Global functional profiling of gene expression

The typical result of a microarray experiment is a list of tens or hundreds of genes found to be differentially regulated in the condition under study. Independent of the methods used to select these genes, the common task faced by any researcher is to translate these lists of genes into a better understanding of the biological phenomena involved. Currently, this is done through a tedious combination of searches through the literature and a number of public databases. We developed Onto-Express (OE) as a novel tool able to automatically translate such lists of differentially regulated genes into functional profiles characterizing the impact of the condition studied. OE constructs functional profiles (using Gene Ontology terms) for the following categories: biochemical function, biological process, cellular role, cellular component, molecular function, and chromosome location. Statistical significance values are calculated for each category. We demonstrate the validity and the utility of this comprehensive global analysis of gene function by analyzing two breast cancer datasets from two separate laboratories. OE was able to identify correctly all biological processes postulated by the original authors, as well as discover novel relevant mechanisms.     

Important processes during differentiation and early development of somatic embryos of Norway spruce as revealed by changes in global gene expression

The aim of this study has been to identify important processes that regulate early stages of embryo development in conifers. Somatic embryogenesis in Picea abies has become a model system for studying embryology in conifers, providing a well-characterized sequence of developmental stages, resembling zygotic embryogeny, which can be synchronized by specific treatments, making it possible to collect a large number of somatic embryos at specific developmental stages. We have used this model to analyze global changes in gene expression during early stages of embryo development by generating an expression profile of 12,536 complementary DNA clones. This has allowed us to identify molecular events regulating putative processes associated with pattern formation during the earliest stages of embryogenesis which have not been identified on the molecular level in conifers before. We recognize notable changes in the expression of genes involved in regulating auxin biosynthesis and auxin response, gibberellin-mediated signaling, signaling between the embryo and the female gametophyte, tissue specification including the formation of boundary regions, and the switch from embryonic to vegetative development. In addition, our results confirm the involvement of previously described processes, including stress, differentiation of a protoderm, and programmed cell death.     

Molecular analysis of early rice stamen development using organ-specific gene expression profiling

Elucidating the regulatory mechanisms of plant organ formation is an important component of plant developmental biology and will be useful for crop improvement applications. Plant organ formation, or organogenesis, occurs when a group of primordial cells differentiates into an organ, through a well-orchestrated series of events, with a given shape, structure and function. Research over the past two decades has elucidated the molecular mechanisms of organ identity and dorsalventral axis determinations. However, little is known about the molecular mechanisms underlying the successive processes. To develop an effective approach for studying organ formation at the molecular level, we generated organ-specific gene expression profiles (GEPs) reflecting early development in rice stamen. In this study, we demonstrated that the GEPs are highly correlated with early stamen development, suggesting that this analysis is useful for dissecting stamen development regulation. Based on the molecular and morphological correlation, we found that over 26 genes, that were preferentially up-regulated during early stamen development, may participate in stamen development regulation. In addition, we found that differentially expressed genes during early stamen development are clustered into two clades, suggesting that stamen development may comprise of two distinct phases of pattern formation and cellular differentiation. Moreover, the organ-specific quantitative changes in gene expression levels may play a critical role for regulating plant organ formation. Electronic Supplementary Material Supplementary material is available for this article at Xiao-Chun Lu, Hua-Qin Gong contributed equally to this work.     

DNA array profiling of gene expression changes during maize embryo development

We are using DNA microarray-based gene expression profiling to classify temporal patterns of gene expression during the development of maize embryos, to understand mRNA-level control of embryogenesis and to dissect metabolic pathways and their interactions in the maize embryo. Genes involved in carbohydrate, fatty acid, and amino acid metabolism, the tricarboxylic acid (TCA) cycle, glycolysis, the pentose phosphate pathway, embryogenesis, membrane transport, signal transduction, cofactor biosynthesis, photosynthesis, oxidative phosphorylation and electron transfer, as well as 600 random complementary DNA (cDNA) clones from maize embryos, were arrayed on glass slides. DNA arrays were hybridized with fluorescent dye-labeled cDNA probes synthesized from kernel and embryo poly(A)⁺RNA from different stages of maize seed development. Several characteristic developmental patterns of expression were identified and correlated with gene function. Patterns of coordinated gene expression in the TCA cycle and glycolysis were analyzed in detail. The steady state level of poly(A)⁺ RNA for many genes varies dramatically during maize embryo development. Expression patterns of genes coding for enzymes of fatty acid biosynthesis and glycolysis are coordinately regulated during development. Genes of unknown function may by assigned a hypothetical role based on their patterns of expression resembling well characterized genes. Electronic supplementary material to this paper can be obtained by using the Springer LINK server located at http://dx.doi.org/10.1007/s10142-002-0046-6. Electronic Publication     

luxS-dependent gene regulation in Escherichia coli K-12 revealed by genomic expression profiling

The bacterial quorum-sensing autoinducer 2 (AI-2) has received intense interest because the gene for its synthase, luxS, is common among a large number of bacterial species. We have identified luxS-controlled genes in Escherichia coli under two different growth conditions using DNA microarrays. Twenty-three genes were affected by luxS deletion in the presence of glucose, and 63 genes were influenced by luxS deletion in the absence of glucose. Minimal overlap among these gene sets suggests the role of luxS is condition dependent. Under the latter condition, the metE gene, the lsrACDBFG operon, and the flanking genes of the lsr operon (lsrR, lsrK, tam, and yneE) were among the most significantly induced genes by luxS. The E. coli lsr operon includes an additional gene, tam, encoding an S-adenosyl-l-methionine-dependent methyltransferase. Also, lsrR and lsrK belong to the same operon, lsrRK, which is positively regulated by the cyclic AMP receptor protein and negatively regulated by LsrR. lsrK is additionally transcribed by a promoter between lsrR and lsrK. Deletion of luxS was also shown to affect genes involved in methionine biosynthesis, methyl transfer reactions, iron uptake, and utilization of carbon. It was surprising, however, that so few genes were affected by luxS deletion in this E. coli K-12 strain under these conditions. Most of the highly induced genes are related to AI-2 production and transport. These data are consistent with the function of LuxS as an important metabolic enzyme but appear not to support the role of AI-2 as a true signal molecule for E. coli W3110 under the investigated conditions.     

Genome-wide annotation and expression profiling of cell cycle regulatory genes in Chlamydomonas reinhardtii

Eukaryotic cell cycles are driven by a set of regulators that have undergone lineage-specific gene loss, duplication, or divergence in different taxa. It is not known to what extent these genomic processes contribute to differences in cell cycle regulatory programs and cell division mechanisms among different taxonomic groups. We have undertaken a genome-wide characterization of the cell cycle genes encoded by Chlamydomonas reinhardtii, a unicellular eukaryote that is part of the green algal/land plant clade. Although Chlamydomonas cells divide by a noncanonical mechanism termed multiple fission, the cell cycle regulatory proteins from Chlamydomonas are remarkably similar to those found in higher plants and metazoans, including the proteins of the RB-E2F pathway that are absent in the fungal kingdom. Unlike in higher plants and vertebrates where cell cycle regulatory genes have undergone extensive duplication, most of the cell cycle regulators in Chlamydomonas have not. The relatively small number of cell cycle genes and growing molecular genetic toolkit position Chlamydomonas to become an important model for higher plant and metazoan cell cycles.     

Stage-dependent gene expression profiles during natural killer cell development

Natural killer (NK) cells develop from hematopoietic stem cells (HSCs) in the bone marrow. To understand the molecular regulation of NK cell development, serial analysis of gene expression (SAGE) was applied to HSCs, NK precursor (pNK) cells, and mature NK cells (mNK) cultured without or with OP9 stromal cells. From 170,464 total individual tags from four SAGE libraries, 35,385 unique genes were identified. A set of genes was expressed in a stage-specific manner: 15 genes in HSCs, 30 genes in pNK cells, and 27 genes in mNK cells. Among them, lipoprotein lipase induced NK cell maturation and cytotoxic activity. Identification of genome-wide profiles of gene expression in different stages of NK cell development affords us a fundamental basis for defining the molecular network during NK cell development.