The fou2 gain-of-function allele and the wild-type allele of Two Pore Channel 1 contribute to different extents or by different mechanisms to defense gene expression in Arabidopsis.

The fatty acid oxygenation up-regulated 2 (fou2) mutant in Arabidopsis thaliana creates a gain-of-function allele in a non-selective cation channel encoded by the Two Pore Channel 1 (TPC1) gene. This mutant genetically implicates cation fluxes in the control of the positive feedback loop whereby jasmonic acid (JA) stimulates its own synthesis. In this study we observed extensive transcriptome reprogramming in healthy fou2 leaves closely resembling that induced by treatment with methyl jasmonate, biotic stresses and the potassium starvation response. Proteomic analysis of fou2 leaves identified increased levels of seven biotic stress- and JA-inducible proteins. In agreement with these analyses, epistasis studies performed by crossing fou2 with aos indicated that elevated levels of JA in fou2 are the major determinant of the mutant phenotype. In addition, generation of fou2 aba1-5, fou2 etr1-1 and fou2 npr1-1 double mutants showed that the fou2 phenotype was only weakly affected by ABA levels and unaffected by mutations in NPR1 and ETR1. The results now suggest possible mechanisms whereby fou2 could induce JA synthesis/signaling early in the wound response. In contrast to fou2, transcriptome analysis of a loss-of-function allele of TPC1, tpc1-2, revealed no differential expression of JA biosynthesis genes in resting leaves. However, the analysis disclosed reduced mRNA levels of the pathogenesis-related genes PDF1.2a and THI2.1 in healthy and diseased tpc1-2 leaves. The results suggest that wild-type TPC1 contributes to their expression by mechanisms somewhat different from those affecting their expression in fou2.

Identification of corticosteroid-regulated genes in cardiomyocytes by serial analysis of gene expression.

Corticosteroids (aldosterone, cortisol/corticosterone) exert direct functional effects on cardiomyocytes. However, gene networks activated by corticosteroids in cardiomyocytes, as well as the involvement of the mineralocorticoid receptor (MR) vs the glucocorticoid receptor (GR) in these effects, remain largely unknown. Here we characterized the corticosteroid-dependent transcriptome in primary culture of neonatal mouse cardiomyocytes treated with 10(-6) M aldosterone, a concentration predicted to occupy both MR and GR. Serial analysis of gene expression revealed 101 aldosterone-regulated genes. The MR/GR specificity was characterized for one regulated transcript, namely ecto-ADP-ribosyltransferase-3 (Art3). Using cardiomyocytes from GR(null/null) or MR(null/null) mice we demonstrate that in GR(null/null) cardiomyocytes the response is abrogated, but it is fully maintained in MR(null/null) cardiomyocytes. We conclude that Art3 expression is regulated exclusively via the GR. Our study identifies a new set of corticosteroid-regulated genes in cardiomyocytes and demonstrates a new approach to studying the selectivity of MR- vs GR-dependent effects.

The effect of translocation-induced nuclear reorganization on gene expression.

Translocations are known to affect the expression of genes at the breakpoints and, in the case of unbalanced translocations, alter the gene copy number. However, a comprehensive understanding of the functional impact of this class of variation is lacking. Here, we have studied the effect of balanced chromosomal rearrangements on gene expression by comparing the transcriptomes of cell lines from controls and individuals with the t(11;22)(q23;q11) translocation. The number of differentially expressed transcripts between translocation-carrying and control cohorts is significantly higher than that observed between control samples alone, suggesting that balanced rearrangements have a greater effect on gene expression than normal variation. Many of the affected genes are located along the length of the derived chromosome 11. We show that this chromosome is concomitantly altered in its spatial organization, occupying a more central position in the nucleus than its nonrearranged counterpart. Derivative 22-mapping chromosome 22 genes, on the other hand, remain in their usual environment. Our results are consistent with recent studies that experimentally altered nuclear organization, and indicated that nuclear position plays a functional role in regulating the expression of some genes in mammalian cells. Our study suggests that chromosomal translocations can result in hitherto unforeseen, large-scale changes in gene expression that are the consequence of alterations in normal chromosome territory positioning. This has consequences for the patterns of gene expression change seen during tumorigenesis-associated genome instability and during the karyotype changes that lead to speciation.

Angiogenesis-related gene expression profile with independent prognostic value in advanced ovarian carcinoma.

BACKGROUND Ovarian carcinoma is the most important cause of gynecological cancer-related mortality in Western societies. Despite the improved median overall survival in patients receiving chemotherapy regimens such as paclitaxel and carboplatin combination, relapse still occurs in most advanced diseased patients. Increased angiogenesis is associated with rapid recurrence and decreased survival in ovarian cancer. This study was planned to identify an angiogenesis-related gene expression profile with prognostic value in advanced ovarian carcinoma patients. METHODOLOGY/PRINCIPAL FINDINGS RNAs were collected from formalin-fixed paraffin-embedded samples of 61 patients with III/IV FIGO stage ovarian cancer who underwent surgical cytoreduction and received a carboplatin plus paclitaxel regimen. Expression levels of 82 angiogenesis related genes were measured by quantitative real-time polymerase chain reaction using TaqMan low-density arrays. A 34-gene-profile which was able to predict the overall survival of ovarian carcinoma patients was identified. After a leave-one-out cross validation, the profile distinguished two groups of patients with different outcomes. Median overall survival and progression-free survival for the high risk group was 28.3 and 15.0 months, respectively, and was not reached by patients in the low risk group at the end of follow-up. Moreover, the profile maintained an independent prognostic value in the multivariate analysis. The hazard ratio for death was 2.3 (95% CI, 1.5 to 3.2; p<0.001). CONCLUSIONS/SIGNIFICANCE It is possible to generate a prognostic model for advanced ovarian carcinoma based on angiogenesis-related genes using formalin-fixed paraffin-embedded samples. The present results are consistent with the increasing weight of angiogenesis genes in the prognosis of ovarian carcinoma.

MicroRNA expression profiling in Imatinib-resistant Chronic Myeloid Leukemia patients without clinically significant ABL1-mutations.

The development of Imatinib Mesylate (IM), the first specific inhibitor of BCR-ABL1, has had a major impact in patients with Chronic Myeloid Leukemia (CML), establishing IM as the standard therapy for CML. Despite the clinical success obtained with the use of IM, primary resistance to IM and molecular evidence of persistent disease has been observed in 20-25% of IM treated patients. The existence of second generation TK inhibitors, which are effective in patients with IM resistance, makes identification of predictors of resistance to IM an important goal in CML. In this study, we have identified a group of 19 miRNAs that may predict clinical resistance to IM in patients with newly diagnosed CML.

Expression profiling of rectal tumors defines response to neoadjuvant treatment related genes.

To date, no effective method exists that predicts the response to preoperative chemoradiation (CRT) in locally advanced rectal cancer (LARC). Nevertheless, identification of patients who have a higher likelihood of responding to preoperative CRT could be crucial in decreasing treatment morbidity and avoiding expensive and time-consuming treatments. The aim of this study was to identify signatures or molecular markers related to response to pre-operative CRT in LARC. We analyzed the gene expression profiles of 26 pre-treatment biopsies of LARC (10 responders and 16 non-responders) without metastasis using Human WG CodeLink microarray platform. Two hundred and fifty seven genes were differentially over-expressed in the responder patient subgroup. Ingenuity Pathway Analysis revealed a significant ratio of differentially expressed genes related to cancer, cellular growth and proliferation pathways, and c-Myc network. We demonstrated that high Gng4, c-Myc, Pola1, and Rrm1 mRNA expression levels was a significant prognostic factor for response to treatment in LARC patients (p<0.05). Using this gene set, we were able to establish a new model for predicting the response to CRT in rectal cancer with a sensitivity of 60% and 100% specificity. Our results reflect the value of gene expression profiling to gain insight about the molecular pathways involved in the response to treatment of LARC patients. These findings could be clinically relevant and support the use of mRNA levels when aiming to identify patients who respond to CRT therapy.

Cross-species and cross-platform gene expression studies with the Bioconductor-compliant R package 'annotationTools'.

Background: The variety of DNA microarray formats and datasets presently available offers an unprecedented opportunity to perform insightful comparisons of heterogeneous data. Cross-species studies, in particular, have the power of identifying conserved, functionally important molecular processes. Validation of discoveries can now often be performed in readily available public data which frequently requires cross-platform studies.Cross-platform and cross-species analyses require matching probes on different microarray formats. This can be achieved using the information in microarray annotations and additional molecular biology databases, such as orthology databases. Although annotations and other biological information are stored using modern database models ( e. g. relational), they are very often distributed and shared as tables in text files, i.e. flat file databases. This common flat database format thus provides a simple and robust solution to flexibly integrate various sources of information and a basis for the combined analysis of heterogeneous gene expression profiles.Results: We provide annotationTools, a Bioconductor-compliant R package to annotate microarray experiments and integrate heterogeneous gene expression profiles using annotation and other molecular biology information available as flat file databases. First, annotationTools contains a specialized set of functions for mining this widely used database format in a systematic manner. It thus offers a straightforward solution for annotating microarray experiments. Second, building on these basic functions and relying on the combination of information from several databases, it provides tools to easily perform cross-species analyses of gene expression data.Here, we present two example applications of annotationTools that are of direct relevance for the analysis of heterogeneous gene expression profiles, namely a cross-platform mapping of probes and a cross-species mapping of orthologous probes using different orthology databases. We also show how to perform an explorative comparison of disease-related transcriptional changes in human patients and in a genetic mouse model.Conclusion: The R package annotationTools provides a simple solution to handle microarray annotation and orthology tables, as well as other flat molecular biology databases. Thereby, it allows easy integration and analysis of heterogeneous microarray experiments across different technological platforms or species.

Genome-wide analysis of cancer/testis gene expression.

Cancer/Testis (CT) genes, normally expressed in germ line cells but also activated in a wide range of cancer types, often encode antigens that are immunogenic in cancer patients, and present potential for use as biomarkers and targets for immunotherapy. Using multiple in silico gene expression analysis technologies, including twice the number of expressed sequence tags used in previous studies, we have performed a comprehensive genome-wide survey of expression for a set of 153 previously described CT genes in normal and cancer expression libraries. We find that although they are generally highly expressed in testis, these genes exhibit heterogeneous gene expression profiles, allowing their classification into testis-restricted (39), testis/brain-restricted (14), and a testis-selective (85) group of genes that show additional expression in somatic tissues. The chromosomal distribution of these genes confirmed the previously observed dominance of X chromosome location, with CT-X genes being significantly more testis-restricted than non-X CT. Applying this core classification in a genome-wide survey we identified >30 CT candidate genes; 3 of them, PEPP-2, OTOA, and AKAP4, were confirmed as testis-restricted or testis-selective using RT-PCR, with variable expression frequencies observed in a panel of cancer cell lines. Our classification provides an objective ranking for potential CT genes, which is useful in guiding further identification and characterization of these potentially important diagnostic and therapeutic targets.

The evolution of gene expression levels in mammalian organs.

Changes in gene expression are thought to underlie many of the phenotypic differences between species. However, large-scale analyses of gene expression evolution were until recently prevented by technological limitations. Here we report the sequencing of polyadenylated RNA from six organs across ten species that represent all major mammalian lineages (placentals, marsupials and monotremes) and birds (the evolutionary outgroup), with the goal of understanding the dynamics of mammalian transcriptome evolution. We show that the rate of gene expression evolution varies among organs, lineages and chromosomes, owing to differences in selective pressures: transcriptome change was slow in nervous tissues and rapid in testes, slower in rodents than in apes and monotremes, and rapid for the X chromosome right after its formation. Although gene expression evolution in mammals was strongly shaped by purifying selection, we identify numerous potentially selectively driven expression switches, which occurred at different rates across lineages and tissues and which probably contributed to the specific organ biology of various mammals.

A gene expression signature that distinguishes desmoid tumours from nodular fasciitis.

Nodular fasciitis (NF) is a rapidly growing cellular mass composed of fibroblasts/myofibroblasts, usually localized in subcutaneous tissues, that typically undergoes fibrosis and almost never recurs. Desmoid tumours (DTs) are rare forms of fibroblastic/myofibroblastic growth that arise in deep soft tissues, display a propensity for local infiltration and recurrence, but fail to metastasize. Given that both entities are primarily fibroblastic/myofibroblastic lesions with overlapping histological features, their gene expression profiles were compared to identify differentially expressed genes that may provide not only potential diagnostic markers, but also clues as to the pathogenesis of each disorder. Differentially expressed transcripts (89 clones displaying increased expression in DTs and 246 clones displaying increased expression in NF) included genes encoding several receptor and non-receptor tyrosine kinases (EPHB3, PTPRF, GNAZ, SYK, LYN, EPHA4, BIRC3), transcription factors (TWIST1, PITX2, EYA2, OAS1, MITF, TCF20), and members of the Wnt signalling pathway (AXIN2, WISP1, SFRP). Remarkably, almost one-quarter of the differentially expressed genes encode proteins associated with inflammation and tissue remodelling, including members of the interferon (IFN), tumour necrosis factor (TNF), and transforming growth factor beta (TGF-beta) signalling pathways as well as metalloproteinases (MMP1, 9, 13, 23), urokinase plasminogen activator (PLAU), and cathepsins. The observations provide the first comparative molecular characterization of desmoid tumours and nodular fasciitis and suggest that selected tyrosine kinases, transcription factors, and members of the Wnt, TGF-beta, IFN, and TNF signalling pathways may be implicated in influencing and distinguishing their fate.

eVOC: a controlled vocabulary for unifying gene expression data.

Expression data contribute significantly to the biological value of the sequenced human genome, providing extensive information about gene structure and the pattern of gene expression. ESTs, together with SAGE libraries and microarray experiment information, provide a broad and rich view of the transcriptome. However, it is difficult to perform large-scale expression mining of the data generated by these diverse experimental approaches. Not only is the data stored in disparate locations, but there is frequent ambiguity in the meaning of terms used to describe the source of the material used in the experiment. Untangling semantic differences between the data provided by different resources is therefore largely reliant on the domain knowledge of a human expert. We present here eVOC, a system which associates labelled target cDNAs for microarray experiments, or cDNA libraries and their associated transcripts with controlled terms in a set of hierarchical vocabularies. eVOC consists of four orthogonal controlled vocabularies suitable for describing the domains of human gene expression data including Anatomical System, Cell Type, Pathology and Developmental Stage. We have curated and annotated 7016 cDNA libraries represented in dbEST, as well as 104 SAGE libraries,with expression information,and provide this as an integrated, public resource that allows the linking of transcripts and libraries with expression terms. Both the vocabularies and the vocabulary-annotated libraries can be retrieved from http://www.sanbi.ac.za/evoc/. Several groups are involved in developing this resource with the aim of unifying transcript expression information.

Reactive electrophile species activate defense gene expression in Arabidopsis.

Compounds containing alpha,beta-unsaturated carbonyl groups are increasingly implicated as potent regulators of gene expression; some are powerful cytotoxins known to accumulate at the site of lesion formation in host-pathogen interactions. We used a robust measurement of photosynthetic efficiency to quantify the toxicity of a variety of lipid derivatives in Arabidopsis leaves. Small alpha,beta-unsaturated carbonyl compounds (e.g. acrolein and methyl vinyl ketone) were highly active and proved to be potent stimulators of expression of the pathogenesis-related gene HEL (PR4). These small volatile electrophiles were far more active than larger alkenal homologs like 2(E)-hexenal, and activated HEL expression in a manner independent of salicylate, ethylene, and jasmonate production/perception. Electrophile treatment massively increased the levels of unesterified cyclopentenone jasmonates, which themselves are electrophiles. Patterns of gene expression in response to electrophile treatment and in response to avirulent bacteria were compared, which revealed strikingly similar transcript profiles. The results broaden the range of known biologic effects of reactive electrophile species to include the activation of a pathogenesis-related gene (HEL) and genes involved in metabolism. Electrophiles can act as mediators of both genetic and biochemical effects on core defense signal transduction.

Regional and cellular gene expression changes in human Huntington's disease brain.

Huntington's disease (HD) pathology is well understood at a histological level but a comprehensive molecular analysis of the effect of the disease in the human brain has not previously been available. To elucidate the molecular phenotype of HD on a genome-wide scale, we compared mRNA profiles from 44 human HD brains with those from 36 unaffected controls using microarray analysis. Four brain regions were analyzed: caudate nucleus, cerebellum, prefrontal association cortex [Brodmann's area 9 (BA9)] and motor cortex [Brodmann's area 4 (BA4)]. The greatest number and magnitude of differentially expressed mRNAs were detected in the caudate nucleus, followed by motor cortex, then cerebellum. Thus, the molecular phenotype of HD generally parallels established neuropathology. Surprisingly, no mRNA changes were detected in prefrontal association cortex, thereby revealing subtleties of pathology not previously disclosed by histological methods. To establish that the observed changes were not simply the result of cell loss, we examined mRNA levels in laser-capture microdissected neurons from Grade 1 HD caudate compared to control. These analyses confirmed changes in expression seen in tissue homogenates; we thus conclude that mRNA changes are not attributable to cell loss alone. These data from bona fide HD brains comprise an important reference for hypotheses related to HD and other neurodegenerative diseases.

Meta-analysis of gene expression profiles in breast cancer: toward a unified understanding of breast cancer subtyping and prognosis signatures.

INTRODUCTION: Breast cancer subtyping and prognosis have been studied extensively by gene expression profiling, resulting in disparate signatures with little overlap in their constituent genes. Although a previous study demonstrated a prognostic concordance among gene expression signatures, it was limited to only one dataset and did not fully elucidate how the different genes were related to one another nor did it examine the contribution of well-known biological processes of breast cancer tumorigenesis to their prognostic performance. METHOD: To address the above issues and to further validate these initial findings, we performed the largest meta-analysis of publicly available breast cancer gene expression and clinical data, which are comprised of 2,833 breast tumors. Gene coexpression modules of three key biological processes in breast cancer (namely, proliferation, estrogen receptor [ER], and HER2 signaling) were used to dissect the role of constituent genes of nine prognostic signatures. RESULTS: Using a meta-analytical approach, we consolidated the signatures associated with ER signaling, ERBB2 amplification, and proliferation. Previously published expression-based nomenclature of breast cancer 'intrinsic' subtypes can be mapped to the three modules, namely, the ER-/HER2- (basal-like), the HER2+ (HER2-like), and the low- and high-proliferation ER+/HER2- subtypes (luminal A and B). We showed that all nine prognostic signatures exhibited a similar prognostic performance in the entire dataset. Their prognostic abilities are due mostly to the detection of proliferation activity. Although ER- status (basal-like) and ERBB2+ expression status correspond to bad outcome, they seem to act through elevated expression of proliferation genes and thus contain only indirect information about prognosis. Clinical variables measuring the extent of tumor progression, such as tumor size and nodal status, still add independent prognostic information to proliferation genes. CONCLUSION: This meta-analysis unifies various results of previous gene expression studies in breast cancer. It reveals connections between traditional prognostic factors, expression-based subtyping, and prognostic signatures, highlighting the important role of proliferation in breast cancer prognosis.