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PubMed2005

Analysis of 101 nuclear transcriptomes reveals 23 distinct regulons and their relationship to metabolism, chromosomal gene distribution and co-ordination of nuclear and plastid gene expression.

Post-endosymbiotic evolution of the proto-chloroplast was characterized by gene transfer to the nucleus. Hence, most chloroplast proteins are nuclear-encoded and the regulation of chloroplast functions includes nuclear transcriptional control. The expression profiles of 3292 nuclear Arabidopsis genes, most of them encoding chloroplast proteins, were determined from 101 different conditions and have been deposited at the GEO database (http://www.ncbi.nih.gov/geo/) under . The 1590 most-regulated genes fell into 23 distinct groups of co-regulated genes (regulons). Genes of some regulons are not evenly distributed among the five Arabidopsis chromosomes and pairs of adjacent, co-expressed genes exist. Except regulons 1 and 2, regulons are heterogeneous and consist of genes coding for proteins with different subcellular locations or contributing to several biochemical functions. This implies that different organelles and/or metabolic pathways are co-ordinated at the nuclear transcriptional level, and a prototype for this is regulon 12 which contains genes with functions in amino acid and carbohydrate metabolism, as well as genes associated with transport or transcription. The co-expression of nuclear genes coding for subunits of the photosystems or encoding proteins involved in the transcription/translation of plastome genes (particularly ribosome polypeptides) (regulons 1 and 2, respectively) implies the existence of a novel mechanism that co-ordinates plastid and nuclear gene expression and involves nuclear control of plastid ribosome abundance. The co-regulation of genes for photosystem and plastid ribosome proteins escapes a previously described general control of nuclear chloroplast proteins imposed by a transcriptional master switch, highlighting a mode of transcriptional regulation of photosynthesis which is different compared to other chloroplast functions. From the evolutionary standpoint, the results provided indicate that functional integration of the proto-chloroplast into the eukaryotic cell was associated with the establishment of different layers of nuclear transcriptional control.

Arabidopsis↗
PubMed2005

Insights on the evolution of metabolic networks of unicellular translationally biased organisms from transcriptomic data and sequence analysis.

Codon bias is related to metabolic functions in translationally biased organisms, and two facts are argued about. First, genes with high codon bias describe in meaningful ways the metabolic characteristics of the organism; important metabolic pathways corresponding to crucial characteristics of the lifestyle of an organism, such as photosynthesis, nitrification, anaerobic versus aerobic respiration, sulfate reduction, methanogenesis, and others, happen to involve especially biased genes. Second, gene transcriptional levels of sets of experiments representing a significant variation of biological conditions strikingly confirm, in the case of Saccharomyces cerevisiae, that metabolic preferences are detectable by purely statistical analysis: the high metabolic activity of yeast during fermentation is encoded in the high bias of enzymes involved in the associated pathways, suggesting that this genome was affected by a strong evolutionary pressure that favored a predominantly fermentative metabolism of yeast in the wild. The ensemble of metabolic pathways involving enzymes with high codon bias is rather well defined and remains consistent across many species, even those that have not been considered as translationally biased, such as Helicobacter pylori, for instance, reveal some weak form of translational bias for this genome. We provide numerical evidence, supported by experimental data, of these facts and conclude that the metabolic networks of translationally biased genomes, observable today as projections of eons of evolutionary pressure, can be analyzed numerically and predictions of the role of specific pathways during evolution can be derived. The new concepts of Comparative Pathway Index, used to compare organisms with respect to their metabolic networks, and Evolutionary Pathway Index, used to detect evolutionarily meaningful bias in the genetic code from transcriptional data, are introduced.

Ammonia↗
PubMed2006

The heterogeneity of human mesenchymal stem cell preparations--evidence from simultaneous analysis of proteomes and transcriptomes.

OBJECTIVE: Mesenchymal stem cells (MSC) raise high hopes in clinical applications. However, the lack of common standards and a precise definition of MSC preparations remains a major obstacle in research and application of MSC. Whereas surface antigen markers have failed to precisely define this population, a combination of proteomic data and microarray data provides a new dimension for the definition of MSC preparations. METHODS: In our continuing effort to characterize MSC, we have analyzed the differential transcriptome and proteome expression profiles of MSC preparations isolated from human bone marrow under two different expansion media (BM-MSC-M1 and BM-MSC-M2). RESULTS: In proteomics, 136 protein spots were unambiguously identified by MALDI-TOF-MS and corresponding cDNA spots were selected on our "Human Transcriptome cDNA Microarray." Combination of datasets revealed a correlation in differential gene expression and protein expression of BM-MSC-M1 vs BM-MSC-M2. Genes involved in metabolism were more highly expressed in BM-MSC-M1, whereas genes involved in development, morphogenesis, extracellular matrix, and differentiation were more highly expressed in BM-MSC-M2. Interchanging culture conditions for 8 days revealed that differential expression was retained in several genes whereas it was altered in others. CONCLUSION: Our results have provided evidence that homogeneous BM-MSC preparations can reproducibly be isolated under standardized conditions, whereas culture conditions exert a prominent impact on transcriptome, proteome, and cellular organization of BM-MSC.

Cell Culture Techniques↗
PubMed2002

Analysis of the mouse transcriptome based on functional annotation of 60,770 full-length cDNAs.

Only a small proportion of the mouse genome is transcribed into mature messenger RNA transcripts. There is an international collaborative effort to identify all full-length mRNA transcripts from the mouse, and to ensure that each is represented in a physical collection of clones. Here we report the manual annotation of 60,770 full-length mouse complementary DNA sequences. These are clustered into 33,409 'transcriptional units', contributing 90.1% of a newly established mouse transcriptome database. Of these transcriptional units, 4,258 are new protein-coding and 11,665 are new non-coding messages, indicating that non-coding RNA is a major component of the transcriptome. 41% of all transcriptional units showed evidence of alternative splicing. In protein-coding transcripts, 79% of splice variations altered the protein product. Whole-transcriptome analyses resulted in the identification of 2,431 sense-antisense pairs. The present work, completely supported by physical clones, provides the most comprehensive survey of a mammalian transcriptome so far, and is a valuable resource for functional genomics.

Alternative Splicing↗
PubMed2003

Analysis of the mouse transcriptome for genes involved in the function of the nervous system.

We analyzed the mouse Representative Transcript and Protein Set for molecules involved in brain function. We found full-length cDNAs of many known brain genes and discovered new members of known brain gene families, including Family 3 G-protein coupled receptors, voltage-gated channels, and connexins. We also identified previously unknown candidates for secreted neuroactive molecules. The existence of a large number of unique brain ESTs suggests an additional molecular complexity that remains to be explored.A list of genes containing CAG stretches in the coding region represents a first step in the potential identification of candidates for hereditary neurological disorders.

Adenine↗
PubMed2003

Negative feedback regulation of dnaK, clpB and lon expression by the DnaK chaperone machine in Streptomyces coelicolor, identified by transcriptome and in vivo DnaK-depletion analysis.

The dnaK operon of Streptomyces coelicolor encodes the DnaK chaperone machine and the negative autoregulator HspR, which confers repression of the operon by binding to several inverted repeat sequences in the promoter region, dnaKp. Previous in vitro studies demonstrated that DnaK forms a specific complex with HspR bound to its operator sequences in dnaKp, and a model was proposed in which DnaK functions as a corepressor of the dnaK operon (Bucca, G., Brassington, A., Schonfeld, H.J., and Smith, C.P. (2000) Mol Microbiol 38: 1093-1103). Here we report in vivo DnaK depletion experiments which demonstrate that DnaK is a negative regulator of the dnaK operon. Cellular depletion of the DnaK chaperone leads to high-level transcription from dnaKp at the normal growth temperature. DNA microarray-based analysis of gene expression in wild-type and hspR-disruption mutant strains has identified a core cluster of genes regulated by HspR: the dnaK and clpB-SCO3660 operons and lon. These three transcription units are considered to be the direct targets of HspR. Significantly, analysis of the entire genome sequence revealed that the promoter regions of dnaK, clpB and lon are the only sequences that contain the HspR consensus binding sequence 5'-TTGAGY-N7-ACTCAA. S1 nuclease mapping confirmed that transcription of both clpB and lon is substantially enhanced at ambient temperature in strains depleted of DnaK, providing further evidence that these genes are members of the DnaK-HspR regulon. From transcriptome analysis, 17 genes were shown to be upregulated more than twofold in an hspR disruption mutant. This included the seven genes encoded by the dnaK, clpB and lon transcription units. Significantly, the other 10 genes are not heat-shock inducible in the wild type and their upregulation in the hspR mutant is considered to be an indirect consequence of enhanced synthesis of one or more components of the HspR regulon (the DnaK chaperone machine, ClpB and Lon protease).

Adenosine Triphosphatases↗
PubMed2005

Profiles of metabolites and gene expression in rats with chemically induced hepatic necrosis.

This study investigated whether integrated analysis of transcriptomics and metabolomics data increased the sensitivity of detection and provided new insight in the mechanisms of hepatotoxicity. Metabolite levels in plasma or urine were analyzed in relation to changes in hepatic gene expression in rats that received bromobenzene to induce acute hepatic centrilobular necrosis. Bromobenzene-induced lesions were only observed after treatment with the highest of 3 dose levels. Multivariate statistical analysis showed that metabolite profiles of blood plasma were largely different from controls when the rats were treated with bromobenzene, also at doses that did not elicit histopathological changes. Changes in levels of genes and metabolites were related to the degree of necrosis, providing putative novel markers of hepatotoxicity. Levels of endogenous metabolites like alanine, lactate, tyrosine and dimethylglycine differed in plasma from treated and control rats. The metabolite profiles of urine were found to be reflective of the exposure levels. This integrated analysis of hepatic transcriptomics and plasma metabolomics was able to more sensitively detect changes related to hepatotoxicity and discover novel markers. The relation between gene expression and metabolite levels was explored and additional insight in the role of various biological pathways in bromobenzene-induced hepatic necrosis was obtained, including the involvement of apoptosis and changes in glycolysis and amino acid metabolism. The complete Table 2 is available as a supplemental file online at http://taylorandfrancis.metapress.com/openurlasp?genre=journal&issn=0192-6233. To access the file, click on the issue link for 33(4), then select this article. A download option appears at the bottom of this abstract. In order to access the full article online, you must either have an individual subscription or a member subscription accessed through www.toxpath.org.

Amino Acids↗
PubMed2026

Multi-omics integration and colocalization analyses prioritize candidate molecular loci associated with hypothermia.

BACKGROUND: Hypothermia is a life-threatening condition lacking specific pharmacological treatments. This study aimed to prioritize genetically supported molecular loci associated with hypothermia and to explore their pharmacological tractability using multi-omics data. METHODS: Initially, 2532 druggable genes were curated from the Drug-Gene Interaction Database and established literature. These were cross-referenced with cis-eQTL and cis-pQTL datasets, encompassing 870,655 and 114,281 SNPs for blood, respectively, alongside 2379 shared SNPs across adipose, skeletal muscle, and heart tissues. Matched instrumental variables were integrated with hypothermia GWAS summary statistics for two-sample Mendelian randomization (MR) and Bayesian colocalization. Transcriptomic differential expression analysis (DEA) was subsequently conducted as an exploratory analysis of cold-exposure-associated expression changes. Database-derived compound annotations were systematically re-evaluated according to target specificity, established pharmacological mechanism, and concordance with the direction of the MR estimates. RESULTS: Among 671 gene-level MR tests, 36 genes reached nominal significance, whereas only ABCC8 remained significant after FDR correction. Colocalization was evaluable for 8 of these 36 genes, and 4 loci (COL18A1, SLC1A7, ADIPOQ, and MERTK) met the prespecified PP.H4>0.90 threshold. The remaining 28 loci were not evaluable because sufficient overlapping regional variants were unavailable after harmonization. Transcriptomic analysis identified altered expression of SLC1A3 and SLCO4A1 under cold exposure, although these findings did not directly validate the colocalization-supported loci. Re-evaluation of database-derived compound annotations did not identify any direct, selective, and directionally concordant drug-repurposing candidate for hypothermia. CONCLUSIONS: COL18A1, SLC1A7, ADIPOQ, and MERTK showed colocalization support among the 8 evaluable nominal MR-associated loci. Because colocalization coverage was limited, these genes should be regarded as preliminary candidate loci rather than established therapeutic targets. The pharmacological annotations were indirect, non-selective, unsupported, or directionally inconsistent and should be interpreted solely as hypothesis-generating information.

Bayesian colocalization↗
PubMed2004

Different gene expression profiles between microsatellite instability-high and microsatellite stable colorectal carcinomas.

Recent molecular genetic studies have revealed that two major types of genomic instabilities, chromosomal instability (CIN) and microsatellite instability (MSI), exist in colorectal carcinomas. In order to clarify the molecular signature related to the CIN and MSI in colorectal carcinomas, we performed transcriptomic expression analysis on eight microsatellite instability-high (MSI-H) colorectal carcinomas and compared the results obtained with that of nine microsatellite stable (MSS) colorectal carcinomas using oligonucleotide microarrays containing 17 334 known genes and 1331 unknown genes or expression sequence tags (ESTs). Unsupervised two-way hierarchical clustering with 5724 genes successfully classified tumors from normal mucosa, and displayed a distinctive MSI-H carcinomas subgroup. Based on intensive filtering, 57 known genes and eight ESTs were found to be highly relevant to the differentiation of MSI-H and MSS colorectal carcinomas. These genes successfully distinguish the new test set of six MSI-H and five MSS colorectal carcinomas. Many up- and downregulated genes in MSI-H colorectal carcinomas were related to the previously reported phenotypic characteristics; increased mucin production and intense peritumoral immune response in MSI-H carcinomas. Some of these differences were confirmed by semiquantitative reverse transcription-PCR and immunohistochemical analysis. Our findings indicate that there are many different genetic and transcriptomic characteristics between MSI-H and MSS colorectal carcinomas, and some of these differently expressed genes can be used as diagnostic or prognostic markers.

Base Sequence↗
PubMed2006

VAMP: visualization and analysis of array-CGH, transcriptome and other molecular profiles.

MOTIVATION: Microarray-based CGH (Comparative Genomic Hybridization), transcriptome arrays and other large-scale genomic technologies are now routinely used to generate a vast amount of genomic profiles. Exploratory analysis of this data is crucial in helping to understand the data and to help form biological hypotheses. This step requires visualization of the data in a meaningful way to visualize the results and to perform first level analyses. RESULTS: We have developed a graphical user interface for visualization and first level analysis of molecular profiles. It is currently in use at the Institut Curie for cancer research projects involving CGH arrays, transcriptome arrays, SNP (single nucleotide polymorphism) arrays, loss of heterozygosity results (LOH), and Chromatin ImmunoPrecipitation arrays (ChIP chips). The interface offers the possibility of studying these different types of information in a consistent way. Several views are proposed, such as the classical CGH karyotype view or genome-wide multi-tumor comparison. Many functionalities for analyzing CGH data are provided by the interface, including looking for recurrent regions of alterations, confrontation to transcriptome data or clinical information, and clustering. Our tool consists of PHP scripts and of an applet written in Java. It can be run on public datasets at http://bioinfo.curie.fr/vamp AVAILABILITY: The VAMP software (Visualization and Analysis of array-CGH,transcriptome and other Molecular Profiles) is available upon request. It can be tested on public datasets at http://bioinfo.curie.fr/vamp. The documentation is available at http://bioinfo.curie.fr/vamp/doc.

Algorithms↗
PubMed2025

Stereo-cell: Spatial enhanced-resolution single-cell sequencing with high-density DNA nanoball-patterned arrays.

Single-cell sequencing technologies have advanced our understanding of cellular heterogeneity and biological complexity. However, existing methods face limitations in throughput, capture uniformity, cell size flexibility, and technical extensibility. We present Stereo-cell, a spatial enhanced-resolution single-cell sequencing platform based on high-density DNA nanoball (DNB)-patterned arrays, which enables scalable and unbiased cell capture at a wide input range and supports high-fidelity transcriptome profiling. Stereo-cell further allows integration with imaging-based modalities and multiomics strategies, including immunofluorescence and epitope profiling. This platform is also compatible with profiling extracellular vesicles, microstructures, and large cells, whereas its spatial resolution facilitates in situ analysis of cell-cell interactions, cellular microenvironments, and subcellular transcript localization. Together, Stereo-cell provides a flexible framework for expanding single-cell research applications.

Animals↗
PubMed2026

Single-cell and spatial transcriptomics define a progenitor subpopulation and fibroinflammatory niche at the leading edge of parathyroid carcinoma.

Parathyroid carcinoma (PC) is a rare but clinically aggressive endocrine malignancy with limited treatment options and a poorly defined tumor microenvironment (TME). To elucidate its cellular heterogeneity and spatial architecture, we integrated single-cell and spatial transcriptomic profiling with whole-exome sequencing and multiplex immunohistochemistry on eight parathyroid neoplasm specimens, including PC, parathyroid adenoma, and atypical parathyroid tumor. We identified a distinct progenitor-like endocrine subpopulation (Ca-1) enriched in CDC73-mutant PC, exhibiting stem-like properties, elevated cell cycle activity, and pronounced genomic instability. Spatial mapping revealed that Ca-1 cells preferentially localize at the leading edge, forming a fibroinflammatory niche characterized by the enrichment of inflammatory cancer-associated fibroblasts (iCAFs) and SPP1+ macrophages. Within this niche, the dipeptidyl peptidase 4 (DPP4) is selectively expressed in Ca-1 cells and iCAFs, implicating a potential paracrine axis driving stromal remodeling and immunosuppression. These findings suggest that a spatially organized ecosystem may promote PC progression through TME remodeling and highlight the DPP4-CXCL2 axis as a candidate pathway for future investigation in aggressive parathyroid neoplasms.

Humans↗
PubMed2004

Intregrated analysis of the human cardiac transcriptome, proteome and phosphoproteome.

Altered expression of different classes of genes has been shown to differentiate between failing and nonfailing human hearts. However, characterization of proteins and the post-translational modifications that regulate their functions is required for understanding both the physiology of cardiac muscle and the mechanisms leading to pathological states associated with cardiac diseases. We present in this paper, an analysis of the human cardiac transcriptome, proteome and phosphoproteome. Data from two sources (i) experiments performed in our laboratory and (ii) bioinformatics searches of public databases (SWISS-PROT, NCBI, Cardiac Gene Expression Knowledge Base, Gene Ontology Consortium and Affymetrix) are reported in a relational database that allows user-designed specific queries. Microarray experiments were performed with Affymetrix Hu95Av2. Cardiac proteins were digested with trypsin. An 11 step cation exchange procedure produced fractions for analysis in separate reversed phase high-performance liquid chromatography-tandem mass spectrometry (MS/MS) experiments. Immobilized metal affinity chromatography was used to select the phosphopeptides from the same tryptic peptide mixture. They were then further investigated by MS/MS. Gel-free approaches were used to detect 267 proteins and 47 phosphopeptides. Our human cardiac database contains 447 entries. We propose the use of this platform, built with data derived from nonfailing hearts, as a template for initiating the effort to characterize the human cardiac proteome and its associated post-translational modifications.

Chromatography, Affinity↗
PubMed2003

[Therapeutic gene clusters as drug action mechanisms].

Pharmacogenomics is defined to identify the therapeutic gene clusters which are involved in determining the responsiveness and to distinguish responders and non-responders to a given drug. Genome sequencing, transcriptome, proteome and metabolome analysis are of particular significance in pharmacogenomics. Sequencing is used to locate polymorphisms, and monitoring of functional gene expression can provide clue about the genomic response to disease and treatment. The transcriptome analysis can be done by methods of random cDNA sequencing, mRNA display(fluorescent differential display et al.) and differential hybridization(cDNA microarray, oligomicrochip et al.). We used transcriptome/proteome/metabolome analysis to identify therapeutic target genes by studying change of gene expression in animal models and human model cells of various diseases and found novel drug target candidates through this pharmacogenomic strategy. The present study describes combined transcriptome and metabolome analysis for therapeutic target validation in hypoxia-induced vascular remodeling. The pharmacogenomic analysis and pharmainformatics has the potential for strategy to define novel therapeutic gene clusters in various diseases and holds the promise that drugs might be tailor-maid for individuals and adapted to each person's own genetic makeup. These techniques provided an excellent strategy for screening and validation of targets.

Animals↗
PubMed2006

Impact of intestinal colonization and invasion on the Entamoeba histolytica transcriptome.

A genome-wide transcriptional analysis of Entamoeba histolytica was performed on trophozoites isolated from the colon of six infected mice and from in vitro culture. An Affymetrix platform gene expression array was designed for this analysis that included probe sets for 9435 open reading frames (ORFs) and 9066 5' and 3' flanking regions. Transcripts were detected for > 80% of all ORFs. A total of 523 transcripts (5.2% of all E. histolytica genes) were significantly changed in amebae isolated from the intestine on Days 1 and 29 after infection: 326 and 109 solely on Days 1 and 29, and 88 on both days. Quantitative real-time reverse transcriptase PCR confirmed these changes in 11/12 genes tested using mRNA isolated from an additional six mice. Adaptation to the intestinal environment was accompanied by increases in a subset of cell signaling genes including transmembrane kinases, ras and rho family GTPases, and calcium binding proteins. Significant decreases in mRNA abundance for genes involved in glycolysis and concomitant increases in lipases were consistent with a change in energy metabolism. Defense against bacteria present in the intestine (but lacking from in vitro culture) was suggested by alterations in mRNA levels of genes similar to the AIG1 plant antibacterial proteins. Decreases in oxygen detoxification pathways were observed as expected in the anaerobic colonic lumen. Of the known virulence factors the most remarkable changes were a 20-35-fold increase in a cysteine proteinase four-like gene, and a 2-3-fold decrease in two members of the Gal/GalNAc lectin light subunit family. Control of the observed changes in mRNA abundance in the intestine might potentially rest with four related proteins with DNA binding domains that were down-regulated 6-16-fold in the intestinal environment. In conclusion, the first genome-wide analysis of the transcriptome of E. histolytica demonstrated that the vast majority of genes are transcribed in trophozoites, and that in the host intestine trophozoites altered the expression of mRNAs for genes implicated in metabolism, oxygen defense, cell signaling, virulence, antibacterial activity, and DNA binding.

Animals↗
PubMed2006

Prognostic testing in uveal melanoma by transcriptomic profiling of fine needle biopsy specimens.

Many uveal melanoma patients die of metastasis despite ocular treatment. Transcriptomic profiling of enucleated tumors can identify patients at high metastatic risk. Because most uveal melanomas do not require enucleation, a biopsy would be required for this analysis. Here, we establish the feasibility of transcriptomic analysis of uveal melanomas from fine needle aspirates. Transcriptomic profiles were analyzed from postenucleation "mock" needle biopsies and matching tumors from eight enucleated eyes and from fine needle aspirates in 17 uveal melanomas before radiotherapy. Predictive accuracy was assessed using a weighted voting classifier optimized for probe set selection using a minimal redundancy/maximum relevance algorithm. Transcriptomic profiles from mock biopsies were highly similar to those from their matching tumor samples (P < 0.0001). Transcriptomic profiles from fine needle aspirates clustered into two classes with discriminating probe sets that overlapped significantly with those for our published classification (P < 0.00001). No loss of predictive accuracy was identified among eight needle aspirates obtained from a distant location. Thus, it is feasible to obtain RNA of adequate quality and quantity to perform transcriptomic analysis on uveal melanoma samples obtained by fine needle biopsy. This method can be applied to specimens obtained from distant geographic locations and can stratify uveal melanoma patients based on metastatic risk.

Adult↗
PubMed2004

Incidence of "quasi-ditags" in catalogs generated by Serial Analysis of Gene Expression (SAGE).

BACKGROUND: Serial Analysis of Gene Expression (SAGE) is a functional genomic technique that quantitatively analyzes the cellular transcriptome. The analysis of SAGE libraries relies on the identification of ditags from sequencing files; however, the software used to examine SAGE libraries cannot distinguish between authentic versus false ditags ("quasi-ditags"). RESULTS: We provide examples of quasi-ditags that originate from cloning and sequencing artifacts (i.e. genomic contamination or random combinations of nucleotides) that are included in SAGE libraries. We have employed a mathematical model to predict the frequency of quasi-ditags in random nucleotide sequences, and our data show that clones containing less than or equal to 2 ditags (which include chromosomal cloning artifacts) should be excluded from the analysis of SAGE catalogs. CONCLUSIONS: Cloning and sequencing artifacts contaminating SAGE libraries could be eliminated using simple pre-screening procedure to increase the reliability of the data.

Animals↗