Hubert Hackl

ClueGO: a Cytoscape plug-in to decipher functionally grouped gene ontology and pathway annotation networks

Summary: We have developed ClueGO, an easy to use Cytoscape plug-in that strongly improves biological interpretation of large lists of genes. ClueGO integrates Gene Ontology (GO) terms as well as KEGG/BioCarta pathways and creates a functionally organized GO/pathway term network. It can analyze one or compare two lists of genes and comprehensively visualizes functionally grouped terms. A one-click update option allows ClueGO to automatically download the most recent GO/KEGG release at any time. ClueGO provides an intuitive representation of the analysis results and can be optionally used in conjunction with the GOlorize plug-in. Availability: http://www.ici.upmc.fr/cluego/cluegoDownload.shtml Contact: jerome.galon@crc.jussieu.fr Supplementary information: Supplementary data are available at Bioinformatics online.

Characterization of the immunophenotypes and antigenomes of colorectal cancers reveals distinct tumor escape mechanisms and novel targets for immunotherapy

BackgroundWhile large-scale cancer genomic projects are comprehensively characterizing the mutational spectrum of various cancers, so far little attention has been devoted to either define the antigenicity of these mutations or to characterize the immune responses they elicit. Here we present a strategy to characterize the immunophenotypes and the antigen-ome of human colorectal cancer.ResultsWe apply our strategy to a large colorectal cancer cohort (n = 598) and show that subpopulations of tumor-infiltrating lymphocytes are associated with distinct molecular phenotypes. The characterization of the antigenome shows that a large number of cancer-germline antigens are expressed in all patients. In contrast, neo-antigens are rarely shared between patients, indicating that cancer vaccination requires individualized strategy. Analysis of the genetic basis of the tumors reveals distinct tumor escape mechanisms for the patient subgroups. Hypermutated tumors are depleted of immunosuppressive cells and show upregulation of immunoinhibitory molecules. Non-hypermutated tumors are enriched with immunosuppressive cells, and the expression of immunoinhibitors and MHC molecules is downregulated. Reconstruction of the interaction network of tumor-infiltrating lymphocytes and immunomodulatory molecules followed by a validation with 11 independent cohorts (n = 1,945) identifies BCMA as a novel druggable target. Finally, linear regression modeling identifies major determinants of tumor immunogenicity, which include well-characterized modulators as well as a novel candidate, CCR8, which is then tested in an orthologous immunodeficient mouse model.ConclusionsThe immunophenotypes of the tumors and the cancer antigenome remain widely unexplored, and our findings represent a step toward the development of personalized cancer immunotherapies.

PathwayExplorer: web service for visualizing high-throughput expression data on biological pathways.

While generation of high-throughput expression data is becoming routine, the fast, easy, and systematic presentation and analysis of these data in a biological context is still an obstacle. To address this need, we have developed PathwayExplorer, which maps expression profiles of genes or proteins simultaneously onto major, currently available regulatory, metabolic and cellular pathways from KEGG, BioCarta and GenMAPP. PathwayExplorer is a platform-independent web server application with an optional standalone Java application using a SOAP (simple object access protocol) interface. Mapped pathways are ranked for the easy selection of the pathway of interest, displaying all available genes of this pathway with their expression profiles in a selectable and intuitive color code. Pathway maps produced can be downloaded as PNG, JPG or as high-resolution vector graphics SVG. The web service is freely available at ; the standalone client can be downloaded at .

Comparative transcriptomics of human multipotent stem cells during adipogenesis and osteoblastogenesis

BackgroundA reciprocal relationship between bone and fat development in osteoporosis is clinically well established. Some of the key molecular regulators involved in this tissue replacement process have been identified. The detailed mechanisms governing the differentiation of mesenchymal stem cells (MSC) – the key cells involved – are however only now beginning to emerge. In an attempt to address the regulation of the adipocyte/osteoblast balance at the level of gene transcription in a comprehensive and unbiased manner, we performed a large-scale gene expression profiling study using a unique cellular model, human multipotent adipose tissue-derived stem cells (hMADS).ResultsThe analysis of 1606 genes that were found to be differentially expressed between adipogenesis and osteoblastogenesis revealed gene repression to be most prevalent prior to commitment in both lineages. Computational analyses suggested that this gene repression is mediated by miRNAs. The transcriptional activation of lineage-specific molecular processes in both cases occurred predominantly after commitment. Analyses of the gene expression data and promoter sequences produced a set of 65 genes that are candidates for genes involved in the process of adipocyte/osteoblast commitment. Four of these genes were studied in more detail: LXRα and phospholipid transfer protein (PLTP) for adipogenesis, the nuclear receptor COUP-TF1 and one uncharacterized gene, TMEM135 for osteoblastogenesis. PLTP was secreted during both early and late time points of hMADS adipocyte differentiation. LXRα, COUP-TF1, and the transmembrane protein TMEM135 were studied in primary cultures of differentiating bone marrow stromal cells from healthy donors and were found to be transcriptionally activated in the corresponding lineages.ConclusionOur results reveal gene repression as a predominant early mechanism before final cell commitment. We were moreover able to identify 65 genes as candidates for genes controlling the adipocyte/osteoblast balance and to further evaluate four of these. Additional studies will explore the precise role of these candidate genes in regulating the adipogenesis/osteoblastogenesis switch.

Grape-seed derived procyanidins interfere with adipogenesis of 3T3-L1 cells at the onset of differentiation

OBJECTIVE:Our groups previous results on the effects of a grape seed procyanidin extract (GSPE) on adipose metabolism showed that peroxisome proliferator-activated receptor-γ (PPARγ) plays a central role in the lipolytic effects of GSPE on adipocytes. Since PPARγ2 is a main regulator of the differentiation process of adipocytes, we investigated whether GSPE affects the adipogenesis of 3T3-L1 cells.DESIGN:We performed a time point screening by treating 3T3-L1 cells with GSPE during the differentiation process for 24 h.MEASUREMENTS:Differentiation markers and differential gene expression due to GSPE treatment (using the microarray technique).RESULTS:Twenty four hour-GSPE treatment at the onset of differentiation reduces adipose-specific markers and maintains the expression of preadipocyte marker preadipocyte factor-1 (Pref-1) significantly elevated. These effects were not found in other time points. Microarray analysis of gene expression after GSPE treatment at the early stage of differentiation showed a modified gene expression profile in which cell cycle and growth-related genes were downregulated by GSPE.CONCLUSION:These results suggest that GSPE affects adipogenesis, mainly at the induction of differentiation, and that procyanidins may have a new role in which they impede the formation of adipose cells.

MARS: Microarray analysis, retrieval, and storage system

BackgroundMicroarray analysis has become a widely used technique for the study of gene-expression patterns on a genomic scale. As more and more laboratories are adopting microarray technology, there is a need for powerful and easy to use microarray databases facilitating array fabrication, labeling, hybridization, and data analysis. The wealth of data generated by this high throughput approach renders adequate database and analysis tools crucial for the pursuit of insights into the transcriptomic behavior of cells.ResultsMARS (Microarray Analysis and Retrieval System) provides a comprehensive MIAME supportive suite for storing, retrieving, and analyzing multi color microarray data. The system comprises a laboratory information management system (LIMS), a quality control management, as well as a sophisticated user management system. MARS is fully integrated into an analytical pipeline of microarray image analysis, normalization, gene expression clustering, and mapping of gene expression data onto biological pathways. The incorporation of ontologies and the use of MAGE-ML enables an export of studies stored in MARS to public repositories and other databases accepting these documents.ConclusionWe have developed an integrated system tailored to serve the specific needs of microarray based research projects using a unique fusion of Web based and standalone applications connected to the latest J2EE application server technology. The presented system is freely available for academic and non-profit institutions. More information can be found at http://genome.tugraz.at.

Computational genomics tools for dissecting tumour-immune cell interactions

Recent breakthroughs in cancer immunotherapy and decreasing costs of high-throughput technologies have sparked intensive research into tumour–immune cell interactions using genomic tools. The wealth of the generated data and the added complexity pose considerable challenges and require computational tools to process, to analyse and to visualize the data. Recently, various tools have been developed and used to mine tumour immunologic and genomic data effectively and to provide novel mechanistic insights. Here, we review computational genomics tools for cancer immunology and provide information on the requirements and functionality in order to assist in the selection of tools and assembly of analytical pipelines.

Early structural and metabolic cardiac remodelling in response to inducible adipose triglyceride lipase ablation

AIMS While chronic alterations in cardiac triacylglycerol (TAG) metabolism and accumulation are associated with cardiomyopathy, it is unclear whether TAG catabolizing enzymes such as adipose triglyceride lipase (ATGL) play a role in acquired cardiomyopathies. Importantly, germline deletion of ATGL leads to marked cardiac steatosis and heart failure in part through reducing peroxisome proliferator-activated receptor α (PPARα) activity and subsequent fatty acid oxidation (FAO). However, whether ATGL deficiency specifically in adult cardiomyocytes contributes to impaired PPARα activity, cardiac function, and metabolism is not known. METHODS AND RESULTS To study the effects of acquired cardiac ATGL deficiency on cardiac PPARα activity, function, and metabolism, we generated adult mice with tamoxifen-inducible cardiomyocyte-specific ATGL deficiency (icAtglKO). Within 4-6 weeks following ATGL ablation, icAtglKO mice had markedly increased myocardial TAG accumulation, fibrotic remodelling, and pathological hypertrophy. Echocardiographic analysis of hearts in vivo revealed that contractile function was moderately reduced in icAtglKO mice. Analysis of energy metabolism in ex vivo perfused working hearts showed diminished FAO rates which was not paralleled by markedly impaired PPARα target gene expression. CONCLUSIONS This study shows that acquired cardiomyocyte-specific ATGL deficiency in adult mice is sufficient to promote fibrotic and hypertrophic cardiomyopathy and impair myocardial FAO in the absence of markedly reduced PPARα signalling.

The Human Placental Sexome Differs between Trophoblast Epithelium and Villous Vessel Endothelium

Molecular mechanisms underlying sexual dimorphism in mammals, fetal sex influences on intrauterine development, and the sex-biased susceptibility for selected diseases in adulthood are novel areas of current research. As importantly, two decades of multifaceted research has established that susceptibility to many adult disorders originates in utero, commonly secondary to the effects of placental dysfunction. We hypothesized that fetal sex influences gene expression and produces functional differences in human placentas. We thus extended previous studies on sexual dimorphism in mammals, which used RNA isolated from whole tissues, to investigate the effects of sex on four cell-phenotypes within a single key tissue, human placental villi. The cells studied included cytotrophoblasts, syncytiotrophoblast, arterial and venous endothelial cells. The cells were isolated from placentas of male or female fetuses and subjected to microarray analysis. We found that fetal sex differentially affected gene expression in a cell-phenotype dependent manner among all four cell-phenotypes. The markedly enriched pathways in males were identified to be signaling pathways for graft-versus-host disease as well as the immune and inflammatory systems that parallel the reported poorer outcome of male fetuses. Our study is the first to compare global gene expression by microarray analysis in purified, characterized, somatic cells from a single human tissue, i.e. placental villi. Importantly, our findings demonstrate that there are cell-phenotype specific, and tissue-specific, sex-biased responses in the human placenta, suggesting fetal sex should be considered as an independent variable in gene expression analysis of human placental villi.

Differential transcriptional modulation of biological processes in adipocyte triglyceride lipase and hormone-sensitive lipase-deficient mice

Adipocyte triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL) are intracellular lipases that mobilize triglycerides, the main energy source in mammals. Deletion of genes encoding ATGL (Pnpla2) or HSL (Lipe) in mice results in striking phenotypic differences, suggesting distinct roles for these lipases. The goal of the present study was to identify the biological processes that are modulated in the metabolic tissues of ATGL- and HSL-deficient mice. DNA microarrays were employed to provide full genome coverage concerning the types of genes that are differentially expressed in wild-type and mutant mice. For both mouse models, transcript signatures were identified in white adipose tissue, brown adipose tissue (BAT), skeletal muscle (SM), cardiac muscle (CM), and liver. Genetic ablation of ATGL and HSL alters the transcript levels of a large number of genes in metabolic tissues. The genes affected in the two models are, however, largely different ones. Indeed, only one biological process was modulated in the same way in both mouse models, namely the down-regulation of fatty acid metabolism in BAT. The most pronounced modulation of biological processes was observed in ATGL-/- CM, in which a concerted down-regulation of transcripts associated with oxidative pathways was observed. In HSL-/- mice, in contrast, the most marked changes were seen in SM, namely, alterations in transcript levels reflecting a change of energy source from lipid to carbohydrate. The transcript signatures also provided novel insights into the metabolic derangements that are characteristic of ATGL-/- mice. Our findings suggest that ATGL and HSL differentially modulate biological processes in metabolic tissues. We hypothesize that the intermediary metabolites of the lipolytic pathways are signaling molecules and activators of a wide range of biochemical and cellular processes in mammals.