Integrative Biology Approach Research Articles

An Integrative Network Biology Approach to Evaluate the Role of Endoplasmic Reticulum Stress Response in Obese Type 2 Diabetes

Extracellular/intracellular stimuli can influence eukaryotic cell function through organelles that regulate critical signaling pathways. The endoplasmic reticulum (ER), for example, impacts cellular processes including protein synthesis, folding and secretion; amino acid transport; apoptosis; cell proliferation; lipid synthesis across major cell types in response to stimuli such as accumulation of misfolded proteins and glucose deprivation. Dysregulated signaling pathways underlying the ER-mediated processes mentioned above have been linked to disease conditions such as diabetes, obesity, and Alzheimer's disease. Our current understanding, however, lacks a detailed network view that integrates organelle-mediated pathway dysregulation with cellular processes and disease pathogenesis. In this report, we introduce an integrative network biology approach that combines ER-stress response pathways with basic cellular processes using data from peer-reviewed literature. As an example, we apply our systems biology approach to study the role of ER stress in pancreatic β cells under obese diabetic conditions, generate testable hypotheses, and provide novel insights into β-cell pathogenesis.

Integrative System Biology Strategies for Disease Biomarker Discovery

Biomarkers are currently widely used to diagnose diseases, monitor treatments, and evaluate potential drug candidates. Research of differential Omics accelerate the advancements of biomarkers' discovery. By extracting biological knowledge from the 'omics' through integration, integrative system biology creates predictive models of cells, organs, biochemical processes and complete organisms, in addition to identifying human disease biomarkers. Recent development in high-throughput methods enables analysis of genome, transcriptome, proteome, and metabolome at an unprecedented scale, thus contributing to the deluge of experimental data in numerous public databases. Several integrative system biology approaches have been developed and applied to the discovery of disease biomarkers from databases. In this review, we highlight several of these approaches and identify future steps in the context of the field of integrative system biology.

Common Genetic Variants and Gene-Expression Changes Associated with Bipolar Disorder Are Over-Represented in Brain Signaling Pathway Genes

Despite high heritability, the genetic variants influencing bipolar disorder (BD) susceptibility remain largely unknown. Low statistical power to detect the small effect-size alleles believed to underlie much of the genetic risk and possible heterogeneity between cohorts are an increasing concern. Integrative biology approaches might offer advantages over genetic analysis alone by combining different genomic datasets at the higher level of biological processes rather than the level of specific genetic variants or genes. We employed this strategy to identify biological processes involved in BD etiopathology. Three genome-wide association studies and a brain gene-expression study were combined with the Human Protein Reference Database protein-protein interaction network data. We used bioinformatic analysis to search for biological networks with evidence of association on the basis of enrichment among both genetic and differential-expression associations with BD. We identified association with gene networks involved in transmission of nerve impulse, Wnt, and Notch signaling. Three features stand out among these genes: 1) they localized to the human postsynaptic density, which is crucial for neuronal function; 2) their mouse knockouts present altered behavioral phenotypes; and 3) some are known targets of the pharmacological treatments for BD. Genetic and gene-expression associations of BD cluster in discrete regions of the protein-protein interaction network. We found replicated evidence for association for networks involving several interlinked signaling pathways. These genes are promising candidates to generate animal models and pharmacological interventions. Our results demonstrate the potential advantage of integrative biology analyses of BD datasets.

Copy Number and Loss of Heterozygosity Detected by SNP Array of Formalin-Fixed Tissues Using Whole-Genome Amplification

The requirement for large amounts of good quality DNA for whole-genome applications prohibits their use for small, laser capture micro-dissected (LCM), and/or rare clinical samples, which are also often formalin-fixed and paraffin-embedded (FFPE). Whole-genome amplification of DNA from these samples could, potentially, overcome these limitations. However, little is known about the artefacts introduced by amplification of FFPE-derived DNA with regard to genotyping, and subsequent copy number and loss of heterozygosity (LOH) analyses. Using a ligation adaptor amplification method, we present data from a total of 22 Affymetrix SNP 6.0 experiments, using matched paired amplified and non-amplified DNA from 10 LCM FFPE normal and dysplastic oral epithelial tissues, and an internal method control. An average of 76.5% of SNPs were called in both matched amplified and non-amplified DNA samples, and concordance was a promising 82.4%. Paired analysis for copy number, LOH, and both combined, showed that copy number changes were reduced in amplified DNA, but were 99.5% concordant when detected, amplifications were the changes most likely to be ‘missed’, only 30% of non-amplified LOH changes were identified in amplified pairs, and when copy number and LOH are combined ∼50% of gene changes detected in the unamplified DNA were also detected in the amplified DNA and within these changes, 86.5% were concordant for both copy number and LOH status. However, there are also changes introduced as ∼20% of changes in the amplified DNA are not detected in the non-amplified DNA. An integrative network biology approach revealed that changes in amplified DNA of dysplastic oral epithelium localize to topologically critical regions of the human protein-protein interaction network, suggesting their functional implication in the pathobiology of this disease. Taken together, our results support the use of amplification of FFPE-derived DNA, provided sufficient samples are used to increase power and compensate for increased error rates.

Investigation of the adaptation of Lactococcus lactis to isoleucine starvation integrating dynamic transcriptome and proteome information

BackgroundAmino acid assimilation is crucial for bacteria and this is particularly true for Lactic Acid Bacteria (LAB) that are generally auxotroph for amino acids. The global response of the LAB model Lactococcus lactis ssp. lactis was characterized during progressive isoleucine starvation in batch culture using a chemically defined medium in which isoleucine concentration was fixed so as to become the sole limiting nutriment. Dynamic analyses were performed using transcriptomic and proteomic approaches and the results were analysed conjointly with fermentation kinetic data.ResultsThe response was first deduced from transcriptomic analysis and corroborated by proteomic results. It occurred progressively and could be divided into three major mechanisms: (i) a global down-regulation of processes linked to bacterial growth and catabolism (transcription, translation, carbon metabolism and transport, pyrimidine and fatty acid metabolism), (ii) a specific positive response related to the limiting nutrient (activation of pathways of carbon or nitrogen metabolism and leading to isoleucine supply) and (iii) an unexpected oxidative stress response (positive regulation of aerobic metabolism, electron transport, thioredoxin metabolism and pyruvate dehydrogenase). The involvement of various regulatory mechanisms during this adaptation was analysed on the basis of transcriptomic data comparisons. The global regulator CodY seemed specifically dedicated to the regulation of isoleucine supply. Other regulations were massively related to growth rate and stringent response.ConclusionThis integrative biology approach provided an overview of the metabolic pathways involved during isoleucine starvation and their regulations. It has extended significantly the physiological understanding of the metabolism of L. lactis ssp. lactis. The approach can be generalised to other conditions and will contribute significantly to the identification of the biological processes involved in complex regulatory networks of micro-organisms.

Abstract 140: Tumor suppressor role of microRNA-1 in prostate cancer

Abstract MicroRNAs (miRNA) are small non-coding RNAs that reduce the translation of target mRNAs via 3’ untranslated region (UTR) binding sites. MicroRNA-1 has been previously shown by genome-wide studies in our lab to be under-expressed in prostate cancer (PCa) tissues and lost in most of the NCI-60 cell lines. We hypothesize that miRNA-1 plays a role in the progression of PCa through tumor suppressor mechanisms. To explore this we used an integrated system biology approach by re-expressing miRNA-1 and studying its targetome by mRNA microarray analysis, miRNA target prediction, pathway analysis, and connectivity mapping. From these analyses we found that miRNA-1 down-regulated transcripts were significantly enriched for predicted targets and genes involved in cell cycle progression, mitosis, and DNA replication/repair. Following this we found evidence that miRNA-1 could increase the number of cells in S-phase while reducing mitotic cell numbers and the protein levels of genes required for S-phase progression including CHK1 and MCM7. Connectivity mapping of miRNA-1 expression profiles revealed that the effects of this miRNA overlap with histone deacetylase inhibitors like Vorinostat and Trichostain (TSA). Supporting this we found that miRNA-1 could be re-expressed when PCa cells were treated with epigenetic drugs such as 5-azadeoxycytidine and TSA. This could implicate a downstream role of miRNA-1 in these epigenetic mechanisms. Moreover, miRNA-1 expression increased radiation mediated levels and resolution of the DNA damage biomarker gamma-H2A.X, while reducing the clonogenic survival of PCa cells to radiation. This indicates that the loss of miRNA-1 could have clinical importance, by leading to an increase in the survivability of PCa cells to radiation therapy. Further, we found that several possible targets of miRNA-1 are actin binding genes that could play roles in both mitosis and migration. Using 3’UTR reporter assays we found that miRNA-1 directly targets several of these genes including, LASP-1, XPO6, NOTCH-3, FN1, MMD, and PTMA. Subsequently, epi-fluorescence microscopy revealed that miRNA-1 was capable of increasing F-actin levels and altering its organization in PCa cells. In conclusion, our findings indicate that miRNA-1 functions as a tumor suppressor in PCa by influencing cell cycle, the DNA damage response, and regulating actin related genes involved in mitosis and cell motility. These studies implicate a possible utility of miRNA-1 expression patterns in the therapeutic outcome of PCa patients. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 140. doi:10.1158/1538-7445.AM2011-140

Abstract 5110: RNA helicase p68 is a new marker for breast cancer heterogeneity

Abstract Two major challenges in diagnosis and therapy of breast cancer lie in its heterogeneity and drug resistance. RNA helicase p68 (DDX5) has been shown to be involved in all aspects of RNA metabolism and serves as a transcriptional co-regulator. However, its functional role in breast cancer remains elusive. We utilized an integrative biology approach strategy to further define the role of p68 in breast cancer. Specifically, we determined the expression pattern of p68 at both mRNA and protein levels for a panel of ∼50 breast cancer cell lines. We performed immunohistochemistry of p68 using tissue microarrays containing a total of over 200 cases of primary human malignant breast cancer (various grades and stages). We knocked down p68 in a panel of representative breast cancer cell lines (e.g., SKBR3 and MDA-MB-231) and investigated the functional consequences such as their proliferation and responses to Lapatinib after the p68 knockdown. Finally, we carried out SILAC-based proteomic profiling of these breast cancer cells after p68 knockdown and identified p68-targeted proteins and networks contributing to the drug responses. We found that p68 expression pattern is distinct among different subtypes of breast cancers. More aggressive basal subtypes have predominantly high p68 protein expression while low p68 expression is predominantly associated with the luminal subtype. Knockdown of p68 inhibits the proliferation of breast cancer cells and sensitizes them to cancer drugs such as Lapatinib. We demonstrated that p68 is a new marker for breast cancer heterogeneity in addition to ER, PR, and Her2. Importantly, we identified p68-regualted cytoskeletal reorganization as a new mechanism in controlling breast cancer proliferation and drug resistance. In summary, RNA helicase p68 may serve as a new marker for breast cancer heterogeneity, as a predictor for drug resistance, and as a target for a combinational therapy to circumvent drug resistance of aggressive tumors. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5110. doi:10.1158/1538-7445.AM2011-5110

A study protocol for the evaluation of occupational mutagenic/carcinogenic risks in subjects exposed to antineoplastic drugs: a multicentric project

BackgroundSome industrial hygiene studies have assessed occupational exposure to antineoplastic drugs; other epidemiological investigations have detected various toxicological effects in exposure groups labeled with the job title. In no research has the same population been studied both environmentally and epidemiologically. The protocol of the epidemiological study presented here uses an integrated environmental and biological monitoring approach. The aim is to assess in hospital nurses preparing and/or administering therapy to cancer patients the current level of occupational exposure to antineoplastic drugs, DNA and chromosome damage as cancer predictive effects, and the association between the two.Methods/DesignAbout 80 healthy non-smoking female nurses, who job it is to prepare or handle antineoplastic drugs, and a reference group of about 80 healthy non-smoking female nurses not occupationally exposed to chemicals will be examined simultaneously in a cross-sectional study. All the workers will be recruited from five hospitals in northern and central Italy after their informed consent has been obtained.Evaluation of surface contamination and dermal exposure to antineoplastic drugs will be assessed by determining cyclophosphamide on selected surfaces (wipes) and on the exposed nurses' clothes (pads). The concentration of unmetabolized cyclophosphamide as a biomarker of internal dose will be measured in end-shift urine samples from exposed nurses.Biomarkers of effect and susceptibility will be assessed in exposed and unexposed nurses: urinary concentration of 8-hydroxy-2-deoxyguanosine; DNA damage detected using the single-cell microgel electrophoresis (comet) assay in peripheral white blood cells; micronuclei and chromosome aberrations in peripheral blood lymphocytes. Genetic polymorphisms for enzymes involved in metabolic detoxification (i.e. glutathione S-transferases) will also be analysed.Using standardized questionnaires, occupational exposure will be determined in exposed nurses only, whereas potential confounders (medicine consumption, lifestyle habits, diet and other non-occupational exposures) will be assessed in both groups of hospital workers.Statistical analysis will be performed to ascertain the association between occupational exposure to antineoplastic drugs and biomarkers of DNA and chromosome damage, after taking into account the effects of individual genetic susceptibility, and the presence of confounding exposures.DiscussionThe findings of the study will be useful in updating prevention procedures for handling antineoplastic drugs.

An integrative biological approach to the analysis of tissue culture data: application to the antitumour agent RHPS4

We describe a mathematical model of cell growth and death and explain how it can be used to integrate data from classic tissue culture experiments on antitumour agents and thus aid the identification of their mechanism of action. Experimental data relating to time- and dose-dependent changes in growth rate, cell cycle distribution, plus apoptotic and senescent fractions, are reinterpreted in terms of modulations to kinetic parameters that describe the rates at which cells transit between phenotypic compartments. The mathematical model is analytical, in the sense that the kinetic parameters are calculated from the experimental data directly, without any fitting process. Since the kinetic parameters are much more directly related to potential molecular targets than are the experimentally measured quantities, this approach can provide a more informative picture of the mechanism of action of the antitumour agent under investigation. We demonstrate the potential value of our model by applying it to data from RHPS4 (3,11-difluoro-6,8,13-trimethyl-8H-quino [4,3,2-kl] acridinium methosulfate). This agent is a DNA-interactive pentacyclic acridine for which at least three potential mechanisms of antitumour activity have been identified. Firstly RHPS4 is a telomerase inhibitor, secondly it is a telomerase-independent destabiliser of telomeres, and thirdly it is a telomere-independent binder to genomic DNA. Each mechanism can induce a separate, but overlapping, pattern of cellular responses, making the interpretation of tissue culture data very complex. Here we study the time- and dose-dependent effects of RHPS4 on the HCT116 cell line, and develop a five-compartment mathematical model to interpret the data. Application of the model to the data suggests that RHPS4 increases the rate at which cells became senescent state but, rather surprisingly, actually inhibits the rate of cell death. As a control, we also apply the model to data describing the time- and dose-dependent effects of doxorubicin since its mechanism of action is better characterised.

Genome-Wide DNA Methylation Maps in Follicular Lymphoma Cells Determined by Methylation-Enriched Bisulfite Sequencing

BackgroundFollicular lymphoma (FL) is a form of non-Hodgkin's lymphoma (NHL) that arises from germinal center (GC) B-cells. Despite the significant advances in immunotherapy, FL is still not curable. Beyond transcriptional profiling and genomics datasets, there currently is no epigenome-scale dataset or integrative biology approach that can adequately model this disease and therefore identify novel mechanisms and targets for successful prevention and treatment of FL.Methodology/Principal FindingsWe performed methylation-enriched genome-wide bisulfite sequencing of FL cells and normal CD19+ B-cells using 454 sequencing technology. The methylated DNA fragments were enriched with methyl-binding proteins, treated with bisulfite, and sequenced using the Roche-454 GS FLX sequencer. The total number of bases covered in the human genome was 18.2 and 49.3 million including 726,003 and 1.3 million CpGs in FL and CD19+ B-cells, respectively. 11,971 and 7,882 methylated regions of interest (MRIs) were identified respectively. The genome-wide distribution of these MRIs displayed significant differences between FL and normal B-cells. A reverse trend in the distribution of MRIs between the promoter and the gene body was observed in FL and CD19+ B-cells. The MRIs identified in FL cells also correlated well with transcriptomic data and ChIP-on-Chip analyses of genome-wide histone modifications such as tri-methyl-H3K27, and tri-methyl-H3K4, indicating a concerted epigenetic alteration in FL cells.Conclusions/SignificanceThis study is the first to provide a large scale and comprehensive analysis of the DNA methylation sequence composition and distribution in the FL epigenome. These integrated approaches have led to the discovery of novel and frequent targets of aberrant epigenetic alterations. The genome-wide bisulfite sequencing approach developed here can be a useful tool for profiling DNA methylation in clinical samples.

Abstract 2016: Role of RNA helicase p68 in breast cancer

Abstract (a) Background and purpose. Two major challenges in diagnosis and therapy of breast cancer lie in its heterogeneity and drug resistance. RNA helicase p68 (DDX5) has been shown to be involved in all aspects of RNA metabolism and serves as a transcriptional co-regulator. Strikingly, a recent study demonstrated a new role of p53 in modulating miRNA processing, probably through the p53-Drosha/p68 interactions. As a result, the gene expression regulated by p53-Drosha/p68 is under intensive investigation. Previous work has shown that p68 is one of the potential 6-gene predictors of breast cancer resistance to Lapatinib, a dual ErbB2/EGFR tyrosine kinase inhibitor. Furthermore, we and others have shown that p68 is involved in a complex protein-protein interacting network (e.g., Ca2+/CaM binding). This study is designed to mechanistically define the role of p68 in human breast cancer using an integrative biology approach. (b) Experimental procedures. We utilized a multifaceted strategy including cellular and molecular biology and various emerging technologies. Specifically, we measured the expression pattern of p68 at both mRNA and protein levels for a panel of ∼50 breast cancer cell lines. We performed immunohistochemistry of p68 using human breast tumor microarrays containing a total of 225 cases of normal and malignant (various grades and stages) tissues of the breast. We knocked down p68 in two representative breast cancer cell lines (SKBR3 and MDA-MB-231) and investigated their proliferation and responses to Lapatinib before and after p68 knockdown. Finally, we carried out SILAC-based proteomic profiling of these breast cancer cells after p68 knockdown and identified p68-targeted proteins and networks contributing to the drug responses. (c) Results. P68 expression pattern is distinct among different subtypes of breast cancers. More aggressive basal A and B subtypes have predominantly high p68 protein expression while low p68 expression is predominantly associated with the luminal subtype. Knockdown of p68 inhibits the proliferation of breast cancer cells and sensitize them to cancer drugs such as Lapatinib. Importantly, we identified cofilin-1, a major actin severing and -depolymerization protein, as a key target of p68 through proteomics studies. Knockdown of p68 increases cofilin expression. Cofilin has been previously implicated in platinum-resistance of ovarian cancer cells, and migration and invasion of breast cancer cells. Therefore, our results suggest that high p68 expression may contribute to drug resistance and metastasis of breast cancer cells through cytoskeletal reorganization by promoting actin polymerization. (d) Conclusions. RNA helicase p68 may contribute to personalized medicine by serving as a new molecular marker to characterize breast cancer subtypes, as a predictor for drug resistance, and as a target for a combinational therapy to circumvent drug resistance of aggressive tumors. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2016.

Analysis and modeling of the integrative response of Medicago truncatula to nitrogen constraints

An integrative biology approach was conducted in Medicago truncatula for: (i) unraveling the coordinated regulation of NO 3 − , NH 4 + and N 2 acquisition by legumes to fulfill the plant N demand; and (ii) modeling the emerging properties occurring at the whole plant level. Upon localized addition of a high level of mineral N, the three N acquisition pathways displayed similar systemic feedback repression to adjust N acquisition capacities to the plant N status. Genes associated to these responses were in contrast rather specific to the N source. Following an N deficit, NO 3 − fed plants maintained efficiently their N status through rapid functional and developmental up regulations while N 2 fed plants responded by long term plasticity of nodule development. Regulatory genes associated with various symbiotic stages were further identified. An ecophysiological model simulating relations between leaf area and roots N retrieval was developed and now furnishes an analysis grid to characterize a spontaneous or induced genetic variability for plant N nutrition. To cite this article: C. Salon et al., C. R. Biologies 332 (2009).

Examination of post-transcriptional regulations in prokaryotes by integrative biology

In cells, mRNA and protein levels are fine-regulated to adjust continuously to cellular needs. Recently, several large-scale studies in prokaryotes showed weak correlations between mRNA and protein abundances highlighting the significant importance of post-transcriptional regulations. Post-transcriptional regulations involve dynamic adaptation of mRNA and protein turnover and also modulation of the efficiency of mRNA translation into protein. mRNA and protein stabilities are function of both sequence determinants and decay processes. Translation efficiency is mainly dependent on ribosome synthesis and activity. Conciliation through an integrative biology approach of large-scale data obtained for each level of regulation is now required to better understand global cell response to different environmental growth conditions. In this review, we report mechanisms involved in mRNA and protein stability and translation regulation in prokaryotes, and their dependence on growth phase and environmental growth conditions is particularly highlighted. To cite this article: F. Picard et al., C. R. Biologies 332 (2009).

Commentaries on Viewpoint: Fatigue mechanisms determining exercise performance: Integrative physiology is systems physiology

Commentaries on Viewpoint: Fatigue mechanisms determining exercise performance: Integrative physiology is systems physiology

Genetics of gene expression and its effect on disease

Common human diseases result from the interplay of many genes and environmental factors. Therefore, a more integrative biology approach is needed to unravel the complexity and causes of such diseases. To elucidate the complexity of common human diseases such as obesity, we have analysed the expression of 23,720 transcripts in large population-based blood and adipose tissue cohorts comprehensively assessed for various phenotypes, including traits related to clinical obesity. In contrast to the blood expression profiles, we observed a marked correlation between gene expression in adipose tissue and obesity-related traits. Genome-wide linkage and association mapping revealed a highly significant genetic component to gene expression traits, including a strong genetic effect of proximal (cis) signals, with 50% of the cis signals overlapping between the two tissues profiled. Here we demonstrate an extensive transcriptional network constructed from the human adipose data that exhibits significant overlap with similar network modules constructed from mouse adipose data. A core network module in humans and mice was identified that is enriched for genes involved in the inflammatory and immune response and has been found to be causally associated to obesity-related traits.

Expression of hepatitis B virus proteins in transgenic mice alters lipid metabolism and induces oxidative stress in the liver

Hepatitis B virus transgenic mice (HBV-Tg mice) have been widely used as animal models in the study of pathogenesis and control of hepatitis B. It is important for the evaluation of such animal models to define the physiological differences between HBV-Tg and wild-type mice. The aim of this research was to investigate whether the integrated system biology approach that combines proteomics and metabonomics describes the physiological changes and provides new insights into the pathogenesis of the early stages of HBV infection. In this study the protein and metabolite profiles of the liver were established based on two-dimensional electrophoresis and HPLC/MS analysis. Several protein molecules, whose expression was altered in HBV-Tg mouse liver, were identified including protective enzymes against oxidative stress and regulatory proteins related to lipid metabolism. Metabonomics confirmed the potential derangement of lipid metabolism by discovering the intermediate and the final products of lipid metabolism that were markedly changed in transgenic mice. This study demonstrated that HBV antigens could impair host cell lipid metabolism and induce modest oxidative stress in vivo.

An Integrated Biological Approach to Nuclear Receptor Signaling in Physiological Control and Disease

The nuclear receptors (NRs)--vitamin D receptor (VDR); peroxisome proliferator-activated receptor (PPAR) alpha, delta, gamma; and pregnane X receptor (PXR)--act as sensors for various molecules encountered by the body on a daily basis. The effects of these ligands can be understood by the fact that numerous genes involved in the cellular processes, such as general homeostasis, growth, and defense against microbes, are under the control of these five NRs. The target gene and protein expression patterns of VDR, PPARs, and PXR; the resulting changes in metabolite levels; and their physiological consequences create a network that can be monitored by high-throughput methods and analyzed by multimodal approaches, such as systems biology. We suggest that the fine regulation of this NR network is specific to each human individual and depends, in part, on the constellation of regulatory small nucleotide polymorphisms (SNPs) in his or her genome. When regulatory SNPs affect NRs response elements, lifetime exposure to food components will have different accumulative consequences on the expression of the respective NR target genes. These differences will influence the individual's susceptibility to aging-related diseases, such as type 2 diabetes, atherosclerosis, cancer, and osteoporosis. Furthermore, it is anticipated that systems biology methods will also help to identify the most critical genes, proteins, or metabolites in the NR network that will serve as biomarkers for the early detection of these diseases.

The Arabidopsis genome: A foundation for plant research

The sequence of the first plant genome was completed and published at the end of 2000. This spawned a series of large-scale projects aimed at discovering the functions of the 25,000+ genes identified in Arabidopsis thaliana (Arabidopsis). This review summarizes progress made in the past five years and speculates about future developments in Arabidopsis research and its implications for crop science. The provision of large populations of gene disruption lines to the research community has greatly accelerated the impact of genomics on many areas of plant science. The tools and community organization required for plant integrative and systems biology approaches are now ready to accomplish the next big step in plant biology--the integration of knowledge and modeling of biological processes. In the future, plant science will continue to be enriched by the alignment of high-quality basic research (generally conducted in Arabidopsis), with strategic objectives in crop plants. The sequence and analysis of an increasing number of crop plant genomes enhance this alignment and provide new insights into genome evolution and crop plant domestication.

Inferring Hypotheses on Functional Relationships of Genes: Analysis of the Arabidopsis thaliana Subtilase Gene Family

The gene family of subtilisin-like serine proteases (subtilases) in Arabidopsis thaliana comprises 56 members, divided into six distinct subfamilies. Whereas the members of five subfamilies are similar to pyrolysins, two genes share stronger similarity to animal kexins. Mutant screens confirmed 144 T-DNA insertion lines with knockouts for 55 out of the 56 subtilases. Apart from SDD1, none of the confirmed homozygous mutants revealed any obvious visible phenotypic alteration during growth under standard conditions. Apart from this specific case, forward genetics gave us no hints about the function of the individual 54 non-characterized subtilase genes. Therefore, the main objective of our work was to overcome the shortcomings of the forward genetic approach and to infer alternative experimental approaches by using an integrative bioinformatics and biological approach. Computational analyses based on transcriptional co-expression and co-response pattern revealed at least two expression networks, suggesting that functional redundancy may exist among subtilases with limited similarity. Furthermore, two hubs were identified, which may be involved in signalling or may represent higher-order regulatory factors involved in responses to environmental cues. A particular enrichment of co-regulated genes with metabolic functions was observed for four subtilases possibly representing late responsive elements of environmental stress. The kexin homologs show stronger associations with genes of transcriptional regulation context. Based on the analyses presented here and in accordance with previously characterized subtilases, we propose three main functions of subtilases: involvement in (i) control of development, (ii) protein turnover, and (iii) action as downstream components of signalling cascades. Supplemental material is available in the Plant Subtilase Database (PSDB) (http://csbdb.mpimp-golm.mpg.de/psdb.html) , as well as from the CSB.DB (http://csbdb.mpimp-golm.mpg.de).

Validation of the Neuroinflammation Cycle as a Drug Discovery Target Using Integrative Chemical Biology and Lead Compound Development with an Alzheimers Disease-Related Mouse Model

The neuroinflammation cycle has been proposed as a potential therapeutic target in the development of new approaches to altering Alzheimer's disease (AD) progression. However, the efficacy and toxicological profile of compounds that focus only on classical NSAID targets have been disappointing to date. Therefore, we recently initiated an unbiased, integrative chemical biology approach that used a hierarchal set of cell-based screens, followed by efficacy analysis in a new AD-relevant animal model that more closely resembles human pathology endpoints in terms of neuroinflammation and neuronal loss. The prior investigations provided a proof of concept that targeting the neuroinflammation cycle may be a viable drug discovery approach for AD. However, recent informatics analyses of the high attrition rate in drug development have identified the need for starting drug development with lead compounds that are well below cut off values in computed molecular properties in order to facilitate late stage medicinal chemistry refinement to improve in vivo functions. We describe here how we are leveraging our novel, unbiased, integrative chemical biology approach for the rapid discovery of potential lead compounds for AD drug discovery. Specifically, we show that orally bioavailable compounds with the desired physical properties and in vivo functions can be identified in focused synthetic libraries composed of chemical diversifications of the inactive but privileged pyridazine molecular fragment.