automatically rearranged to present information about genes common to multiple groups, or to contrast the differences between the lists. The combination of these methods has improved our understanding of infant leukemia. While the complicated reality of the biology dashed our initial, optimistic hopes for simple answers from

  19. Modeling gene expression in time and space.

    PubMed

    Rué, Pau; Garcia-Ojalvo, Jordi

    2013-01-01

    Cell populations rarely exhibit gene-expression profiles that are homogeneous in time and space. In the temporal domain, dynamical behaviors such as oscillations and pulses of protein production pervade cell biology, underlying phenomena as diverse as circadian rhythmicity, cell cycle control, stress and damage responses, and stem-cell pluripotency. In multicellular populations, spatial heterogeneities are crucial for decision making and development, among many other functions. Cells need to exquisitely coordinate this temporal and spatial variation to survive. Although the spatiotemporal character of gene expression is challenging to quantify experimentally at the level of individual cells, it is beneficial from the modeling viewpoint, because it provides strong constraints that can be probed by theoretically analyzing mathematical models of candidate gene and protein circuits. Here, we review recent examples of temporal dynamics and spatial patterning in gene expression to show how modeling such phenomenology can help us unravel the molecular mechanisms of cellular function.

  20. Chemically regulated gene expression in plants.

    PubMed

    Padidam, Malla

    2003-04-01

    Chemically inducible systems that activate or inactivate gene expression have many potential applications in the determination of gene function and in plant biotechnology. The precise timing and control of gene expression are important aspects of chemically inducible systems. Several systems have been developed and used to analyze gene function, marker-free plant transformation, site-specific DNA excision, activation tagging, conditional genetic complementation, and restoration of male fertility. Chemicals that are used to regulate transgene expression include the antibiotic tetracycline, the steroids dexamethasone and estradiol, copper, ethanol, the inducer of pathogen-related proteins benzothiadiazol, herbicide safeners, and the insecticide methoxyfenozide. Systems that are suitable for field application are particularly useful for experimental systems and have potential applications in biotechnology.

  1. CIRCADIAN CLOCK AND CELL CYCLE GENE EXPRESSION

    PubMed Central

    Metz, Richard P.; Qu, Xiaoyu; Laffin, Brian; Earnest, David; Porter, Weston W.

    2009-01-01

    Mouse mammary epithelial cells (HC-11) and mammary tissues were analyzed for developmental changes in circadian clock, cellular proliferation and differentiation marker genes. Expression of the clock genes, Per1 and Bmal1, were elevated in differentiated HC-11 cells whereas Per2 mRNA levels were higher in undifferentiated cells. This differentiation-dependent profile of clock gene expression was consistent with that observed in mouse mammary glands as Per1 and Bmal1 mRNA levels were elevated in late pregnant and lactating mammary tissues, while Per2 expression was higher in proliferating virgin and early pregnant glands. In both HC-11 cells and mammary glands, elevated Per2 expression was positively correlated with c-Myc and Cyclin D1 mRNA levels while Per1 and Bmal1 expression changed in conjunction with ß-casein mRNA levels. Interestingly, developmental stage had differential effects on rhythms of clock gene expression in the mammary gland. These data suggest that circadian clock genes may play a role in mouse mammary gland development and differentiation. PMID:16261617

  2. Paternally expressed genes predominate in the placenta.

    PubMed

    Wang, Xu; Miller, Donald C; Harman, Rebecca; Antczak, Douglas F; Clark, Andrew G

    2013-06-25

    The discovery of genomic imprinting through studies of manipulated mouse embryos indicated that the paternal genome has a major influence on placental development. However, previous research has not demonstrated paternal bias in imprinted genes. We applied RNA sequencing to trophoblast tissue from reciprocal hybrids of horse and donkey, where genotypic differences allowed parent-of-origin identification of most expressed genes. Using this approach, we identified a core group of 15 ancient imprinted genes, of which 10 were paternally expressed. An additional 78 candidate imprinted genes identified by RNA sequencing also showed paternal bias. Pyrosequencing was used to confirm the imprinting status of six of the genes, including the insulin receptor (INSR), which may play a role in growth regulation with its reciprocally imprinted ligand, histone acetyltransferase-1 (HAT1), a gene involved in chromatin modification, and lymphocyte antigen 6 complex, locus G6C, a newly identified imprinted gene in the major histocompatibility complex. The 78 candidate imprinted genes displayed parent-of-origin expression bias in placenta but not fetus, and most showed less than 100% silencing of the imprinted allele. Some displayed variability in imprinting status among individuals. This variability results in a unique epigenetic signature for each placenta that contributes to variation in the intrauterine environment and thus presents the opportunity for natural selection to operate on parent-of-origin differential regulation. Taken together, these features highlight the plasticity of imprinting in mammals and the central importance of the placenta as a target tissue for genomic imprinting.

  3. Hepatic Xenobiotic Metabolizing Enzyme Gene Expression ...

    EPA Pesticide Factsheets

    BACKGROUND: Differences in responses to environmental chemicals and drugs between life stages are likely due in part to differences in the expression of xenobiotic metabolizing enzymes and transporters (XMETs). No comprehensive analysis of the mRNA expression of XMETs has been carried out through life stages in any species. RESULTS: Using full-genome arrays, the mRNA expression of all XMETs and their regulatory proteins was examined during fetal (gestation day (GD) 19), neonatal (postnatal day (PND) 7), prepubescent (PND32), middle age (12 months), and old age (18 and 24 months) in the C57BL/6J (C57) mouse liver and compared to adults. Fetal and neonatal life stages exhibited dramatic differences in XMET mRNA expression compared to the relatively minor effects of old age. The total number of XMET probe sets that differed from adults was 636, 500, 84, 5, 43, and 102 for GD19, PND7, PND32, 12 months, 18 months and 24 months, respectively. At all life stages except PND32, under-expressed genes outnumbered over-expressed genes. The altered XMETs included those in all of the major metabolic and transport phases including introduction of reactive or polar groups (Phase I), conjugation (Phase II) and excretion (Phase III). In the fetus and neonate, parallel increases in expression were noted in the dioxin receptor, Nrf2 components and their regulated genes while nuclear receptors and regulated genes were generally down-regulated. Suppression of male-specific XMETs w

  4. Three gene expression vector sets for concurrently expressing multiple genes in Saccharomyces cerevisiae.

    PubMed

    Ishii, Jun; Kondo, Takashi; Makino, Harumi; Ogura, Akira; Matsuda, Fumio; Kondo, Akihiko

    2014-05-01

    Yeast has the potential to be used in bulk-scale fermentative production of fuels and chemicals due to its tolerance for low pH and robustness for autolysis. However, expression of multiple external genes in one host yeast strain is considerably labor-intensive due to the lack of polycistronic transcription. To promote the metabolic engineering of yeast, we generated systematic and convenient genetic engineering tools to express multiple genes in Saccharomyces cerevisiae. We constructed a series of multi-copy and integration vector sets for concurrently expressing two or three genes in S. cerevisiae by embedding three classical promoters. The comparative expression capabilities of the constructed vectors were monitored with green fluorescent protein, and the concurrent expression of genes was monitored with three different fluorescent proteins. Our multiple gene expression tool will be helpful to the advanced construction of genetically engineered yeast strains in a variety of research fields other than metabolic engineering.

  5. Strain-Dependent Genomic Factors Affect Allergen-Induced Airway Hyperresponsiveness in Mice

    PubMed Central

    Kelada, Samir N. P.; Wilson, Mark S.; Tavarez, Urraca; Kubalanza, Kari; Borate, Bhavesh; Whitehead, Greg S.; Maruoka, Shuichiro; Roy, Michelle G.; Olive, Michelle; Carpenter, Danielle E.; Brass, David M.; Wynn, Thomas A.; Cook, Donald N.; Evans, Christopher M.; Schwartz, David A.

    2011-01-01

    Asthma is etiologically and clinically heterogeneous, making the genomic basis of asthma difficult to identify. We exploited the strain-dependence of a murine model of allergic airway disease to identify different genomic responses in the lung. BALB/cJ and C57BL/6J mice were sensitized with the immunodominant allergen from the Dermatophagoides pteronyssinus species of house dust mite (Der p 1), without exogenous adjuvant, and the mice then underwent a single challenge with Der p 1. Allergic inflammation, serum antibody titers, mucous metaplasia, and airway hyperresponsiveness were evaluated 72 hours after airway challenge. Whole-lung gene expression analyses were conducted to identify genomic responses to allergen challenge. Der p 1–challenged BALB/cJ mice produced all the key features of allergic airway disease. In comparison, C57BL/6J mice produced exaggerated Th2-biased responses and inflammation, but exhibited an unexpected decrease in airway hyperresponsiveness compared with control mice. Lung gene expression analysis revealed genes that were shared by both strains and a set of down-regulated genes unique to C57BL/6J mice, including several G-protein–coupled receptors involved in airway smooth muscle contraction, most notably the M2 muscarinic receptor, which we show is expressed in airway smooth muscle and was decreased at the protein level after challenge with Der p 1. Murine strain–dependent genomic responses in the lung offer insights into the different biological pathways that develop after allergen challenge. This study of two different murine strains demonstrates that inflammation and airway hyperresponsiveness can be decoupled, and suggests that the down-modulation of expression of G-protein–coupled receptors involved in regulating airway smooth muscle contraction may contribute to this dissociation. PMID:21378263

  6. SEGEL: A Web Server for Visualization of Smoking Effects on Human Lung Gene Expression.

    PubMed

    Xu, Yan; Hu, Brian; Alnajm, Sammy S; Lu, Yin; Huang, Yangxin; Allen-Gipson, Diane; Cheng, Feng

    2015-01-01

    Cigarette smoking is a major cause of death worldwide resulting in over six million deaths per year. Cigarette smoke contains complex mixtures of chemicals that are harmful to nearly all organs of the human body, especially the lungs. Cigarette smoking is considered the major risk factor for many lung diseases, particularly chronic obstructive pulmonary diseases (COPD) and lung cancer. However, the underlying molecular mechanisms of smoking-induced lung injury associated with these lung diseases still remain largely unknown. Expression microarray techniques have been widely applied to detect the effects of smoking on gene expression in different human cells in the lungs. These projects have provided a lot of useful information for researchers to understand the potential molecular mechanism(s) of smoke-induced pathogenesis. However, a user-friendly web server that would allow scientists to fast query these data sets and compare the smoking effects on gene expression across different cells had not yet been established. For that reason, we have integrated eight public expression microarray data sets from trachea epithelial cells, large airway epithelial cells, small airway epithelial cells, and alveolar macrophage into an online web server called SEGEL (Smoking Effects on Gene Expression of Lung). Users can query gene expression patterns across these cells from smokers and nonsmokers by gene symbols, and find the effects of smoking on the gene expression of lungs from this web server. Sex difference in response to smoking is also shown. The relationship between the gene expression and cigarette smoking consumption were calculated and are shown in the server. The current version of SEGEL web server contains 42,400 annotated gene probe sets represented on the Affymetrix Human Genome U133 Plus 2.0 platform. SEGEL will be an invaluable resource for researchers interested in the effects of smoking on gene expression in the lungs. The server also provides useful information

  7. Expression of myriapod pair rule gene orthologs

    PubMed Central

    2011-01-01

    Background Segmentation is a hallmark of the arthropods; most knowledge about the molecular basis of arthropod segmentation comes from work on the fly Drosophila melanogaster. In this species a hierarchic cascade of segmentation genes subdivides the blastoderm stepwise into single segment wide regions. However, segmentation in the fly is a derived feature since all segments form virtually simultaneously. Conversely, in the vast majority of arthropods the posterior segments form one at a time from a posterior pre-segmental zone. The pair rule genes (PRGs) comprise an important level of the Drosophila segmentation gene cascade and are indeed the first genes that are expressed in typical transverse stripes in the early embryo. Information on expression and function of PRGs outside the insects, however, is scarce. Results Here we present the expression of the pair rule gene orthologs in the pill millipede Glomeris marginata (Myriapoda: Diplopoda). We find evidence that these genes are involved in segmentation and that components of the hierarchic interaction of the gene network as found in insects may be conserved. We further provide evidence that segments are formed in a single-segment periodicity rather than in pairs of two like in another myriapod, the centipede Strigamia maritima. Finally we show that decoupling of dorsal and ventral segmentation in Glomeris appears already at the level of the PRGs. Conclusions Although the pair rule gene network is partially conserved among insects and myriapods, some aspects of PRG interaction are, as suggested by expression pattern analysis, convergent, even within the Myriapoda. Conserved expression patterns of PRGs in insects and myriapods, however, may represent ancestral features involved in segmenting the arthropod ancestor. PMID:21352542

  8. Selective prostacyclin receptor agonism augments glucocorticoid-induced gene expression in human bronchial epithelial cells.

    PubMed

    Wilson, Sylvia M; Shen, Pamela; Rider, Christopher F; Traves, Suzanne L; Proud, David; Newton, Robert; Giembycz, Mark A

    2009-11-15

    Prostacyclin receptor (IP-receptor) agonists display anti-inflammatory and antiviral activity in cell-based assays and in preclinical models of asthma and chronic obstructive pulmonary disease. In this study, we have extended these observations by demonstrating that IP-receptor activation also can enhance the ability of glucocorticoids to induce genes with anti-inflammatory activity. BEAS-2B bronchial epithelial cells stably transfected with a glucocorticoid response element (GRE) luciferase reporter were activated in a concentration-dependent manner by the glucocorticoid dexamethasone. An IP-receptor agonist, taprostene, increased cAMP in these cells and augmented luciferase expression at all concentrations of dexamethasone examined. Analysis of the concentration-response relationship that described this effect showed that taprostene increased the magnitude of transcription without affecting the potency of dexamethasone and was, thus, steroid-sparing in this simple system. RO3244794, an IP-receptor antagonist, and oligonucleotides that selectively silenced the IP-receptor gene, PTGIR, abolished these effects of taprostene. Infection of BEAS-2B GRE reporter cells with an adenovirus vector encoding a highly selective inhibitor of cAMP-dependent protein kinase (PKA) also prevented taprostene from enhancing GRE-dependent transcription. In BEAS-2B cells and primary cultures of human airway epithelial cells, taprostene and dexamethasone interacted either additively or cooperatively in the expression of three glucocorticoid-inducible genes (GILZ, MKP-1, and p57(kip2)) that have anti-inflammatory potential. Collectively, these data show that IP-receptor agonists can augment the ability of glucocorticoids to induce anti-inflammatory genes in human airway epithelial cells by activating a cAMP/PKA-dependent mechanism. This observation may have clinical relevance in the treatment of airway inflammatory diseases that are either refractory or respond suboptimally to

  9. Human AZU-1 gene, variants thereof and expressed gene products

    DOEpatents

    Chen, Huei-Mei; Bissell, Mina

    2004-06-22

    A human AZU-1 gene, mutants, variants and fragments thereof. Protein products encoded by the AZU-1 gene and homologs encoded by the variants of AZU-1 gene acting as tumor suppressors or markers of malignancy progression and tumorigenicity reversion. Identification, isolation and characterization of AZU-1 and AZU-2 genes localized to a tumor suppressive locus at chromosome 10q26, highly expressed in nonmalignant and premalignant cells derived from a human breast tumor progression model. A recombinant full length protein sequences encoded by the AZU-1 gene and nucleotide sequences of AZU-1 and AZU-2 genes and variant and fragments thereof. Monoclonal or polyclonal antibodies specific to AZU-1, AZU-2 encoded protein and to AZU-1, or AZU-2 encoded protein homologs.

  10. Rubisco gene expression in C4 plants.

    PubMed

    Patel, Minesh; Berry, James O

    2008-01-01

    In leaves of most C(4) plants, ribulose 1,5 bisphosphate carboxylase (Rubisco) accumulates only in bundle sheath (bs) cells that surround the vascular centres, and not in mesophyll (mp) cells. It has been shown previously that in the C(4) dicots amaranth and Flaveria bidentis, post-transcriptional control of mRNA translation and stability mediate the C(4) expression patterns of genes encoding the large and small Rubisco subunits (chloroplast rbcL and nuclear RbcS, respectively). Translational control appears to regulate bs cell-specific Rubisco gene expression during early dicot leaf development, while control of mRNA stability appears to mediate bs-specific accumulation of RbcS and rbcL transcripts in mature leaves. Post-transcriptional control is also involved in the regulation of Rubisco gene expression by light, and in response to photosynthetic activity. Transgenic and transient expression studies in F. bidentis provide direct evidence for post-transcriptional control of bs cell-specific RbcS expression, which is mediated by the 5' and 3' untranslated regions (UTRs) of the mRNA. Comparisons of Rubisco gene expression in these dicots and in the monocot maize indicates possible commonalities in the regulation of RbcS and rbcL genes in these divergent C(4) species. Now that the role of post-transcriptional regulation in C(4) gene expression has been established, it is likely that future studies of mRNA-protein interactions will address long-standing questions about the establishment and maintenance of cell type-specificity in these plants. Some of these regulatory mechanisms may have ancestral origins in C(3) species, through modification of pre-existing factors, or by the acquisition of novel C(4) processes.

  11. Alternative-splicing-mediated gene expression

    NASA Astrophysics Data System (ADS)

    Wang, Qianliang; Zhou, Tianshou

    2014-01-01

    Alternative splicing (AS) is a fundamental process during gene expression and has been found to be ubiquitous in eukaryotes. However, how AS impacts gene expression levels both quantitatively and qualitatively remains to be fully explored. Here, we analyze two common models of gene expression, each incorporating a simple splice mechanism that a pre-mRNA is spliced into two mature mRNA isoforms in a probabilistic manner. In the constitutive expression case, we show that the steady-state molecular numbers of two mature mRNA isoforms follow mutually independent Poisson distributions. In the bursting expression case, we demonstrate that the tail decay of the steady-state distribution for both mature mRNA isoforms that in general are not mutually independent can be characterized by the product of mean burst size and splicing probability. In both cases, we find that AS can efficiently modulate both the variability (measured by variance) and the noise level of the total mature mRNA, and in particular, the latter is always lower than the noise level of the pre-mRNA, implying that AS always reduces the noise. These results altogether reveal that AS is a mechanism of efficiently controlling the gene expression noise.

  12. The ΔF508-CFTR mutation inhibits wild-type CFTR processing and function when co-expressed in human airway epithelia and in mouse nasal mucosa

    PubMed Central

    2012-01-01

    Background Rescue or correction of CFTR function in native epithelia is the ultimate goal of CF therapeutics development. Wild-type (WT) CFTR introduction and replacement is also of particular interest. Such therapies may be complicated by possible CFTR self-assembly into an oligomer or multimer. Results Surprisingly, functional CFTR assays in native airway epithelia showed that the most common CFTR mutant, ΔF508-CFTR (ΔF-CFTR), inhibits WT-CFTR when both forms are co-expressed. To examine more mechanistically, both forms of CFTR were transfected transiently in varying amounts into IB3-1 CF human airway epithelial cells and HEK-293 human embryonic kidney cells null for endogenous CFTR protein expression. Increasing amounts of ΔF-CFTR inhibited WT-CFTR protein processing and function in CF human airway epithelial cells but not in heterologous HEK-293 cells. Stably expressed ΔF-CFTR in clones of the non-CF human airway epithelial cell line, CALU-3, also showed reduction in cAMP-stimulated anion secretion and in WT-CFTR processing. An ultimate test of this dominant negative-like effect of ΔF-CFTR on WT-CFTR was the parallel study of two different CF mouse models: the ΔF-CFTR mouse and the bitransgenic CFTR mouse corrected in the gut but null in the lung and airways. WT/ΔF heterozygotes had an intermediate phenotype with regard to CFTR agonist responses in in vivo nasal potential difference (NPD) recordings and in Ussing chamber recordings of short-circuit current (ISC) in vitro on primary tracheal epithelial cells isolated from the same mice. In contrast, CFTR bitransgenic +/− heterozygotes had no difference in their responses versus +/+ wild-type mice. Conclusions Taken altogether, these data suggest that ΔF-CFTR and WT-CFTR co-assemble into an oligomeric macromolecular complex in native epithelia and share protein processing machinery and regulation at the level of the endoplasmic reticulum (ER). As a consequence, ΔF-CFTR slows WT-CFTR protein processing

  13. Airway arginase expression and Nω-hydroxy-nor-arginine effect on methacholine-induced bronchoconstriction differentiate Lewis and Fischer rat strains.

    PubMed

    Risse, Paul-André; Lavoie-Lamoureux, Anouk; Jo, Taisuke; Tsuchiya, Kimitake; Siddiqui, Sana; Martin, James G

    2014-03-15

    Innate airway hyperresponsiveness (AHR) is well modeled by two strains of rat, the hyperresponsive Fischer 344 rat and the normoresponsive Lewis rat. Arginase has been implicated in AHR associated with allergic asthma models. We addressed the role of arginase in innate AHR using the Fischer-Lewis model. In vivo arginase inhibition with N(ω)-hydroxy-nor-arginine (nor-NOHA) was evaluated on methacholine-induced bronchoconstriction in the Fischer and the Lewis rats. Arginase activity and mRNA expression were quantified in structural and resident cells of the proximal airway tree. The effect of nor-NOHA was evaluated on cultured tracheal smooth muscle proliferation. Fischer rats exhibited significantly greater changes in respiratory resistance and elastance in response to methacholine compared with Lewis rats. nor-NOHA reduced the methacholine-induced bronchoconstriction in the central airways of Lewis rats, while it did not change the innate AHR of Fischer rats. Lewis rats exhibited greater arginase activity in tracheal smooth muscle but a lower proliferation rate compared with Fischer rats. Smooth muscle proliferation was not affected by nor-NOHA in either strain of rats. The strain-specific arginase expression in the smooth muscle may contribute to the differences in sensitivity of the methacholine challenged airways of Lewis and Fischer rats to inhibition of arginase.

  14. Gene expression profiles in irradiated cancer cells

    NASA Astrophysics Data System (ADS)

    Minafra, L.; Bravatà, V.; Russo, G.; Ripamonti, M.; Gilardi, M. C.

    2013-07-01

    Knowledge of the molecular and genetic mechanisms underlying cellular response to radiation may provide new avenues to develop innovative predictive tests of radiosensitivity of tumours and normal tissues and to improve individual therapy. Nowadays very few studies describe molecular changes induced by hadrontherapy treatments, therefore this field has to be explored and clarified. High-throughput methodologies, such as DNA microarray, allow us to analyse mRNA expression of thousands of genes simultaneously in order to discover new genes and pathways as targets of response to hadrontherapy. Our aim is to elucidate the molecular networks involved in the sensitivity/resistance of cancer cell lines subjected to hadrontherapy treatments with a genomewide approach by using cDNA microarray technology to identify gene expression profiles and candidate genes responsible of differential cellular responses.

  15. Gene expression profiles in irradiated cancer cells

    SciTech Connect

    Minafra, L.; Bravatà, V.; Russo, G.; Ripamonti, M.; Gilardi, M. C.

    2013-07-26

    Knowledge of the molecular and genetic mechanisms underlying cellular response to radiation may provide new avenues to develop innovative predictive tests of radiosensitivity of tumours and normal tissues and to improve individual therapy. Nowadays very few studies describe molecular changes induced by hadrontherapy treatments, therefore this field has to be explored and clarified. High-throughput methodologies, such as DNA microarray, allow us to analyse mRNA expression of thousands of genes simultaneously in order to discover new genes and pathways as targets of response to hadrontherapy. Our aim is to elucidate the molecular networks involved in the sensitivity/resistance of cancer cell lines subjected to hadrontherapy treatments with a genomewide approach by using cDNA microarray technology to identify gene expression profiles and candidate genes responsible of differential cellular responses.

  16. Trachea Epithelium as a “Canary” for Cigarette Smoking-induced Biologic Phenotype of the Small Airway Epithelium*

    PubMed Central

    Turetz, Meredith L.; O’Connor, Timothy P.; Tilley, Ann E.; Strulovici-Barel, Yael; Salit, Jacqueline; Dang, David; Teater, Matthew; Mezey, Jason; Clark, Andrew G.; Crystal, Ronald G.

    2013-01-01

    The initial site of smoking-induced lung disease is the small airway epithelium, which is difficult and time consuming to sample by fiberoptic bronchoscopy. We developed a rapid, office-based procedure to obtain trachea epithelium without conscious sedation from healthy nonsmokers (n=26) and healthy smokers (n=19, 27 ± 15 pack-yr). Gene expression differences (fold-change >1.5, p<0.01, Benjamini-Hochberg correction) were assessed with Affymetrix microarrays. 1,057 probe sets were differentially expressed in healthy smokers vs nonsmokers, representing >500 genes. Trachea gene expression was compared to an independent group of small airway epithelial samples (n=23 healthy nonsmokers, n=19 healthy smokers, 25 ± 12 pack-yr). The trachea epithelium is more sensitive to smoking, responding with 3-fold more differentially-expressed genes than small airway epithelium. The trachea transcriptome paralleled the small airway epithelium, with 156 of 167 (93%) genes that are significantly upand down-regulated by smoking in the small airway epithelium showing similar direction and magnitude of response to smoking in the trachea. Trachea epithelium can be obtained without conscious sedation, representing a less invasive surrogate “canary” for smoking-induced changes in the small airway epithelium. This should prove useful in epidemiologic studies correlating gene expression with clinical outcome in assessing smoking-induced lung disease. PMID:20443905

  17. Leukotriene E4 is a full functional agonist for human cysteinyl leukotriene type 1 receptor-dependent gene expression

    PubMed Central

    Foster, Holly R.; Fuerst, Elisabeth; Branchett, William; Lee, Tak H.; Cousins, David J.; Woszczek, Grzegorz

    2016-01-01

    Leukotriene E4 (LTE4) the most stable of the cysteinyl leukotrienes (cysLTs) binds poorly to classical type 1 (CysLT1) and 2 (CysLT2) receptors although it induces potent responses in human airways in vivo, such as bronchoconstriction, airway hyperresponsiveness and inflammatory cell influx suggesting the presence of a novel receptor that preferentially responds to LTE4. To identify such a receptor two human mast cell lines, LAD2 and LUVA, were selected that differentially responded to LTE4 when analysed by intracellular signalling and gene expression. Comparative transcriptome analysis and recombinant gene overexpression experiments revealed CysLT1 as a receptor responsible for potent LTE4-induced response in LAD2 but not in LUVA cells, an observation confirmed further by gene knockdown and selective inhibitors. Lentiviral overexpression of CysLT1 in LUVA cells augmented intracellular calcium signalling induced by LTE4 but did not restore full agonist responses at the gene expression level. Our data support a model where both an increased expression of Gαq-coupled CysLT1, and sustained intracellular calcium mobilisation and extracellular signal-regulated kinase (Erk) activation, are required for LTE4-mediated regulation of gene expression in human cells. Our study shows for the first time that CysLT1 expression is critically important for responsiveness to LTE4 within a human cell system. PMID:26830450

  18. Visualizing Gene Expression In Situ

    SciTech Connect

    Burlage, R.S.

    1998-11-02

    Visualizing bacterial cells and describing their responses to the environment are difficult tasks. Their small size is the chief reason for the difficulty, which means that we must often use many millions of cells in a sample in order to determine what the average response of the bacteria is. However, an average response can sometimes mask important events in bacterial physiology, which means that our understanding of these organisms will suffer. We have used a variety of instruments to visualize bacterial cells, all of which tell us something different about the sample. We use a fluorescence activated cell sorter to sort cells based on the fluorescence provided by bioreporter genes, and these can be used to select for particular genetic mutations. Cells can be visualized by epifluorescent microscopy, and sensitive photodetectors can be added that allow us to find a single bacterial cell that is fluorescent or bioluminescent. We have also used standard photomultipliers to examine cell aggregates as field bioreporter microorganisms. Examples of each of these instruments show how our understanding of bacterial physiology has changed with the technology.

  19. Gene expression profile in pelvic organ prolapse†

    PubMed Central

    Brizzolara, S.S.; Killeen, J.; Urschitz, J.

    2009-01-01

    It was hypothesized that the processes contributing to pelvic organ prolapse (POP) may be identified by transcriptional profiling of pelvic connective tissue in conjunction with light microscopy. In order to test this, we performed a frequency-matched case–control study of women undergoing hysterectomy for POP and controls. Total RNA, extracted from uterosacral and round ligament samples used to generate labeled cRNA, was hybridized to microarrays and analyzed for the expression of 32 878 genes. Significance Analysis of Microarrays (Stanford University, CA, USA) identified differentially expressed genes used for ontoanalysis. Quantitative PCR (qPCR) confirmed results. Light microscopy confirmed the tissue type and assessed inflammatory infiltration. The analysis of 34 arrays revealed 249 differentially expressed genes with fold changes (FC) larger than 1.5 and false discovery rates ≤5.2%. Immunity and defense was the most significant biological process differentially expressed in POP. qPCR confirmed the elevated steady-state mRNA levels for four genes: interleukin-6 (FC 9.8), thrombospondin 1 (FC 3.5) and prostaglandin-endoperoxide synthase 2 (FC 2.4) and activating transcription factor 3 (FC 2.6). Light microscopy showed all the samples were composed of fibromuscular connective tissue with no inflammatory infiltrates. In conclusion, genes enriched for ‘immunity and defense’ contribute to POP independent of inflammatory infiltrates. PMID:19056808

  20. Clustering of High Throughput Gene Expression Data

    PubMed Central

    Pirim, Harun; Ekşioğlu, Burak; Perkins, Andy; Yüceer, Çetin

    2012-01-01

    High throughput biological data need to be processed, analyzed, and interpreted to address problems in life sciences. Bioinformatics, computational biology, and systems biology deal with biological problems using computational methods. Clustering is one of the methods used to gain insight into biological processes, particularly at the genomics level. Clearly, clustering can be used in many areas of biological data analysis. However, this paper presents a review of the current clustering algorithms designed especially for analyzing gene expression data. It is also intended to introduce one of the main problems in bioinformatics - clustering gene expression data - to the operations research community. PMID:23144527

  1. Facilitated diffusion buffers noise in gene expression.

    PubMed

    Schoech, Armin P; Zabet, Nicolae Radu

    2014-09-01

    Transcription factors perform facilitated diffusion [three-dimensional (3D) diffusion in the cytosol and 1D diffusion on the DNA] when binding to their target sites to regulate gene expression. Here, we investigated the influence of this binding mechanism on the noise in gene expression. Our results showed that, for biologically relevant parameters, the binding process can be represented by a two-state Markov model and that the accelerated target finding due to facilitated diffusion leads to a reduction in both the mRNA and the protein noise.

  2. Facilitated diffusion buffers noise in gene expression

    NASA Astrophysics Data System (ADS)

    Schoech, Armin P.; Zabet, Nicolae Radu

    2014-09-01

    Transcription factors perform facilitated diffusion [three-dimensional (3D) diffusion in the cytosol and 1D diffusion on the DNA] when binding to their target sites to regulate gene expression. Here, we investigated the influence of this binding mechanism on the noise in gene expression. Our results showed that, for biologically relevant parameters, the binding process can be represented by a two-state Markov model and that the accelerated target finding due to facilitated diffusion leads to a reduction in both the mRNA and the protein noise.

  3. Objective and subjective probability in gene expression.

    PubMed

    Velasco, Joel D

    2012-09-01

    In this paper I address the question of whether the probabilities that appear in models of stochastic gene expression are objective or subjective. I argue that while our best models of the phenomena in question are stochastic models, this fact should not lead us to automatically assume that the processes are inherently stochastic. After distinguishing between models and reality, I give a brief introduction to the philosophical problem of the interpretation of probability statements. I argue that the objective vs. subjective distinction is a false dichotomy and is an unhelpful distinction in this case. Instead, the probabilities in our models of gene expression exhibit standard features of both objectivity and subjectivity.

  4. Genomic signatures of germline gene expression.

    PubMed

    McVicker, Graham; Green, Phil

    2010-11-01

    Transcribed regions in the human genome differ from adjacent intergenic regions in transposable element density, crossover rates, and asymmetric substitution and sequence composition patterns. We tested whether these differences reflect selection or are instead a byproduct of germline transcription, using publicly available gene expression data from a variety of germline and somatic tissues. Crossover rate shows a strong negative correlation with gene expression in meiotic tissues, suggesting that crossover is inhibited by transcription. Strand-biased composition (G+T content) and A → G versus T → C substitution asymmetry are both positively correlated with germline gene expression. We find no evidence for a strand bias in allele frequency data, implying that the substitution asymmetry reflects a mutation rather than a fixation bias. The density of transposable elements is positively correlated with germline expression, suggesting that such elements preferentially insert into regions that are actively transcribed. For each of the features examined, our analyses favor a nonselective explanation for the observed trends and point to the role of germline gene expression in shaping the mammalian genome.

  5. [Imprinting genes and it's expression in Arabidopsis].

    PubMed

    Zhang, Hong-Yu; Xu, Pei-Zhou; Yang, Hua; Wu, Xian-Jun

    2010-07-01

    Genomic imprinting refers to the phenomenon that the expression of a gene copy depends on its parent of origin. The Arabidopsis imprinted FIS (Fertilisation-independent seed) genes, mea, fis2, and fie, play essential roles in the repression of central cell and the regulation of early endosperm development. fis mutants display two phenotypes: autonomous diploid endosperm development when fertilization is absent and un-cellularised endosperm formation when fertilization occurs. The FIS Polycomb protein complex including the above three FIS proteins catalyzes histone H3 K27 tri-methylation on target loci. DME (DEMETER), a DNA glycosylase, and AtMET1 (Methyltransferase1), a DNA methyltransferase, are involved in the regulation of imprinted expression of both mea and fis2. This review summarizes the studies on the Arabidopsis imprinted FIS genes and other related genes. Recent works have shown that the insertion of transposons may affect nearby gene expression, which may be the main driving force behind the evolution of genomic imprinting. This summary covers the achievements on Arabidopsis imprinted genes will provide important information for studies on genomic imprinting in the important crops such as rice and maize.

  6. Induction of ANGPTL4 expression in human airway smooth muscle cells by PMA through activation of PKC and MAPK pathways.

    PubMed

    Stapleton, Cliona M; Joo, Joung Hyuck; Kim, Yong-Sik; Liao, Grace; Panettieri, Reynold A; Jetten, Anton M

    2010-02-15

    In this study, we demonstrate that protein kinase C (PKC) activators, including phorbol-12-myristate-13-acetate (PMA), 1,2-dioctanoyl-sn-glycerol (DOG), and platelet-derived growth factor alpha are potent inducers of angiopoietin-like protein 4 (ANGPTL4) expression in several normal lung cell types and carcinoma cell lines. In human airway smooth muscle (HASM) cells induction of ANGPTL4 expression is observed as early as 2 h after the addition of PMA. PMA also increases the level of ANGPTL4 protein released in the medium. PKC inhibitors Ro31-8820 and Gö6983 greatly inhibit the induction of ANGPTL4 mRNA by PMA suggesting that this up-regulation involves activation of PKC. Knockdown of several PKCs by corresponding siRNAs suggest a role for PKCalpha. PMA does not activate MAPK p38 and p38 inhibitors have little effect on the induction of ANGPTL4 indicating that p38 is not involved in the regulation of ANGPTL4 by PMA. In contrast, treatment of HASM by PMA induces phosphorylation and activation of Ra, MEK1/2, ERK1/2, JNK, Elk-1, and c-Jun. The Ras inhibitor manumycin A, the MEK1/2 inhibitor U0126, and the JNK inhibitor SP600125, greatly reduce the increase in ANGPTL4 expression by PMA. Knockdown of MEK1/2 and JNK1/2 expression by corresponding siRNAs inhibits the induction of ANGPTL4. Our observations suggest that the induction of ANGPTL4 by PMA in HASM involves the activation of PKC, ERK, and JNK pathways. This induction may play a role in tissue remodeling during lung injury and be implicated in several lung pathologies.

  7. Sequence and gene expression evolution of paralogous genes in willows

    PubMed Central

    Harikrishnan, Srilakshmy L.; Pucholt, Pascal; Berlin, Sofia

    2015-01-01

    Whole genome duplications (WGD) have had strong impacts on species diversification by triggering evolutionary novelties, however, relatively little is known about the balance between gene loss and forces involved in the retention of duplicated genes originating from a WGD. We analyzed putative Salicoid duplicates in willows, originating from the Salicoid WGD, which took place more than 45 Mya. Contigs were constructed by de novo assembly of RNA-seq data derived from leaves and roots from two genotypes. Among the 48,508 contigs, 3,778 pairs were, based on fourfold synonymous third-codon transversion rates and syntenic positions, predicted to be Salicoid duplicates. Both copies were in most cases expressed in both tissues and 74% were significantly differentially expressed. Mean Ka/Ks was 0.23, suggesting that the Salicoid duplicates are evolving by purifying selection. Gene Ontology enrichment analyses showed that functions related to DNA- and nucleic acid binding were over-represented among the non-differentially expressed Salicoid duplicates, while functions related to biosynthesis and metabolism were over-represented among the differentially expressed Salicoid duplicates. We propose that the differentially expressed Salicoid duplicates are regulatory neo- and/or subfunctionalized, while the non-differentially expressed are dose sensitive, hence, functionally conserved. Multiple evolutionary processes, thus drive the retention of Salicoid duplicates in willows. PMID:26689951

  8. Sequence and gene expression evolution of paralogous genes in willows.

    PubMed

    Harikrishnan, Srilakshmy L; Pucholt, Pascal; Berlin, Sofia

    2015-12-22

    Whole genome duplications (WGD) have had strong impacts on species diversification by triggering evolutionary novelties, however, relatively little is known about the balance between gene loss and forces involved in the retention of duplicated genes originating from a WGD. We analyzed putative Salicoid duplicates in willows, originating from the Salicoid WGD, which took place more than 45 Mya. Contigs were constructed by de novo assembly of RNA-seq data derived from leaves and roots from two genotypes. Among the 48,508 contigs, 3,778 pairs were, based on fourfold synonymous third-codon transversion rates and syntenic positions, predicted to be Salicoid duplicates. Both copies were in most cases expressed in both tissues and 74% were significantly differentially expressed. Mean Ka/Ks was 0.23, suggesting that the Salicoid duplicates are evolving by purifying selection. Gene Ontology enrichment analyses showed that functions related to DNA- and nucleic acid binding were over-represented among the non-differentially expressed Salicoid duplicates, while functions related to biosynthesis and metabolism were over-represented among the differentially expressed Salicoid duplicates. We propose that the differentially expressed Salicoid duplicates are regulatory neo- and/or subfunctionalized, while the non-differentially expressed are dose sensitive, hence, functionally conserved. Multiple evolutionary processes, thus drive the retention of Salicoid duplicates in willows.

  9. The TRANSFAC system on gene expression regulation.

    PubMed

    Wingender, E; Chen, X; Fricke, E; Geffers, R; Hehl, R; Liebich, I; Krull, M; Matys, V; Michael, H; Ohnhäuser, R; Prüss, M; Schacherer, F; Thiele, S; Urbach, S

    2001-01-01

    The TRANSFAC database on transcription factors and their DNA-binding sites and profiles (http://www.gene-regulation.de/) has been quantitatively extended and supplemented by a number of modules. These modules give information about pathologically relevant mutations in regulatory regions and transcription factor genes (PathoDB), scaffold/matrix attached regions (S/MARt DB), signal transduction (TRANSPATH) and gene expression sources (CYTOMER). Altogether, these distinct database modules constitute the TRANSFAC system. They are accompanied by a number of program routines for identifying potential transcription factor binding sites or for localizing individual components in the regulatory network of a cell.

  10. Marker gene tethering by nucleoporins affects gene expression in plants.

    PubMed

    Smith, Sarah; Galinha, Carla; Desset, Sophie; Tolmie, Frances; Evans, David; Tatout, Christophe; Graumann, Katja

    2015-01-01

    In non-plant systems, chromatin association with the nuclear periphery affects gene expression, where interactions with nuclear envelope proteins can repress and interactions with nucleoporins can enhance transcription. In plants, both hetero- and euchromatin can localize at the nuclear periphery, but the effect of proximity to the nuclear periphery on gene expression remains largely unknown. This study explores the putative function of Seh1 and Nup50a nucleoporins on gene expression by using the Lac Operator / Lac Repressor (LacI-LacO) system adapted to Arabidopsis thaliana. We used LacO fused to the luciferase reporter gene (LacO:Luc) to investigate whether binding of the LacO:Luc transgene to nucleoporin:LacI protein fusions alters luciferase expression. Two separate nucleoporin-LacI-YFP fusions were introduced into single insert, homozygous LacO:Luc Arabidopsis plants. Homozygous plants carrying LacO:Luc and a single insert of either Seh1-LacI-YFP or Nup50a-LacI-YFP were tested for luciferase activity and compared to plants containing LacO:Luc only. Seh1-LacI-YFP increased, while Nup50a-LacI-YFP decreased luciferase activity. Seh1-LacI-YFP accumulated at the nuclear periphery as expected, while Nup50a-LacI-YFP was nucleoplasmic and was not selected for further study. Protein and RNA levels of luciferase were quantified by western blotting and RT-qPCR, respectively. Increased luciferase activity in LacO:Luc+Seh1-LacI-YFP plants was correlated with increased luciferase protein and RNA levels. This change of luciferase expression was abolished by disruption of LacI-LacO binding by treating with IPTG in young seedlings, rosette leaves and inflorescences. This study suggests that association with the nuclear periphery is involved in the regulation of gene expression in plants.

  11. Transgenic control of perforin gene expression

    SciTech Connect

    Lichtenheld, M.G.; Podack, E.R.; Levy, R.B.

    1995-03-01

    Perforin is a pore-forming effector molecule of CTL and NK cells. To characterize perforin gene expression and its transcriptional control mechanisms in vivo, expression of a cell surface tag, i.e., human CD4, was driven by 5.1 kb of the murin perforin 5{prime} flanking and promoter region in transgenic mice. Six out of seven transgenic lines expressed the perforin-tag hybrid gene at low to intermediate levels, depending on the integration site. Transgene expression occurred in all cells that physiologically are able to express perforin. At the whole organ level, significant amounts of transgenic mRNA and endogenous perforin mRNA were co-expressed in the lymphoid organs, as well as in the lung, the ileum, the oviduct/uterus, and the bone marrow. At the single cell level, the perforin tag was present on NK cells and on CD8{sup +}, as well as on CD4{sup +} cells. Also targeted were Thy-1.2{sup +} {gamma}{delta} T cells, but not Thy-1.2{sup -} {gamma}{delta} T cells, B cells, nor monocytes. During thymic T cell development, transgene expression occurred in double negative (CD4{sup -}CD8{sup -}) thymocytes and was detected at all subsequent stages, but exceeded the expression levels of the endogenous gene in the thymus. In conclusion, the analyzed perforin 5{prime} flanking and promoter region contains important cis-acting sequences that restrict perforin expression to T cells and NK cells, and therefore provides a unique tool for manipulating T cell and/or Nk cell-mediated immune responses in transgenic mice. On the other hand, the normal control of perforin gene expression involves at least one additional negative control mechanism that was not mediated by the transgenic promoter and upstream region. This control restricts perforin gene expression in thymically developing T cells and in most resting peripheral T cells, but can be released upon T cell activation. 43 refs., 7 figs., 1 tab.

  12. Organization and expression of hair follicle genes.

    PubMed

    Rogers, G E; Powell, B C

    1993-07-01

    Several families of proteins are expressed in the growth of hair and an estimated 50-100 proteins constitute the final hair fiber. The cumbersome nomenclature for naming these different proteins has led to a proposal to modify that which is currently used for epidermal keratins. Investigations of the organization of hair genes indicate that the members of each family are clustered in the genome and their expression could be under some general control. Interestingly, the protein called trichohyalin, markedly distinct from the hair proteins, is produced in the inner root sheath cells and the gene for it has been found to be located at the same human chromosome locus as the genes for profilaggrin, involucrin, and loricrin. A mainstream objective is to identify controls responsible for the production in the hair cortex of keratin intermediate filaments (IFs) and two large groups of keratin-associated proteins (KAPs) rich in the amino acids cysteine or glycine/tyrosine. A specific family of cysteine-rich proteins is expressed in the hair cuticle. Comparisons of promoter regions of IF genes and KAP genes, including a recently characterized gene for a glycine/tyrosine-rich protein, have revealed putative hair-specific motifs in addition to known elements that regulate gene expression. In the sheep, the patterns of expression in hair differentiation are particularly interesting insofar as there are distinct segments of para- and orthocortical type cells that have significantly different pathways of expression. The testing of candidate hair-specific regulatory sequences by mouse transgenesis has produced several interesting hair phenotypes. Transgenic sheep over-expressing keratin genes but showing no hair growth change have been obtained and compared with the equivalent transgenic hair-loss mice. Studies of the effects of amino acid supply on the rate of hair growth have demonstrated that with cysteine supplementation of sheep a perturbation occurs in which there is a

  13. Laser capture microdissection reveals dose-response of gene expression in situ consequent to asbestos exposure.

    PubMed

    Yin, Qi; Brody, Arnold R; Sullivan, Deborah E

    2007-12-01

    The genes that mediate fibroproliferative lung disease remain to be defined. Prior studies from our laboratory showed by in situ hybridization and immunohistochemistry that the genes coding for tumour necrosis factor alpha, transforming growth factor beta, the platelet-derived growth factor A and B isoforms, and alpha-1 pro-collagen are expressed in fibroproliferative lesions that develop quickly after asbestos inhalation. These five genes, along with matrix metalloproteinase 9, a collagenase found to be increased in several lung diseases, are known to control matrix production and cell proliferation in humans and animals. Here we show by laser capture microdissection that (i) The six genes are expressed at significantly higher levels in the asbestos-exposed mice when comparing the same anatomic regions 'captured' in unexposed mice. (ii) The bronchiolar-alveolar duct (BAD) junctions, where the greatest number of fibres initially deposit, were always significantly higher than the other anatomic regions for each gene. The first alveolar duct bifurcation (ADB) generally was higher than the second ADB, the ADBs were always significantly higher than the airway walls and pleura, and the airway walls and pleura were generally higher than the unexposed tissues. (iii) Animals exposed for 3 days always exhibited significantly higher levels of gene expression at the BAD junctions and ADBs than animals exposed for 2 days. To our knowledge, this is the first demonstration of a dose-response to a toxic particle in situ, and this response appears to be dependent on the number of fibres that deposits at the individual anatomic site.

  14. Regulation of Calreticulin Gene Expression by Calcium

    PubMed Central

    Waser, Mathilde; Mesaeli, Nasrin; Spencer, Charlotte; Michalak, Marek

    1997-01-01

    We have isolated and characterized a 12-kb mouse genomic DNA fragment containing the entire calreticulin gene and 2.14 kb of the promoter region. The mouse calreticulin gene consists of nine exons and eight introns, and it spans 4.2 kb of genomic DNA. A 1.8-kb fragment of the calreticulin promoter was subcloned into a reporter gene plasmid containing chloramphenicol acetyltransferase. This construct was then used in transient and stable transfection of NIH/ 3T3 cells. Treatment of transfected cells either with the Ca2+ ionophore A23187, or with the ER Ca2+-ATPase inhibitor thapsigargin, resulted in a five- to sevenfold increase of the expression of chloramphenicol acetyltransferase protein. Transactivation of the calreticulin promoter was also increased by fourfold in NIH/3T3 cells treated with bradykinin, a hormone that induces Ca2+ release from the intracellular Ca2+ stores. Analysis of the promoter deletion constructs revealed that A23187- and thapsigargin-responsive regions are confined to two regions (−115 to −260 and −685 to −1,763) in the calreticulin promoter that contain the CCAAT nucleotide sequences. Northern blot analysis of cells treated with A23187, or with thapsigargin, revealed a fivefold increase in calreticulin mRNA levels. Thapsigargin also induced a fourfold increase in calreticulun protein levels. Importantly, we show by nuclear run-on transcription analysis that calreticulin gene transcription is increased in NIH/3T3 cells treated with A23187 and thapsigargin in vivo. This increase in gene expression required over 4 h of continuous incubation with the drugs and was also sensitive to treatment with cycloheximide, suggesting that it is dependent on protein synthesis. Changes in the concentration of extracellular and cytoplasmic Ca2+ did not affect the increased expression of the calreticulin gene. These studies suggest that stress response to the depletion of intracellular Ca2+ stores induces expression of the calreticulin gene in vitro

  15. Decreased S100A9 Expression Promoted Rat Airway Smooth Muscle Cell Proliferation by Stimulating ROS Generation and Inhibiting p38 MAPK

    PubMed Central

    Yin, Lei-Miao; Han, Xiao-Jie; Duan, Ting-Ting; Xu, Yu-Dong; Ulloa, Luis

    2016-01-01

    Background. Asthma is a disease with a core abnormality in airway smooth muscle function, and the proliferation of airway smooth muscle cells (ASMCs) plays a pivotal role in asthma airway remodeling. Our previous study showed that S100A9 (S100 calcium-binding protein A9; 400 and 800 ng/mL) significantly inhibited rat ASMCs proliferation at 48 h, and 50–800 ng/mL S100A9 (50, 100, 200, 400, and 800 ng/mL) also induced a lasting effect by significantly inhibiting rat ASMCs proliferation at 72 h in a dose-dependent manner. However, the intracellular effects of S100A9 on ASMCs proliferation remain unknown. Methods. Rat ASMCs with stable S100A9 knockdown were generated using short hairpin RNA. The effects of decreased S100A9 expression on cellular proliferation, the production of reactive oxygen species (ROS), and p38 MAPK pathway protein expression were examined. Results. Decreased intracellular S100A9 expression significantly promoted platelet-derived growth factor-induced rat ASMCs proliferation and increased ROS production. The antioxidative agent N-acetylcysteine significantly inhibited rat ASMCs proliferation. Western blot results showed that the decreased intracellular S100A9 expression significantly inhibited p38 MAPK phosphorylation. Conclusion. Decreased S100A9 expression promoted rat ASMCs proliferation by stimulating ROS generation and inhibiting p38 MAPK. Our study may provide novel insights into the regulation of asthma airway remodeling. PMID:28050155

  16. Wheatgrass Extract Ameliorates Hypoxia-induced Mucin Gene Expression in A549 cells

    PubMed Central

    Sim, Ju hwan; Choi, Moon-Hee; Shin, Hyun-Jae; Lee, Ji-Eun

    2017-01-01

    Background: Wheatgrass is known to have antioxidant, antiaging, and anti-inflammatory effect. However, its protective effect against hypoxia is not yet evaluated. Objective: In this study, we evaluated the protective and anti-inflammatory effect of wheatgrass against the hypoxia in airway epithelial cells. Materials and Methods: A549 human lung adenocarcinoma cells were incubated in a hypoxic condition (CO2 5%/O2 1%) for 24 hr in the presence of different concentration of wheatgrass 50, 75, 100, and 150 μg/mL, and the magnitude of each immunologic response produced by the A549 cells was compared. The mRNA expression level of mucin gene (MUC), 5A, 5B, 8, GM-CSF, TNF-α, and VEGF were evaluated by using real-time polymerase chain reaction. The MUC proteins level before and after knocking out the hypoxia-inducible factor (hif)-1α via short interfering (si) RNA transfection were assessed by immunoblot analysis. Accordingly, the involved cell signaling pathway was evaluated by immunoblot analysis. Results: The inflammatory cytokines (GM-CSF, TNF- α) and the expressions of MUC 5A, 5B, and 8 were augmented by hypoxia. The augmented MUC expression was decreased by the wheatgrass extract administration. Hif-1α gene expression after hypoxia exposure was decreased by wheatgrass. Knockdown of hif-1α by siRNA reduced the mucin gene expression and which was more enhanced by wheatgrass extract. Conclusion: Theses results suggest that wheatgrass may be useful in the treatment of sinonasal disease by inhibiting mucus hypersecretion in airway epithelium. SUMMARY Wheatgrass extract decreases the hypoxia-induced MUC 5A, 5B and 8 expression.Hif-1α gene expression after hypoxia exposure was decreased by wheatgrass.Wheatgrass inhibits p44/42 phosphorylation in hypoxia-exposed airway epithelial cells. Abbreviations used: A549: human lung adenocarcinoma cells, GM-CSF: granulocyte-macrophage colony stimulating factor, HIF: hypoxia inducible factor, IL: interleukin, MUC: mucin, MTT: 3

  17. The frustrated gene: origins of eukaryotic gene expression

    PubMed Central

    Madhani, Hiten D.

    2014-01-01

    Eukarytotic gene expression is frustrated by a series of steps that are generally not observed in prokaryotes and are therefore not essential for the basic chemistry of transcription and translation. Their evolution may have been driven by the need to defend against parasitic nucleic acids. PMID:24209615

  18. Cystic Fibrosis Transmembrane Conductance Regulator in Sarcoplasmic Reticulum of Airway Smooth Muscle. Implications for Airway Contractility

    PubMed Central

    Cook, Daniel P.; Rector, Michael V.; Bouzek, Drake C.; Michalski, Andrew S.; Gansemer, Nicholas D.; Reznikov, Leah R.; Li, Xiaopeng; Stroik, Mallory R.; Ostedgaard, Lynda S.; Abou Alaiwa, Mahmoud H.; Thompson, Michael A.; Prakash, Y. S.; Krishnan, Ramaswamy; Meyerholz, David K.; Seow, Chun Y.

    2016-01-01

    Rationale: An asthma-like airway phenotype has been described in people with cystic fibrosis (CF). Whether these findings are directly caused by loss of CF transmembrane conductance regulator (CFTR) function or secondary to chronic airway infection and/or inflammation has been difficult to determine. Objectives: Airway contractility is primarily determined by airway smooth muscle. We tested the hypothesis that CFTR is expressed in airway smooth muscle and directly affects airway smooth muscle contractility. Methods: Newborn pigs, both wild type and with CF (before the onset of airway infection and inflammation), were used in this study. High-resolution immunofluorescence was used to identify the subcellular localization of CFTR in airway smooth muscle. Airway smooth muscle function was determined with tissue myography, intracellular calcium measurements, and regulatory myosin light chain phosphorylation status. Precision-cut lung slices were used to investigate the therapeutic potential of CFTR modulation on airway reactivity. Measurements and Main Results: We found that CFTR localizes to the sarcoplasmic reticulum compartment of airway smooth muscle and regulates airway smooth muscle tone. Loss of CFTR function led to delayed calcium reuptake following cholinergic stimulation and increased myosin light chain phosphorylation. CFTR potentiation with ivacaftor decreased airway reactivity in precision-cut lung slices following cholinergic stimulation. Conclusions: Loss of CFTR alters porcine airway smooth muscle function and may contribute to the airflow obstruction phenotype observed in human CF. Airway smooth muscle CFTR may represent a therapeutic target in CF and other diseases of airway narrowing. PMID:26488271

  19. Trigger finger, tendinosis, and intratendinous gene expression.

    PubMed

    Lundin, A-C; Aspenberg, P; Eliasson, P

    2014-04-01

    The pathogenesis of trigger finger has generally been ascribed to primary changes in the first annular ligament. In contrast, we recently found histological changes in the tendons, similar to the findings in Achilles tendinosis or tendinopathy. We therefore hypothesized that trigger finger tendons would show differences in gene expression in comparison to normal tendons in a pattern similar to what is published for Achilles tendinosis. We performed quantitative real-time polymerase chain reaction on biopsies from finger flexor tendons, 13 trigger fingers and 13 apparently healthy control tendons, to assess the expression of 10 genes which have been described to be differently expressed in tendinosis (collagen type 1a1, collagen 3a1, MMP-2, MMP-3, ADAMTS-5, TIMP-3, aggrecan, biglycan, decorin, and versican). In trigger finger tendons, collagen types 1a1 and 3a1, aggrecan and biglycan were all up-regulated, and MMP-3and TIMP-3 were down-regulated. These changes were statistically significant and have been previously described for Achilles tendinosis. The remaining four genes were not significantly altered. The changes in gene expression support the hypothesis that trigger finger is a form of tendinosis. Because trigger finger is a common condition, often treated surgically, it could provide opportunities for clinical research on tendinosis.

  20. The low noise limit in gene expression

    SciTech Connect

    Dar, Roy D.; Weinberger, Leor S.; Cox, Chris D.; Simpson, Michael L.; Razooky, Brandon S.

    2015-10-21

    Protein noise measurements are increasingly used to elucidate biophysical parameters. Unfortunately noise analyses are often at odds with directly measured parameters. Here we show that these inconsistencies arise from two problematic analytical choices: (i) the assumption that protein translation rate is invariant for different proteins of different abundances, which has inadvertently led to (ii) the assumption that a large constitutive extrinsic noise sets the low noise limit in gene expression. While growing evidence suggests that transcriptional bursting may set the low noise limit, variability in translational bursting has been largely ignored. We show that genome-wide systematic variation in translational efficiency can-and in the case of E. coli does-control the low noise limit in gene expression. Therefore constitutive extrinsic noise is small and only plays a role in the absence of a systematic variation in translational efficiency. Lastly, these results show the existence of two distinct expression noise patterns: (1) a global noise floor uniformly imposed on all genes by expression bursting; and (2) high noise distributed to only a select group of genes.

  1. The Low Noise Limit in Gene Expression

    PubMed Central

    Dar, Roy D.; Razooky, Brandon S.; Weinberger, Leor S.; Cox, Chris D.; Simpson, Michael L.

    2015-01-01

    Protein noise measurements are increasingly used to elucidate biophysical parameters. Unfortunately noise analyses are often at odds with directly measured parameters. Here we show that these inconsistencies arise from two problematic analytical choices: (i) the assumption that protein translation rate is invariant for different proteins of different abundances, which has inadvertently led to (ii) the assumption that a large constitutive extrinsic noise sets the low noise limit in gene expression. While growing evidence suggests that transcriptional bursting may set the low noise limit, variability in translational bursting has been largely ignored. We show that genome-wide systematic variation in translational efficiency can–and in the case of E. coli does–control the low noise limit in gene expression. Therefore constitutive extrinsic noise is small and only plays a role in the absence of a systematic variation in translational efficiency. These results show the existence of two distinct expression noise patterns: (1) a global noise floor uniformly imposed on all genes by expression bursting; and (2) high noise distributed to only a select group of genes. PMID:26488303

  2. Digital gene expression signatures for maize development

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Genome-wide expression signatures detect specific perturbations in developmental programs and contribute to functional resolution of key regulatory networks. In maize (Zea mays) inflorescences, mutations in the RAMOSA (RA) genes affect determinacy of axillary meristems and thus alter branching patt...

  3. Analysis of baseline gene expression levels from ...

    EPA Pesticide Factsheets

    The use of gene expression profiling to predict chemical mode of action would be enhanced by better characterization of variance due to individual, environmental, and technical factors. Meta-analysis of microarray data from untreated or vehicle-treated animals within the control arm of toxicogenomics studies has yielded useful information on baseline fluctuations in gene expression. A dataset of control animal microarray expression data was assembled by a working group of the Health and Environmental Sciences Institute's Technical Committee on the Application of Genomics in Mechanism Based Risk Assessment in order to provide a public resource for assessments of variability in baseline gene expression. Data from over 500 Affymetrix microarrays from control rat liver and kidney were collected from 16 different institutions. Thirty-five biological and technical factors were obtained for each animal, describing a wide range of study characteristics, and a subset were evaluated in detail for their contribution to total variability using multivariate statistical and graphical techniques. The study factors that emerged as key sources of variability included gender, organ section, strain, and fasting state. These and other study factors were identified as key descriptors that should be included in the minimal information about a toxicogenomics study needed for interpretation of results by an independent source. Genes that are the most and least variable, gender-selectiv

  4. Multiple Stochastic Point Processes in Gene Expression

    NASA Astrophysics Data System (ADS)

    Murugan, Rajamanickam

    2008-04-01

    We generalize the idea of multiple-stochasticity in chemical reaction systems to gene expression. Using Chemical Langevin Equation approach we investigate how this multiple-stochasticity can influence the overall molecular number fluctuations. We show that the main sources of this multiple-stochasticity in gene expression could be the randomness in transcription and translation initiation times which in turn originates from the underlying bio-macromolecular recognition processes such as the site-specific DNA-protein interactions and therefore can be internally regulated by the supra-molecular structural factors such as the condensation/super-coiling of DNA. Our theory predicts that (1) in case of gene expression system, the variances ( φ) introduced by the randomness in transcription and translation initiation-times approximately scales with the degree of condensation ( s) of DNA or mRNA as φ ∝ s -6. From the theoretical analysis of the Fano factor as well as coefficient of variation associated with the protein number fluctuations we predict that (2) unlike the singly-stochastic case where the Fano factor has been shown to be a monotonous function of translation rate, in case of multiple-stochastic gene expression the Fano factor is a turn over function with a definite minimum. This in turn suggests that the multiple-stochastic processes can also be well tuned to behave like a singly-stochastic point processes by adjusting the rate parameters.

  5. The low noise limit in gene expression

    DOE PAGES

    Dar, Roy D.; Weinberger, Leor S.; Cox, Chris D.; ...

    2015-10-21

    Protein noise measurements are increasingly used to elucidate biophysical parameters. Unfortunately noise analyses are often at odds with directly measured parameters. Here we show that these inconsistencies arise from two problematic analytical choices: (i) the assumption that protein translation rate is invariant for different proteins of different abundances, which has inadvertently led to (ii) the assumption that a large constitutive extrinsic noise sets the low noise limit in gene expression. While growing evidence suggests that transcriptional bursting may set the low noise limit, variability in translational bursting has been largely ignored. We show that genome-wide systematic variation in translational efficiencymore » can-and in the case of E. coli does-control the low noise limit in gene expression. Therefore constitutive extrinsic noise is small and only plays a role in the absence of a systematic variation in translational efficiency. Lastly, these results show the existence of two distinct expression noise patterns: (1) a global noise floor uniformly imposed on all genes by expression bursting; and (2) high noise distributed to only a select group of genes.« less

  6. Relationships among CFTR expression, HCO3− secretion, and host defense may inform gene- and cell-based cystic fibrosis therapies

    PubMed Central

    Shah, Viral S.; Ernst, Sarah; Tang, Xiao Xiao; Karp, Philip H.; Parker, Connor P.; Ostedgaard, Lynda S.; Welsh, Michael J.

    2016-01-01

    Cystic fibrosis (CF) is caused by mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR) anion channel. Airway disease is the major source of morbidity and mortality. Successful implementation of gene- and cell-based therapies for CF airway disease requires knowledge of relationships among percentages of targeted cells, levels of CFTR expression, correction of electrolyte transport, and rescue of host defense defects. Previous studies suggested that, when ∼10–50% of airway epithelial cells expressed CFTR, they generated nearly wild-type levels of Cl− secretion; overexpressing CFTR offered no advantage compared with endogenous expression levels. However, recent discoveries focused attention on CFTR-mediated HCO3− secretion and airway surface liquid (ASL) pH as critical for host defense and CF pathogenesis. Therefore, we generated porcine airway epithelia with varying ratios of CF and wild-type cells. Epithelia with a 50:50 mix secreted HCO3− at half the rate of wild-type epithelia. Likewise, heterozygous epithelia (CFTR+/− or CFTR+/∆F508) expressed CFTR and secreted HCO3− at ∼50% of wild-type values. ASL pH, antimicrobial activity, and viscosity showed similar relationships to the amount of CFTR. Overexpressing CFTR increased HCO3− secretion to rates greater than wild type, but ASL pH did not exceed wild-type values. Thus, in contrast to Cl− secretion, the amount of CFTR is rate-limiting for HCO3− secretion and for correcting host defense abnormalities. In addition, overexpressing CFTR might produce a greater benefit than expressing CFTR at wild-type levels when targeting small fractions of cells. These findings may also explain the risk of airway disease in CF carriers. PMID:27114540

  7. Expression of mouse metallothionein genes in tobacco

    SciTech Connect

    Maiti, I.B.; Yeargan, R.; Wagner, G.J.; Hunt, A.G. )

    1990-05-01

    We have expressed a mouse metallothionein (NT) gene in tobacco under control of the cauliflower mosaic virus (CaMV) 35S promoter and a pea ribulose-1,5-bisphosphate carboxylase small subunit (rbcS) gene promoter. Seedlings in which MT gene expression is driven by the 35S promoter are resistant to toxic levels of cadmium. Mature plants carrying the 35S-MT gene accumulate less Cd in their leaves when exposed to low levels of Cd in laboratory growth conditions. Plants with the rbcS-MT construction express this gene in a light-regulated and tissue-specific manner, as expected. Moreover, the MT levels in leaves in these plants are about 20% of those seen in 35S-MT plants. These plants are currently being tested for Cd resistance. In addition, a small field evaluation of 35S-MT lines for Cd levels is being evaluated. These experiments will address the possibility of using MTs to alter Cd levels in crop species.

  8. Regulation of methane genes and genome expression

    SciTech Connect

    John N. Reeve

    2009-09-09

    At the start of this project, it was known that methanogens were Archaeabacteria (now Archaea) and were therefore predicted to have gene expression and regulatory systems different from Bacteria, but few of the molecular biology details were established. The goals were then to establish the structures and organizations of genes in methanogens, and to develop the genetic technologies needed to investigate and dissect methanogen gene expression and regulation in vivo. By cloning and sequencing, we established the gene and operon structures of all of the “methane” genes that encode the enzymes that catalyze methane biosynthesis from carbon dioxide and hydrogen. This work identified unique sequences in the methane gene that we designated mcrA, that encodes the largest subunit of methyl-coenzyme M reductase, that could be used to identify methanogen DNA and establish methanogen phylogenetic relationships. McrA sequences are now the accepted standard and used extensively as hybridization probes to identify and quantify methanogens in environmental research. With the methane genes in hand, we used northern blot and then later whole-genome microarray hybridization analyses to establish how growth phase and substrate availability regulated methane gene expression in Methanobacterium thermautotrophicus ΔH (now Methanothermobacter thermautotrophicus). Isoenzymes or pairs of functionally equivalent enzymes catalyze several steps in the hydrogen-dependent reduction of carbon dioxide to methane. We established that hydrogen availability determine which of these pairs of methane genes is expressed and therefore which of the alternative enzymes is employed to catalyze methane biosynthesis under different environmental conditions. As were unable to establish a reliable genetic system for M. thermautotrophicus, we developed in vitro transcription as an alternative system to investigate methanogen gene expression and regulation. This led to the discovery that an archaeal protein

  9. Fluid Mechanics, Arterial Disease, and Gene Expression

    PubMed Central

    Tarbell, John M.; Shi, Zhong-Dong; Dunn, Jessilyn; Jo, Hanjoong

    2014-01-01

    This review places modern research developments in vascular mechanobiology in the context of hemodynamic phenomena in the cardiovascular system and the discrete localization of vascular disease. The modern origins of this field are traced, beginning in the 1960s when associations between flow characteristics, particularly blood flow–induced wall shear stress, and the localization of atherosclerotic plaques were uncovered, and continuing to fluid shear stress effects on the vascular lining endothelial) cells (ECs), including their effects on EC morphology, biochemical production, and gene expression. The earliest single-gene studies and genome-wide analyses are considered. The final section moves from the ECs lining the vessel wall to the smooth muscle cells and fibroblasts within the wall that are fluid me chanically activated by interstitial flow that imposes shear stresses on their surfaces comparable with those of flowing blood on EC surfaces. Interstitial flow stimulates biochemical production and gene expression, much like blood flow on ECs. PMID:25360054

  10. Fluid Mechanics, Arterial Disease, and Gene Expression.

    PubMed

    Tarbell, John M; Shi, Zhong-Dong; Dunn, Jessilyn; Jo, Hanjoong

    2014-01-01

    This review places modern research developments in vascular mechanobiology in the context of hemodynamic phenomena in the cardiovascular system and the discrete localization of vascular disease. The modern origins of this field are traced, beginning in the 1960s when associations between flow characteristics, particularly blood flow-induced wall shear stress, and the localization of atherosclerotic plaques were uncovered, and continuing to fluid shear stress effects on the vascular lining endothelial) cells (ECs), including their effects on EC morphology, biochemical production, and gene expression. The earliest single-gene studies and genome-wide analyses are considered. The final section moves from the ECs lining the vessel wall to the smooth muscle cells and fibroblasts within the wall that are fluid me chanically activated by interstitial flow that imposes shear stresses on their surfaces comparable with those of flowing blood on EC surfaces. Interstitial flow stimulates biochemical production and gene expression, much like blood flow on ECs.

  11. Gene expression profiling of human ovarian tumours

    PubMed Central

    Biade, S; Marinucci, M; Schick, J; Roberts, D; Workman, G; Sage, E H; O'Dwyer, P J; LiVolsi, V A; Johnson, S W

    2006-01-01

    There is currently a lack of reliable diagnostic and prognostic markers for ovarian cancer. We established gene expression profiles for 120 human ovarian tumours to identify determinants of histologic subtype, grade and degree of malignancy. Unsupervised cluster analysis of the most variable set of expression data resulted in three major tumour groups. One consisted predominantly of benign tumours, one contained mostly malignant tumours, and one was comprised of a mixture of borderline and malignant tumours. Using two supervised approaches, we identified a set of genes that distinguished the benign, borderline and malignant phenotypes. These algorithms were unable to establish profiles for histologic subtype or grade. To validate these findings, the expression of 21 candidate genes selected from these analyses was measured by quantitative RT–PCR using an independent set of tumour samples. Hierarchical clustering of these data resulted in two major groups, one benign and one malignant, with the borderline tumours interspersed between the two groups. These results indicate that borderline ovarian tumours may be classified as either benign or malignant, and that this classifier could be useful for predicting the clinical course of borderline tumours. Immunohistochemical analysis also demonstrated increased expression of CD24 antigen in malignant versus benign tumour tissue. The data that we have generated will contribute to a growing body of expression data that more accurately define the biologic and clinical characteristics of ovarian cancers. PMID:16969345

  12. Gene expression profiling of human ovarian tumours.

    PubMed

    Biade, S; Marinucci, M; Schick, J; Roberts, D; Workman, G; Sage, E H; O'Dwyer, P J; Livolsi, V A; Johnson, S W

    2006-10-23

    There is currently a lack of reliable diagnostic and prognostic markers for ovarian cancer. We established gene expression profiles for 120 human ovarian tumours to identify determinants of histologic subtype, grade and degree of malignancy. Unsupervised cluster analysis of the most variable set of expression data resulted in three major tumour groups. One consisted predominantly of benign tumours, one contained mostly malignant tumours, and one was comprised of a mixture of borderline and malignant tumours. Using two supervised approaches, we identified a set of genes that distinguished the benign, borderline and malignant phenotypes. These algorithms were unable to establish profiles for histologic subtype or grade. To validate these findings, the expression of 21 candidate genes selected from these analyses was measured by quantitative RT-PCR using an independent set of tumour samples. Hierarchical clustering of these data resulted in two major groups, one benign and one malignant, with the borderline tumours interspersed between the two groups. These results indicate that borderline ovarian tumours may be classified as either benign or malignant, and that this classifier could be useful for predicting the clinical course of borderline tumours. Immunohistochemical analysis also demonstrated increased expression of CD24 antigen in malignant versus benign tumour tissue. The data that we have generated will contribute to a growing body of expression data that more accurately define the biologic and clinical characteristics of ovarian cancers.

  13. Repression of gene expression by oxidative stress.

    PubMed Central

    Morel, Y; Barouki, R

    1999-01-01

    Gene expression is modulated by both physiological signals (hormones, cytokines, etc.) and environmental stimuli (physical parameters, xenobiotics, etc.). Oxidative stress appears to be a key pleiotropic modulator which may be involved in either pathway. Indeed, reactive oxygen species (ROS) have been described as second messengers for several growth factors and cytokines, but have also been shown to rise following cellular insults such as xenobiotic metabolism or enzymic deficiency. Extensive studies on the induction of stress-response genes by oxidative stress have been reported. In contrast, owing to the historical focus on gene induction, less attention has been paid to gene repression by ROS. However, a growing number of studies have shown that moderate (i.e. non-cytotoxic) oxidative stress specifically down-regulates the expression of various genes. In this review, we describe the alteration of several physiological functions resulting from oxidative-stress-mediated inhibition of gene transcription. We will then focus on the repressive oxidative modulation of various transcription factors elicited by ROS. PMID:10477257

  14. [Structure and expression of thyroglobulin gene].

    PubMed

    Vassart, G; Brocas, H; Christophe, D; de Martynoff, G; Leriche, A; Mercken, L; Pohl, V; Van Heuverswyn, B

    1982-01-01

    Thyroglobulin is composed of two 300000 dalton polypeptide chains, translated from an 8000 base mRNA. Preparation of a full length cDNA and its cloning in E. coli have lead to the demonstration that the polypeptides of thyroglobulin protomers were identical. Used as molecular probes, the cloned cDNA allowed the isolation of a fragment of thyroglobulin gene. Electron microscopic studies have demonstrated that this gene contains more than 90% intronic material separating small size exons (less than 200 bp). Sequencing of bovine thyroglobulin structural gene is in progress. Preliminary results show evidence for the existence of repetitive segments. Availability of cloned DNA complementary to bovine and human thyroglobulin mRNA allows the study of genetic defects of thyroglobulin gene expression in the human and in various animal models.

  15. Exposure to PM2.5 induces aberrant activation of NF-κB in human airway epithelial cells by downregulating miR-331 expression.

    PubMed

    Song, Lei; Li, Dan; Li, Xiaoping; Ma, Lianjun; Bai, Xiaoxue; Wen, Zhongmei; Zhang, Xiufang; Chen, Dong; Peng, Liping

    2017-03-01

    Exposure to particulate matter (PM) with an aerodynamic diameter≤2.5μm (PM2.5) induces reactive oxygen species (ROS) and pro-inflammatory cytokine production, leading to airway epithelial injury. However, the mechanisms underlying the toxicity of PM2.5 have not been clarified. Here, we show that exposure to PM2.5 induces sustained activation of the nuclear factor kappa B (NF-κB) signaling in human airway epithelial Beas-2B (B2B) cells. In addition, PM2.5 exposure significantly decreased miR-331 expression in B2B cells, which was abrogated by inhibition of ROS or phosphoinositide 3-kinase (PI3K)/Akt pathway. Induction of miR-331 overexpression attenuated the PM2.5 exposure-induced NF-kBp65 nuclear translocation, IL-6 and IL-8 expression in B2B cells. Furthermore, miR-331 targeted the inhibitor of NF-κB kinase beta (IKK-β) by down-regulating the IKK-β-regulated luciferase activity in HEK293 cells. Moreover, induction of miR-331 over-expression inhibited IKK-β expression while induction of IKK-β over-expression prevented the inhibition of miR-331 on the PM2.5 exposure-induced NF-kBp65 nuclear translocation, IL-6 and IL-8 expression in B2B cells. Therefore, PM2.5 exposure decreased miR-331 expression via the ROS/PI3K/Akt pathway, resulting in an increase in the IKK-β expression and sustained NF-κB activation in human airway epithelial cells. Our findings may provide new insights into the molecular mechanisms underlying the toxicity of PM2.5 exposure and aid in design of new therapeutic strategies to prevent PM2.5-induced toxicity.

  16. Determining Physical Mechanisms of Gene Expression Regulation from Single Cell Gene Expression Data

    PubMed Central

    Moignard, Victoria; Göttgens, Berthold; Adryan, Boris

    2016-01-01

    Many genes are expressed in bursts, which can contribute to cell-to-cell heterogeneity. It is now possible to measure this heterogeneity with high throughput single cell gene expression assays (single cell qPCR and RNA-seq). These experimental approaches generate gene expression distributions which can be used to estimate the kinetic parameters of gene expression bursting, namely the rate that genes turn on, the rate that genes turn off, and the rate of transcription. We construct a complete pipeline for the analysis of single cell qPCR data that uses the mathematics behind bursty expression to develop more accurate and robust algorithms for analyzing the origin of heterogeneity in experimental samples, specifically an algorithm for clustering cells by their bursting behavior (Simulated Annealing for Bursty Expression Clustering, SABEC) and a statistical tool for comparing the kinetic parameters of bursty expression across populations of cells (Estimation of Parameter changes in Kinetics, EPiK). We applied these methods to hematopoiesis, including a new single cell dataset in which transcription factors (TFs) involved in the earliest branchpoint of blood differentiation were individually up- and down-regulated. We could identify two unique sub-populations within a seemingly homogenous group of hematopoietic stem cells. In addition, we could predict regulatory mechanisms controlling the expression levels of eighteen key hematopoietic transcription factors throughout differentiation. Detailed information about gene regulatory mechanisms can therefore be obtained simply from high throughput single cell gene expression data, which should be widely applicable given the rapid expansion of single cell genomics. PMID:27551778

  17. Coevolution of gene expression among interacting proteins

    SciTech Connect

    Fraser, Hunter B.; Hirsh, Aaron E.; Wall, Dennis P.; Eisen,Michael B.

    2004-03-01

    Physically interacting proteins or parts of proteins are expected to evolve in a coordinated manner that preserves proper interactions. Such coevolution at the amino acid-sequence level is well documented and has been used to predict interacting proteins, domains, and amino acids. Interacting proteins are also often precisely coexpressed with one another, presumably to maintain proper stoichiometry among interacting components. Here, we show that the expression levels of physically interacting proteins coevolve. We estimate average expression levels of genes from four closely related fungi of the genus Saccharomyces using the codon adaptation index and show that expression levels of interacting proteins exhibit coordinated changes in these different species. We find that this coevolution of expression is a more powerful predictor of physical interaction than is coevolution of amino acid sequence. These results demonstrate previously uncharacterized coevolution of gene expression, adding a different dimension to the study of the coevolution of interacting proteins and underscoring the importance of maintaining coexpression of interacting proteins over evolutionary time. Our results also suggest that expression coevolution can be used for computational prediction of protein protein interactions.

  18. Differential var gene expression in children with malaria and antidromic effects on host gene expression.

    PubMed

    Kalmbach, Yvonne; Rottmann, Matthias; Kombila, Maryvonne; Kremsner, Peter G; Beck, Hans-Peter; Kun, Jürgen F J

    2010-07-15

    Among 62 children with mild malaria, cerebral malaria, or severe malarial anemia, we analyzed the transcription of different var gene types. There was no difference in parasitemia level or body temperature between groups. However, a significantly different expression pattern was observed in children with cerebral malaria, compared with that in patients in the other 2 groups: children with cerebral malaria had lower expression of the upsA subtype but higher expression of the upsB and upsC subtypes. Furthermore, expression of human genes responsive to tumor necrosis factor and hypoxia correlated with distinct ups types.

  19. Transcriptome-Level Signatures in Gene Expression and Gene Expression Variability during Bacterial Adaptive Evolution

    PubMed Central

    Erickson, Keesha E.; Otoupal, Peter B.

    2017-01-01

    ABSTRACT Antibiotic-resistant bacteria are an increasingly serious public health concern, as strains emerge that demonstrate resistance to almost all available treatments. One factor that contributes to the crisis is the adaptive ability of bacteria, which exhibit remarkable phenotypic and gene expression heterogeneity in order to gain a survival advantage in damaging environments. This high degree of variability in gene expression across biological populations makes it a challenging task to identify key regulators of bacterial adaptation. Here, we research the regulation of adaptive resistance by investigating transcriptome profiles of Escherichia coli upon adaptation to disparate toxins, including antibiotics and biofuels. We locate potential target genes via conventional gene expression analysis as well as using a new analysis technique examining differential gene expression variability. By investigating trends across the diverse adaptation conditions, we identify a focused set of genes with conserved behavior, including those involved in cell motility, metabolism, membrane structure, and transport, and several genes of unknown function. To validate the biological relevance of the observed changes, we synthetically perturb gene expression using clustered regularly interspaced short palindromic repeat (CRISPR)-dCas9. Manipulation of select genes in combination with antibiotic treatment promotes adaptive resistance as demonstrated by an increased degree of antibiotic tolerance and heterogeneity in MICs. We study the mechanisms by which identified genes influence adaptation and find that select differentially variable genes have the potential to impact metabolic rates, mutation rates, and motility. Overall, this work provides evidence for a complex nongenetic response, encompassing shifts in gene expression and gene expression variability, which underlies adaptive resistance. IMPORTANCE Even initially sensitive bacteria can rapidly thwart antibiotic treatment

  20. Transcriptome-Level Signatures in Gene Expression and Gene Expression Variability during Bacterial Adaptive Evolution.

    PubMed

    Erickson, Keesha E; Otoupal, Peter B; Chatterjee, Anushree

    2017-01-01

    Antibiotic-resistant bacteria are an increasingly serious public health concern, as strains emerge that demonstrate resistance to almost all available treatments. One factor that contributes to the crisis is the adaptive ability of bacteria, which exhibit remarkable phenotypic and gene expression heterogeneity in order to gain a survival advantage in damaging environments. This high degree of variability in gene expression across biological populations makes it a challenging task to identify key regulators of bacterial adaptation. Here, we research the regulation of adaptive resistance by investigating transcriptome profiles of Escherichia coli upon adaptation to disparate toxins, including antibiotics and biofuels. We locate potential target genes via conventional gene expression analysis as well as using a new analysis technique examining differential gene expression variability. By investigating trends across the diverse adaptation conditions, we identify a focused set of genes with conserved behavior, including those involved in cell motility, metabolism, membrane structure, and transport, and several genes of unknown function. To validate the biological relevance of the observed changes, we synthetically perturb gene expression using clustered regularly interspaced short palindromic repeat (CRISPR)-dCas9. Manipulation of select genes in combination with antibiotic treatment promotes adaptive resistance as demonstrated by an increased degree of antibiotic tolerance and heterogeneity in MICs. We study the mechanisms by which identified genes influence adaptation and find that select differentially variable genes have the potential to impact metabolic rates, mutation rates, and motility. Overall, this work provides evidence for a complex nongenetic response, encompassing shifts in gene expression and gene expression variability, which underlies adaptive resistance. IMPORTANCE Even initially sensitive bacteria can rapidly thwart antibiotic treatment through stress

  1. Gene expression regulation in roots under drought.

    PubMed

    Janiak, Agnieszka; Kwaśniewski, Mirosław; Szarejko, Iwona

    2016-02-01

    Stress signalling and regulatory networks controlling expression of target genes are the basis of plant response to drought. Roots are the first organs exposed to water deficiency in the soil and are the place of drought sensing. Signalling cascades transfer chemical signals toward the shoot and initiate molecular responses that lead to the biochemical and morphological changes that allow plants to be protected against water loss and to tolerate stress conditions. Here, we present an overview of signalling network and gene expression regulation pathways that are actively induced in roots under drought stress. In particular, the role of several transcription factor (TF) families, including DREB, AP2/ERF, NAC, bZIP, MYC, CAMTA, Alfin-like and Q-type ZFP, in the regulation of root response to drought are highlighted. The information provided includes available data on mutual interactions between these TFs together with their regulation by plant hormones and other signalling molecules. The most significant downstream target genes and molecular processes that are controlled by the regulatory factors are given. These data are also coupled with information about the influence of the described regulatory networks on root traits and root development which may translate to enhanced drought tolerance. This is the first literature survey demonstrating the gene expression regulatory machinery that is induced by drought stress, presented from the perspective of roots.

  2. Expression of bacterial genes in plant cells.

    PubMed Central

    Fraley, R T; Rogers, S G; Horsch, R B; Sanders, P R; Flick, J S; Adams, S P; Bittner, M L; Brand, L A; Fink, C L; Fry, J S; Galluppi, G R; Goldberg, S B; Hoffmann, N L; Woo, S C

    1983-01-01

    Chimeric bacterial genes conferring resistance to aminoglycoside antibiotics have been inserted into the Agrobacterium tumefaciens tumor-inducing (Ti) plasmid and introduced into plant cells by in vitro transformation techniques. The chimeric genes contain the nopaline synthase 5' and 3' regulatory regions joined to the genes for neomycin phosphotransferase type I or type II. The chimeric genes were cloned into an intermediate vector, pMON120, and inserted into pTiB6S3 by recombination and then introduced into petunia and tobacco cells by cocultivating A. tumefaciens cells with protoplast-derived cells. Southern hybridization was used to confirm the presence of the chimeric genes in the transformed plant tissues. Expression of the chimeric genes was determined by the ability of the transformed cells to proliferate on medium containing normally inhibitory levels of kanamycin (50 micrograms/ml) or other aminoglycoside antibiotics. Plant cells transformed by wild-type pTiB6S3 or derivatives carrying the bacterial neomycin phosphotransferase genes with their own promoters failed to grow under these conditions. The significance of these results for plant genetic engineering is discussed. Images PMID:6308651

  3. Transient gene expression in electroporated Solanum protoplasts.

    PubMed

    Jones, H; Ooms, G; Jones, M G

    1989-11-01

    Electroporation was used to evaluate parameters important in transient gene expression in potato protoplasts. The protoplasts were from leaves of wild potato Solanum brevidens, and from leaves, tubers and suspension cells of cultivated Solanum tuberosum cv. Désirée. Reporter enzyme activity, chloramphenicol acetyl transferase (CAT) under the control of the cauliflower mosaic virus (CaMV) 35S promoter, depended on the field strength and the pulse duration used for electroporation. Using field pulses of 85 ms duration, the optimum field strengths for maximum CAT activity were: S. brevidens mesophyll protoplasts--250 V/cm; Désirée mesophyll protoplasts--225 V/cm; Désirée suspension culture protoplasts--225 V/cm; and Désirée tuber protoplasts--150 V/cm. The optimum field strengths correlated inversely with the size of the protoplasts electroporated; this is consistent with biophysical theory. In time courses, maximum CAT activity (in Désirée mesophyll protoplasts) occurred 36-48 h after electroporation. Examination at optimised conditions of a chimaeric gene consisting of a class II patatin promoter linked to the beta-glucuronidase (gus) gene, showed expression (at DNA concentrations between 0-10 pmol/ml) comparable to the CaMV 35S promoter in both tuber and mesophyll protoplasts. At higher DNA concentrations (20-30 pmol/ml) the patatin promoter directed 4-5 times higher levels of gus expression. Implications and potential contributions towards studying gene expression, in particular of homologous genes in potato, are discussed.

  4. Exposure to Mexicali PM10 Induces IL-8 Expression Through an Alternative NFKB Mechanism In Human Airway Epithelial Cells

    EPA Science Inventory

    Studies have shown associations between exposure to ambient air particulate matter (PM) and increased rates of cardio-pulmonary morbidity and mortality. The aim of this study was to examine the signaling events involved in the expression of inflammatory genes in cultured human ai...

  5. NF-kappaB Signaling in Chronic Inflammatory Airway Disease

    PubMed Central

    Schuliga, Michael

    2015-01-01

    Asthma and chronic obstructive pulmonary disease (COPD) are obstructive airway disorders which differ in their underlying causes and phenotypes but overlap in patterns of pharmacological treatments. In both asthma and COPD, oxidative stress contributes to airway inflammation by inducing inflammatory gene expression. The redox-sensitive transcription factor, nuclear factor (NF)-kappaB (NF-κB), is an important participant in a broad spectrum of inflammatory networks that regulate cytokine activity in airway pathology. The anti-inflammatory actions of glucocorticoids (GCs), a mainstay treatment for asthma, involve inhibition of NF-κB induced gene transcription. Ligand bound GC receptors (GRs) bind NF-κB to suppress the transcription of NF-κB responsive genes (i.e., transrepression). However, in severe asthma and COPD, the transrepression of NF-κB by GCs is negated as a consequence of post-translational changes to GR and histones involved in chromatin remodeling. Therapeutics which target NF-κB activation, including inhibitors of IκB kinases (IKKs) are potential treatments for asthma and COPD. Furthermore, reversing GR/histone acetylation shows promise as a strategy to treat steroid refractory airway disease by augmenting NF-κB transrepression. This review examines NF-κB signaling in airway inflammation and its potential as target for treatment of asthma and COPD. PMID:26131974

  6. Interferon gamma and interleukin 4 stimulate prolonged expression of inducible nitric oxide synthase in human airway epithelium through synthesis of soluble mediators.

    PubMed Central

    Guo, F H; Uetani, K; Haque, S J; Williams, B R; Dweik, R A; Thunnissen, F B; Calhoun, W; Erzurum, S C

    1997-01-01

    Human respiratory epithelium expresses inducible nitric oxide synthase (iNOS) continuously in vivo, however mechanisms responsible for maintenance of expression are not known. We show that IFNgamma is sufficient for induction of iNOS in primary human airway epithelial cells (HAEC) in vitro, and IL-4 potentiates IFNgamma-induced iNOS expression in HAEC through stabilization of iNOS mRNA. IFNgamma/IL-4- induced iNOS expression in HAEC was delayed in onset and prolonged with expression up to 1 wk. Removal of overlying culture media resulted in loss of expression, while transfer of conditioned media induced iNOS mRNA in other HAEC. IFNgamma and IL-4 stimulation activated STAT1 and STAT6 in HAEC, but conditioned media transfer to HAEC produced even higher levels of STAT1 activation than achieved by direct addition of cytokines. Although cytokine induction of iNOS was dependent on new protein synthesis, conditioned media induction of iNOS in HAEC was not. Further, removal of overlying culture media from cells at different times after cytokine stimulation demonstrated that mediator synthesis and/or secretion important for induction and maintenance of iNOS occurs early after cytokine stimulation. In conclusion, a combination of IFNgamma/ IL-4, which occurs naturally in the lung epithelial lining fluid, leads to maintenance of iNOS expression in human airway epithelium through production of soluble mediators and stabilization of mRNA. PMID:9259582

  7. Hypoxia Potentiates Allergen Induction of HIF-1α, Chemokines, Airway Inflammation, TGF-β1, and Airway Remodeling in a mouse model

    PubMed Central

    Baek, Kwang Je; Cho, Jae Youn; Rosenthal, Peter; Alexander, Laura E. Crotty; Nizet, Victor; Broide, David H.

    2013-01-01

    Whether hypoxia contributes to airway inflammation and remodeling in asthma is unknown. In this study we used mice exposed to a hypoxic environment during allergen challenge (simulating hypoxia during an asthma exacerbation) to investigate the contribution of hypoxia to airway inflammation and remodeling. Although neither hypoxia alone, nor OVA allergen alone, induced significant neutrophil influx into the lung, the combination of OVA and hypoxia induced a synergistic 27 fold increase in peribronchial neutrophils, enhanced expression of HIF-1α and one of its target genes, the CXC-family neutrophil chemokine KC. The combination of hypoxia and OVA allergen increased eotaxin-1, peribronchial eosinophils, lung TGB-β1 expression, and indices of airway remodeling (fibrosis and smooth muscle) compared to either stimulus alone. As hypoxia is present in >90% of severe asthma exacerbations, these findings underscore the potential of hypoxia to potentiate the airway inflammatory response, remodeling, and accelerate the decline of lung function in asthma exacerbations. PMID:23499929

  8. Toward stable gene expression in CHO cells

    PubMed Central

    Mariati; Koh, Esther YC; Yeo, Jessna HM; Ho, Steven CL; Yang, Yuansheng

    2014-01-01

    Maintaining high gene expression level during long-term culture is critical when producing therapeutic recombinant proteins using mammalian cells. Transcriptional silencing of promoters, most likely due to epigenetic events such as DNA methylation and histone modifications, is one of the major mechanisms causing production instability. Previous studies demonstrated that the core CpG island element (IE) from the hamster adenine phosphoribosyltransferase gene is effective to prevent DNA methylation. We generated one set of modified human cytomegalovirus (hCMV) promoters by insertion of one or two copies of IE in either forward or reverse orientations into different locations of the hCMV promoter. The modified hCMV with one copy of IE inserted between the hCMV enhancer and core promoter in reverse orientation (MR1) was most effective at enhancing expression stability in CHO cells without comprising expression level when compared with the wild type hCMV. We also found that insertion of IE into a chimeric murine CMV (mCMV) enhancer and human elongation factor-1α core (hEF) promoter in reverse orientation did not enhance expression stability, indicating that the effect of IE on expression stability is possibly promoter specific. PMID:25482237

  9. Engineering Genes for Predictable Protein Expression

    PubMed Central

    Gustafsson, Claes; Minshull, Jeremy; Govindarajan, Sridhar; Ness, Jon; Villalobos, Alan; Welch, Mark

    2013-01-01

    The DNA sequence used to encode a polypeptide can have dramatic effects on its expression. Lack of readily available tools has until recently inhibited meaningful experimental investigation of this phenomenon. Advances in synthetic biology and the application of modern engineering approaches now provide the tools for systematic analysis of the sequence variables affecting heterologous expression of recombinant proteins. We here discuss how these new tools are being applied and how they circumvent the constraints of previous approaches, highlighting some of the surprising and promising results emerging from the developing field of gene engineering. PMID:22425659

  10. Engineering genes for predictable protein expression.

    PubMed

    Gustafsson, Claes; Minshull, Jeremy; Govindarajan, Sridhar; Ness, Jon; Villalobos, Alan; Welch, Mark

    2012-05-01

    The DNA sequence used to encode a polypeptide can have dramatic effects on its expression. Lack of readily available tools has until recently inhibited meaningful experimental investigation of this phenomenon. Advances in synthetic biology and the application of modern engineering approaches now provide the tools for systematic analysis of the sequence variables affecting heterologous expression of recombinant proteins. We here discuss how these new tools are being applied and how they circumvent the constraints of previous approaches, highlighting some of the surprising and promising results emerging from the developing field of gene engineering.

  11. Cancer outlier differential gene expression detection.

    PubMed

    Wu, Baolin

    2007-07-01

    We study statistical methods to detect cancer genes that are over- or down-expressed in some but not all samples in a disease group. This has proven useful in cancer studies where oncogenes are activated only in a small subset of samples. We propose the outlier robust t-statistic (ORT), which is intuitively motivated from the t-statistic, the most commonly used differential gene expression detection method. Using real and simulation studies, we compare the ORT to the recently proposed cancer outlier profile analysis (Tomlins and others, 2005) and the outlier sum statistic of Tibshirani and Hastie (2006). The proposed method often has more detection power and smaller false discovery rates. Supplementary information can be found at http://www.biostat.umn.edu/~baolin/research/ort.html.

  12. Programming gene expression with combinatorial promoters

    PubMed Central

    Cox, Robert Sidney; Surette, Michael G; Elowitz, Michael B

    2007-01-01

    Promoters control the expression of genes in response to one or more transcription factors (TFs). The architecture of a promoter is the arrangement and type of binding sites within it. To understand natural genetic circuits and to design promoters for synthetic biology, it is essential to understand the relationship between promoter function and architecture. We constructed a combinatorial library of random promoter architectures. We characterized 288 promoters in Escherichia coli, each containing up to three inputs from four different TFs. The library design allowed for multiple −10 and −35 boxes, and we observed varied promoter strength over five decades. To further analyze the functional repertoire, we defined a representation of promoter function in terms of regulatory range, logic type, and symmetry. Using these results, we identified heuristic rules for programming gene expression with combinatorial promoters. PMID:18004278

  13. Combinatorial engineering for heterologous gene expression.

    PubMed

    Zwick, Friederike; Lale, Rahmi; Valla, Svein

    2013-01-01

    Tools for strain engineering with predictable outcome are of crucial importance for the nascent field of synthetic biology. The success of combining different DNA biological parts is often restricted by poorly understood factors deriving from the complexity of the systems. We have previously identified variants for different regulatory elements of the expression cassette XylS/Pm. When such elements are combined they act in a manner consistent with their individual behavior, as long as they affect different functions, such as transcription and translation. Interestingly, sequence context does not seem to influence the final outcome significantly. Expression of reporter gene bla could be increased up to 75 times at the protein level by combining three variants in one cassette. For other tested reporter genes similar results were obtained, except that the stimulatory effect was quantitatively less. Combination of individually characterized DNA parts thus stands as suitable method to achieve a desired phenotype.

  14. Structure, expression and functions of MTA genes.

    PubMed

    Kumar, Rakesh; Wang, Rui-An

    2016-05-15

    Metastatic associated proteins (MTA) are integrators of upstream regulatory signals with the ability to act as master coregulators for modifying gene transcriptional activity. The MTA family includes three genes and multiple alternatively spliced variants. The MTA proteins neither have their own enzymatic activity nor have been shown to directly interact with DNA. However, MTA proteins interact with a variety of chromatin remodeling factors and complexes with enzymatic activities for modulating the plasticity of nucleosomes, leading to the repression or derepression of target genes or other extra-nuclear and nucleosome remodeling and histone deacetylase (NuRD)-complex independent activities. The functions of MTA family members are driven by the steady state levels and subcellular localization of MTA proteins, the dynamic nature of modifying signals and enzymes, the structural features and post-translational modification of protein domains, interactions with binding proteins, and the nature of the engaged and resulting features of nucleosomes in the proximity of target genes. In general, MTA1 and MTA2 are the most upregulated genes in human cancer and correlate well with aggressive phenotypes, therapeutic resistance, poor prognosis and ultimately, unfavorable survival of cancer patients. Here we will discuss the structure, expression and functions of the MTA family of genes in the context of cancer cells.

  15. Identifying driver genes in cancer by triangulating gene expression, gene location, and survival data.

    PubMed

    Rouam, Sigrid; Miller, Lance D; Karuturi, R Krishna Murthy

    2014-01-01

    Driver genes are directly responsible for oncogenesis and identifying them is essential in order to fully understand the mechanisms of cancer. However, it is difficult to delineate them from the larger pool of genes that are deregulated in cancer (ie, passenger genes). In order to address this problem, we developed an approach called TRIAngulating Gene Expression (TRIAGE through clinico-genomic intersects). Here, we present a refinement of this approach incorporating a new scoring methodology to identify putative driver genes that are deregulated in cancer. TRIAGE triangulates - or integrates - three levels of information: gene expression, gene location, and patient survival. First, TRIAGE identifies regions of deregulated expression (ie, expression footprints) by deriving a newly established measure called the Local Singular Value Decomposition (LSVD) score for each locus. Driver genes are then distinguished from passenger genes using dual survival analyses. Incorporating measurements of gene expression and weighting them according to the LSVD weight of each tumor, these analyses are performed using the genes located in significant expression footprints. Here, we first use simulated data to characterize the newly established LSVD score. We then present the results of our application of this refined version of TRIAGE to gene expression data from five cancer types. This refined version of TRIAGE not only allowed us to identify known prominent driver genes, such as MMP1, IL8, and COL1A2, but it also led us to identify several novel ones. These results illustrate that TRIAGE complements existing tools, allows for the identification of genes that drive cancer and could perhaps elucidate potential future targets of novel anticancer therapeutics.

  16. Gene expression during normal and malignant differentiation

    SciTech Connect

    Andersson, L.C.; Gahmberg, C.G.; Ekblom, P.

    1985-01-01

    This book contains 18 selections. Some of the titles are: Exploring Carcinogenesis with Retroviral and Cellular Oncogenes; Retroviruses, Oncogenes and Evolution; HTLV and Human Neoplasi; Modes of Activation of cMyc Oncogene in B and T Lymphoid Tumors; The Structure and Function of the Epidermal Growth Factor Receptor: Its Relationship to the Protein Product of the V-ERB-B Oncogene; and Expression of Human Retrovirus Genes in Normal and Neoplastic Epithelial Cells.

  17. Nonreplicating vaccinia vector efficiently expresses recombinant genes.

    PubMed

    Sutter, G; Moss, B

    1992-11-15

    Modified vaccinia Ankara (MVA), a highly attenuated vaccinia virus strain that has been safety tested in humans, was evaluated for use as an expression vector. MVA has multiple genomic deletions and is severely host cell restricted: it grows well in avian cells but is unable to multiply in human and most other mammalian cells tested. Nevertheless, we found that replication of viral DNA appeared normal and that both early and late viral proteins were synthesized in human cells. Proteolytic processing of viral structural proteins was inhibited, however, and only immature virus particles were detected by electron microscopy. We constructed an insertion plasmid with the Escherichia coli lacZ gene under the control of the vaccinia virus late promoter P11, flanked by sequences of MVA DNA, to allow homologous recombination at the site of a naturally occurring 3500-base-pair deletion within the MVA genome. MVA recombinants were isolated and propagated in permissive avian cells and shown to express the enzyme beta-galactosidase upon infection of nonpermissive human cells. The amount of enzyme made was similar to that produced by a recombinant of vaccinia virus strain Western Reserve, which also had the lacZ gene under control of the P11 promoter, but multiplied to high titers. Since recombinant gene expression is unimpaired in nonpermissive human cells, MVA may serve as a highly efficient and exceptionally safe vector.

  18. A gene expression biomarker accurately predicts estrogen ...

    EPA Pesticide Factsheets

    The EPA’s vision for the Endocrine Disruptor Screening Program (EDSP) in the 21st Century (EDSP21) includes utilization of high-throughput screening (HTS) assays coupled with computational modeling to prioritize chemicals with the goal of eventually replacing current Tier 1 screening tests. The ToxCast program currently includes 18 HTS in vitro assays that evaluate the ability of chemicals to modulate estrogen receptor α (ERα), an important endocrine target. We propose microarray-based gene expression profiling as a complementary approach to predict ERα modulation and have developed computational methods to identify ERα modulators in an existing database of whole-genome microarray data. The ERα biomarker consisted of 46 ERα-regulated genes with consistent expression patterns across 7 known ER agonists and 3 known ER antagonists. The biomarker was evaluated as a predictive tool using the fold-change rank-based Running Fisher algorithm by comparison to annotated gene expression data sets from experiments in MCF-7 cells. Using 141 comparisons from chemical- and hormone-treated cells, the biomarker gave a balanced accuracy for prediction of ERα activation or suppression of 94% or 93%, respectively. The biomarker was able to correctly classify 18 out of 21 (86%) OECD ER reference chemicals including “very weak” agonists and replicated predictions based on 18 in vitro ER-associated HTS assays. For 114 chemicals present in both the HTS data and the MCF-7 c

  19. Expression of foreign genes in filamentous cyanobacteria

    SciTech Connect

    Kuritz, T.; Wolk, C.P. )

    1993-06-01

    Several advantages make cyanobacteria attractive hosts for biodegradative genes and possibly for other exogenous genes that have practical uses. The authors have obtained expression in Anabaena sp. strain PCC 7120 and Nostoc ellipsosporum of a dechlorination operon, fcbAB, from Arthrobacter globiformis, and have also developed a simple method for qualitative assessment of dechlorination by microorganisms, such as cyanobacteria, whose metabolism is dependent on the presence of chloride in the medium. Transcription of fcbAB under the control of a variety of promoters was monitored by placing luxAB (encoding luciferase) downstream from fcbAB, and by measuring light emission from luciferase. They believe that the system that they have described has value as a means to screen for factors influencing transcription of foreign genes in cyanobacteria.

  20. Upper airway loading induces growth retardation and change in local chondrocyte IGF-I expression is reversed by stimulation of GH release in juvenile rats.

    PubMed

    Segev, Yael; Berdugo-Boura, Nilly; Porati, Orit; Tarasiuk, Ariel

    2008-11-01

    Chronic resistive airway loading (CAL) impairs growth in juvenile rats. The effects of CAL on epiphyseal growth plate (EGP) structure and insulin-like growth factor (IGF)-I gene expression have not been explored. Little is known about whether stimulants of endogenous growth hormone (GH) secretion can normalize this growth impairment. This study explored the effect of CAL on circulating and EGP GH/IGF-I pathway GH and the effect of ritanserin (endogenous GH stimulant) on somatic growth and the GH/IGF-I axis. We hypothesized that CAL would lead to a decrease in body temperature (Tb) and alterations of GH/IGF-I pathways, consequently leading to growth retardation. The tracheae of 22-day-old male rats were obstructed by tracheal banding (38 sham-operated control, 42 CAL). Tibial EGP morphometry, liver and EGP IGF mRNA, and serum GH and IGF-I levels were analyzed with quantitative real-time PCR and ELISA. Tb and locomotion activity (MA) were measured with telemetric transmitters inserted into the abdominal cavity. CAL animals had lower Tb and MA despite preserved food consumption. CAL impaired both tibial and tail length gains. Tail and tibial length gains inversely correlated with tracheal resistance. Circulating GH and IGF-I, liver and EGP IGF-I mRNA, and EGP width were decreased in the CAL group. Ritanserin administration to CAL animals normalized circulating and local EGP GH and IGF-I levels and minimized the longitudinal growth impairment. We conclude that CAL causes growth delay associated with alterations in the GH/IGF-I axis. Stimulation of GH release by ritanserin restored both global and local GH/IGF-I pathways, yet growth parameters were only partially restored.

  1. GeneTIER: prioritization of candidate disease genes using tissue-specific gene expression profiles

    PubMed Central

    Antanaviciute, Agne; Daly, Catherine; Crinnion, Laura A.; Markham, Alexander F.; Watson, Christopher M.; Bonthron, David T.; Carr, Ian M.

    2015-01-01

    Motivation: In attempts to determine the genetic causes of human disease, researchers are often faced with a large number of candidate genes. Linkage studies can point to a genomic region containing hundreds of genes, while the high-throughput sequencing approach will often identify a great number of non-synonymous genetic variants. Since systematic experimental verification of each such candidate gene is not feasible, a method is needed to decide which genes are worth investigating further. Computational gene prioritization presents itself as a solution to this problem, systematically analyzing and sorting each gene from the most to least likely to be the disease-causing gene, in a fraction of the time it would take a researcher to perform such queries manually. Results: Here, we present Gene TIssue Expression Ranker (GeneTIER), a new web-based application for candidate gene prioritization. GeneTIER replaces knowledge-based inference traditionally used in candidate disease gene prioritization applications with experimental data from tissue-specific gene expression datasets and thus largely overcomes the bias toward the better characterized genes/diseases that commonly afflict other methods. We show that our approach is capable of accurate candidate gene prioritization and illustrate its strengths and weaknesses using case study examples. Availability and Implementation: Freely available on the web at http://dna.leeds.ac.uk/GeneTIER/. Contact: umaan@leeds.ac.uk Supplementary information: Supplementary data are available at Bioinformatics online. PMID:25861967

  2. Reduced expression of Autographa californica nucleopolyhedrovirus ORF34, an essential gene, enhances heterologous gene expression

    SciTech Connect

    Salem, Tamer Z.; Zhang, Fengrui; Thiem, Suzanne M.

    2013-01-20

    Autographa californica multiple nucleopolyhedrovirus ORF34 is part of a transcriptional unit that includes ORF32, encoding a viral fibroblast growth factor (FGF) and ORF33. We identified ORF34 as a candidate for deletion to improve protein expression in the baculovirus expression system based on enhanced reporter gene expression in an RNAi screen of virus genes. However, ORF34 was shown to be an essential gene. To explore ORF34 function, deletion (KO34) and rescue bacmids were constructed and characterized. Infection did not spread from primary KO34 transfected cells and supernatants from KO34 transfected cells could not infect fresh Sf21 cells whereas the supernatant from the rescue bacmids transfection could recover the infection. In addition, budded viruses were not observed in KO34 transfected cells by electron microscopy, nor were viral proteins detected from the transfection supernatants by western blots. These demonstrate that ORF34 is an essential gene with a possible role in infectious virus production.

  3. BLUNTING AIRWAYS EOSINOPHILIC INFLAMMATION RESULTS IN A DECREASED AIRWAY NEUTROPHIL RESPONSE TO INHALED LPS IN ATOPIC ASTHMATICS A ROLE FOR CD-14

    EPA Science Inventory

    Recent data demonstrate that atopic inflammation might enhance airway responses to inhaled LPS in individuals with atopic asthma by increasing CD14 expression on airway macrophages. We sought to determine whether blunting airway eosinophilic inflammation decreases CD14 expressio...

  4. Screening of differentially expressed genes in pathological scar tissues using expression microarray.

    PubMed

    Huang, L P; Mao, Z; Zhang, L; Liu, X X; Huang, C; Jia, Z S

    2015-09-09

    Pathological scar tissues and normal skin tissues were differentiated by screening for differentially expressed genes in pathologic scar tissues via gene expression microarray. The differentially expressed gene data was analyzed by gene ontology and pathway analyses. There were 5001 up- or down-regulated genes in 2-fold differentially expressed genes, 956 up- or down-regulated genes in 5-fold differentially expressed genes, and 114 up- or down-regulated genes in 20-fold differentially expressed genes. Therefore, significant differences were observed in the gene expression in pathological scar tissues and normal foreskin tissues. The development of pathological scar tissues has been correlated to changes in multiple genes and pathways, which are believed to form a dynamic network connection.

  5. Gene expression and IG-DMR hypomethylation of maternally expressed gene 3 in developing corticospinal neurons.

    PubMed

    Qu, Chunsheng; Jiang, Tian; Li, Yong; Wang, Xiongwei; Cao, Huateng; Xu, Hongping; Qu, Jia; Chen, Jie-Guang

    2013-01-01

    The mammalian cerebral cortex plays a central role in higher cognitive functions and in the complex task of motor control. Maternally expressed gene 3 (Meg3) appears to play a role in cortical development and neurodegeneration, but the expression and regulation of Meg3 in the cortex is not clear. In this study, we examined the expression of transcript variants of Meg3 in the developing mouse cerebral cortex. By in situ hybridization, we found that a novel transcript variant of Meg3 with 8 small exons was expressed in the developing cortex, whereas the long isoforms of Meg3 (~11 kb) were enriched in corticospinal neurons (CSNs) in layer V of the cortex. No transcript variants of Meg3 were found in the neural progenitors at E12.5, when the intergenic differential methylation region (IG-DMR) near Meg3 was highly methylated. IG-DMR became demethylated at E15.5 and remained hypomethylated in early CSNs isolated from Fezf2-EGFP transgenic mice. The expression of Meg3 transcript variant 1 was inversely correlated with the IG-DMR methylation level during development. Moreover, expression of paternally expressed gene Peg11 was limited to the upper layers, consistent with the idea that the maternally expressed gene may be preferentially transcribed in the lower layers of the cortex. The spatiotemporal expression pattern of Meg3 suggests that it may participate in the early development of CSNs and contribute to cortical malfunctions related to aberrant imprinting in Meg3.

  6. Gravity-Induced Gene Expression in Plants.

    NASA Astrophysics Data System (ADS)

    Sederoff, Heike; Heber, Steffen; Howard, Brian; Myburg-Nichols, Henrietta; Hammond, Rebecca; Salinas-Mondragon, Raul; Brown, Christopher S.

    Plants sense changes in their orientation towards the vector of gravity and respond with directional growth. Several metabolites in the signal transduction cascade have been identified. However, very little is known about the interaction between these sensing and signal transduction events and even less is known about their role in the differential growth response. Gravity induced changes in transcript abundance have been identified in Arabidopsis whole seedlings and root apices (Moseyko et al. 2002; Kimbrough et al. 2004). Gravity induced transcript abundance changes can be observed within less than 1 min after stimulation (Salinas-Mondragon et al. 2005). Gene expression however requires not only transcription but also translation of the mRNA. Translation can only occur when mRNA is associated with ribosomes, even though not all mRNA associated with ribosomes is actively translated. To approximate translational capacity we quantified whole genome transcript abundances in corn stem pulvini during the first hour after gravity stimulation in total and poly-ribosomal fractions. As in Arabidopsis root apices, transcript abundances of several clusters of genes responded to gravity stimulation. The vast majority of these transcripts were also found to associate with polyribosomes in the same temporal and quantitative pattern. These genes are transcriptionally regulated by gravity stimulation, but do not exhibit translational regulation. However, a small group of genes showed increased transcriptional regulation after gravity stimulation, but no association with polysomes. These transcripts likely are translationally repressed. The mechanism of translational repression for these transcripts is unknown. Based on the hypothesis that the genes essential for gravitropic responses should be expressed in most or all species, we compared the temporal gravity induced expression pattern of all orthologs identified between maize and Arabidopsis. A small group of genes showed high

  7. X chromosome regulation of autosomal gene expression in bovine blastocysts.

    PubMed

    Itoh, Yuichiro; Arnold, Arthur P

    2014-10-01

    Although X chromosome inactivation in female mammals evolved to balance the expression of X chromosome and autosomal genes in the two sexes, female embryos pass through developmental stages in which both X chromosomes are active in somatic cells. Bovine blastocysts show higher expression of many X genes in XX than XY embryos, suggesting that X inactivation is not complete. Here, we reanalyzed bovine blastocyst microarray expression data from a network perspective with a focus on interactions between X chromosome and autosomal genes. Whereas male-to-female ratios of expression of autosomal genes were distributed around a mean of 1, X chromosome genes were clearly shifted towards higher expression in females. We generated gene coexpression networks and identified a major module of genes with correlated gene expression that includes female-biased X genes and sexually dimorphic autosomal genes for which the sexual dimorphism is likely driven by the X genes. In this module, expression of X chromosome genes correlates with autosome genes, more than the expression of autosomal genes with each other. Our study identifies correlated patterns of autosomal and X-linked genes that are likely influenced by the sexual imbalance of X gene expression when X inactivation is inefficient.

  8. Glucocorticoid receptor and histone deacetylase-2 mediate dexamethasone-induced repression of MUC5AC gene expression.

    PubMed

    Chen, Yajun; Watson, Alan M; Williamson, Chad D; Rahimi, Michael; Liang, Chong; Colberg-Poley, Anamaris M; Rose, Mary C

    2012-11-01

    Airway occlusion in obstructive airway diseases is caused in part by the overproduction of secretory mucin glycoproteins through the up-regulation of mucin (MUC) genes by inflammatory mediators. Some pharmacological agents, including the glucocorticoid dexamethasone (Dex), repress mucin concentrations in lung epithelial cancer cells. Here, we show that Dex reduces the expression of MUC5AC, a major airway mucin gene, in primary differentiated normal human bronchial epithelial (NHBE) cells in a dose-dependent and time-dependent manner, and that the Dex-induced repression is mediated by the glucocorticoid receptor (GR) and two glucocorticoid response elements (GREs) in the MUC5AC promoter. The pre-exposure of cells to RU486, a GR antagonist, and mutations in either the GRE3 or GRE5 cis-sites abolished the Dex-induced repression. Chromatin immunoprecipitation (ChIP) assays showed a rapid temporal recruitment of GR to the GRE3 and GRE5 cis-elements in the MUC5AC promoter in NHBE and in A549 cells. Immunofluorescence showed nuclear colocalization of GR and histone deacetylase-2 (HDAC2) in MUC5AC-expressing NHBE cells. ChIP also showed a rapid temporal recruitment of HDAC2 to the GRE3 and GRE5 cis-elements in the MUC5AC promoter in both cell types. The knockdown of HDAC2 by HDAC2-specific short interfering RNA prevented the Dex-induced repression of MUC5AC in NHBE and A549 cells. These data demonstrate that GR and HDAC2 are recruited to the GRE3 and GRE5 cis-sites in the MUC5AC promoter and mediate the Dex-induced cis repression of MUC5AC gene expression. A better understanding of the mechanisms whereby glucocorticoids repress MUC5AC gene expression may be useful in formulating therapeutic interventions in chronic lung diseases.

  9. Gene expression and cAMP.

    PubMed Central

    Nagamine, Y; Reich, E

    1985-01-01

    By comparing the 5'-flanking region of the porcine gene for the urokinase form of plasminogen activator with those of other cAMP-regulated genes, we identify a 29-nucleotide sequence that is tentatively proposed as the cAMP-regulatory unit. Homologous sequences are present (i) in the cAMP-regulated rat tyrosine aminotransferase, prolactin, and phosphoenolpyruvate carboxykinase genes and (ii) 5' to the transcription initiation sites of cAMP-regulated Escherichia coli genes. From this we conclude that the expression of cAMP-responsive genes in higher eukaryotes may be controlled, as in E. coli, by proteins that form complexes with cAMP and then show sequence-specific DNA-binding properties. The complex formed by cAMP and the regulatory subunit of the type II mammalian protein kinase might be one candidate for this function. Based on several homologies we suggest that this subunit may have retained both the DNA-binding specificity and transcription-regulating properties in addition to the nucleotide-binding domains of the bacterial cAMP-binding protein. If this were so, dissociation of protein kinase by cAMP would activate two processes: (i) protein phosphorylation by the catalytic subunit and (ii) transcription regulation by the regulatory subunit. PMID:2991882

  10. Differential expression of the ras gene family in mice.

    PubMed Central

    Leon, J; Guerrero, I; Pellicer, A

    1987-01-01

    We compared the expression of the ras gene family (H-ras, K-ras, and N-ras) in adult mouse tissues and during development. We found substantial variations in expression among different organs and in the amounts of the different transcripts originating from each gene, especially for the N-ras gene. The expression patterns were consistent with the reported preferential tissue activation of ras genes and suggested different cellular functions for each of the ras genes. Images PMID:3600635

  11. Influence of β2-Integrin Adhesion Molecule Expression and Pulmonary Infection with Pasteurella haemolytica on Cytokine Gene Expression in Cattle

    PubMed Central

    Lee, Haa-Yung; Kehrli, Marcus E.; Brogden, Kim A.; Gallup, Jack M.; Ackermann, Mark R.

    2000-01-01

    β2-Integrins are leukocyte adhesion molecules composed of alpha (CD11a, -b, -c, or -d) and beta (CD18) subunit heterodimers. Genetic CD18 deficiency results in impaired neutrophil egress into tissues that varies between conducting airways and alveoli of the lung. In this study, we investigated whether CD18 deficiency in cattle affects proinflammatory cytokine (PIC) expression in pulmonary tissue after respiratory infection with Pasteurella haemolytica. Cattle were infected with P. haemolytica via fiberoptic deposition of organisms into the posterior part of the right cranial lung lobe. Animals were euthanized at 2 or 4 h postinoculation (p.i.), and tissues were collected to assess PIC gene expression using antisense RNA probes specific for bovine interleukin-1α (IL-1α), IL-1β, IL-6, gamma interferon (IFN-γ), and tumor necrosis factor alpha (TNF-α) along with the β-actin (β-Act) housekeeping gene. Expression of PIC was induced at 2 h p.i. in P. haemolytica-infected cattle and continued to 4 h p.i. At 2 h p.i., induction of gene expression and increase of cells that expressed PIC were observed both in CD18+ and CD18− cattle after inoculation of P. haemolytica. The induction of gene expression with P. haemolytica inoculation was more prominent in CD18− cattle than in CD18+ cattle by comparison to pyrogen-free saline (PFS)-inoculated control animals. At 4 h p.i., however, the induction of PIC, especially IL-1α, IL-6, and IFN-γ, in the lungs of CD18+ cattle inoculated with P. haemolytica was greater than that in lungs of the CD18− cattle. IFN-γ and TNF-α genes were not increased in P. haemolytica-inoculated CD18− cattle lungs compared to the PFS-inoculated control lungs at 4 h p.i. In PFS-inoculated lungs, we generally observed a higher percentage of cells and higher level of gene expression in the lungs of CD18− cattle than in the lungs of CD18+ cattle, especially at 4 h p.i. The rate of neutrophil infiltration into the lungs of CD18− cattle at

  12. Studying the complex expression dependences between sets of coexpressed genes.

    PubMed

    Huerta, Mario; Casanova, Oriol; Barchino, Roberto; Flores, Jose; Querol, Enrique; Cedano, Juan

    2014-01-01

    Organisms simplify the orchestration of gene expression by coregulating genes whose products function together in the cell. The use of clustering methods to obtain sets of coexpressed genes from expression arrays is very common; nevertheless there are no appropriate tools to study the expression networks among these sets of coexpressed genes. The aim of the developed tools is to allow studying the complex expression dependences that exist between sets of coexpressed genes. For this purpose, we start detecting the nonlinear expression relationships between pairs of genes, plus the coexpressed genes. Next, we form networks among sets of coexpressed genes that maintain nonlinear expression dependences between all of them. The expression relationship between the sets of coexpressed genes is defined by the expression relationship between the skeletons of these sets, where this skeleton represents the coexpressed genes with a well-defined nonlinear expression relationship with the skeleton of the other sets. As a result, we can study the nonlinear expression relationships between a target gene and other sets of coexpressed genes, or start the study from the skeleton of the sets, to study the complex relationships of activation and deactivation between the sets of coexpressed genes that carry out the different cellular processes present in the expression experiments.

  13. Covariance Structure Models for Gene Expression Microarray Data

    ERIC Educational Resources Information Center

    Xie, Jun; Bentler, Peter M.

    2003-01-01

    Covariance structure models are applied to gene expression data using a factor model, a path model, and their combination. The factor model is based on a few factors that capture most of the expression information. A common factor of a group of genes may represent a common protein factor for the transcript of the co-expressed genes, and hence, it…

  14. Gene Expression Omnibus: NCBI gene expression and hybridization array data repository.

    PubMed

    Edgar, Ron; Domrachev, Michael; Lash, Alex E

    2002-01-01

    The Gene Expression Omnibus (GEO) project was initiated in response to the growing demand for a public repository for high-throughput gene expression data. GEO provides a flexible and open design that facilitates submission, storage and retrieval of heterogeneous data sets from high-throughput gene expression and genomic hybridization experiments. GEO is not intended to replace in house gene expression databases that benefit from coherent data sets, and which are constructed to facilitate a particular analytic method, but rather complement these by acting as a tertiary, central data distribution hub. The three central data entities of GEO are platforms, samples and series, and were designed with gene expression and genomic hybridization experiments in mind. A platform is, essentially, a list of probes that define what set of molecules may be detected. A sample describes the set of molecules that are being probed and references a single platform used to generate its molecular abundance data. A series organizes samples into the meaningful data sets which make up an experiment. The GEO repository is publicly accessible through the World Wide Web at http://www.ncbi.nlm.nih.gov/geo.

  15. Control of Neurotransmission by NaV1.7 in Human, Guinea Pig, and Mouse Airway Parasympathetic Nerves.

    PubMed

    Kocmalova, Michaela; Kollarik, Marian; Canning, Brendan J; Ru, Fei; Adam Herbstsomer, R; Meeker, Sonya; Fonquerna, Silvia; Aparici, Monica; Miralpeix, Montserrat; Chi, Xian Xuan; Li, Baolin; Wilenkin, Ben; McDermott, Jeff; Nisenbaum, Eric; Krajewski, Jeffrey L; Undem, Bradley J

    2017-04-01

    Little is known about the neuronal voltage-gated sodium channels (NaVs) that control neurotransmission in the parasympathetic nervous system. We evaluated the expression of the α subunits of each of the nine NaVs in human, guinea pig, and mouse airway parasympathetic ganglia. We combined this information with a pharmacological analysis of selective NaV blockers on parasympathetic contractions of isolated airway smooth muscle. As would be expected from previous studies, tetrodotoxin potently blocked the parasympathetic responses in the airways of each species. Gene expression analysis showed that that NaV 1.7 was virtually the only tetrodotoxin-sensitive NaV1 gene expressed in guinea pig and human airway parasympathetic ganglia, where mouse ganglia expressed NaV1.1, 1.3, and 1.7. Using selective pharmacological blockers supported the gene expression results, showing that blocking NaV1.7 alone can abolish the responses in guinea pig and human bronchi, but not in mouse airways. To block the responses in mouse airways requires that NaV1.7 along with NaV1.1 and/or NaV1.3 is blocked. These results may suggest novel indications for NaV1.7-blocking drugs, in which there is an overactive parasympathetic drive, such as in asthma. The data also raise the potential concern of antiparasympathetic side effects for systemic NaV1.7 blockers.

  16. Novel recombinant papillomavirus genomes expressing selectable genes

    PubMed Central

    Van Doorslaer, Koenraad; Porter, Samuel; McKinney, Caleb; Stepp, Wesley H.; McBride, Alison A.

    2016-01-01

    Papillomaviruses infect and replicate in keratinocytes, but viral proteins are initially expressed at low levels and there is no effective and quantitative method to determine the efficiency of infection on a cell-to-cell basis. Here we describe human papillomavirus (HPV) genomes that express marker proteins (antibiotic resistance genes and Green Fluorescent Protein), and can be used to elucidate early stages in HPV infection of primary keratinocytes. To generate these recombinant genomes, the late region of the oncogenic HPV18 genome was replaced by CpG free marker genes. Insertion of these exogenous genes did not affect early replication, and had only minimal effects on early viral transcription. When introduced into primary keratinocytes, the recombinant marker genomes gave rise to drug-resistant keratinocyte colonies and cell lines, which maintained the extrachromosomal recombinant genome long-term. Furthermore, the HPV18 “marker” genomes could be packaged into viral particles (quasivirions) and used to infect primary human keratinocytes in culture. This resulted in the outgrowth of drug-resistant keratinocyte colonies containing replicating HPV18 genomes. In summary, we describe HPV18 marker genomes that can be used to quantitatively investigate many aspects of the viral life cycle. PMID:27892937

  17. Nuclear AXIN2 represses MYC gene expression

    SciTech Connect

    Rennoll, Sherri A.; Konsavage, Wesley M.; Yochum, Gregory S.

    2014-01-03

    Highlights: •AXIN2 localizes to cytoplasmic and nuclear compartments in colorectal cancer cells. •Nuclear AXIN2 represses the activity of Wnt-responsive luciferase reporters. •β-Catenin bridges AXIN2 to TCF transcription factors. •AXIN2 binds the MYC promoter and represses MYC gene expression. -- Abstract: The β-catenin transcriptional coactivator is the key mediator of the canonical Wnt signaling pathway. In the absence of Wnt, β-catenin associates with a cytosolic and multi-protein destruction complex where it is phosphorylated and targeted for proteasomal degradation. In the presence of Wnt, the destruction complex is inactivated and β-catenin translocates into the nucleus. In the nucleus, β-catenin binds T-cell factor (TCF) transcription factors to activate expression of c-MYC (MYC) and Axis inhibition protein 2 (AXIN2). AXIN2 is a member of the destruction complex and, thus, serves in a negative feedback loop to control Wnt/β-catenin signaling. AXIN2 is also present in the nucleus, but its function within this compartment is unknown. Here, we demonstrate that AXIN2 localizes to the nuclei of epithelial cells within normal and colonic tumor tissues as well as colorectal cancer cell lines. In the nucleus, AXIN2 represses expression of Wnt/β-catenin-responsive luciferase reporters and forms a complex with β-catenin and TCF. We demonstrate that AXIN2 co-occupies β-catenin/TCF complexes at the MYC promoter region. When constitutively localized to the nucleus, AXIN2 alters the chromatin structure at the MYC promoter and directly represses MYC gene expression. These findings suggest that nuclear AXIN2 functions as a rheostat to control MYC expression in response to Wnt/β-catenin signaling.

  18. Inducible gene expression systems and plant biotechnology.

    PubMed

    Corrado, Giandomenico; Karali, Marianthi

    2009-01-01

    Plant biotechnology relies heavily on the genetic manipulation of crops. Almost invariantly, the gene of interest is expressed in a constitutive fashion, although this may not be strictly necessary for several applications. Currently, there are several regulatable expression systems for the temporal, spatial and quantitative control of transgene activity. These molecular switches are based on components derived from different organisms, which range from viruses to higher eukaryotes. Many inducible systems have been designed for fundamental and applied research and since their initial development, they have become increasingly popular in plant molecular biology. This review covers a broad number of inducible expression systems examining their properties and relevance for plant biotechnology in its various guises, from molecular breeding to pharmaceutical and industrial applications. For each system, we examine some advantages and limitations, also in relation to the strategy on which they rely. Besides being necessary to control useful genes that may negatively affect crop yield and quality, we discuss that inducible systems can be also used to increase public acceptance of GMOs, reducing some of the most common concerns. Finally, we suggest some directions and future developments for their further diffusion in agriculture and biotechnology.

  19. Nonatopic asthma: in vivo airway hyperreactivity adoptively transferred to naive mice by THY-1(+) and B220(+) antigen-specific cells that lack surface expression of CD3.

    PubMed Central

    Geba, G P; Wegner, C D; Wolyniec, W W; Li, Y; Askenase, P W

    1997-01-01

    To investigate the cellular immune events contributing to airway hyperreactivity (AHR), we studied an in vivo mouse model induced by the hapten picryl (trinitrophenyl) chloride (PCl). Mice were immunized by cutaneous contact sensitization with PCl and airway challenged subsequently with picryl sulfonic acid (PSA) antigen (Ag). Increased airway resistance was produced late (24 h) after Ag challenge, disappeared by 48 h, and was associated with no decrease in diffusion capacity. AHR could be produced in PCl immune/ PSA challenged mice on day 7 or even, with challenge, as early as 1 d after contact sensitization, after adoptive transfer of immune cells lacking CD3(+) contact sensitivity effector T cells, or after transfer of Ag-specific lymphoid cells depleted of conventional T lymphocytes with surface determinants for CD3, CD4, CD8, TCR-beta, or TCR-delta molecules. Further experiments showed that development of AHR depended upon transfer of immune cells expressing surface membrane Thy-1 and B220 (CD45RA) determinants. We concluded that a novel population of Ag-specific lymphoid cells with a defined surface phenotype (Thy-1(+), CD3(-), CD4(-), CD8(-), TCR-alphabeta-, TCR-gammadelta-, and CD45RA+) is required in a mouse model for the development of AHR. PMID:9241124

  20. Combined clustering models for the analysis of gene expression

    SciTech Connect

    Angelova, M. Ellman, J.

    2010-02-15

    Clustering has become one of the fundamental tools for analyzing gene expression and producing gene classifications. Clustering models enable finding patterns of similarity in order to understand gene function, gene regulation, cellular processes and sub-types of cells. The clustering results however have to be combined with sequence data or knowledge about gene functionality in order to make biologically meaningful conclusions. In this work, we explore a new model that integrates gene expression with sequence or text information.

  1. Using PCR to Target Misconceptions about Gene Expression

    PubMed Central

    Wright, Leslie K.; Newman, Dina L.

    2013-01-01

    We present a PCR-based laboratory exercise that can be used with first- or second-year biology students to help overcome common misconceptions about gene expression. Biology students typically do not have a clear understanding of the difference between genes (DNA) and gene expression (mRNA/protein) and often believe that genes exist in an organism or cell only when they are expressed. This laboratory exercise allows students to carry out a PCR-based experiment designed to challenge their misunderstanding of the difference between genes and gene expression. Students first transform E. coli with an inducible GFP gene containing plasmid and observe induced and un-induced colonies. The following exercise creates cognitive dissonance when actual PCR results contradict their initial (incorrect) predictions of the presence of the GFP gene in transformed cells. Field testing of this laboratory exercise resulted in learning gains on both knowledge and application questions on concepts related to genes and gene expression. PMID:23858358

  2. Hyperbaric oxygen treatment induces antioxidant gene expression.

    PubMed

    Godman, Cassandra A; Joshi, Rashmi; Giardina, Charles; Perdrizet, George; Hightower, Lawrence E

    2010-06-01

    Although the underlying molecular causes of aging are not entirely clear, hormetic agents like exercise, heat, and calorie restriction may generate a mild pro-oxidant stress that induces cell protective responses to promote healthy aging. As an individual ages, many cellular and physiological processes decline, including wound healing and reparative angiogenesis. This is particularly critical in patients with chronic non-healing wounds who tend to be older. We are interested in the potential beneficial effects of hyperbaric oxygen as a mild hormetic stress on human microvascular endothelial cells. We analyzed global gene expression changes in human endothelial cells following a hyperbaric exposure comparable to a clinical treatment. Our analysis revealed an upregulation of antioxidant, cytoprotective, and immediate early genes. This increase coincided with an increased resistance to a lethal oxidative stress. Our data indicate that hyperbaric oxygen can induce protection against oxidative insults in endothelial cells and may provide an easily administered hormetic treatment to help promote healthy aging.

  3. Expressing exogenous genes in newts by transgenesis.

    PubMed

    Casco-Robles, Martin Miguel; Yamada, Shouta; Miura, Tomoya; Nakamura, Kenta; Haynes, Tracy; Maki, Nobuyasu; Del Rio-Tsonis, Katia; Tsonis, Panagiotis A; Chiba, Chikafumi

    2011-05-01

    The great regenerative abilities of newts provide the impetus for studies at the molecular level. However, efficient methods for gene regulation have historically been quite limited. Here we describe a protocol for transgenically expressing exogenous genes in the newt Cynops pyrrhogaster. This method is simple: a reaction mixture of I-SceI meganuclease and a plasmid DNA carrying a transgene cassette flanked by the enzyme recognition sites is directly injected into fertilized eggs. The protocol achieves a high efficiency of transgenesis, comparable to protocols used in other animal systems, and it provides a practical number of transgenic newts (∼20% of injected embryos) that survive beyond metamorphosis and that can be applied to regenerative studies. The entire protocol for obtaining transgenic adult newts takes 4-5 months.

  4. Gene expression signatures in lymphoid tumours.

    PubMed

    Kees, Ursula R

    2004-04-01

    Lymphoid tumours comprise the acute and chronic leukaemias, the broad spectrum of lymphomas, including Hodgkin's disease, and multiple myeloma. The subdivision of the acute leukaemias according to the proliferating type of white blood cells has had a major impact on the care of these patients. More recently, specific chromosomal translocations have been used to identify patients who may benefit from more intensive therapies. The novel high-throughput genomic technologies, such as microarrays, provide new avenues for the molecular diagnosis of the haematological malignancies. Rapid advances in genome sequencing and gene expression profiling provide unprecedented opportunities to identify specific genes involved in complex biological processes, including tumorigenesis. The features of microarray technology and the variety of experimental approaches to elucidate lymphoid malignancies are discussed. Microarray technology has the potential to lead to more accurate prognostic assessment for patients and is expected to ultimately allow the clinician to select therapies optimally suited to each patient.

  5. Retrotransposons as regulators of gene expression.

    PubMed

    Elbarbary, Reyad A; Lucas, Bronwyn A; Maquat, Lynne E

    2016-02-12

    Transposable elements (TEs) are both a boon and a bane to eukaryotic organisms, depending on where they integrate into the genome and how their sequences function once integrated. We focus on two types of TEs: long interspersed elements (LINEs) and short interspersed elements (SINEs). LINEs and SINEs are retrotransposons; that is, they transpose via an RNA intermediate. We discuss how LINEs and SINEs have expanded in eukaryotic genomes and contribute to genome evolution. An emerging body of evidence indicates that LINEs and SINEs function to regulate gene expression by affecting chromatin structure, gene transcription, pre-mRNA processing, or aspects of mRNA metabolism. We also describe how adenosine-to-inosine editing influences SINE function and how ongoing retrotransposition is countered by the body's defense mechanisms.

  6. Gene expression-targeted isoflavone therapy.

    PubMed

    Węgrzyn, Alicja

    2012-04-01

    Lysosomal storage diseases (LSD) form a group of inherited metabolic disorders caused by dysfunction of one of the lysosomal proteins, resulting in the accumulation of certain compounds. Although these disorders are among first genetic diseases for which specific treatments were proposed, there are still serious unsolved problems that require development of novel therapeutic procedures. An example is neuronopathy, which develops in most of LSD and cannot be treated efficiently by currently approved therapies. Recently, a new potential therapy, called gene expression-targeted isoflavone therapy (GET IT), has been proposed for a group of LSD named mucopolysaccharidoses (MPS), in which storage of incompletely degraded glycosaminoglycans (GAGs) results in severe symptoms of virtually all tissues and organs, including central nervous system. The idea of this therapy is to inhibit synthesis of GAGs by modulating expression of genes coding for enzymes involved in synthesis of these compounds. Such a modulation is possible by using isoflavones, particularly genistein, which interfere with a signal transduction process necessary for stimulation of expression of certain genes. Results of in vitro experiments and studies on animal models indicated a high efficiency of GET IT, including correction of behavior of affected mice. However, clinical trials, performed with soy isoflavone extracts, revealed only limited efficacy. This caused a controversy about GET IT as a potential, effective treatment of patients suffering from MPS, especially neuronopathic forms of these diseases. It this critical review, I present possible molecular mechanisms of therapeutic action of isoflavones (particularly genistein) and suggest that efficacy of GET IT might be sufficiently high when using relatively high doses of synthetic genistein (which was employed in experiments on cell cultures and mouse models) rather than low doses of soy isoflavone extracts (which were used in clinical trials). This

  7. Maternal diet programs embryonic kidney gene expression.

    PubMed

    Welham, Simon J M; Riley, Paul R; Wade, Angie; Hubank, Mike; Woolf, Adrian S

    2005-06-16

    Human epidemiological data associating birth weight with adult disease suggest that organogenesis is "programmed" by maternal diet. In rats, protein restriction in pregnancy produces offspring with fewer renal glomeruli and higher systemic blood pressures than controls. We tested the hypothesis that maternal diet alters gene expression in the metanephros, the precursor of the definitive mammalian kidney. We demonstrated that maternal low-protein diet initiated when pregnancy starts and maintained to embryonic day 13, when the metanephros consists of mesenchyme surrounding a once-branched ureteric bud, is sufficient to significantly reduce glomerular numbers in offspring by about 20%. As assessed by representational difference analyses and real-time quantitative polymerase chain reactions, low-protein diet modulated gene expression in embryonic day 13 metanephroi. In particular, levels of prox-1, the ortholog of Drosophila transcription factor prospero, and cofilin-1, a regulator of the actin cytoskeleton, were reduced. During normal metanephrogenesis, prox-1 protein was first detected in mesenchymal cells around the ureteric tree and thereafter in nascent nephron epithelia, whereas cofilin-1 immunolocalized to bud derivatives and condensing mesenchyme. Previously, we reported that low-protein diets increased mesenchymal apoptosis cells when metanephrogenesis began and thereafter reduced numbers of precursor cells. Collectively, these studies prove that the maternal diet programs the embryonic kidney, altering cell turnover and gene expression at a time when nephrons and glomeruli have yet to form. The human implication is that the maternal diet ingested between conception and 5- 6-wk gestation contributes to the variation in glomerular numbers that are known to occur between healthy and hypertensive populations.

  8. Pathway network inference from gene expression data

    PubMed Central

    2014-01-01

    Background The development of high-throughput omics technologies enabled genome-wide measurements of the activity of cellular elements and provides the analytical resources for the progress of the Systems Biology discipline. Analysis and interpretation of gene expression data has evolved from the gene to the pathway and interaction level, i.e. from the detection of differentially expressed genes, to the establishment of gene interaction networks and the identification of enriched functional categories. Still, the understanding of biological systems requires a further level of analysis that addresses the characterization of the interaction between functional modules. Results We present a novel computational methodology to study the functional interconnections among the molecular elements of a biological system. The PANA approach uses high-throughput genomics measurements and a functional annotation scheme to extract an activity profile from each functional block -or pathway- followed by machine-learning methods to infer the relationships between these functional profiles. The result is a global, interconnected network of pathways that represents the functional cross-talk within the molecular system. We have applied this approach to describe the functional transcriptional connections during the yeast cell cycle and to identify pathways that change their connectivity in a disease condition using an Alzheimer example. Conclusions PANA is a useful tool to deepen in our understanding of the functional interdependences that operate within complex biological systems. We show the approach is algorithmically consistent and the inferred network is well supported by the available functional data. The method allows the dissection of the molecular basis of the functional connections and we describe the different regulatory mechanisms that explain the network's topology obtained for the yeast cell cycle data. PMID:25032889

  9. Altered gene expression correlates with DNA structure.

    PubMed

    Kohwi, Y; Kohwi-Shigematsu, T

    1991-12-01

    We examined the participation of triplex DNA structure in gene regulation using a poly(dG)-poly(dC) sequence as a model. We show that a poly(dG)-poly(dC) sequence, which can adopt an intramolecular dG.dG.dC triplex under superhelical strain, strongly augments gene expression when placed 5' to a promoter. The activity of this sequence exhibits a striking length dependency: dG tracts of 27-30 bp augment the expression of a reporter gene to a level comparable to that observed with the polyoma enhancer in mouse LTK- cells, whereas tracts of 35 bp and longer have virtually no effect. A supercoiled plasmid containing a dG tract of 30 bp competes in vivo for a trans-acting factor as revealed by reduction in the reporter gene transcription driven by the (dG)29/promoter of the test plasmid, while dGs of 35 bp and longer in the competition plasmid failed to compete. In purified supercoiled plasmid DNA at a superhelical density of -0.05, dG tracts of 32 bp and longer form a triplex, whereas those of 30 bp and shorter remain double-stranded under a PBS solution. These results suggest that a localized superhelical strain can exist, at least transiently, in mouse LTK- cells, and before being relaxed by topoisomerases this rapidly induces dG tracts of 35 bp and longer to adopt a triplex preventing the factor from binding. Thus, these data suggest that a poly(dG)-poly(dC) sequence can function as a negative regulator by adopting an intramolecular triple helix structure in vivo.

  10. Dynamics of single-cell gene expression

    PubMed Central

    Longo, Diane; Hasty, Jeff

    2006-01-01

    Cellular behavior has traditionally been investigated by utilizing bulk-scale methods that measure average values for a population of cells. Such population-wide studies mask the behavior of individual cells and are often insufficient for characterizing biological processes in which cellular heterogeneity plays a key role. A unifying theme of many recent studies has been a focus on the development and utilization of single-cell experimental techniques that are capable of probing key biological phenomena in individual living cells. Recently, novel information about gene expression dynamics has been obtained from single-cell experiments that draw upon the unique capabilities of fluorescent reporter proteins. PMID:17130866

  11. Solid state nanopores for gene expression profiling

    NASA Astrophysics Data System (ADS)

    Mussi, V.; Fanzio, P.; Repetto, L.; Firpo, G.; Valbusa, U.; Scaruffi, P.; Stigliani, S.; Tonini, G. P.

    2009-07-01

    Recently, nanopore technology has been introduced for genome analysis. Here we show results related to the possibility of preparing "engineered solid state nanopores". The nanopores were fabricated on a suspended Si 3N 4 membrane by Focused Ion Beam (FIB) drilling and chemically functionalized in order to covalently bind oligonucleotides (probes) on their surface. Our data show the stable effect of DNA attachment on the ionic current measured through the nanopore, making it possible to conceive and develop a selective biosensor for gene expression profiling.

  12. Clinical diagnostic gene expression thyroid testing.

    PubMed

    Steward, David L; Kloos, Richard T

    2014-08-01

    Thyroid fine-needle aspiration biopsies are cytologically indeterminate in 15% to 30% of cases. When cytologically indeterminate thyroid nodules undergo diagnostic surgery, approximately three-quarters prove to be histologically benign. A negative predictive value of more than or equal to 94% for the Afirma Gene Expression Classifier (GEC) is achieved for indeterminate nodules. Most Afirma GEC benign nodules can be clinically observed, as suggested by the National Comprehensive Cancer Network Thyroid Carcinoma Guideline. More than half of the benign nodules with indeterminate cytology (Bethesda categories III/IV) can be identified as GEC benign and removed from the surgical pool to prevent unnecessary diagnostic surgery.

  13. Clustering gene expression data using graph separators.

    PubMed

    Kaba, Bangaly; Pinet, Nicolas; Lelandais, Gaëlle; Sigayret, Alain; Berry, Anne

    2007-01-01

    Recent work has used graphs to modelize expression data from microarray experiments, in view of partitioning the genes into clusters. In this paper, we introduce the use of a decomposition by clique separators. Our aim is to improve the classical clustering methods in two ways: first we want to allow an overlap between clusters, as this seems biologically sound, and second we want to be guided by the structure of the graph to define the number of clusters. We test this approach with a well-known yeast database (Saccharomyces cerevisiae). Our results are good, as the expression profiles of the clusters we find are very coherent. Moreover, we are able to organize into another graph the clusters we find, and order them in a fashion which turns out to respect the chronological order defined by the the sporulation process.

  14. Gene expression during the life cycle of Drosophila melanogaster.

    PubMed

    Arbeitman, Michelle N; Furlong, Eileen E M; Imam, Farhad; Johnson, Eric; Null, Brian H; Baker, Bruce S; Krasnow, Mark A; Scott, Matthew P; Davis, Ronald W; White, Kevin P

    2002-09-27

    Molecular genetic studies of Drosophila melanogaster have led to profound advances in understanding the regulation of development. Here we report gene expression patterns for nearly one-third of all Drosophila genes during a complete time course of development. Mutations that eliminate eye or germline tissue were used to further analyze tissue-specific gene expression programs. These studies define major characteristics of the transcriptional programs that underlie the life cycle, compare development in males and females, and show that large-scale gene expression data collected from whole animals can be used to identify genes expressed in particular tissues and organs or genes involved in specific biological and biochemical processes.

  15. Gene Expression During the Life Cycle of Drosophila melanogaster

    NASA Astrophysics Data System (ADS)

    Arbeitman, Michelle N.; Furlong, Eileen E. M.; Imam, Farhad; Johnson, Eric; Null, Brian H.; Baker, Bruce S.; Krasnow, Mark A.; Scott, Matthew P.; Davis, Ronald W.; White, Kevin P.

    2002-09-01

    Molecular genetic studies of Drosophila melanogaster have led to profound advances in understanding the regulation of development. Here we report gene expression patterns for nearly one-third of all Drosophila genes during a complete time course of development. Mutations that eliminate eye or germline tissue were used to further analyze tissue-specific gene expression programs. These studies define major characteristics of the transcriptional programs that underlie the life cycle, compare development in males and females, and show that large-scale gene expression data collected from whole animals can be used to identify genes expressed in particular tissues and organs or genes involved in specific biological and biochemical processes.

  16. An extensive network of coupling among gene expression machines.

    PubMed

    Maniatis, Tom; Reed, Robin

    2002-04-04

    Gene expression in eukaryotes requires several multi-component cellular machines. Each machine carries out a separate step in the gene expression pathway, which includes transcription, several pre-messenger RNA processing steps and the export of mature mRNA to the cytoplasm. Recent studies lead to the view that, in contrast to a simple linear assembly line, a complex and extensively coupled network has evolved to coordinate the activities of the gene expression machines. The extensive coupling is consistent with a model in which the machines are tethered to each other to form 'gene expression factories' that maximize the efficiency and specificity of each step in gene expression.

  17. ZN2+-INDUCED IL-8 EXPRESSION INVOLVES AP-1, JNK, AND ERK ACTIVITIES IN HUMAN AIRWAY EPITHELIAL CELLS

    EPA Science Inventory

    Exposure to zinc-laden particulate matter (PM) in ambient and occupational settings has been associated with proinflammatory responses in the lung. IL-8 is an important proinflammatory cytokine in the human lung and is induced in human airway epithelial cells exposed to zin...

  18. Soluble ADAM33 initiates airway remodeling to promote susceptibility for allergic asthma in early life

    PubMed Central

    Davies, Elizabeth R.; Kelly, Joanne F.C.; Howarth, Peter H.; Wilson, David I.; Holgate, Stephen T.; Davies, Donna E.; Whitsett, Jeffrey A.

    2016-01-01

    Asthma is a chronic inflammatory airways disease that usually begins in early life and involves gene-environment interactions. Although most asthma exhibits allergic inflammation, many allergic individuals do not have asthma. Here, we report how the asthma gene a disintegrin and metalloprotease 33 (ADAM33) acts as local tissue susceptibility gene that promotes allergic asthma. We show that enzymatically active soluble ADAM33 (sADAM33) is increased in asthmatic airways and plays a role in airway remodeling, independent of inflammation. Furthermore, remodeling and inflammation are both suppressed in Adam33-null mice after allergen challenge. When induced in utero or added ex vivo, sADAM33 causes structural remodeling of the airways, which enhances postnatal airway eosinophilia and bronchial hyperresponsiveness following subthreshold challenge with an aeroallergen. This substantial gene-environment interaction helps to explain the end-organ expression of allergic asthma in genetically susceptible individuals. Finally, we show that sADAM33-induced airway remodeling is reversible, highlighting the therapeutic potential of targeting ADAM33 in asthma. PMID:27489884

  19. A Double Selection Approach to Achieve Specific Expression of Toxin Genes for Ovarian Cancer Gene Therapy

    DTIC Science & Technology

    2006-11-01

    specific expression of toxin genes for ovarian cancer gene therapy PRINCIPAL INVESTIGATOR: David T. Curiel, M.D., Ph.D. Gene Siegal...A double selection approach to achieve specific expression of toxin genes for ovarian cancer gene therapy 5b. GRANT NUMBER W81XWH-05-1-0035...cancer. This system should result in highly efficient and specific expression of toxin encoding genes in tumor cells, enabling these cells to be

  20. Repeated Delivery of Adeno-Associated Virus Vectors to the Rabbit Airway

    PubMed Central

    Beck, Suzanne E.; Jones, Lori A.; Chesnut, Kye; Walsh, Scott M.; Reynolds, Thomas C.; Carter, Barrie J.; Askin, Frederic B.; Flotte, Terence R.; Guggino, William B.

    1999-01-01

    Efficient local expression from recombinant adeno-associated virus (rAAV)-cystic fibrosis (CF) transmembrane conductance regulator (CFTR) vectors has been observed in the airways of rabbits and monkeys for up to 6 months following a single bronchoscopic delivery. However, it is likely that repeated administrations of rAAV vectors will be necessary for sustained correction of the CF defect in the airways. The current study was designed to test the feasibility of repeated airway delivery of rAAV vectors in the rabbit lung. After two doses of rAAV-CFTR to the airways, rabbits generated high titers of serum anti-AAV neutralizing antibodies. Rabbits then received a third dose of a rAAV vector containing the green fluorescent protein (GFP) reporter gene packaged in either AAV serotype 2 (AAV2) or serotype 3 (AAV3) capsids. Each dose consisted of 1 ml containing 5 × 109 DNase-resistant particles of rAAV vector, having no detectable replication-competent AAV or adenovirus. Three weeks later, GFP expression was observed in airway epithelial cells despite high anti-AAV neutralizing titers at the time of delivery. There was no significant difference in the efficiency of DNA transfer or expression between the rAAV3 and rAAV2 groups. No significant inflammatory responses to either repeated airway exposure to rAAV2-CFTR vectors or to GFP expression were observed. These experiments demonstrate that serum anti-AAV neutralizing antibody titers do not predict airway neutralization in vivo and that repeated airway delivery rAAV allows for safe and effective gene transfer. PMID:10516053

  1. Effects of transient receptor potential canonical 1 (TRPC1) on the mechanical stretch-induced expression of airway remodeling-associated factors in human bronchial epithelioid cells.

    PubMed

    Yu, Qian; Li, Minchao

    2017-01-25

    Research has shown that mechanical stress stimulation can cause airway remodeling. We investigate the effects of mechanical stretch on the expression of the airway remodeling-associated factors interleukin-13 (IL-13) and matrix metalloprotein-9 (MMP-9) and signaling pathways in human bronchial epithelioid (16HBE) cells under mechanical stretch. A Flexcell FX-4000 Tension System with a flexible substrate was applied to stretch 16HBE cells at a 15% elongation amplitude and 1Hz frequency, with stretching for 0.5h, 1h, 1.5h and 2h. The experimental group with higher IL-13, MMP-9, and TRPC1 expression and higher Ca(2+) levels was selected for performing intervention experiment. These cells were pretreated with the transient receptor potential canonical 1 (TRPC1) channel antagonist SKF96365 and TRPC1-specific siRNA, and then mechanical stretch was applied. Our results provided evidences that mechanical pressure significantly increased IL-13, MMP-9, and TRPC1 protein and mRNA expression levels and intracellular Ca(2+) fluorescence intensity at 4 time points compared with the control group. The peak IL-13, MMP-9, and TRPC1 expression levels were observed at 0.5h after exposure to mechanical pressure. IL-13 and MMP-9 expression levels and Ca(2+) fluorescence intensity in the stretch+SKF96365 group and in the stretch+TRPC1 siRNA group were significantly lower than those were in the mechanical stretch group. By incubating the cells with the intracellular calcium chelator BAPTA-AM, the expression of IL-13 and MMP9 was significantly decreased, and the expression level of TRPC1 remained unchanged. These observations suggest that mechanical stretch may induce an influx of Ca(2+) and up-regulation of IL-13 and MMP-9 expression in 16HBE cells via activation of TRPC1.

  2. Differential gene expression in anatomical compartments of the human eye

    PubMed Central

    Diehn, Jennifer J; Diehn, Maximilian; Marmor, Michael F; Brown, Patrick O

    2005-01-01

    Background The human eye is composed of multiple compartments, diverse in form, function, and embryologic origin, that work in concert to provide us with our sense of sight. We set out to systematically characterize the global gene expression patterns that specify the distinctive characteristics of the various eye compartments. Results We used DNA microarrays representing approximately 30,000 human genes to analyze gene expression in the cornea, lens, iris, ciliary body, retina, and optic nerve. The distinctive patterns of expression in each compartment could be interpreted in relation to the physiology and cellular composition of each tissue. Notably, the sets of genes selectively expressed in the retina and in the lens were particularly large and diverse. Genes with roles in immune defense, particularly complement components, were expressed at especially high levels in the anterior segment tissues. We also found consistent differences between the gene expression patterns of the macula and peripheral retina, paralleling the differences in cell layer densities between these regions. Based on the hypothesis that genes responsible for diseases that affect a particular eye compartment are likely to be selectively expressed in that compartment, we compared our gene expression signatures with genetic mapping studies to identify candidate genes for diseases affecting the cornea, lens, and retina. Conclusion Through genome-scale gene expression profiling, we were able to discover distinct gene expression 'signatures' for each eye compartment and identified candidate disease genes that can serve as a reference database for investigating the physiology and pathophysiology of the eye. PMID:16168081

  3. Identification of human HK genes and gene expression regulation study in cancer from transcriptomics data analysis.

    PubMed

    Chen, Meili; Xiao, Jingfa; Zhang, Zhang; Liu, Jingxing; Wu, Jiayan; Yu, Jun

    2013-01-01

    The regulation of gene expression is essential for eukaryotes, as it drives the processes of cellular differentiation and morphogenesis, leading to the creation of different cell types in multicellular organisms. RNA-Sequencing (RNA-Seq) provides researchers with a powerful toolbox for characterization and quantification of transcriptome. Many different human tissue/cell transcriptome datasets coming from RNA-Seq technology are available on public data resource. The fundamental issue here is how to develop an effective analysis method to estimate expression pattern similarities between different tumor tissues and their corresponding normal tissues. We define the gene expression pattern from three directions: 1) expression breadth, which reflects gene expression on/off status, and mainly concerns ubiquitously expressed genes; 2) low/high or constant/variable expression genes, based on gene expression level and variation; and 3) the regulation of gene expression at the gene structure level. The cluster analysis indicates that gene expression pattern is higher related to physiological condition rather than tissue spatial distance. Two sets of human housekeeping (HK) genes are defined according to cell/tissue types, respectively. To characterize the gene expression pattern in gene expression level and variation, we firstly apply improved K-means algorithm and a gene expression variance model. We find that cancer-associated HK genes (a HK gene is specific in cancer group, while not in normal group) are expressed higher and more variable in cancer condition than in normal condition. Cancer-associated HK genes prefer to AT-rich genes, and they are enriched in cell cycle regulation related functions and constitute some cancer signatures. The expression of large genes is also avoided in cancer group. These studies will help us understand which cell type-specific patterns of gene expression differ among different cell types, and particularly for cancer.

  4. Airway mucus: From production to secretion.

    PubMed

    Williams, Olatunji W; Sharafkhaneh, Amir; Kim, Victor; Dickey, Burton F; Evans, Christopher M

    2006-05-01

    Mucus hypersecretion is a phenotype associated with multiple obstructive lung diseases. However, in spite of its nefarious reputation under pathologic conditions, there are significant benefits to having low levels of mucus present in the airways at baseline, such as the ability to trap and eliminate inhaled particles and to prevent desiccation of airway surfaces. Mucins are high-molecular-weight glycoproteins that are the chief components that render viscoelastic and gel-forming properties to mucus. Recent advances in animal models and in vitro systems have provided a wealth of information regarding the identification of the mucin genes that are expressed in the lungs, the signal transduction pathways that regulate the expression of these mucins, and the secretory pathways that mediate their release into the airways. In addition, the clinical and pathologic literature has corroborated many of the basic laboratory findings. As a result, mucin overproduction and hypersecretion are moving away from being markers of disease and toward being testable as functional components of lung disease processes.

  5. Gut microbiota, host gene expression, and aging.

    PubMed

    Patrignani, Paola; Tacconelli, Stefania; Bruno, Annalisa

    2014-01-01

    Novel concepts of disease susceptibility and development suggest an important role of gastrointestinal microbiota and microbial pathogens. They can contribute to physiological systems and disease processes, even outside of the gastrointestinal tract. There is increasing evidence that genetics of the host influence and interact with gut microbiota. Moreover, aging-associated oxidative stress may cause morphologic alterations of bacterial cells, thus influencing the aggressive potential and virulence markers of an anaerobic bacterium and finally the type of interaction with the host. At the same time, microbiota may influence host gene expression and it is becoming apparent that it may occur through the regulation of microRNAs. They are short single-stranded noncoding RNAs that regulate posttranscriptional gene expression by affecting mRNA stability and/or translational repression of their target mRNAs. The introduction of -omics approaches (such as metagenomics, metaproteomics, and metatranscriptomics) in microbiota research will certainly advance our knowledge of this area. This will lead to greatly deepen our understanding of the molecular targets in the homeostatic interaction between the gut microbiota and the host and, thereby, promises to reveal new ways to treat diseases and maintain health.

  6. Increased oxidative DNA damage and decreased expression of base excision repair proteins in airway epithelial cells of women who cook with biomass fuels.

    PubMed

    Mukherjee, Bidisha; Bindhani, Banani; Saha, Hirak; Ray, Manas Ranjan

    2014-09-01

    To investigate whether biomass burning causes oxidative DNA damage and alters the expression of DNA base excision repair (BER) proteins in airway cells, sputum samples were collected from 80 premenopausal rural biomass-users and 70 age-matched control women who cooked with liquefied petroleum gas. Compared with control the airway cells of biomass-users showed increased DNA damage in alkaline comet assay. Biomass-users showed higher percentage of cells expressing oxidative DNA damage marker 8-oxoguanine and lower percentages of BER proteins OGG1 and APE1 by immunocytochemical staining. Reactive oxygen species (ROS) generation was doubled and level of superoxide dismutase was depleted significantly among biomass-users. The concentrations of particulate matters were higher in biomass-using households which positively correlated with ROS generation and negatively with BER proteins expressions. ROS generation was positively correlated with 8-oxoguanine and negatively with BER proteins suggesting cooking with biomass is a risk for genotoxicity among rural women in their child-bearing age.

  7. Posttranscriptional Control of Gene Expression in Yeast

    PubMed Central

    McCarthy, John E. G.

    1998-01-01

    Studies of the budding yeast Saccharomyces cerevisiae have greatly advanced our understanding of the posttranscriptional steps of eukaryotic gene expression. Given the wide range of experimental tools applicable to S. cerevisiae and the recent determination of its complete genomic sequence, many of the key challenges of the posttranscriptional control field can be tackled particularly effectively by using this organism. This article reviews the current knowledge of the cellular components and mechanisms related to translation and mRNA decay, with the emphasis on the molecular basis for rate control and gene regulation. Recent progress in characterizing translation factors and their protein-protein and RNA-protein interactions has been rapid. Against the background of a growing body of structural information, the review discusses the thermodynamic and kinetic principles that govern the translation process. As in prokaryotic systems, translational initiation is a key point of control. Modulation of the activities of translational initiation factors imposes global regulation in the cell, while structural features of particular 5′ untranslated regions, such as upstream open reading frames and effector binding sites, allow for gene-specific regulation. Recent data have revealed many new details of the molecular mechanisms involved while providing insight into the functional overlaps and molecular networking that are apparently a key feature of evolving cellular systems. An overall picture of the mechanisms governing mRNA decay has only very recently begun to develop. The latest work has revealed new information about the mRNA decay pathways, the components of the mRNA degradation machinery, and the way in which these might relate to the translation apparatus. Overall, major challenges still to be addressed include the task of relating principles of posttranscriptional control to cellular compartmentalization and polysome structure and the role of molecular channelling

  8. Social regulation of cortisol receptor gene expression

    PubMed Central

    Korzan, Wayne J.; Grone, Brian P.; Fernald, Russell D.

    2014-01-01

    In many social species, individuals influence the reproductive capacity of conspecifics. In a well-studied African cichlid fish species, Astatotilapia burtoni, males are either dominant (D) and reproductively competent or non-dominant (ND) and reproductively suppressed as evidenced by reduced gonadotropin releasing hormone (GnRH1) release, regressed gonads, lower levels of androgens and elevated levels of cortisol. Here, we asked whether androgen and cortisol levels might regulate this reproductive suppression. Astatotilapia burtoni has four glucocorticoid receptors (GR1a, GR1b, GR2 and MR), encoded by three genes, and two androgen receptors (ARα and ARβ), encoded by two genes. We previously showed that ARα and ARβ are expressed in GnRH1 neurons in the preoptic area (POA), which regulates reproduction, and that the mRNA levels of these receptors are regulated by social status. Here, we show that GR1, GR2 and MR mRNAs are also expressed in GnRH1 neurons in the POA, revealing potential mechanisms for both androgens and cortisol to influence reproductive capacity. We measured AR, MR and GR mRNA expression levels in a microdissected region of the POA containing GnRH1 neurons, comparing D and ND males. Using quantitative PCR (qPCR), we found D males had higher mRNA levels of ARα, MR, total GR1a and GR2 in the POA compared with ND males. In contrast, ND males had significantly higher levels of GR1b mRNA, a receptor subtype with a reduced transcriptional response to cortisol. Through this novel regulation of receptor type, neurons in the POA of an ND male will be less affected by the higher levels of cortisol typical of low status, suggesting GR receptor type change as a potential adaptive mechanism to mediate high cortisol levels during social suppression. PMID:25013108

  9. Expressing genes do not forget their LINEs: transposable elements and gene expression.

    PubMed

    Kines, Kristine J; Belancio, Victoria P

    2012-01-01

    Historically the accumulated mass of mammalian transposable elements (TEs), particularly those located within gene boundaries, was viewed as a genetic burden potentially detrimental to the genomic landscape. This notion has been strengthened by the discovery that transposable sequences can alter the architecture of the transcriptome, not only through insertion, but also long after the integration process is completed. Insertions previously considered harmless are now known to impact the expression of host genes via modification of the transcript quality or quantity, transcriptional interference, or by the control of pathways that affect the mRNA life-cycle. Conversely, several examples of the evolutionary advantageous impact of TEs on the host gene structure that diversified the cellular transcriptome are reported. TE-induced changes in gene expression can be tissue- or disease-specific, raising the possibility that the impact of TE sequences may vary during development, among normal cell types, and between normal and disease-affected tissues. The understanding of the rules and abundance of TE-interference with gene expression is in its infancy, and its contribution to human disease and/or evolution remains largely unexplored.

  10. [Mechanism on differential gene expression and heterosis formation].

    PubMed

    Xu, Chen-Lu; Sun, Xiao-Mei; Zhang, Shou-Gong

    2013-06-01

    Despite the rediscovery of heterosis about a century ago and the suggestion of various genetic models to explain this phenomenon, little consensus has yet been reached about the genetic basis of heterosis. Following the genome organization variation and gene effects, an understanding of gene differential expression in hybrids and its parents provides a new opportunity to speculate on mechanisms that might lead to heterosis. Investigation on allele-specific gene expression in hybrid and gene differential expression between hybrids and its parents might contribute to improve our understanding of the molecular basis of heterosis and eventually guide breeding practices. In this review, we discussed the recent researches on allelic-specific expression in hybrid which was frequently observed in recent studies and analyzed its regulatory mechanism. All possible modes of gene action, including additivity, high- and low-parent dominance, underdominance, and over-dominance, were observed when investigating gene differential expression between hybrids and its parents. Data from transcriptomic studies screened several heterosis-associated genes and highlighted the importance of certain key biochemical pathways that may prove to be quintessential for the manifestation of heterosis. So far, no uniform global expression pat-terns were observed in these gene expression studies. Most heterosis-associated gene expression analyses have not revealed a predominant functional category to which differentially expressed genes belong. However, these gene expression profiling studies represent a first step towards the definition of the complex gene expression networks that might be relevant in the context of heterosis. New technique on gene expression profile and advancements in bioinformatics will facilitate our understanding of the genetic basis of heterosis at the gene-expression level.

  11. Correspondence between resting state activity and brain gene expression

    PubMed Central

    Wang, Guang-Zhong; Belgard, T. Grant; Mao, Deng; Chen, Leslie; Berto, Stefano; Preuss, Todd M.; Lu, Hanzhang; Geschwind, Daniel H.; Konopka, Genevieve

    2015-01-01

    SUMMARY The relationship between functional brain activity and gene expression has not been fully explored in the human brain. Here, we identify significant correlations between gene expression in the brain and functional activity by comparing fractional Amplitude of Low Frequency Fluctuations (fALFF) from two independent human fMRI resting state datasets to regional cortical gene expression from a newly generated RNA-seq dataset and two additional gene expression datasets to obtain robust and reproducible correlations. We find significantly more genes correlated with fALFF than expected by chance, and identify specific genes correlated with the imaging signals in multiple expression datasets in the default mode network. Together, these data support a population-level relationship between regional steady state brain gene expression and resting state brain activity. PMID:26590343

  12. Homologous versus heterologous gene expression in the yeast, Saccharomyces cerevisiae.

    PubMed Central

    Chen, C Y; Oppermann, H; Hitzeman, R A

    1984-01-01

    DNA sequences normally flanking the highly expressed yeast 3-phosphoglycerate kinase (PGK) gene have been placed adjacent to heterologous mammalian genes on high copy number plasmid vectors and used for expression experiments in yeast. For many genes thus far expressed with this system, expression has been 15-50 times lower than the expression of the natural homologous PGK gene on the same plasmid. We have extensively investigated this dramatic difference and have found that in most cases it is directly proportional to the steady-state levels of mRNAs. We demonstrate this phenomenon and suggest possible causes for this effect on mRNA levels. Images PMID:6096814

  13. Sequence determinants of prokaryotic gene expression level under heat stress.

    PubMed

    Xiong, Heng; Yang, Yi; Hu, Xiao-Pan; He, Yi-Ming; Ma, Bin-Guang

    2014-11-01

    Prokaryotic gene expression is environment-dependent and temperature plays an important role in shaping the gene expression profile. Revealing the regulation mechanisms of gene expression pertaining to temperature has attracted tremendous efforts in recent years particularly owning to the yielding of transcriptome and proteome data by high-throughput techniques. However, most of the previous works concentrated on the characterization of the gene expression profile of individual organism and little effort has been made to disclose the commonality among organisms, especially for the gene sequence features. In this report, we collected the transcriptome and proteome data measured under heat stress condition from recently published literature and studied the sequence determinants for the expression level of heat-responsive genes on multiple layers. Our results showed that there indeed exist commonness and consistent patterns of the sequence features among organisms for the differentially expressed genes under heat stress condition. Some features are attributed to the requirement of thermostability while some are dominated by gene function. The revealed sequence determinants of bacterial gene expression level under heat stress complement the knowledge about the regulation factors of prokaryotic gene expression responding to the change of environmental conditions. Furthermore, comparisons to thermophilic adaption have been performed to reveal the similarity and dissimilarity of the sequence determinants for the response to heat stress and for the adaption to high habitat temperature, which elucidates the complex landscape of gene expression related to the same physical factor of temperature.

  14. NOTCH1 is required for regeneration of Clara cells during repair of airway injury.

    PubMed

    Xing, Yiming; Li, Aimin; Borok, Zea; Li, Changgong; Minoo, Parviz

    2012-05-01

    The airways of the mammalian lung are lined with highly specialized epithelial cell types that are the targets of airborne toxicants and injury. Notch signaling plays an important role in the ontogeny of airway epithelial cells, but its contributions to recruitment, expansion or differentiation of resident progenitor/stem cells, and repair and re-establishment of the normal composition of airway epithelium following injury have not been addressed. In this study, the role of a specific Notch receptor, Notch1, was investigated by targeted inactivation in the embryonic lung epithelium using the epithelial-specific Gata5-Cre driver line. Notch1-deficient mice are viable without discernible defects in pulmonary epithelial cell-fate determination and differentiation. However, in an experimental model of airway injury, activity of Notch1 is found to be required for normal repair of the airway epithelium. Absence of Notch1 reduced the ability of a population of cells distinguished by expression of PGP9.5, otherwise a marker of pulmonary neuroendocrine cells, which appears to serve as a reservoir for regeneration of Clara cells. Hairy/enhancer of split-5 (Hes5) and paired-box-containing gene 6 (Pax6) were found to be downstream targets of Notch1. Both Hes5 and Pax6 expressions were significantly increased in association with Clara cell regeneration in wild-type lungs. Ablation of Notch1 reduced Hes5 and Pax6 and inhibited airway epithelial repair. Thus, although dispensable in developmental ontogeny of airway epithelial cells, normal activity of Notch1 is required for repair of the airway epithelium. The signaling pathway by which Notch1 regulates the repair process includes stimulation of Hes5 and Pax6 gene expression.

  15. Gene Expression patterns in cryogenically stored Arabidopsis thaliana shoot tips

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The genes expressed in response to cryostress in plant shoot tips are not known. In this project we compared the gene expression patterns in untreated, cryoprotectant-treated, and recovering shoot tips using differential display methods. This project identified two genes that appeared to be differ...

  16. Indications for distinct pathogenic mechanisms of asbestos and silica through gene expression profiling of the response of lung epithelial cells

    PubMed Central

    Perkins, Timothy N.; Peeters, Paul M.; Shukla, Arti; Arijs, Ingrid; Dragon, Julie; Wouters, Emiel F.M.; Reynaert, Niki L.; Mossman, Brooke T.

    2015-01-01

    Occupational and environmental exposures to airborne asbestos and silica are associated with the development of lung fibrosis in the forms of asbestosis and silicosis, respectively. However, both diseases display distinct pathologic presentations, likely associated with differences in gene expression induced by different mineral structures, composition and bio-persistent properties. We hypothesized that effects of mineral exposure in the airway epithelium may dictate deviating molecular events that may explain the different pathologies of asbestosis versus silicosis. Using robust gene expression-profiling in conjunction with in-depth pathway analysis, we assessed early (24 h) alterations in gene expression associated with crocidolite asbestos or cristobalite silica exposures in primary human bronchial epithelial cells (NHBEs). Observations were confirmed in an immortalized line (BEAS-2B) by QRT-PCR and protein assays. Utilization of overall gene expression, unsupervised hierarchical cluster analysis and integrated pathway analysis revealed gene alterations that were common to both minerals or unique to either mineral. Our findings reveal that both minerals had potent effects on genes governing cell adhesion/migration, inflammation, and cellular stress, key features of fibrosis. Asbestos exposure was most specifically associated with aberrant cell proliferation and carcinogenesis, whereas silica exposure was highly associated with additional inflammatory responses, as well as pattern recognition, and fibrogenesis. These findings illustrate the use of gene-profiling as a means to determine early molecular events that may dictate pathological processes induced by exogenous cellular insults. In addition, it is a useful approach for predicting the pathogenicity of potentially harmful materials. PMID:25351596

  17. Gene expression profiling in male genital lichen sclerosus

    PubMed Central

    Edmonds, Emma; Barton, Geraint; Buisson, Sandrine; Francis, Nick; Gotch, Frances; Game, Laurence; Haddad, Munther; Dinneen, Michael; Bunker, Chris

    2011-01-01

    Male genital lichen sclerosus (MGLSc) has a bimodal distribution in boys and men. It is associated with squamous cell carcinoma (SCC). The pathogenesis of MGLSc is unknown. HPV and autoimmune mechanisms have been mooted. Anti extracellular matrix protein (ECM)1 antibodies have been identified in women with GLSc. The gene expression pattern of LSc is unknown. Using DNA microarrays we studied differences in gene expression in healthy and diseased prepuces obtained at circumcision in adult males with MGLSc (n = 4), paediatric LSc (n = 2) and normal healthy paediatric foreskin (n = 4). In adult samples 51 genes with significantly increased expression and 87 genes with significantly reduced expression were identified; paediatric samples revealed 190 genes with significantly increased expression and 148 genes with significantly reduced expression. Concordance of expression profiles between adult and paediatric samples indicates the same disease process. Functional analysis revealed increased expression in the adult and child MGSLc samples in the immune response/cellular defence gene ontology (GO) category and reduced expression in other categories including genes related to squamous cancer. No specific HPV, autoimmune or squamous carcinogenesis-associated gene expression patterns were found. ECM1 and CABLES1 expression were significantly reduced in paediatric and adult samples respectively. PMID:21718371

  18. CD4+ T cells enhance the unloaded shortening velocity of airway smooth muscle by altering the contractile protein expression.

    PubMed

    Matusovsky, Oleg S; Nakada, Emily M; Kachmar, Linda; Fixman, Elizabeth D; Lauzon, Anne-Marie

    2014-07-15

    Abundant data indicate that pathogenesis in allergic airways disease is orchestrated by an aberrant T-helper 2 (Th2) inflammatory response. CD4(+) T cells have been localized to airway smooth muscle (ASM) in both human asthmatics and in rodent models of allergic airways disease, where they have been implicated in proliferative responses of ASM. Whether CD4(+) T cells also alter ASM contractility has not been addressed. We established an in vitro system to assess the ability of antigen-stimulated CD4(+) T cells to modify contractile responses of the Brown Norway rat trachealis muscle. Our data demonstrated that the unloaded velocity of shortening (Vmax) of ASM was significantly increased upon 24 h co-incubation with antigen-stimulated CD4(+) T cells, while stress did not change. Enhanced Vmax was dependent upon contact between the CD4(+) T cells and the ASM and correlated with increased levels of the fast (+)insert smooth muscle myosin heavy chain isoform. The levels of myosin light chain kinase and myosin light chain phosphorylation were also increased within the muscle. The alterations in mechanics and in the levels of contractile proteins were transient, both declining to control levels after 48 h of co-incubation. More permanent alterations in muscle phenotype might be attainable when several inflammatory cells and mediators interact together or after repeated antigenic challenges. Further studies will await new tissue culture methodologies that preserve the muscle properties over longer periods of time. In conclusion, our data suggest that inflammatory cells promote ASM hypercontractility in airway hyper-responsiveness and asthma.

  19. Evolution of Gene Expression Balance Among Homeologs of Natural Polyploids

    PubMed Central

    Mutti, Jasdeep S.; Bhullar, Ramanjot K.; Gill, Kulvinder S.

    2017-01-01

    Polyploidy is a major evolutionary process in eukaryotes, yet the expression balance of homeologs in natural polyploids is largely unknown. To study this expression balance, the expression patterns of 2180 structurally well-characterized genes of wheat were studied, of which 813 had the expected three copies and 375 had less than three. Copy numbers of the remaining 992 ranged from 4 to 14, including homeologs, orthologs, and paralogs. Of the genes with three structural copies corresponding to homeologs, 55% expressed from all three, 38% from two, and the remaining 7% expressed from only one of the three copies. Homeologs of 76–87% of the genes showed differential expression patterns in different tissues, thus have evolved different gene expression controls, possibly resulting in novel functions. Homeologs of 55% of the genes showed tissue-specific expression, with the largest percentage (14%) in the anthers and the smallest (7%) in the pistils. The highest number (1.72/3) of homeologs/gene expression was in the roots and the lowest (1.03/3) in the anthers. As the expression of homeologs changed with changes in structural copy number, about 30% of the genes showed dosage dependence. Chromosomal location also impacted expression pattern as a significantly higher proportion of genes in the proximal regions showed expression from all three copies compared to that present in the distal regions. PMID:28193629

  20. Evaluating Fumonisin Gene Expression in Fusarium verticillioides.

    PubMed

    Scala, Valeria; Visentin, Ivan; Cardinale, Francesca

    2017-01-01

    Transcript levels of key genes in a biosynthetic pathway are often taken as a proxy for metabolite production. This is the case of FUM1, encoding the first dedicated enzyme in the metabolic pathway leading to the production of the mycotoxins Fumonisins by fungal species belonging to the genus Fusarium. FUM1 expression can be quantified by different methods; here, we detail a protocol based on quantitative reverse transcriptase polymerase chain reaction (RT-qPCR), by which relative or absolute transcript abundance can be estimated in Fusaria grown in vitro or in planta. As very seldom commercial kits for RNA extraction and cDNA synthesis are optimized for fungal samples, we developed a protocol tailored for these organisms, which stands alone but can be also easily integrated with specific reagents and kits commercially available.

  1. Monoallelic expression of the human FOXP2 speech gene

    PubMed Central

    Adegbola, Abidemi A.; Cox, Gerald F.; Bradshaw, Elizabeth M.; Hafler, David A.; Gimelbrant, Alexander; Chess, Andrew

    2015-01-01

    The recent descriptions of widespread random monoallelic expression (RMAE) of genes distributed throughout the autosomal genome indicate that there are more genes subject to RMAE on autosomes than the number of genes on the X chromosome where X-inactivation dictates RMAE of X-linked genes. Several of the autosomal genes that undergo RMAE have independently been implicated in human Mendelian disorders. Thus, parsing the relationship between allele-specific expression of these genes and disease is of interest. Mutations in the human forkhead box P2 gene, FOXP2, cause developmental verbal dyspraxia with profound speech and language deficits. Here, we show that the human FOXP2 gene undergoes RMAE. Studying an individual with developmental verbal dyspraxia, we identify a deletion 3 Mb away from the FOXP2 gene, which impacts FOXP2 gene expression in cis. Together these data suggest the intriguing possibility that RMAE impacts the haploinsufficiency phenotypes observed for FOXP2 mutations. PMID:25422445

  2. Monoallelic expression of the human FOXP2 speech gene.

    PubMed

    Adegbola, Abidemi A; Cox, Gerald F; Bradshaw, Elizabeth M; Hafler, David A; Gimelbrant, Alexander; Chess, Andrew

    2015-06-02

    The recent descriptions of widespread random monoallelic expression (RMAE) of genes distributed throughout the autosomal genome indicate that there are more genes subject to RMAE on autosomes than the number of genes on the X chromosome where X-inactivation dictates RMAE of X-linked genes. Several of the autosomal genes that undergo RMAE have independently been implicated in human Mendelian disorders. Thus, parsing the relationship between allele-specific expression of these genes and disease is of interest. Mutations in the human forkhead box P2 gene, FOXP2, cause developmental verbal dyspraxia with profound speech and language deficits. Here, we show that the human FOXP2 gene undergoes RMAE. Studying an individual with developmental verbal dyspraxia, we identify a deletion 3 Mb away from the FOXP2 gene, which impacts FOXP2 gene expression in cis. Together these data suggest the intriguing possibility that RMAE impacts the haploinsufficiency phenotypes observed for FOXP2 mutations.

  3. Phenotypic plasticity and divergence in gene expression.

    PubMed

    Healy, Timothy M; Schulte, Patricia M

    2015-07-01

    The extent to which phenotypic plasticity, or the ability of a single genotype to produce different phenotypes in different environments, impedes or promotes genetic divergence has been a matter of debate within evolutionary biology for many decades (see, for example, Ghalambor et al. ; Pfennig et al. ). Similarly, the role of evolution in shaping phenotypic plasticity remains poorly understood (Pigliucci ). In this issue of Molecular Ecology, Dayan et al. () provide empirical data relevant to these questions by assessing the extent of plasticity and divergence in the expression levels of 2272 genes in muscle tissue from killifish (genus Fundulus) exposed to different temperatures. F. heteroclitus (Fig. A) and F. grandis are minnows that inhabit estuarine marshes (Fig. B) along the coasts of the Atlantic Ocean and Gulf of Mexico in North America. These habitats undergo large variations in temperature both daily and seasonally, and these fish are known to demonstrate substantial phenotypic plasticity in response to temperature change (e.g. Fangue et al. ). Furthermore, the range of F. heteroclitus spans a large latitudinal gradient of temperatures, such that northern populations experience temperatures that are on average ~10°C colder than do southern populations (Schulte ). By comparing gene expression patterns between populations of these fish from different thermal habitats held in the laboratory at three different temperatures, Dayan et al. () address two important questions regarding the interacting effects of plasticity and evolution: (i) How does phenotypic plasticity affect adaptive divergence? and (ii) How does adaptive divergence affect plasticity?

  4. Modulation of R-gene expression across environments

    PubMed Central

    MacQueen, Alice; Bergelson, Joy

    2016-01-01

    Some environments are more conducive to pathogen growth than others, and, as a consequence, plants might be expected to invest more in resistance when pathogen growth is favored. Resistance (R-) genes in Arabidopsis thaliana have unusually extensive variation in basal expression when comparing the same R-gene among accessions collected from different environments. R-gene expression variation was characterized to explore whether R-gene expression is up-regulated in environments favoring pathogen proliferation and down-regulated when risks of infection are low; down-regulation would follow if costs of R-gene expression negatively impact plant fitness in the absence of disease. Quantitative reverse transcription–PCR was used to quantify the expression of 13 R-gene loci in plants grown in eight environmental conditions for each of 12 A. thaliana accessions, and large effects of the environment on R-gene expression were found. Surprisingly, almost every change in the environment—be it a change in biotic or abiotic conditions—led to an increase in R-gene expression, a response that was distinct from the average transcriptome response and from that of other stress response genes. These changes in expression are functional in that environmental change prior to infection affected levels of specific disease resistance to isolates of Pseudomonas syringae. In addition, there are strong latitudinal clines in basal R-gene expression and clines in R-gene expression plasticity correlated with drought and high temperatures. These results suggest that variation in R-gene expression across environments may be shaped by natural selection to reduce fitness costs of R-gene expression in permissive or predictable environments. PMID:26983577

  5. Modulation of R-gene expression across environments.

    PubMed

    MacQueen, Alice; Bergelson, Joy

    2016-03-01

    Some environments are more conducive to pathogen growth than others, and, as a consequence, plants might be expected to invest more in resistance when pathogen growth is favored. Resistance (R-) genes in Arabidopsis thaliana have unusually extensive variation in basal expression when comparing the same R-gene among accessions collected from different environments. R-gene expression variation was characterized to explore whether R-gene expression is up-regulated in environments favoring pathogen proliferation and down-regulated when risks of infection are low; down-regulation would follow if costs of R-gene expression negatively impact plant fitness in the absence of disease. Quantitative reverse transcription-PCR was used to quantify the expression of 13 R-gene loci in plants grown in eight environmental conditions for each of 12 A. thaliana accessions, and large effects of the environment on R-gene expression were found. Surprisingly, almost every change in the environment--be it a change in biotic or abiotic conditions--led to an increase in R-gene expression, a response that was distinct from the average transcriptome response and from that of other stress response genes. These changes in expression are functional in that environmental change prior to infection affected levels of specific disease resistance to isolates of Pseudomonas syringae. In addition, there are strong latitudinal clines in basal R-gene expression and clines in R-gene expression plasticity correlated with drought and high temperatures. These results suggest that variation in R-gene expression across environments may be shaped by natural selection to reduce fitness costs of R-gene expression in permissive or predictable environments.

  6. Genomic Expression Analysis Reveals Strategies of Burkholderia cenocepacia to Adapt to Cystic Fibrosis Patients' Airways and Antimicrobial Therapy

    PubMed Central

    Mira, Nuno P.; Madeira, Andreia; Moreira, Ana Sílvia; Coutinho, Carla P.; Sá-Correia, Isabel

    2011-01-01

    Pulmonary colonization of cystic fibrosis (CF) patients with Burkholderia cenocepacia or other bacteria of the Burkholderia cepacia complex (Bcc) is associated with worse prognosis and increased risk of death. During colonization, the bacteria may evolve under the stressing selection pressures exerted in the CF lung, in particular, those resulting from challenges of the host immune defenses, antimicrobial therapy, nutrient availability and oxygen limitation. Understanding the adaptive mechanisms that promote successful colonization and long-term survival of B. cenocepacia in the CF lung is essential for an improved therapeutic outcome of chronic infections. To get mechanistic insights into these adaptive strategies a transcriptomic analysis, based on DNA microarrays, was explored in this study. The genomic expression levels in two clonal variants isolated during long-term colonization of a CF patient who died from the cepacia syndrome were compared. One of the isolates examined, IST439, is the first B. cenocepacia isolate retrieved from the patient and the other isolate, IST4113, was obtained three years later and is more resistant to different classes of antimicrobials. Approximately 1000 genes were found to be differently expressed in the two clonal variants reflecting a marked reprogramming of genomic expression. The up-regulated genes in IST4113 include those involved in translation, iron uptake (in particular, in ornibactin biosynthesis), efflux of drugs and in adhesion to epithelial lung tissue and to mucin. Alterations related with adaptation to the nutritional environment of the CF lung and to an oxygen-limited environment are also suggested to be a key feature of transcriptional reprogramming occurring during long-term colonization, antibiotic therapy and the progression of the disease. PMID:22216120

  7. Phosphodiesterase 4 inhibitors augment the ability of formoterol to enhance glucocorticoid-dependent gene transcription in human airway epithelial cells: a novel mechanism for the clinical efficacy of roflumilast in severe chronic obstructive pulmonary disease.

    PubMed

    Moodley, Thunicia; Wilson, Sylvia M; Joshi, Taruna; Rider, Christopher F; Sharma, Pawan; Yan, Dong; Newton, Robert; Giembycz, Mark A

    2013-04-01

    Post-hoc analysis of two phase III clinical studies found that the phosphodiesterase 4 (PDE4) inhibitor, roflumilast, reduced exacerbation frequency in patients with severe chronic obstructive pulmonary disease (COPD) who were taking inhaled corticosteroids (ICS) concomitantly, whereas patients not taking ICS derived no such benefit. In contrast, in two different trials also performed in patients with severe COPD, roflumilast reduced exacerbation rates in the absence of ICS, indicating that PDE4 inhibition alone is sufficient for therapeutic activity to be realized. Given that roflumilast is recommended as an "add-on" medication to patients with severe disease who will inevitably be taking a long-acting β2-adrenoceptor agonist (LABA)/ICS combination therapy, we tested the hypothesis that roflumilast augments the ability of glucocorticoids to induce genes with anti-inflammatory activity. Using a glucocorticoid response element (GRE) luciferase reporter transfected into human airway epithelial cells [both bronchial epithelium + adenovirus 12 - SV40 hybrid (BEAS-2B) cells and primary cultures], roflumilast enhanced fluticasone propionate-induced GRE-dependent transcription. Roflumilast also produced a sinistral displacement of the concentration-response curves that described the augmentation of GRE-dependent transcription by the LABA formoterol. In BEAS-2B cells and primary airway epithelia, roflumilast interacted with formoterol in a positive cooperative manner to enhance the expression of several glucocorticoid-inducible genes that have anti-inflammatory potential. We suggest that the ability of roflumilast and formoterol to interact in this way supports the concept that these drugs together may impart clinical benefit beyond that achievable by an ICS alone, a PDE4 inhibitor alone, or an ICS/LABA combination therapy. Roflumilast may, therefore, be especially effective in patients with severe COPD.

  8. Random Monoallelic Gene Expression Increases upon Embryonic Stem Cell Differentiation

    PubMed Central

    Eckersley-Maslin, Mélanie A.; Thybert, David; Bergmann, Jan H.; Marioni, John C.; Flicek, Paul; Spector, David L.

    2014-01-01

    Summary Random autosomal monoallelic gene expression refers to the transcription of a gene from one of two homologous alleles. We assessed the dynamics of monoallelic expression during development through an allele-specific RNA sequencing screen in clonal populations of hybrid mouse embryonic stem cells (ESCs) and neural progenitor cells (NPCs). We identified 67 and 376 inheritable autosomal random monoallelically expressed genes in ESCs and NPCs respectively, a 5.6-fold increase upon differentiation. While DNA methylation and nuclear positioning did not distinguish the active and inactive alleles, specific histone modifications were differentially enriched between the two alleles. Interestingly, expression levels of 8% of the monoallelically expressed genes remained similar between monoallelic and biallelic clones. These results support a model in which random monoallelic expression occurs stochastically during differentiation, and for some genes is compensated for by the cell to maintain the required transcriptional output of these genes. PMID:24576421

  9. Noise in gene expression: origins, consequences, and control.

    PubMed

    Raser, Jonathan M; O'Shea, Erin K

    2005-09-23

    Genetically identical cells and organisms exhibit remarkable diversity even when they have identical histories of environmental exposure. Noise, or variation, in the process of gene expression may contribute to this phenotypic variability. Recent studies suggest that this noise has multiple sources, including the stochastic or inherently random nature of the biochemical reactions of gene expression. In this review, we summarize noise terminology and comment on recent investigations into the sources, consequences, and control of noise in gene expression.

  10. Carcinogen-induced trans activation of gene expression.

    PubMed Central

    Kleinberger, T; Flint, Y B; Blank, M; Etkin, S; Lavi, S

    1988-01-01

    We report a new mechanism of carcinogen action by which the expression of several genes was concomitantly enhanced. This mechanism involved the altered activity of cellular factors which modulate the expression of genes under their control. The increased expression was regulated at least in part on the transcriptional level and did not require amplification of the overexpressed genes. This phenomenon was transient; it was apparent as early as 24 h after carcinogen treatment and declined a few days later. Images PMID:2835673

  11. Carcinogen-induced trans activation of gene expression

    SciTech Connect

    Kleinberger, T.; Flint, Y.B.; Blank, M.; Etkin, S.; Lavi, S.

    1988-03-01

    The authors report a new mechanism of carcinogen action by which the expression of several genes was concomitantly enhanced. This mechanism involved the altered activity of cellular factors which modulate the expression of genes under their control. The increased expression was regulated at least in part on the transcriptional level and did not require amplification of the overexpressed genes. This phenomenon was transient; it was apparent as early as 24 h after carcinogen treatment and declined a few days later.

  12. cell type–specific gene expression differences in complex tissues

    PubMed Central

    Shen-Orr, Shai S; Tibshirani, Robert; Khatri, Purvesh; Bodian, Dale L; Staedtler, Frank; Perry, Nicholas M; Hastie, Trevor; Sarwal, Minnie M; Davis, Mark M; Butte, Atul J

    2013-01-01

    We describe cell type–specific significance analysis of microarrays (cssam) for analyzing differential gene expression for each cell type in a biological sample from microarray data and relative cell-type frequencies. first, we validated cssam with predesigned mixtures and then applied it to whole-blood gene expression datasets from stable post-transplant kidney transplant recipients and those experiencing acute transplant rejection, which revealed hundreds of differentially expressed genes that were otherwise undetectable. PMID:20208531

  13. Acanthamoeba protease activity promotes allergic airway inflammation via protease-activated receptor 2.

    PubMed

    Park, Mi Kyung; Cho, Min Kyoung; Kang, Shin Ae; Park, Hye-Kyung; Kim, Dong-Hee; Yu, Hak Sun

    2014-01-01

    Acanthamoeba is a free-living amoeba commonly present in the environment and often found in human airway cavities. Acanthamoeba possesses strong proteases that can elicit allergic airway inflammation. To our knowledge, the aeroallergenicity of Acanthamoeba has not been reported. We repeatedly inoculated mice with Acanthamoeba trophozoites or excretory-secretory (ES) proteins intra-nasally and evaluated symptoms and airway immune responses. Acanthamoeba trophozoites or ES proteins elicited immune responses in mice that resembled allergic airway inflammation. ES proteins had strong protease activity and activated the expression of several chemokine genes (CCL11, CCL17, CCL22, TSLP, and IL-25) in mouse lung epithelial cells. The serine protease inhibitor phenyl-methane-sulfonyl fluoride (PMSF) inhibited ES protein activity. ES proteins also stimulated dendritic cells and enhanced the differentiation of naive T cells into IL-4-secreting T cells. After repeated inoculation of the protease-activated receptor 2 knockout mouse with ES proteins, airway inflammation and Th2 immune responses were markedly reduced, but not to basal levels. Furthermore, asthma patients had higher Acanthamoeba-specific IgE titers than healthy controls and we found Acanthamoeba specific antigen from house dust in typical living room. Our findings suggest that Acanthamoeba elicits allergic airway symptoms in mice via a protease allergen. In addition, it is possible that Acanthamoeba may be one of the triggers human airway allergic disease.

  14. Clustering cancer gene expression data by projective clustering ensemble

    PubMed Central

    Yu, Xianxue; Yu, Guoxian

    2017-01-01

    Gene expression data analysis has paramount implications for gene treatments, cancer diagnosis and other domains. Clustering is an important and promising tool to analyze gene expression data. Gene expression data is often characterized by a large amount of genes but with limited samples, thus various projective clustering techniques and ensemble techniques have been suggested to combat with these challenges. However, it is rather challenging to synergy these two kinds of techniques together to avoid the curse of dimensionality problem and to boost the performance of gene expression data clustering. In this paper, we employ a projective clustering ensemble (PCE) to integrate the advantages of projective clustering and ensemble clustering, and to avoid the dilemma of combining multiple projective clusterings. Our experimental results on publicly available cancer gene expression data show PCE can improve the quality of clustering gene expression data by at least 4.5% (on average) than other related techniques, including dimensionality reduction based single clustering and ensemble approaches. The empirical study demonstrates that, to further boost the performance of clustering cancer gene expression data, it is necessary and promising to synergy projective clustering with ensemble clustering. PCE can serve as an effective alternative technique for clustering gene expression data. PMID:28234920

  15. Association of tissue lineage and gene expression: conservatively and differentially expressed genes define common and special functions of tissues

    PubMed Central

    2010-01-01

    Background Embryogenesis is the process by which the embryo is formed, develops, and establishes developmental hierarchies of tissues. The recent advance in microarray technology made it possible to investigate the tissue specific patterns of gene expression and their relationship with tissue lineages. This study is focused on how tissue specific functions, tissue lineage, and cell differentiation are correlated, which is essential to understand embryonic development and organism complexity. Results We performed individual gene and gene set based analysis on multiple tissue expression data, in association with the classic topology of mammalian fate maps of embryogenesis. For each sub-group of tissues on the fate map, conservatively, differentially and correlatively expressed genes or gene sets were identified. Tissue distance was found to correlate with gene expression divergence. Tissues of the ectoderm or mesoderm origins from the same segments on the fate map shared more similar expression pattern than those from different origins. Conservatively expressed genes or gene sets define common functions in a tissue group and are related to tissue specific diseases, which is supported by results from Gene Ontology and KEGG pathway analysis. Gene expression divergence is larger in certain human tissues than in the mouse homologous tissues. Conclusion The results from tissue lineage and gene expression analysis indicate that common function features of neighbor tissue groups were defined by the conservatively expressed genes and were related to tissue specific diseases, and differentially expressed genes contribute to the functional divergence of tissues. The difference of gene expression divergence in human and mouse homologous tissues reflected the organism complexity, i.e. distinct neural development levels and different body sizes. PMID:21172044

  16. Sex-specific gene expression in the BXD mouse liver.

    PubMed

    Gatti, Daniel M; Zhao, Ni; Chesler, Elissa J; Bradford, Blair U; Shabalin, Andrey A; Yordanova, Roumyana; Lu, Lu; Rusyn, Ivan

    2010-08-01

    Differences in clinical phenotypes between the sexes are well documented and have their roots in differential gene expression. While sex has a major effect on gene expression, transcription is also influenced by complex interactions between individual genetic variation and environmental stimuli. In this study, we sought to understand how genetic variation affects sex-related differences in liver gene expression by performing genetic mapping of genomewide liver mRNA expression data in a genetically defined population of naive male and female mice from C57BL/6J, DBA/2J, B6D2F1, and 37 C57BL/6J x DBA/2J (BXD) recombinant inbred strains. As expected, we found that many genes important to xenobiotic metabolism and other important pathways exhibit sexually dimorphic expression. We also performed gene expression quantitative trait locus mapping in this panel and report that the most significant loci that appear to regulate a larger number of genes than expected by chance are largely sex independent. Importantly, we found that the degree of correlation within gene expression networks differs substantially between the sexes. Finally, we compare our results to a recently released human liver gene expression data set and report on important similarities in sexually dimorphic liver gene expression between mouse and human. This study enhances our understanding of sex differences at the genome level and between species, as well as increasing our knowledge of the molecular underpinnings of sex differences in responses to xenobiotics.

  17. Analysis of HOX gene expression patterns in human breast cancer.

    PubMed

    Hur, Ho; Lee, Ji-Yeon; Yun, Hyo Jung; Park, Byeong Woo; Kim, Myoung Hee

    2014-01-01

    HOX genes are highly conserved transcription factors that determine the identity of cells and tissues along the anterior-posterior body axis in developing embryos. Aberrations in HOX gene expression have been shown in various tumors. However, the correlation of HOX gene expression patterns with tumorigenesis and cancer progression has not been fully characterized. Here, to analyze putative candidate HOX genes involved in breast cancer tumorigenesis and progression, the expression patterns of 39 HOX genes were analyzed using breast cancer cell lines and patient-derived breast tissues. In vitro analysis revealed that HOXA and HOXB gene expression occurred in a subtype-specific manner in breast cancer cell lines, whereas most HOXC genes were strongly expressed in most cell lines. Among the 39 HOX genes analyzed, 25 were chosen for further analysis in malignant and non-malignant tissues. Fourteen genes, encoding HOXA6, A13, B2, B4, B5, B6, B7, B8, B9, C5, C9, C13, D1, and D8, out of 25 showed statistically significant differential expression patterns between non-malignant and malignant breast tissues and are putative candidates associated with the development and malignant progression of breast cancer. Our data provide a valuable resource for furthering our understanding of HOX gene expression in breast cancer and the possible involvement of HOX genes in tumor progression.

  18. The complexity of gene expression dynamics revealed by permutation entropy

    PubMed Central

    2010-01-01

    Background High complexity is considered a hallmark of living systems. Here we investigate the complexity of temporal gene expression patterns using the concept of Permutation Entropy (PE) first introduced in dynamical systems theory. The analysis of gene expression data has so far focused primarily on the identification of differentially expressed genes, or on the elucidation of pathway and regulatory relationships. We aim to study gene expression time series data from the viewpoint of complexity. Results Applying the PE complexity metric to abiotic stress response time series data in Arabidopsis thaliana, genes involved in stress response and signaling were found to be associated with the highest complexity not only under stress, but surprisingly, also under reference, non-stress conditions. Genes with house-keeping functions exhibited lower PE complexity. Compared to reference conditions, the PE of temporal gene expression patterns generally increased upon stress exposure. High-complexity genes were found to have longer upstream intergenic regions and more cis-regulatory motifs in their promoter regions indicative of a more complex regulatory apparatus needed to orchestrate their expression, and to be associated with higher correlation network connectivity degree. Arabidopsis genes also present in other plant species were observed to exhibit decreased PE complexity compared to Arabidopsis specific genes. Conclusions We show that Permutation Entropy is a simple yet robust and powerful approach to identify temporal gene expression profiles of varying complexity that is equally applicable to other types of molecular profile data. PMID:21176199

  19. The Phillips airway.

    PubMed

    Haridas, R P; Wilkinson, D J

    2012-07-01

    The Phillips airway was developed by George Ramsay Phillips. There is no known original description of the airway and the earliest known reference to it is from 1919. The airway and its modifications are described.

  20. Transposable element influences on gene expression in plants.

    PubMed

    Hirsch, Cory D; Springer, Nathan M

    2017-01-01

    Transposable elements (TEs) comprise a major portion of many plant genomes and bursts of TE movements cause novel genomic variation within species. In order to maintain proper gene function, plant genomes have evolved a variety of mechanisms to tolerate the presence of TEs within or near genes. Here, we review our understanding of the interactions between TEs and gene expression in plants by assessing three ways that transposons can influence gene expression. First, there is growing evidence that TE insertions within introns or untranslated regions of genes are often tolerated and have minimal impact on expression level or splicing. However, there are examples in which TE insertions within genes can result in aberrant or novel transcripts. Second, TEs can provide novel alternative promoters, which can lead to new expression patterns or original coding potential of an alternate transcript. Third, TE insertions near genes can influence regulation of gene expression through a variety of mechanisms. For example, TEs may provide novel cis-acting regulatory sites behaving as enhancers or insert within existing enhancers to influence transcript production. Alternatively, TEs may change chromatin modifications in regions near genes, which in turn can influence gene expression levels. Together, the interactions of genes and TEs provide abundant evidence for the role of TEs in changing basic functions within plant genomes beyond acting as latent genomic elements or as simple insertional mutagens. This article is part of a Special Issue entitled: Plant Gene Regulatory Mechanisms and Networks, edited by Dr. Erich Grotewold and Dr. Nathan Springer.

  1. Method for 3D Airway Topology Extraction

    PubMed Central

    Grothausmann, Roman; Kellner, Manuela; Heidrich, Marko; Lorbeer, Raoul-Amadeus; Ripken, Tammo; Meyer, Heiko; Kuehnel, Mark P.; Ochs, Matthias; Rosenhahn, Bodo

    2015-01-01

    In lungs the number of conducting airway generations as well as bifurcation patterns varies across species and shows specific characteristics relating to illnesses or gene variations. A method to characterize the topology of the mouse airway tree using scanning laser optical tomography (SLOT) tomograms is presented in this paper. It is used to test discrimination between two types of mice based on detected differences in their conducting airway pattern. Based on segmentations of the airways in these tomograms, the main spanning tree of the volume skeleton is computed. The resulting graph structure is used to distinguish between wild type and surfactant protein (SP-D) deficient knock-out mice. PMID:25767561

  2. Blockage of upper airway

    MedlinePlus

    ... Airway obstruction - acute upper Images Throat anatomy Choking Respiratory system References Cukor J, Manno M. Pediatric respiratory emergencies: upper airway obstruction and infections. In: Marx ...

  3. Tensor decomposition for multi-tissue gene expression experiments

    PubMed Central

    Hore, Victoria; Viñuela, Ana; Buil, Alfonso; Knight, Julian; McCarthy, Mark I; Small, Kerrin; Marchini, Jonathan

    2016-01-01

    Genome wide association studies of gene expression traits and other cellular phenotypes have been successful in revealing links between genetic variation and biological processes. The majority of discoveries have uncovered cis eQTL effects via mass univariate testing of SNPs against gene expression in single tissues. We present a Bayesian method for multi-tissue experiments focusing on uncovering gene networks linked to genetic variation. Our method decomposes the 3D array (or tensor) of gene expression measurements into a set of latent components. We identify sparse gene networks, which can then be tested for association against genetic variation genome-wide. We apply our method to a dataset of 845 individuals from the TwinsUK cohort with gene expression measured via RNA sequencing in adipose, LCLs and skin. We uncover several gene networks with a genetic basis and clear biological and statistical significance. Extensions of this approach will allow integration of multi-omic, environmental and phenotypic datasets. PMID:27479908

  4. Optimization of transient gene expression system in Gerbera jemosonii petals.

    PubMed

    Hussein, Gihan M; Abu El-Heba, Ghada A; Abdou, Sara M; Abdallah, Naglaa A

    2013-01-01

    Low transformation efficiency and long generation time for production of transgenic Gerbera jemosonii plants leads to vulnerable gene function studies. Thus, transient expression of genes would be an efficient alternative. In this investigation, a transient expression system for gerbera petals based on the Agrobacterium infiltration protocol was developed using the reporter genes β-glucuronidase (gus) and green florescence protein (gfp). Results revealed the incapability of using the gfp gene as a reporter gene for transient expression study in gerbera flowers due to the detection of green fluorescent color in the non-infiltrated gerbera flower petals. However, the gus reporter gene was successfully utilized for optimizing and obtaining the suitable agroinfiltration system in gerbera flowers. The expression of GUS was detectable after three days of agroinfiltration in gerbera cultivars "Express" and "White Grizzly" with dark pink and white flower colors, respectively. The vacuum agroinfiltration protocol has been applied on the cultivar "Express" for evaluating the transient expression of the two genes involved in the anthocyanin pathway (iris-dfr and petunia-f3' 5'h), which is responsible for the color in flowers. In comparison to the control, transient expression results showed change in the anthocyanin pigment in all infiltrated flowers with color genes. Additionally, blue color was detected in the stigma and pollen grains in the infiltrated flowers. Moreover, blue colors with variant intensities were observed in produced calli during the routine work of stable transformation with f3' 5'h gene.

  5. Robust PCA based method for discovering differentially expressed genes.

    PubMed

    Liu, Jin-Xing; Wang, Yu-Tian; Zheng, Chun-Hou; Sha, Wen; Mi, Jian-Xun; Xu, Yong

    2013-01-01

    How to identify a set of genes that are relevant to a key biological process is an important issue in current molecular biology. In this paper, we propose a novel method to discover differentially expressed genes based on robust principal component analysis (RPCA). In our method, we treat the differentially and non-differentially expressed genes as perturbation signals S and low-rank matrix A, respectively. Perturbation signals S can be recovered from the gene expression data by using RPCA. To discover the differentially expressed genes associated with special biological progresses or functions, the scheme is given as follows. Firstly, the matrix D of expression data is decomposed into two adding matrices A and S by using RPCA. Secondly, the differentially expressed genes are identified based on matrix S. Finally, the differentially expressed genes are evaluated by the tools based on Gene Ontology. A larger number of experiments on hypothetical and real gene expression data are also provided and the experimental results show that our method is efficient and effective.

  6. Gene expression profile analyses of mice livers injured by Leigongteng

    PubMed Central

    Chen, Yong; Zhang, Xiao-Ming; Han, Feng-Mei; Du, Peng; Xia, Qi-Song

    2007-01-01

    AIM: To analyze the gene expression profiles of mice livers injured by Leigongteng and explore the relationship between the differentially expressed genes and liver damage. METHODS: The experimental mice were randomly divided into a control group and a liver-injured group in which the mice were administrated 33 μγ of triptolide/kg per day for 30 d. Liver mRNAs were extracted from animals in both groups and were reverse-transcribed to cDNA with dUTP labeled by different fluorescence (Cy3, Cy5) as hybridization probes. The mixed probes were hybridized with oligonucleotide microarray chips. The fluorescent signal results were acquired by scanner and analyzed with software. RESULTS: Among the 35852 target genes, 29 genes were found to be significantly differentially expressed, with 20 genes up-regulated and 9 genes down-regulated. The reliability of the differentially expressed genes was validated by RT-PCR experiments of 5 randomly selected differentially expressed genes. CONCLUSION: Based on the biological functions of the differentially expressed genes, it is obvious that the occurrence and development of liver damage induced by Leigongteng in mice are highly associated with immune response, metabolism, apoptosis and the cell skeleton of liver cells. This might be important for elucidating the regulatory network of gene expression associated with liver damage and it may also be important for discovering the pathogenic mechanisms of liver damage induced by Leigongteng. PMID:17659714

  7. Aromatase gene expression in the stallion.

    PubMed

    Lemazurier, E; Sourdaine, P; Nativelle, C; Plainfossé, B; Séralini, G

    2001-06-10

    Adult stallion secretes very high estrogen levels in its testicular vein and semen, and the responsible enzyme cytochrome P450 aromatase (P450 arom) is known to be present mainly in Leydig cells. We studied in further details the distribution of equine aromatase in various adult tissues including the brain (hypothalamic area), liver, kidney, small intestine, muscle, bulbourethral gland and testes. The aromatase mRNA was essentially detected by RT-PCR in testis (169+/-14 amol of aromatase mRNA per microg of total RNA) and was barely detectable in brain, or below 0.1 amol/microg RNA in other tissues. This range of expression was confirmed by ELISA (50+/-7 pg/microg total protein) in the testis, and by immunoblot, evidencing a 53 kDA specific protein band in testis and brain only. The corresponding aromatase activity was well detected, by 3H(2)O release from 1beta, 2beta(3)H-androstenedione, in testis and brain (200+/-23 and 25+/-6 pmol/min per mg, respectively) and below 3 pmol product formed/min per mg in other tissues. This study indicates that the testis, among the tissues analyzed, is the major source of aromatase in the adult stallion, and that the aromatase gene expression is specifically enhanced at this level, and is responsible for the high estrogen synthesis observed. Moreover, the study of aromatase in one colt testis has shown lower levels of transcripts, protein and enzyme activity, evidencing that aromatase is regulated during the development and may serve as a useful marker of testicular function. As the second organ where aromatase mRNA and activity are both well detected is brain, this study also underlines the possible role of neurosteroids in stallion on behaviour, brain function or central endocrine control.

  8. Genes, environment and gene expression in colon tissue: a pathway approach to determining functionality.

    PubMed

    Slattery, Martha L; Pellatt, Daniel F; Wolff, Roger K; Lundgreen, Abbie

    2016-01-01

    Genetic and environmental factors have been shown to work together to alter cancer risk. In this study we evaluate previously identified gene and lifestyle interactions in a candidate pathway that were associated with colon cancer risk to see if these interactions altered gene expression. We analyzed non-tumor RNA-seq data from 144 colon cancer patients who had genotype, recent cigarette smoking, diet, body mass index (BMI), and recent aspirin/non-steroidal anti-inflammatory use data. Using a false discovery rate of 0.1, we evaluated differential gene expression between high and low levels of lifestyle exposure and genotypes using DESeq2. Thirteen pathway genes and 17 SNPs within those genes were associated with altered expression of other genes in the pathway. BMI, NSAIDs use and dietary components of the oxidative balance score (OBS) also were associated with altered gene expression. SNPs previously identified as interacting with these lifestyle factors, altered expression of pathway genes. NSAIDs interacted with 10 genes (15 SNPs) within those genes to alter expression of 28 pathway genes; recent cigarette smoking interacted with seven genes (nine SNPs) to alter expression of 27 genes. BMI interacted with FLT1, KDR, SEPN1, TERT, TXNRD2, and VEGFA to alter expression of eight genes. Three genes (five SNPs) interacted with OBS to alter expression of 12 genes. These data provide support for previously identified lifestyle and gene interactions associated with colon cancer in that they altered expression of key pathway genes. The need to consider lifestyle factors in conjunction with genetic factors is illustrated.

  9. Population and sex differences in Drosophila melanogaster brain gene expression

    PubMed Central

    2012-01-01

    Background Changes in gene regulation are thought to be crucial for the adaptation of organisms to their environment. Transcriptome analyses can be used to identify candidate genes for ecological adaptation, but can be complicated by variation in gene expression between tissues, sexes, or individuals. Here we use high-throughput RNA sequencing of a single Drosophila melanogaster tissue to detect brain-specific differences in gene expression between the sexes and between two populations, one from the ancestral species range in sub-Saharan Africa and one from the recently colonized species range in Europe. Results Relatively few genes (<100) displayed sexually dimorphic expression in the brain, but there was an enrichment of sex-biased genes, especially male-biased genes, on the X chromosome. Over 340 genes differed in brain expression between flies from the African and European populations, with the inter-population divergence being highly correlated between males and females. The differentially expressed genes included those involved in stress response, olfaction, and detoxification. Expression differences were associated with transposable element insertions at two genes implicated in insecticide resistance (Cyp6g1 and CHKov1). Conclusions Analysis of the brain transcriptome revealed many genes differing in expression between populations that were not detected in previous studies using whole flies. There was little evidence for sex-specific regulatory adaptation in the brain, as most expression differences between populations were observed in both males and females. The enrichment of genes with sexually dimorphic expression on the X chromosome is consistent with dosage compensation mechanisms affecting sex-biased expression in somatic tissues. PMID:23170910

  10. Social Regulation of Gene Expression in Threespine Sticklebacks

    PubMed Central

    Greenwood, Anna K.; Peichel, Catherine L.

    2015-01-01

    Identifying genes that are differentially expressed in response to social interactions is informative for understanding the molecular basis of social behavior. To address this question, we described changes in gene expression as a result of differences in the extent of social interactions. We housed threespine stickleback (Gasterosteus aculeatus) females in either group conditions or individually for one week, then measured levels of gene expression in three brain regions using RNA-sequencing. We found that numerous genes in the hindbrain/cerebellum had altered expression in response to group or individual housing. However, relatively few genes were differentially expressed in either the diencephalon or telencephalon. The list of genes upregulated in fish from social groups included many genes related to neural development and cell adhesion as well as genes with functions in sensory signaling, stress, and social and reproductive behavior. The list of genes expressed at higher levels in individually-housed fish included several genes previously identified as regulated by social interactions in other animals. The identified genes are interesting targets for future research on the molecular mechanisms of normal social interactions. PMID:26367311

  11. Large Scale Gene Expression Meta-Analysis Reveals Tissue-Specific, Sex-Biased Gene Expression in Humans

    PubMed Central

    Mayne, Benjamin T.; Bianco-Miotto, Tina; Buckberry, Sam; Breen, James; Clifton, Vicki; Shoubridge, Cheryl; Roberts, Claire T.

    2016-01-01

    The severity and prevalence of many diseases are known to differ between the sexes. Organ specific sex-biased gene expression may underpin these and other sexually dimorphic traits. To further our understanding of sex differences in transcriptional regulation, we performed meta-analyses of sex biased gene expression in multiple human tissues. We analyzed 22 publicly available human gene expression microarray data sets including over 2500 samples from 15 different tissues and 9 different organs. Briefly, by using an inverse-variance method we determined the effect size difference of gene expression between males and females. We found the greatest sex differences in gene expression in the brain, specifically in the anterior cingulate cortex, (1818 genes), followed by the heart (375 genes), kidney (224 genes), colon (218 genes), and thyroid (163 genes). More interestingly, we found different parts of the brain with varying numbers and identity of sex-biased genes, indicating that specific cortical regions may influence sexually dimorphic traits. The majority of sex-biased genes in other tissues such as the bladder, liver, lungs, and pancreas were on the sex chromosomes or involved in sex hormone production. On average in each tissue, 32% of autosomal genes that were expressed in a sex-biased fashion contained androgen or estrogen hormone response elements. Interestingly, across all tissues, we found approximately two-thirds of autosomal genes that were sex-biased were not under direct influence of sex hormones. To our knowledge this is the largest analysis of sex-biased gene expression in human tissues to date. We identified many sex-biased genes that were not under the direct influence of sex chromosome genes or sex hormones. These may provide targets for future development of sex-specific treatments for diseases. PMID:27790248

  12. Unstable Expression of Commonly Used Reference Genes in Rat Pancreatic Islets Early after Isolation Affects Results of Gene Expression Studies.

    PubMed

    Kosinová, Lucie; Cahová, Monika; Fábryová, Eva; Týcová, Irena; Koblas, Tomáš; Leontovyč, Ivan; Saudek, František; Kříž, Jan

    2016-01-01

    The use of RT-qPCR provides a powerful tool for gene expression studies; however, the proper interpretation of the obtained data is crucially dependent on accurate normalization based on stable reference genes. Recently, strong evidence has been shown indicating that the expression of many commonly used reference genes may vary significantly due to diverse experimental conditions. The isolation of pancreatic islets is a complicated procedure which creates severe mechanical and metabolic stress leading possibly to cellular damage and alteration of gene expression. Despite of this, freshly isolated islets frequently serve as a control in various gene expression and intervention studies. The aim of our study was to determine expression of 16 candidate reference genes and one gene of interest (F3) in isolated rat pancreatic islets during short-term cultivation in order to find a suitable endogenous control for gene expression studies. We compared the expression stability of the most commonly used reference genes and evaluated the reliability of relative and absolute quantification using RT-qPCR during 0-120 hrs after isolation. In freshly isolated islets, the expression of all tested genes was markedly depressed and it increased several times throughout the first 48 hrs of cultivation. We observed significant variability among samples at 0 and 24 hrs but substantial stabilization from 48 hrs onwards. During the first 48 hrs, relative quantification failed to reflect the real changes in respective mRNA concentrations while in the interval 48-120 hrs, the relative expression generally paralleled the results determined by absolute quantification. Thus, our data call into question the suitability of relative quantification for gene expression analysis in pancreatic islets during the first 48 hrs of cultivation, as the results may be significantly affected by unstable expression of reference genes. However, this method could provide reliable information from 48 hrs onwards.

  13. Gene expression within a dynamic nuclear landscape

    PubMed Central

    Shav-Tal, Yaron; Darzacq, Xavier; Singer, Robert H

    2006-01-01

    Molecular imaging in living cells or organisms now allows us to observe macromolecular assemblies with a time resolution sufficient to address cause-and-effect relationships on specific molecules. These emerging technologies have gained much interest from the scientific community since they have been able to reveal novel concepts in cell biology, thereby changing our vision of the cell. One main paradigm is that cells stochastically vary, thus implying that population analysis may be misleading. In fact, cells should be analyzed within time-resolved single-cell experiments rather than being compared to other cells within a population. Technological imaging developments as well as the stochastic events present in gene expression have been reviewed. Here, we discuss how the structural organization of the nucleus is revealed using noninvasive single-cell approaches, which ultimately lead to the resolution required for the analysis of highly controlled molecular processes taking place within live cells. We also describe the efforts being made towards physiological approaches within the context of living organisms. PMID:16900099

  14. Cell cycle gene expression under clinorotation

    NASA Astrophysics Data System (ADS)

    Artemenko, Olga

    2016-07-01

    Cyclins and cyclin-dependent kinase (CDK) are main regulators of the cell cycle of eukaryotes. It's assumes a significant change of their level in cells under microgravity conditions and by other physical factors actions. The clinorotation use enables to determine the influence of gravity on simulated events in the cell during the cell cycle - exit from the state of quiet stage and promotion presynthetic phase (G1) and DNA synthesis phase (S) of the cell cycle. For the clinorotation effect study on cell proliferation activity is the necessary studies of molecular mechanisms of cell cycle regulation and development of plants under altered gravity condition. The activity of cyclin D, which is responsible for the events of the cell cycle in presynthetic phase can be controlled by the action of endogenous as well as exogenous factors, but clinorotation is one of the factors that influence on genes expression that regulate the cell cycle.These data can be used as a model for further research of cyclin - CDK complex for study of molecular mechanisms regulation of growth and proliferation. In this investigation we tried to summarize and analyze known literature and own data we obtained relatively the main regulators of the cell cycle in altered gravity condition.

  15. Induction of regulator of G-protein signaling 2 expression by long-acting β2-adrenoceptor agonists and glucocorticoids in human airway epithelial cells.

    PubMed

    Holden, Neil S; George, Tresa; Rider, Christopher F; Chandrasekhar, Ambika; Shah, Suharsh; Kaur, Manminder; Johnson, Malcolm; Siderovski, David P; Leigh, Richard; Giembycz, Mark A; Newton, Robert

    2014-01-01

    In asthma and chronic obstructive pulmonary disease (COPD) multiple mediators act on Gαq-linked G-protein-coupled receptors (GPCRs) to cause bronchoconstriction. However, acting on the airway epithelium, such mediators may also elicit inflammatory responses. In human bronchial epithelial BEAS-2B cells (bronchial epithelium + adenovirus 12-SV40 hybrid), regulator of G-protein signaling (RGS) 2 mRNA and protein were synergistically induced in response to combinations of long-acting β2-adrenoceptor agonist (LABA) (salmeterol, formoterol) plus glucocorticoid (dexamethasone, fluticasone propionate, budesonide). Equivalent responses occurred in primary human bronchial epithelial cells. Concentrations of glucocorticoid plus LABA required to induce RGS2 expression in BEAS-2B cells were consistent with the levels achieved therapeutically in the lungs. As RGS2 is a GTPase-activating protein that switches off Gαq, intracellular free calcium ([Ca(2+)]i) flux was used as a surrogate of responses induced by histamine, methacholine, and the thromboxane receptor agonist U46619 [(Z)-7-[(1S,4R,5R,6S)-5-[(E,3S)-3-hydroxyoct-1-enyl]-3-oxabicyclo[2.2.1]heptan-6-yl]hept-5-enoic acid]. This was significantly attenuated by salmeterol plus dexamethasone pretreatment, or RGS2 overexpression, and the protective effect of salmeterol plus dexamethasone was abolished by RGS2 RNA silencing. Although methacholine and U46619 induced interleukin-8 (IL-8) release and this was inhibited by RGS2 overexpression, the repression of U46619-induced IL-8 release by salmeterol plus dexamethasone was unaffected by RGS2 knockdown. Given a role for Gαq-mediated pathways in inducing IL-8 release, we propose that RGS2 acts redundantly with other effector processes to repress IL-8 expression. Thus, RGS2 expression is a novel effector mechanism in the airway epithelium that is induced by glucocorticoid/LABA combinations. This could contribute to the efficacy of glucocorticoid/LABA combinations in asthma and

  16. T-cell-intrinsic Tif1α/Trim24 regulates IL-1R expression on TH2 cells and TH2 cell-mediated airway allergy.

    PubMed

    Perez-Lloret, Jimena; Okoye, Isobel S; Guidi, Riccardo; Kannan, Yashaswini; Coomes, Stephanie M; Czieso, Stephanie; Mengus, Gabrielle; Davidson, Irwin; Wilson, Mark S

    2016-02-02

    There is a paucity of new therapeutic targets to control allergic reactions and forestall the rising trend of allergic diseases. Although a variety of immune cells contribute to allergy, cytokine-secreting αβ(+)CD4(+) T-helper 2 (TH2) cells orchestrate the type-2-driven immune response in a large proportion of atopic asthmatics. To identify previously unidentified putative targets in pathogenic TH2 cells, we performed in silico analyses of recently published transcriptional data from a wide variety of pathogenic TH cells [Okoye IS, et al. (2014) Proc Natl Acad Sci USA 111(30):E3081-E3090] and identified that transcription intermediary factor 1 regulator-alpha (Tif1α)/tripartite motif-containing 24 (Trim24) was predicted to be active in house dust mite (HDM)- and helminth-elicited Il4(gfp+)αβ(+)CD4(+) TH2 cells but not in TH1, TH17, or Treg cells. Testing this prediction, we restricted Trim24 deficiency to T cells by using a mixed bone marrow chimera system and found that T-cell-intrinsic Trim24 is essential for HDM-mediated airway allergy and antihelminth immunity. Mechanistically, HDM-elicited Trim24(-/-) T cells have reduced expression of many TH2 cytokines and chemokines and were predicted to have compromised IL-1-regulated signaling. Following this prediction, we found that Trim24(-/-) T cells have reduced IL-1 receptor (IL-1R) expression, are refractory to IL-1β-mediated activation in vitro and in vivo, and fail to respond to IL-1β-exacerbated airway allergy. Collectively, these data identify a previously unappreciated Trim24-dependent requirement for IL-1R expression on TH2 cells and an important nonredundant role for T-cell-intrinsic Trim24 in TH2-mediated allergy and antihelminth immunity.

  17. T-cell–intrinsic Tif1α/Trim24 regulates IL-1R expression on TH2 cells and TH2 cell-mediated airway allergy

    PubMed Central

    Perez-Lloret, Jimena; Okoye, Isobel S.; Guidi, Riccardo; Kannan, Yashaswini; Coomes, Stephanie M.; Czieso, Stephanie; Mengus, Gabrielle; Davidson, Irwin; Wilson, Mark S.

    2016-01-01

    There is a paucity of new therapeutic targets to control allergic reactions and forestall the rising trend of allergic diseases. Although a variety of immune cells contribute to allergy, cytokine-secreting αβ+CD4+ T-helper 2 (TH2) cells orchestrate the type-2–driven immune response in a large proportion of atopic asthmatics. To identify previously unidentified putative targets in pathogenic TH2 cells, we performed in silico analyses of recently published transcriptional data from a wide variety of pathogenic TH cells [Okoye IS, et al. (2014) Proc Natl Acad Sci USA 111(30):E3081–E3090] and identified that transcription intermediary factor 1 regulator-alpha (Tif1α)/tripartite motif-containing 24 (Trim24) was predicted to be active in house dust mite (HDM)- and helminth-elicited Il4gfp+αβ+CD4+ TH2 cells but not in TH1, TH17, or Treg cells. Testing this prediction, we restricted Trim24 deficiency to T cells by using a mixed bone marrow chimera system and found that T-cell–intrinsic Trim24 is essential for HDM-mediated airway allergy and antihelminth immunity. Mechanistically, HDM-elicited Trim24−/− T cells have reduced expression of many TH2 cytokines and chemokines and were predicted to have compromised IL-1–regulated signaling. Following this prediction, we found that Trim24−/− T cells have reduced IL-1 receptor (IL-1R) expression, are refractory to IL-1β–mediated activation in vitro and in vivo, and fail to respond to IL-1β–exacerbated airway allergy. Collectively, these data identify a previously unappreciated Trim24-dependent requirement for IL-1R expression on TH2 cells and an important nonredundant role for T-cell–intrinsic Trim24 in TH2-mediated allergy and antihelminth immunity. PMID:26787865

  18. The Role of Multiple Transcription Factors In Archaeal Gene Expression

    SciTech Connect

    Charles J. Daniels

    2008-09-23

    Since the inception of this research program, the project has focused on two central questions: What is the relationship between the 'eukaryal-like' transcription machinery of archaeal cells and its counterparts in eukaryal cells? And, how does the archaeal cell control gene expression using its mosaic of eukaryal core transcription machinery and its bacterial-like transcription regulatory proteins? During the grant period we have addressed these questions using a variety of in vivo approaches and have sought to specifically define the roles of the multiple TATA binding protein (TBP) and TFIIB-like (TFB) proteins in controlling gene expression in Haloferax volcanii. H. volcanii was initially chosen as a model for the Archaea based on the availability of suitable genetic tools; however, later studies showed that all haloarchaea possessed multiple tbp and tfb genes, which led to the proposal that multiple TBP and TFB proteins may function in a manner similar to alternative sigma factors in bacterial cells. In vivo transcription and promoter analysis established a clear relationship between the promoter requirements of haloarchaeal genes and those of the eukaryal RNA polymerase II promoter. Studies on heat shock gene promoters, and the demonstration that specific tfb genes were induced by heat shock, provided the first indication that TFB proteins may direct expression of specific gene families. The construction of strains lacking tbp or tfb genes, coupled with the finding that many of these genes are differentially expressed under varying growth conditions, provided further support for this model. Genetic tools were also developed that led to the construction of insertion and deletion mutants, and a novel gene expression scheme was designed that allowed the controlled expression of these genes in vivo. More recent studies have used a whole genome array to examine the expression of these genes and we have established a linkage between the expression of specific tfb

  19. Arabidopsis gene expression patterns are altered during spaceflight

    NASA Astrophysics Data System (ADS)

    Paul, Anna-Lisa; Popp, Michael P.; Gurley, William B.; Guy, Charles; Norwood, Kelly L.; Ferl, Robert J.

    The exposure of Arabidopsis thaliana (Arabidopsis) plants to spaceflight environments results in differential gene expression. A 5-day mission on orbiter Columbia in 1999 (STS-93) carried transgenic Arabidopsis plants engineered with a transgene composed of the alcohol dehydrogenase (Adh) gene promoter linked to the β-Glucuronidase (GUS) reporter gene. The plants were used to evaluate the effects of spaceflight on gene expression patterns initially by using the Adh/GUS transgene to address specifically the possibility that spaceflight induces a hypoxic stress response (Paul, A.L., Daugherty, C.J., Bihn, E.A., Chapman, D.K., Norwood, K.L., Ferl, R.J., 2001. Transgene expression patterns indicate that spaceflight affects stress signal perception and transduction in arabidopsis, Plant Physiol. 126, 613-621). As a follow-on to the reporter gene analysis, we report here the evaluation of genome-wide patterns of native gene expression within Arabidopsis shoots utilizing the Agilent DNA array of 21,000 Arabidopsis genes. As a control for the veracity of the array analyses, a selection of genes was further characterized with quantitative Real-Time RT PCR (ABI - Taqman®). Comparison of the patterns of expression for arrays probed with RNA isolated from plants exposed to spaceflight compared to RNA isolated from ground control plants revealed 182 genes that were differentially expressed in response to the spaceflight mission by more than 4-fold, and of those only 50 genes were expressed at levels chosen to support a conservative change call. None of the genes that are hallmarks of hypoxic stress were induced to this level. However, genes related to heat shock were dramatically induced - but in a pattern and under growth conditions that are not easily explained by elevated temperatures. These gene expression data are discussed in light of current models for plant responses to the spaceflight environment and with regard to potential future spaceflight experiment

  20. Gene expression profiling of mouse embryos with microarrays

    PubMed Central

    Sharov, Alexei A.; Piao, Yulan; Ko, Minoru S. H.

    2011-01-01

    Global expression profiling by DNA microarrays provides a snapshot of cell and tissue status and becomes an essential tool in biological and medical sciences. Typical questions that can be addressed by microarray analysis in developmental biology include: (1) to find a set of genes expressed in a specific cell type; (2) to identify genes expressed commonly in multiple cell types; (3) to follow the time-course changes of gene expression patterns; (4) to demonstrate cell’s identity by showing similarities or differences among two or multiple cell types; (5) to find regulatory pathways and/or networks affected by gene manipulations, such as overexpression or repression of gene expression; (6) to find downstream target genes of transcription factors; (7) to find downstream target genes of cell signaling; (8) to examine the effects of environmental manipulation of cells on gene expression patterns; and (9) to find the effects of genetic manipulation in embryos and adults. Here we describe strategies for executing these experiments and monitoring changes of cell state with gene expression microarrays in application to mouse embryology. Both statistical assessment and interpretation of data are discussed. We also present a protocol for performing microarray analysis on a small amount of embryonic materials. PMID:20699157

  1. Stably Expressed Genes Involved in Basic Cellular Functions

    PubMed Central

    Wang, Kejian; Fuscoe, James C.

    2017-01-01

    Stably Expressed Genes (SEGs) whose expression varies within a narrow range may be involved in core cellular processes necessary for basic functions. To identify such genes, we re-analyzed existing RNA-Seq gene expression profiles across 11 organs at 4 developmental stages (from immature to old age) in both sexes of F344 rats (n = 4/group; 320 samples). Expression changes (calculated as the maximum expression / minimum expression for each gene) of >19000 genes across organs, ages, and sexes ranged from 2.35 to >109-fold, with a median of 165-fold. The expression of 278 SEGs was found to vary ≤4-fold and these genes were significantly involved in protein catabolism (proteasome and ubiquitination), RNA transport, protein processing, and the spliceosome. Such stability of expression was further validated in human samples where the expression variability of the homologous human SEGs was significantly lower than that of other genes in the human genome. It was also found that the homologous human SEGs were generally less subject to non-synonymous mutation than other genes, as would be expected of stably expressed genes. We also found that knockout of SEG homologs in mouse models was more likely to cause complete preweaning lethality than non-SEG homologs, corroborating the fundamental roles played by SEGs in biological development. Such stably expressed genes and pathways across life-stages suggest that tight control of these processes is important in basic cellular functions and that perturbation by endogenous (e.g., genetics) or exogenous agents (e.g., drugs, environmental factors) may cause serious adverse effects. PMID:28125669

  2. Expression and mapping of anthocyanin biosynthesis genes in carrot

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Anthocyanin gene expression has been extensively studied in leaves, fruits and flowers of numerous plants. Little, however, is known about anthocyanin accumulation in roots, or in carrots or other Apiaceae. We quantified expression of six anthocyanin biosynthetic genes (phenylalanine ammonia-lyase (...

  3. An Exercise to Estimate Differential Gene Expression in Human Cells

    ERIC Educational Resources Information Center

    Chaudhry, M. Ahmad

    2006-01-01

    The expression of genes in cells of various tissue types varies considerably and is correlated with the function of a particular organ. The pattern of gene expression changes in diseased tissues, in response to therapy or infection and exposure to environmental mutagens, chemicals, ultraviolet light, and ionizing radiation. To better understand…

  4. Bioinformatic Analysis of Gene Expression for Melanoma Treatment

    PubMed Central

    Kawakami, Akinori; Fisher, David E.

    2016-01-01

    Bioinformatic analysis of genome-wide gene expression allows us to characterize cells, including melanomas. Gene expression profiles have been generated in various stages of melanomas and analyzed by researchers in unique ways. Lauss et al. compared their melanoma subtypes with those of The Cancer Genome Atlas Network and found consistency between the two studies. PMID:27884291

  5. MEPD: medaka expression pattern database, genes and more

    PubMed Central

    Alonso-Barba, Juan I.; Rahman, Raza-Ur; Wittbrodt, Joachim; Mateo, Juan L.

    2016-01-01

    The Medaka Expression Pattern Database (MEPD; http://mepd.cos.uni-heidelberg.de/) is designed as a repository of medaka expression data for the scientific community. In this update we present two main improvements. First, we have changed the previous clone-centric view for in situ data to a gene-centric view. This is possible because now we have linked all the data present in MEPD to the medaka gene annotation in ENSEMBL. In addition, we have also connected the medaka genes in MEPD to their corresponding orthologous gene in zebrafish, again using the ENSEMBL database. Based on this, we provide a link to the Zebrafish Model Organism Database (ZFIN) to allow researches to compare expression data between these two fish model organisms. As a second major improvement, we have modified the design of the database to enable it to host regulatory elements, promoters or enhancers, expression patterns in addition to gene expression. The combination of gene expression, by traditional in situ, and regulatory element expression, typically by fluorescence reporter gene, within the same platform assures consistency in terms of annotation. In our opinion, this will allow researchers to uncover new insights between the expression domain of genes and their regulatory landscape. PMID:26450962

  6. Digital Gene Expression Tag Profiling Analysis of the Gene Expression Patterns Regulating the Early Stage of Mouse Spermatogenesis

    PubMed Central

    Meng, Lijun; Liu, Meiling; Zhao, Lina; Hu, Fen; Ding, Cunbao; Wang, Yang; He, Baoling; Pan, Yuxin; Fang, Wei; Chen, Jing; Hu, Songnian; Jia, Mengchun

    2013-01-01

    Detailed characterization of the gene expression patterns in spermatogonia and primary spermatocytes is critical to understand the processes which occur prior to meiosis during normal spermatogenesis. The genome-wide expression profiles of mouse type B spermatogonia and primary spermatocytes were investigated using the Solexa/Illumina digital gene expression (DGE) system, a tag based high-throughput transcriptome sequencing method, and the developmental processes which occur during early spermatogenesis were systematically analyzed. Gene expression patterns vary significantly between mouse type B spermatogonia and primary spermatocytes. The functional analysis revealed that genes related to junction assembly, regulation of the actin cytoskeleton and pluripotency were most significantly differently expressed. Pathway analysis indicated that the Wnt non-canonical signaling pathway played a central role and interacted with the actin filament organization pathway during the development of spermatogonia. This study provides a foundation for further analysis of the gene expression patterns and signaling pathways which regulate the molecular mechanisms of early spermatogenesis. PMID:23554914

  7. Heme Oxygenase-1-Expressing Dendritic Cells Promote Foxp3+ Regulatory T Cell Differentiation and Induce Less Severe Airway Inflammation in Murine Models

    PubMed Central

    Gau, Rung-Jiun; Yen, Jeng-Hsien; Suen, Jau-Ling

    2016-01-01

    Dendritic cells (DCs) are critical for instructing immune responses toward inflammatory or anti-inflammatory status. Heme oxygenase-1 (HO-1) is known for its cytoprotective effect against oxidative stress and inflammation, suggesting its immune regulatory role in allergic lung inflammation. HO-1 has been implicated in affecting DC maturation; however, its role in DC-mediated T-cell differentiation is unclear. In this study, we demonstrated that HO-1-expressing bone marrow-derived dendritic cells (BM-DCs) displayed tolerogenic phenotypes, including their resistance to lipopolysaccharide (LPS)-induced maturation, high level expression of IL-10, and low T-cell stimulatory activity. In addition, HO-1-expressing DCs were able to induce antigen-specific Foxp3+ regulatory T cells (Treg) differentiation in vitro and in vivo. Also, HO-1-expressing DCs modulated the severity of lung inflammatory responses in two murine models of airway inflammation. This study provided evidence supporting the role of HO-1-expressing DCs in tolerance induction and as a potential therapeutic target for allergic asthma as well as other inflammatory diseases. PMID:28033400

  8. Gene Expression Measurement Module (GEMM) - a fully automated, miniaturized instrument for measuring gene expression in space

    NASA Astrophysics Data System (ADS)

    Karouia, Fathi; Ricco, Antonio; Pohorille, Andrew; Peyvan, Kianoosh

    2012-07-01

    The capability to measure gene expression on board spacecrafts opens the doors to a large number of experiments on the influence of space environment on biological systems that will profoundly impact our ability to conduct safe and effective space travel, and might also shed light on terrestrial physiology or biological function and human disease and aging processes. Measurements of gene expression will help us to understand adaptation of terrestrial life to conditions beyond the planet of origin, identify deleterious effects of the space environment on a wide range of organisms from microbes to humans, develop effective countermeasures against these effects, determine metabolic basis of microbial pathogenicity and drug resistance, test our ability to sustain and grow in space organisms that can be used for life support and in situ resource utilization during long-duration space exploration, and monitor both the spacecraft environment and crew health. These and other applications hold significant potential for discoveries in space biology, biotechnology and medicine. Accordingly, supported by funding from the NASA Astrobiology Science and Technology Instrument Development Program, we are developing a fully automated, miniaturized, integrated fluidic system for small spacecraft capable of in-situ measuring microbial expression of thousands of genes from multiple samples. The instrument will be capable of (1) lysing bacterial cell walls, (2) extracting and purifying RNA released from cells, (3) hybridizing it on a microarray and (4) providing electrochemical readout, all in a microfluidics cartridge. The prototype under development is suitable for deployment on nanosatellite platforms developed by the NASA Small Spacecraft Office. The first target application is to cultivate and measure gene expression of the photosynthetic bacterium Synechococcus elongatus, i.e. a cyanobacterium known to exhibit remarkable metabolic diversity and resilience to adverse conditions

  9. Identification of Scedosporium boydii catalase A1 gene, a reactive oxygen species detoxification factor highly expressed in response to oxidative stress and phagocytic cells.

    PubMed

    Mina, Sara; Staerck, Cindy; d'Almeida, Sènan M; Marot, Agnès; Delneste, Yves; Calenda, Alphonse; Tabiasco, Julie; Bouchara, Jean-Philippe; Fleury, Maxime J J

    2015-12-01

    Scedosporium boydii is an opportunistic filamentous fungus which may be responsible for a large variety of infections in both immunocompetent and immunocompromised individuals. This fungus belongs to the Scedosporium apiospermum species complex which usually ranks second among the filamentous fungi colonizing the airways of patients with cystic fibrosis (CF). Species of the S. apiospermum complex are able to chronically colonize the CF airways suggesting pathogenic mechanisms allowing persistence and growth of these fungi in the respiratory tract. Few putative virulence factors have been purified and characterized so far in the S. apiospermum complex including a cytosolic Cu,Zn-superoxide dismutase (SOD) and a monofunctional catalase (catalase A1). Upon microbial infection, host phagocytes release reactive oxygen species (ROS), such as hydrogen peroxide, as part of the antimicrobial response. Catalases are known to protect pathogens against ROS by degradation of the hydrogen peroxide. Here, we identified the S. boydii catalase A1 gene (CATA1) and investigated its expression in response to the environmental conditions encountered in the CF airways and to the oxidative stress. Results showed that S. boydii CATA1 gene expression is not affected by hypoxia, hypercapnia or pH changes. In contrast, CATA1 gene was overexpressed in response to a chemically induced oxidative stress with a relative gene expression 37-fold higher in the presence of 250 μM H(2)O(2), 20-fold higher with 250 μM menadione and 5-fold higher with 2 mM paraquat. Moreover, S. boydii CATA1 gene expression progressively increased upon exposure to activated THP-1-derived macrophages, reaching a maximum after 12 h (26 fold). Activated HL60-derived neutrophils and activated human peripheral blood neutrophils more rapidly induced S. boydii CATA1 gene overexpression, a maximum gene expression level being reached at 75 min (17 fold) and 60 min (15 fold), respectively. In contrast expression of the gene

  10. The effect of negative autoregulation on eukaryotic gene expression

    NASA Astrophysics Data System (ADS)

    Nevozhay, Dmitry; Adams, Rhys; Murphy, Kevin; Josic, Kresimir; Balázsi, G. Ábor

    2009-03-01

    Negative autoregulation is a frequent motif in gene regulatory networks, which has been studied extensively in prokaryotes. Nevertheless, some effects of negative feedback on gene expression in eukaryotic transcriptional networks remain unknown. We studied how the strength of negative feedback regulation affects the characteristics of gene expression in yeast cells carrying synthetic transcriptional cascades. We observed a drastic reduction of gene expression noise and a change in the shape of the dose-response curve. We explained these experimentally observed effects by stochastic simulations and a simple set of algebraic equations.

  11. Features of Gene Expression of Bacillus pumilus Metalloendopeptidase.

    PubMed

    Rudakova, N L; Sabirova, A R; Balaban, N P; Tikhonova, A O; Sharipova, M R

    2016-08-01

    Features of gene expression of the secreted Bacillus pumilus metalloendopeptidase belonging to the adamalysin/reprolysin family were investigated. In the regulatory region of the gene, we identified hypothetical binding sites for transcription factors CcpA and TnrA. We found that the expression of the metalloendopeptidase gene is controlled by mechanisms of carbon and nitrogen catabolite repression. In experiments involving nitrogen metabolism regulatory protein mutant strains, we found that the control of the metalloendopeptidase gene expression involves proteins of ammonium transport GlnK and AmtB interacting with the TnrA-regulator.

  12. Direct Introduction of Genes into Rats and Expression of the Genes

    NASA Astrophysics Data System (ADS)

    Benvenisty, Nissim; Reshef, Lea

    1986-12-01

    A method of introducing actively expressed genes into intact mammals is described. DNA precipitated with calcium phosphate has been injected intraperitoneally into newborn rats. The injected genes have been taken up and expressed by the animal tissues. To examine the generality of the method we have injected newborn rats with the chloramphenicol acetyltransferase prokaryotic gene fused with various viral and cellular gene promoters and the gene for hepatitis B surface antigen, and we observed appearance of chloramphenicol acetyltransferase activity and hepatitis B surface antigen in liver and spleen. In addition, administration of genes coding for hormones (insulin or growth hormone) resulted in their expression.

  13. Effects of G-gene Deletion and Replacement on Rabies Virus Vector Gene Expression

    PubMed Central

    Sato, Sho; Ohara, Shinya; Tsutsui, Ken-Ichiro; Iijima, Toshio

    2015-01-01

    The glycoprotein-gene (G gene) -deleted rabies virus (RV) vector is a powerful tool to examine the function and structure of neural circuits. We previously reported that the deletion of the G gene enhances the transgene expression level of the RV vector. However, the mechanism of this enhancement remains to be clarified. We presume that there are two possible factors for this enhancement. The first factor is the glycoprotein of RV, which shows cytotoxicity; thus, may cause a dysfunction in the translation process of infected cells. The second possible factor is the enhanced expression of the L gene, which encodes viral RNA polymerase. In the RV, it is known that the gene expression level is altered depending on the position of the gene. Since G-gene deletion displaces the L gene in the genome, the expression of the L gene and viral transcription may be enhanced. In this study, we compared the transgene expression level and viral transcription of three recombinant RV vectors. The effect of glycoprotein was examined by comparing the viral gene expression of G-gene-intact RV and G-gene-replaced RV. Despite the fact that the L-gene transcription level of these two RV vectors was similar, the G-gene-replaced RV vector showed higher viral transcription and transgene expression level than the G-gene-intact RV vector. To examine the effect of the position of the L gene, we compared the viral gene expression of the G-gene-deleted RV and G-gene-replaced RV. The G-gene-deleted RV vector showed higher L-gene transcription, viral transcription, and transgene expression level than the G-gene-replaced RV vector. These results indicate that G-gene deletion enhances the transgene expression level through at least two factors, the absence of glycoprotein and enhancement of L-gene expression. These findings enable investigators to design a useful viral vector that shows a controlled desirable transgene expression level in applications. PMID:26023771

  14. In Utero Cigarette Smoke Affects Allergic Airway Disease But Does Not Alter the Lung Methylome

    PubMed Central

    Eyring, Kenneth R.; Pedersen, Brent S.; Yang, Ivana V.; Schwartz, David A.

    2015-01-01

    Prenatal and postnatal cigarette smoke exposure enhances the risk of developing asthma. Despite this as well as other smoking related risks, 11% of women still smoke during pregnancy. We hypothesized that cigarette smoke exposure during prenatal development generates long lasting differential methylation altering transcriptional activity that correlates with disease. In a house dust mite (HDM) model of allergic airway disease, we measured airway hyperresponsiveness (AHR) and airway inflammation between mice exposed prenatally to cigarette smoke (CS) or filtered air (FA). DNA methylation and gene expression were then measured in lung tissue. We demonstrate that HDM-treated CS mice develop a more severe allergic airway disease compared to HDM-treated FA mice including increased AHR and airway inflammation. While DNA methylation changes between the two HDM-treated groups failed to reach genome-wide significance, 99 DMRs had an uncorrected p-value < 0.001. 6 of these 99 DMRs were selected for validation, based on the immune function of adjacent genes, and only 2 of the 6 DMRs confirmed the bisulfite sequencing data. Additionally, genes near these 6 DMRs (Lif, Il27ra, Tle4, Ptk7, Nfatc2, and Runx3) are differentially expressed between HDM-treated CS mice and HDM-treated FA mice. Our findings confirm that prenatal exposure to cigarette smoke is sufficient to modify allergic airway disease; however, it is unlikely that specific methylation changes account for the exposure-response relationship. These findings highlight the important role in utero cigarette smoke exposure plays in the development of allergic airway disease. PMID:26642056

  15. Key aspects of analyzing microarray gene-expression data.

    PubMed

    Chen, James J

    2007-05-01

    One major challenge with the use of microarray technology is the analysis of massive amounts of gene-expression data for various applications. This review addresses the key aspects of the microarray gene-expression data analysis for the two most common objectives: class comparison and class prediction. Class comparison mainly aims to select which genes are differentially expressed across experimental conditions. Gene selection is separated into two steps: gene ranking and assigning a significance level. Class prediction uses expression profiling analysis to develop a prediction model for patient selection, diagnostic prediction or prognostic classification. Development of a prediction model involves two components: model building and performance assessment. It also describes two additional data analysis methods: gene-class testing and multiple ordering criteria.

  16. A predictive approach to identify genes differentially expressed

    NASA Astrophysics Data System (ADS)

    Saraiva, Erlandson F.; Louzada, Francisco; Milan, Luís A.; Meira, Silvana; Cobre, Juliana

    2012-10-01

    The main objective of gene expression data analysis is to identify genes that present significant changes in expression levels between a treatment and a control biological condition. In this paper, we propose a Bayesian approach to identify genes differentially expressed calculating credibility intervals from predictive densities which are constructed using sampled mean treatment effect from all genes in study excluding the treatment effect of genes previously identified with statistical evidence for difference. We compare our Bayesian approach with the standard ones based on the use of the t-test and modified t-tests via a simulation study, using small sample sizes which are common in gene expression data analysis. Results obtained indicate that the proposed approach performs better than standard ones, especially for cases with mean differences and increases in treatment variance in relation to control variance. We also apply the methodologies to a publicly available data set on Escherichia coli bacteria.

  17. Identification of Development and Pathogenicity Related Gene in Botrytis cinerea via Digital Gene Expression Profile

    PubMed Central

    Zhao, Bin; Si, He Long; Sun, Zhi Ying; Xu, Zheng; Chen, Zhan; Zhang, Jin lin; Xing, Ji Hong; Dong, Jin Gao

    2015-01-01

    Background: Botrytis cinerea, a haploid Euascomycete fungus that infects numerous crops, has been used as a model system for studying molecular phytopathology. Botrytis cinerea adopts various modes of infection, which are mediated by a number of pathogenicity and virulence-related genes. Many of these genes have not been reported previously. Objectives: This study aimed to investigate development and pathogenicity-related genes between a novel nonpathogenic mutant and the Wild Type (WT) in B. cinerea. Materials and Methods: Digital Gene Expression (DGE) tag profiling can reveal novel genes that may be involved in development and pathogenicity of plant pathogen. A large volume of B. cinerea tag-seq was generated to identify differential expressed genes by the Illumina DGE tag profiling technology. Results: A total of 4,182,944 and 4,182,021 clean tags were obtained from the WT and a nonpathogenic mutant stain (BCt89), respectively, and 10,410 differentially expressed genes were identified. In addition, 84 genes were expressed in the WT only while 34 genes were expressed in the mutant only. A total of 664 differentially expressed genes were involved in 91 Kyoto Encyclopedia of Genes and Genome pathways, including signaling and metabolic pathways. Conclusions: Expression levels of 1,426 genes were significantly up-regulated in the mutant compared to WT. Furthermore, 301 genes were down-regulated with False Discovery Rates (FDR) of < 0.001 and absolute value of log2 Ratio of ≥ 1. PMID:26034553

  18. Fundamental principles of energy consumption for gene expression

    NASA Astrophysics Data System (ADS)

    Huang, Lifang; Yuan, Zhanjiang; Yu, Jianshe; Zhou, Tianshou

    2015-12-01

    How energy is consumed in gene expression is largely unknown mainly due to complexity of non-equilibrium mechanisms affecting expression levels. Here, by analyzing a representative gene model that considers complexity of gene expression, we show that negative feedback increases energy consumption but positive feedback has an opposite effect; promoter leakage always reduces energy consumption; generating more bursts needs to consume more energy; and the speed of promoter switching is at the cost of energy consumption. We also find that the relationship between energy consumption and expression noise is multi-mode, depending on both the type of feedback and the speed of promoter switching. Altogether, these results constitute fundamental principles of energy consumption for gene expression, which lay a foundation for designing biologically reasonable gene modules. In addition, we discuss possible biological implications of these principles by combining experimental facts.

  19. Airway Secretory microRNAome Changes during Rhinovirus Infection in Early Childhood

    PubMed Central

    Gutierrez, Maria J.; Gomez, Jose L.; Perez, Geovanny F.; Pancham, Krishna; Val, Stephanie; Pillai, Dinesh K.; Giri, Mamta; Ferrante, Sarah; Freishtat, Robert; Rose, Mary C.; Preciado, Diego; Nino, Gustavo

    2016-01-01

    Background Innate immune responses are fine-tuned by small noncoding RNA molecules termed microRNAs (miRs) that modify gene expression in response to the environment. During acute infections, miRs can be secreted in extracellular vesicles (EV) to facilitate cell-to-cell genetic communication. The purpose of this study was to characterize the baseline population of miRs secreted in EVs in the airways of young children (airway secretory microRNAome) and examine the changes during rhinovirus (RV) infection, the most common cause of asthma exacerbations and the most important early risk factor for the development of asthma beyond childhood. Methods Nasal airway secretions were obtained from children (≤3 yrs. old) during PCR-confirmed RV infections (n = 10) and age-matched controls (n = 10). Nasal EVs were isolated with polymer-based precipitation and global miR profiles generated using NanoString microarrays. We validated our in vivo airway secretory miR data in an in vitro airway epithelium model using apical secretions from primary human bronchial epithelial cells (HBEC) differentiated at air-liquid interface (ALI). Bioinformatics tools were used to determine the unified (nasal and bronchial) signature airway secretory miRNAome and changes during RV infection in children. Results Multiscale analysis identified four signature miRs comprising the baseline airway secretory miRNAome: hsa-miR-630, hsa-miR-302d-3p, hsa- miR-320e, hsa-miR-612. We identified hsa-miR-155 as the main change in the baseline miRNAome during RV infection in young children. We investigated the potential biological relevance of the airway secretion of hsa-mir-155 using in silico models derived from gene datasets of experimental in vivo human RV infection. These analyses confirmed that hsa-miR-155 targetome is an overrepresented pathway in the upper airways of individuals infected with RV. Conclusions Comparative analysis of the airway secretory microRNAome in children indicates that RV infection

  20. The effects of in utero vitamin D deficiency on airway smooth muscle mass and lung function.

    PubMed

    Foong, Rachel E; Bosco, Anthony; Jones, Anya C; Gout, Alex; Gorman, Shelley; Hart, Prue H; Zosky, Graeme R

    2015-11-01

    We have previously demonstrated increased airway smooth muscle (ASM) mass and airway hyperresponsiveness in whole-life vitamin D-deficient female mice. In this study, we aimed to uncover the molecular mechanisms contributing to altered lung structure and function. RNA was extracted from lung tissue of whole-life vitamin D-deficient and -replete female mice, and gene expression patterns were profiled by RNA sequencing. The data showed that genes involved in embryonic organ development, pattern formation, branching morphogenesis, Wingless/Int signaling, and inflammation were differentially expressed in vitamin D-deficient mice. Network analysis suggested that differentially expressed genes were connected by the hubs matrix metallopeptidase 9; NF-κ light polypeptide gene enhancer in B cells inhibitor, α; epidermal growth factor receptor; and E1A binding protein p300. Given our findings that developmental pathways may be altered, we investigated if the timing of vitamin D exposure (in utero vs. postnatal) had an impact on lung health outcomes. Gene expression was measured in in utero or postnatal vitamin D-deficient mice, as well as whole-life vitamin D-deficient and -replete mice at 8 weeks of age. Baseline lung function, airway hyperresponsiveness, and airway inflammation were measured and lungs fixed for lung structure assessment using stereological methods and quantification of ASM mass. In utero vitamin D deficiency was sufficient to increase ASM mass and baseline airway resistance and alter lung structure. There were increased neutrophils but decreased lymphocytes in bronchoalveolar lavage. Expression of inflammatory molecules S100A9 and S100A8 was mainly increased in postnatal vitamin D-deficient mice. These observations suggest that in utero vitamin D deficiency can alter lung structure and function and increase inflammation, contributing to symptoms in chronic diseases, such as asthma.

  1. Dimensionality of Data Matrices with Applications to Gene Expression Profiles

    ERIC Educational Resources Information Center

    Feng, Xingdong

    2009-01-01

    Probe-level microarray data are usually stored in matrices. Take a given probe set (gene), for example, each row of the matrix corresponds to an array, and each column corresponds to a probe. Often, people summarize each array by the gene expression level. Is one number sufficient to summarize a whole probe set for a specific gene in an array?…

  2. Sources of stochasticity in constitutive and autoregulated gene expression

    NASA Astrophysics Data System (ADS)

    Marathe, Rahul; Gomez, David; Klumpp, Stefan

    2012-11-01

    Gene expression is inherently noisy as many steps in the read-out of the genetic information are stochastic. To disentangle the effect of different sources of stochasticity in such systems, we consider various models that describe some processes as stochastic and others as deterministic. We review earlier results for unregulated (constitutive) gene expression and present new results for a gene controlled by negative autoregulation with cell growth modeled by linear volume growth.

  3. Chamber Specific Gene Expression Landscape of the Zebrafish Heart

    PubMed Central

    Singh, Angom Ramcharan; Sivadas, Ambily; Sabharwal, Ankit; Vellarikal, Shamsudheen Karuthedath; Jayarajan, Rijith; Verma, Ankit; Kapoor, Shruti; Joshi, Adita; Scaria, Vinod; Sivasubbu, Sridhar

    2016-01-01

    The organization of structure and function of cardiac chambers in vertebrates is defined by chamber-specific distinct gene expression. This peculiarity and uniqueness of the genetic signatures demonstrates functional resolution attributed to the different chambers of the heart. Altered expression of the cardiac chamber genes can lead to individual chamber related dysfunctions and disease patho-physiologies. Information on transcriptional repertoire of cardiac compartments is important to understand the spectrum of chamber specific anomalies. We have carried out a genome wide transcriptome profiling study of the three cardiac chambers in the zebrafish heart using RNA sequencing. We have captured the gene expression patterns of 13,396 protein coding genes in the three cardiac chambers—atrium, ventricle and bulbus arteriosus. Of these, 7,260 known protein coding genes are highly expressed (≥10 FPKM) in the zebrafish heart. Thus, this study represents nearly an all-inclusive information on the zebrafish cardiac transcriptome. In this study, a total of 96 differentially expressed genes across the three cardiac chambers in zebrafish were identified. The atrium, ventricle and bulbus arteriosus displayed 20, 32 and 44 uniquely expressing genes respectively. We validated the expression of predicted chamber-restricted genes using independent semi-quantitative and qualitative experimental techniques. In addition, we identified 23 putative novel protein coding genes that are specifically restricted to the ventricle and not in the atrium or bulbus arteriosus. In our knowledge, these 23 novel genes have either not been investigated in detail or are sparsely studied. The transcriptome identified in this study includes 68 differentially expressing zebrafish cardiac chamber genes that have a human ortholog. We also carried out spatiotemporal gene expression profiling of the 96 differentially expressed genes throughout the three cardiac chambers in 11 developmental stages and 6

  4. Gene Expression Profiling in the Type 1 Diabetes Rat Diaphragm

    PubMed Central

    van Lunteren, Erik; Moyer, Michelle

    2009-01-01

    Background Respiratory muscle contractile performance is impaired by diabetes, mechanisms of which included altered carbohydrate and lipid metabolism, oxidative stress and changes in membrane electrophysiology. The present study examined to what extent these cellular perturbations involve changes in gene expression. Methodology/Principal Findings Diaphragm muscle from streptozotocin-diabetic rats was analyzed with Affymetrix gene expression arrays. Diaphragm from diabetic rats had 105 genes with at least ±2-fold significantly changed expression (55 increased, 50 decreased), and these were assigned to gene ontology groups based on over-representation analysis using DAVID software. There was increased expression of genes involved in palmitoyl-CoA hydrolase activity (a component of lipid metabolism) (P = 0.037, n = 2 genes, fold change 4.2 to 27.5) and reduced expression of genes related to carbohydrate metabolism (P = 0.000061, n = 8 genes, fold change −2.0 to −8.5). Other gene ontology groups among upregulated genes were protein ubiquitination (P = 0.0053, n = 4, fold change 2.2 to 3.4), oxidoreductase activity (P = 0.024, n = 8, fold change 2.1 to 6.0), and morphogenesis (P = 0.012, n = 10, fold change 2.1 to 4.3). Other downregulated gene groups were extracellular region (including extracellular matrix and collagen) (P = 0.00032, n = 13, fold change −2.2 to −3.7) and organogenesis (P = 0.032, n = 7, fold change −2.1 to −3.7). Real-time PCR confirmed the directionality of changes in gene expression for 30 of 31 genes tested. Conclusions/Significance These data indicate that in diaphragm muscle type 1 diabetes increases expression of genes involved in lipid energetics, oxidative stress and protein ubiquitination, decreases expression of genes involved in carbohydrate metabolism, and has little effect on expression of ion channel genes. Reciprocal changes in expression of genes involved in

  5. Temporal Changes in Gene Expression Profile during Mature Adipocyte Dedifferentiation

    PubMed Central

    Côté, Julie Anne; Guénard, Frédéric; Lessard, Julie; Lapointe, Marc; Biron, Simon

    2017-01-01

    Objective. To characterize changes in gene expression profile during human mature adipocyte dedifferentiation in ceiling culture. Methods. Subcutaneous (SC) and omental (OM) adipose tissue samples were obtained from 4 participants paired for age and BMI. Isolated adipocytes were dedifferentiated in ceiling culture. Gene expression analysis at days 0, 4, 7, and 12 of the cultures was performed using Affymetrix Human Gene 2.0 STvi arrays. Hierarchical clustering according to similarity of expression changes was used to identify overrepresented functions. Results. Four clusters gathered genes with similar expression between day 4 to day 7 but decreasing expression from day 7 to day 12. Most of these genes coded for proteins involved in adipocyte functions (LIPE, PLIN1, DGAT2, PNPLA2, ADIPOQ, CEBPA, LPL, FABP4, SCD, INSR, and LEP). Expression of several genes coding for proteins implicated in cellular proliferation and growth or cell cycle increased significantly from day 7 to day 12 (WNT5A, KITLG, and FGF5). Genes coding for extracellular matrix proteins were differentially expressed between days 0, 4, 7, and 12 (COL1A1, COL1A2, and COL6A3, MMP1, and TGFB1). Conclusion. Dedifferentiation is associated with downregulation of transcripts encoding proteins involved in mature adipocyte functions and upregulation of genes involved in matrix remodeling, cellular development, and cell cycle.

  6. Regulation of mitochondrial gene expression, the epigenetic enigma.

    PubMed

    Mposhi, Archibold; Van der Wijst, Monique Gp; Faber, Klaas Nico; Rots, Marianne G

    2017-03-01

    Epigenetics provides an important layer of information on top of the DNA sequence and is essential for establishing gene expression profiles. Extensive studies have shown that nuclear DNA methylation and histone modifications influence nuclear gene expression. However, it remains unclear whether mitochondrial DNA (mtDNA) undergoes similar epigenetic changes to regulate mitochondrial gene expression. Recently, it has been shown that mtDNA is differentially methylated in various diseases such as diabetes and colorectal cancer. Interestingly, this differential methylation was often associated with altered mitochondrial gene expression. However, the direct role of mtDNA methylation on gene expression remains elusive. Alternatively, the activity of the mitochondrial transcription factor A (TFAM), a protein involved in mtDNA packaging, might also influence gene expression. This review discusses the role of mtDNA methylation and potential epigenetic-like modifications of TFAM with respect to mtDNA transcription and replication. We suggest three mechanisms: (1) methylation within the non-coding D-loop, (2) methylation at gene start sites (GSS) and (3) post-translational modifications (PTMs) of TFAM. Unraveling mitochondrial gene expression regulation could open new therapeutic avenues for mitochondrial diseases.

  7. With Reference to Reference Genes: A Systematic Review of Endogenous Controls in Gene Expression Studies

    PubMed Central

    Chapman, Joanne R.; Waldenström, Jonas

    2015-01-01

    The choice of reference genes that are stably expressed amongst treatment groups is a crucial step in real-time quantitative PCR gene expression studies. Recent guidelines have specified that a minimum of two validated reference genes should be used for normalisation. However, a quantitative review of the literature showed that the average number of reference genes used across all studies was 1.2. Thus, the vast majority of studies continue to use a single gene, with β-actin (ACTB) and/or glyceraldehyde 3-phosphate dehydrogenase (GAPDH) being commonly selected in studies of vertebrate gene expression. Few studies (15%) tested a panel of potential reference genes for stability of expression before using them to normalise data. Amongst studies specifically testing reference gene stability, few found ACTB or GAPDH to be optimal, whereby these genes were significantly less likely to be chosen when larger panels of potential reference genes were screened. Fewer reference genes were tested for stability in non-model organisms, presumably owing to a dearth of available primers in less well characterised species. Furthermore, the experimental conditions under which real-time quantitative PCR analyses were conducted had a large influence on the choice of reference genes, whereby different studies of rat brain tissue showed different reference genes to be the most stable. These results highlight the importance of validating the choice of normalising reference genes before conducting gene expression studies. PMID:26555275

  8. With Reference to Reference Genes: A Systematic Review of Endogenous Controls in Gene Expression Studies.

    PubMed

    Chapman, Joanne R; Waldenström, Jonas

    2015-01-01

    The choice of reference genes that are stably expressed amongst treatment groups is a crucial step in real-time quantitative PCR gene expression studies. Recent guidelines have specified that a minimum of two validated reference genes should be used for normalisation. However, a quantitative review of the literature showed that the average number of reference genes used across all studies was 1.2. Thus, the vast majority of studies continue to use a single gene, with β-actin (ACTB) and/or glyceraldehyde 3-phosphate dehydrogenase (GAPDH) being commonly selected in studies of vertebrate gene expression. Few studies (15%) tested a panel of potential reference genes for stability of expression before using them to normalise data. Amongst studies specifically testing reference gene stability, few found ACTB or GAPDH to be optimal, whereby these genes were significantly less likely to be chosen when larger panels of potential reference genes were screened. Fewer reference genes were tested for stability in non-model organisms, presumably owing to a dearth of available primers in less well characterised species. Furthermore, the experimental conditions under which real-time quantitative PCR analyses were conducted had a large influence on the choice of reference genes, whereby different studies of rat brain tissue showed different reference genes to be the most stable. These results highlight the importance of validating the choice of normalising reference genes before conducting gene expression studies.

  9. Clustering Algorithms: Their Application to Gene Expression Data

    PubMed Central

    Oyelade, Jelili; Isewon, Itunuoluwa; Oladipupo, Funke; Aromolaran, Olufemi; Uwoghiren, Efosa; Ameh, Faridah; Achas, Moses; Adebiyi, Ezekiel

    2016-01-01

    Gene expression data hide vital information required to understand the biological process that takes place in a particular organism in relation to its environment. Deciphering the hidden patterns in gene expression data proffers a prodigious preference to strengthen the understanding of functional genomics. The complexity of biological networks and the volume of genes present increase the challenges of comprehending and interpretation of the resulting mass of data, which consists of millions of measurements; these data also inhibit vagueness, imprecision, and noise. Therefore, the use of clustering techniques is a first step toward addressing these challenges, which is essential in the data mining process to reveal natural structures and identify interesting patterns in the underlying data. The clustering of gene expression data has been proven to be useful in making known the natural structure inherent in gene expression data, understanding gene functions, cellular processes, and subtypes of cells, mining useful information from noisy data, and understanding gene regulation. The other benefit of clustering gene expression data is the identification of homology, which is very important in vaccine design. This review examines the various clustering algorithms applicable to the gene expression data in order to discover and provide useful knowledge of the appropriate clustering technique that will guarantee stability and high degree of accuracy in its analysis procedure. PMID:27932867

  10. Modulation and Expression of Tumor Suppressor Genes by Environmental Agents.

    DTIC Science & Technology

    1996-12-01

    MAMA produced other tumors in medaka (e.g. liver) and Rb expression is altered in many human tumors , the capability of examining the pathology of all...AD GRANT NUMBER DAMDI7-93-J-3011 TITLE: Modulation and Expression of Tumor Suppressor Genes by Environmental Agents PRINCIPAL INVESTIGATOR: Gary K...SUBTITLE 5. FUNDING NUMBERS Modulation and Expression of Tumor Suppressor Genes by Environmental Agents DAMDl7-93- J-3011 6. AUTHOR(S) Gary K

  11. Gene Expression by Mouse Inner Ear Hair Cells during Development

    PubMed Central

    Scheffer, Déborah I.; Shen, Jun

    2015-01-01

    Hair cells of the inner ear are essential for hearing and balance. As a consequence, pathogenic variants in genes specifically expressed in hair cells often cause hereditary deafness. Hair cells are few in number and not easily isolated from the adjacent supporting cells, so the biochemistry and molecular biology of hair cells can be difficult to study. To study gene expression in hair cells, we developed a protocol for hair cell isolation by FACS. With nearly pure hair cells and surrounding cells, from cochlea and utricle and from E16 to P7, we performed a comprehensive cell type-specific RNA-Seq study of gene expression during mouse inner ear development. Expression profiling revealed new hair cell genes with distinct expression patterns: some are specific for vestibular hair cells, others for cochlear hair cells, and some are expressed just before or after maturation of mechanosensitivity. We found that many of the known hereditary deafness genes are much more highly expressed in hair cells than surrounding cells, suggesting that genes preferentially expressed in hair cells are good candidates for unknown deafness genes. PMID:25904789

  12. An atlas of gene expression and gene co-regulation in the human retina

    PubMed Central

    Pinelli, Michele; Carissimo, Annamaria; Cutillo, Luisa; Lai, Ching-Hung; Mutarelli, Margherita; Moretti, Maria Nicoletta; Singh, Marwah Veer; Karali, Marianthi; Carrella, Diego; Pizzo, Mariateresa; Russo, Francesco; Ferrari, Stefano; Ponzin, Diego; Angelini, Claudia; Banfi, Sandro; di Bernardo, Diego

    2016-01-01

    The human retina is a specialized tissue involved in light stimulus transduction. Despite its unique biology, an accurate reference transcriptome is still missing. Here, we performed gene expression analysis (RNA-seq) of 50 retinal samples from non-visually impaired post-mortem donors. We identified novel transcripts with high confidence (Observed Transcriptome (ObsT)) and quantified the expression level of known transcripts (Reference Transcriptome (RefT)). The ObsT included 77 623 transcripts (23 960 genes) covering 137 Mb (35 Mb new transcribed genome). Most of the transcripts (92%) were multi-exonic: 81% with known isoforms, 16% with new isoforms and 3% belonging to new genes. The RefT included 13 792 genes across 94 521 known transcripts. Mitochondrial genes were among the most highly expressed, accounting for about 10% of the reads. Of all the protein-coding genes in Gencode, 65% are expressed in the retina. We exploited inter-individual variability in gene expression to infer a gene co-expression network and to identify genes specifically expressed in photoreceptor cells. We experimentally validated the photoreceptors localization of three genes in human retina that had not been previously reported. RNA-seq data and the gene co-expression network are available online (http://retina.tigem.it). PMID:27235414

  13. Validation of housekeeping genes for studying differential gene expression in the bovine myometrium.

    PubMed

    Rekawiecki, Robert; Kowalik, Magdalena K; Kotwica, Jan

    2013-12-01

    The aim of this study was to determine the steady-state expression of 13 selected housekeeping genes in the myometrium of cyclic and pregnant cows. Cells taken from bovine myometrium on days 1-5, 6-10, 11-16 and 17-20 of the oestrous cycle and in weeks 3-5, 6-8 and 9-12 of pregnancy were used. Reverse transcribed RNA was amplified in real-time PCR using designed primers. Reaction efficiency was determined with the Linreg programme. The geNorm and NormFinder programmes were used to select the best housekeeping genes. They calculate the expression stability factor for each used housekeeping gene with the smallest value for most stably expressed genes. According to geNorm, the most stable housekeeping genes in the myometrium were C2orf29, TPB and TUBB2B, while the least stably expressed genes were 18S RNA, HPRT1 and GAPDH. NormFinder identified the best genes in the myometrium as C2orf29, MRPL12 and TBP, while the worst genes were 18S RNA, B2M and SF3A1. Differences in stability factors between the two programmes may also indicate that the physiological status of the female, e.g. pregnancy, affects the stability of expression of housekeeping genes. The different expression stability of housekeeping genes did not affect progesterone receptor expression but it could be important if small differences in gene expression were measured between studies.

  14. BPH gene expression profile associated to prostate gland volume.

    PubMed

    Descazeaud, Aurelien; Rubin, Mark A; Hofer, Matthias; Setlur, Sunita; Nikolaief, Nathalie; Vacherot, Francis; Soyeux, Pascale; Kheuang, Laurence; Abbou, Claude C; Allory, Yves; de la Taille, Alexandre

    2008-12-01

    The aim of the current study was to analyze gene expression profiles in benign prostatic hyperplasia and to compare them with phenotypic properties. Thirty-seven specimens of benign prostatic hyperplasia were obtained from symptomatic patients undergoing surgery. RNA was extracted and hybridized to Affymetrix Chips containing 54,000 gene expression probes. Gene expression profiles were analyzed using cluster, TreeView, and significance analysis of microarrays softwares. In an initial unsupervised analysis, our 37 samples clustered hierarchically in 2 groups of 18 and 19 samples, respectively. Five clinical parameters were statistically different between the 2 groups: in group 1 compared with group 2, patients had larger prostate glands, had higher prostate specific antigen levels, were more likely to be treated by alpha blockers, to be operated by prostatectomy, and to have major irritative symptoms. The sole independent parameter associated with this dichotome clustering, however, was the prostate gland volume. Therefore, the role of prostate volume was explored in a supervised analysis. Gene expression of prostate glands <60 mL and >60 mL were compared using significance analysis of microarrays and 227 genes were found differentially expressed between the 2 groups (>2 change and false discovery rate of <5%). Several specific pathways including growth factors genes, cell cycle genes, apoptose genes, inflammation genes, and androgen regulated genes, displayed major differences between small and large prostate glands.

  15. Gene expression profile analysis of type 2 diabetic mouse liver.

    PubMed

    Zhang, Fang; Xu, Xiang; Zhang, Yi; Zhou, Ben; He, Zhishui; Zhai, Qiwei

    2013-01-01

    Liver plays a key role in glucose metabolism and homeostasis, and impaired hepatic glucose metabolism contributes to the development of type 2 diabetes. However, the precise gene expression profile of diabetic liver and its association with diabetes and related diseases are yet to be further elucidated. In this study, we detected the gene expression profile by high-throughput sequencing in 9-week-old normal and type 2 diabetic db/db mouse liver. Totally 12132 genes were detected, and 2627 genes were significantly changed in diabetic mouse liver. Biological process analysis showed that the upregulated genes in diabetic mouse liver were mainly enriched in metabolic processes. Surprisingly, the downregulated genes in diabetic mouse liver were mainly enriched in immune-related processes, although all the altered genes were still mainly enriched in metabolic processes. Similarly, KEGG pathway analysis showed that metabolic pathways were the major pathways altered in diabetic mouse liver, and downregulated genes were enriched in immune and cancer pathways. Analysis of the key enzyme genes in fatty acid and glucose metabolism showed that some key enzyme genes were significantly increased and none of the detected key enzyme genes were decreased. In addition, FunDo analysis showed that liver cancer and hepatitis were most likely to be associated with diabetes. Taken together, this study provides the digital gene expression profile of diabetic mouse liver, and demonstrates the main diabetes-associated hepatic biological processes, pathways, key enzyme genes in fatty acid and glucose metabolism and potential hepatic diseases.

  16. An internal regulatory element controls troponin I gene expression

    SciTech Connect

    Yutzey, K.E.; Kline, R.L.; Konieczmy, S.F. . Dept. of Biological Sciences)

    1989-04-01

    During skeletal myogenesis, approximately 20 contractile proteins and related gene products temporally accumulate as the cells fuse to form multinucleated muscle fibers. In most instances, the contractile protein genes are regulated transcriptionally, which suggests that a common molecular mechanism may coordinate the expression of this diverse and evolutionarily unrelated gene set. Recent studies have examined the muscle-specific cis-acting elements associated with numerous contractile protein genes. All of the identified regulatory elements are positioned in the 5'-flanking regions, usually within 1,500 base pairs of the transcription start site. Surprisingly, a DNA consensus sequence that is common to each contractile protein gene has not been identified. In contrast to the results of these earlier studies, the authors have found that the 5'-flanking region of the quail troponin I (TnI) gene is not sufficient to permit the normal myofiber transcriptional activation of the gene. Instead, the TnI gene utilizes a unique internal regulatory element that is responsible for the correct myofiber-specific expression pattern associated with the TnI gene. This is the first example in which a contractile protein gene has been shown to rely primarily on an internal regulatory element to elicit transcriptional activation during myogenesis. The diversity of regulatory elements associated with the contractile protein genes suggests that the temporal expression of the genes may involve individual cis-trans regulatory components specific for each gene.

  17. Effects of carbocisteine on sialyl-Lewis x expression in an airway carcinoma cell line stimulated with tumor necrosis factor-alpha.

    PubMed

    Ishibashi, Yuji; Imai, Shigeru; Inouye, Yoshio; Okano, Teruo; Taniguchi, Akiyoshi

    2006-01-20

    Carbocisteine is a mucoregulatory drug normalizing sialic acid and fucose contents in mucins through the regulation of glycosyltransferase activities. Tumor necrosis factor (TNF)-alpha-induced overexpression of sialyl-Lewis x epitopes, containing sialic acid and fucose, in mucins were previously reported to be regulated by glycosyltransferase mRNAs expression through phosphatidyl inositol-specific phospholipase C (PI-PLC) signaling pathways [Ishibashi, Y., Inouye, Y., Okano, T., Taniguchi, A., 2005. Regulation of sialyl-Lewis x epitope expression by TNF-alpha and EGF in an airway carcinoma cell line. Glycoconj. J. 22, 53-62]. To investigate the mechanism behind the mucoregulatory action of carbocisteine, the present study evaluated the effects of carbocisteine on TNF-alpha-induced overexpression of sialyl-Lewis x epitopes in NCI-H292 cells. 100 mug/ml of carbocisteine was able to inhibit the TNF-alpha-induced expression of hST3GallV mRNA, FUT3 mRNA, C2/4GnT mRNA and sialyl-Lewis x epitopes as well as the TNF-alpha-induced activity of PI-PLC in NCI-H292 cells. These findings suggest that carbocisteine may normalize the sialyl-Lewis x epitopes expression in mucins through the inhibition of cellular PI-PLC activity in vivo.

  18. Cell Cycle Programs of Gene Expression Control Morphogenetic Protein Localization

    PubMed Central

    Lord, Matthew; Yang, Melody C.; Mischke, Michelle; Chant, John

    2000-01-01

    Genomic studies in yeast have revealed that one eighth of genes are cell cycle regulated in their expression. Almost without exception, the significance of cell cycle periodic gene expression has not been tested. Given that many such genes are critical to cellular morphogenesis, we wanted to examine the importance of periodic gene expression to this process. The expression profiles of two genes required for the axial pattern of cell division, BUD3 and BUD10/AXL2/SRO4, are strongly cell cycle regulated. BUD3 is expressed close to the onset of mitosis. BUD10 is expressed in late G1. Through promotor-swap experiments, the expression profile of each gene was altered and the consequences examined. We found that an S/G2 pulse of BUD3 expression controls the timing of Bud3p localization, but that this timing is not critical to Bud3p function. In contrast, a G1 pulse of BUD10 expression plays a direct role in Bud10p localization and function. Bud10p, a membrane protein, relies on the polarized secretory machinery specific to G1 to be delivered to its proper location. Such a secretion-based targeting mechanism for membrane proteins provides cells with flexibility in remodeling their architecture or evolving new forms. PMID:11134078

  19. Expression of heat shock protein genes in insect stress responses

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The heat shock proteins (HSPs) that are abundantly expressed in insects are important modulators of insect survival. Expression of HSP genes in insects is not only developmentally regulated, but also induced by various stressors in order to confer protection against such stressors. The expression o...

  20. Green Fluorescent Protein as a Marker for Gene Expression

    NASA Astrophysics Data System (ADS)

    Chalfie, Martin; Tu, Yuan; Euskirchen, Ghia; Ward, William W.; Prasher, Douglas C.

    1994-02-01

    A complementary DNA for the Aequorea victoria green fluorescent protein (GFP) produces a fluorescent product when expressed in prokaryotic (Escherichia coli) or eukaryotic (Caenorhabditis elegans) cells. Because exogenous substrates and cofactors are not required for this fluorescence, GFP expression can be used to monitor gene expression and protein localization in living organisms.

  1. Gene expression profile analysis of ventilator-associated pneumonia

    PubMed Central

    XU, XIAOLI; YUAN, BO; LIANG, QUAN; HUANG, HUIMIN; YIN, XIANGYI; SHENG, XIAOYUE; NIE, NIUYAN; FANG, HONGMEI

    2015-01-01

    Based on the gene expression profile of patients with ventilator-associated pneumonia (VAP) and patients not affected by the disease, the present study aimed to enhance the current understanding of VAP development using bioinformatics methods. The expression profile GSE30385 was downloaded from the Gene Expression Omnibus database. The Linear Models for Microarray Data package in R language was used to screen and identify differentially expressed genes (DEGs), which were grouped as up- and down-regulated genes. The up- and downregulated genes were functionally enriched using the Database for Annotation, Visualization and Integrated Discovery system and then annotated according to TRANSFAC, Tumor Suppressor Gene and Tumor Associated Gene databases. Subsequently, the protein-protein interaction (PPI) network was constructed, followed by module analysis using CFinder software. A total of 69 DEGs, including 33 up- and 36 downregulated genes were screened out in patients with VAP. Upregulated genes were mainly enriched in functions and pathways associated with the immune response (including the genes ELANE and LTF) and the mitogen-activated protein kinase (MAPK) signaling pathway (including MAPK14). The PPI network comprised 64 PPI pairs and 44 nodes. The top two modules were enriched in different pathways, including the MAPK signaling pathway. Genes including ELANE, LTF and MAPK14 may have important roles in the development of VAP via altering the immune response and the MAPK signaling pathway. PMID:26459786

  2. Building predictive gene signatures through simultaneous assessment of transcription factor activation and gene expression.

    EPA Science Inventory

    Building predictive gene signatures through simultaneous assessment of transcription factor activation and gene expression Exposure to many drugs and environmentally-relevant chemicals can cause adverse outcomes. These adverse outcomes, such as cancer, have been linked to mol...

  3. IDENTIFICATION OF BIOLOGICALLY RELEVANT GENES USING A DATABASE OF RAT LIVER AND KIDNEY BASELINE GENE EXPRESSION

    EPA Science Inventory

    Microarray data from independent labs and studies can be compared to potentially identify toxicologically and biologically relevant genes. The Baseline Animal Database working group of HESI was formed to assess baseline gene expression from microarray data derived from control or...

  4. Meta-analysis of gene expression data identifies causal genes for prostate cancer.

    PubMed

    Wang, Xiang-Yang; Hao, Jian-Wei; Zhou, Rui-Jin; Zhang, Xiang-Sheng; Yan, Tian-Zhong; Ding, De-Gang; Shan, Lei

    2013-01-01

    Prostate cancer is a leading cause of death in male populations across the globe. With the advent of gene expression arrays, many microarray studies have been conducted in prostate cancer, but the results have varied across different studies. To better understand the genetic and biologic mechanisms of prostate cancer, we conducted a meta-analysis of two studies on prostate cancer. Eight key genes were identified to be differentially expressed with progression. After gene co-expression analysis based on data from the GEO database, we obtained a co- expressed gene list which included 725 genes. Gene Ontology analysis revealed that these genes are involved in actin filament-based processes, locomotion and cell morphogenesis. Further analysis of the gene list should provide important clues for developing new prognostic markers and therapeutic targets.

  5. Identifying the optimal gene and gene set in hepatocellular carcinoma based on differential expression and differential co-expression algorithm.

    PubMed

    Dong, Li-Yang; Zhou, Wei-Zhong; Ni, Jun-Wei; Xiang, Wei; Hu, Wen-Hao; Yu, Chang; Li, Hai-Yan

    2017-02-01

    The objective of this study was to identify the optimal gene and gene set for hepatocellular carcinoma (HCC) utilizing differential expression and differential co-expression (DEDC) algorithm. The DEDC algorithm consisted of four parts: calculating differential expression (DE) by absolute t-value in t-statistics; computing differential co-expression (DC) based on Z-test; determining optimal thresholds on the basis of Chi-squared (χ2) maximization and the corresponding gene was the optimal gene; and evaluating functional relevance of genes categorized into different partitions to determine the optimal gene set with highest mean minimum functional information (FI) gain (Δ*G). The optimal thresholds divided genes into four partitions, high DE and high DC (HDE-HDC), high DE and low DC (HDE-LDC), low DE and high DC (LDE‑HDC), and low DE and low DC (LDE-LDC). In addition, the optimal gene was validated by conducting reverse transcription-polymerase chain reaction (RT-PCR) assay. The optimal threshold for DC and DE were 1.032 and 1.911, respectively. Using the optimal gene, the genes were divided into four partitions including: HDE-HDC (2,053 genes), HED-LDC (2,822 genes), LDE-HDC (2,622 genes), and LDE-LDC (6,169 genes). The optimal gene was microtubule‑associated protein RP/EB family member 1 (MAPRE1), and RT-PCR assay validated the significant difference between the HCC and normal state. The optimal gene set was nucleoside metabolic process (GO\\GO:0009116) with Δ*G = 18.681 and 24 HDE-HDC partitions in total. In conclusion, we successfully investigated the optimal gene, MAPRE1, and gene set, nucleoside metabolic process, which may be potential biomarkers for targeted therapy and provide significant insight for revealing the pathological mechanism underlying HCC.