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Results for “Functional Genomics”

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At least 1,153 records · Page 64Linked to original sources
PubMed2002

Integration of genomic datasets to predict protein complexes in yeast.

The ultimate goal of functional genomics is to define the function of all the genes in the genome of an organism. A large body of information of the biological roles of genes has been accumulated and aggregated in the past decades of research, both from traditional experiments detailing the role of individual genes and proteins, and from newer experimental strategies that aim to characterize gene function on a genomic scale. It is clear that the goal of functional genomics can only be achieved by integrating information and data sources from the variety of these different experiments. Integration of different data is thus an important challenge for bioinformatics. The integration of different data sources often helps to uncover non-obvious relationships between genes, but there are also two further benefits. First, it is likely that whenever information from multiple independent sources agrees, it should be more valid and reliable. Secondly, by looking at the union of multiple sources, one can cover larger parts of the genome. This is obvious for integrating results from multiple single gene or protein experiments, but also necessary for many of the results from genome-wide experiments since they are often confined to certain (although sizable) subsets of the genome. In this paper, we explore an example of such a data integration procedure. We focus on the prediction of membership in protein complexes for individual genes. For this, we recruit six different data sources that include expression profiles, interaction data, essentiality and localization information. Each of these data sources individually contains some weakly predictive information with respect to protein complexes, but we show how this prediction can be improved by combining all of them. Supplementary information is available at http:// bioinfo.mbb.yale.edu/integrate/interactions/.

Cell Cycle↗
PubMed1999

Selection of antibodies for intracellular function using a two-hybrid in vivo system.

Expression of antibodies inside cells has been used successfully to ablate protein function. This finding suggests that the technology should have an impact on disease treatment and in functional genomics where proteins of unknown function are predicted from genomic sequences. A major hindrance is the paucity of antibodies that function in eukaryotic cells, presumably because the antibodies fold incorrectly in the cytoplasm. To overcome this problem, we have developed an in vivo assay for functional intracellular antibodies using a two-hybrid approach. In this assay, antibody, as single-chain Fv (scFv) linked to a transcriptional transactivation domain, can interact with a target antigen, linked to a LexA-DNA binding domain, and thereby activate a reporter gene. We find that several characterized antibodies can bind their target antigen in eukaryotic cells in this two-hybrid format, and we have been able to isolate intracellular binders from among sets of scFv that can bind antigen in vitro. Furthermore, we show a model selection in which a single scFv was isolated from a mixture of half a million clones, indicating that this is a robust procedure that should facilitate capture of antibody specificities from complex mixtures. The approach can provide the basis for de novo selection of intracellular scFv from libraries, such as those made from spleen RNA after immunization with antigen, for intracellular analysis of protein function based only on genomic or cDNA sequences.

Animals↗
PubMed2002

Generating new marine cell lines and transgenic species--conference summary.

Marine species offer a tremendous diversity of life histories, physiologies, genetics, behaviors, and biologies, reflecting myriad adaptations to the water environment. Historically, marine vertebrates, particularly fish, have played significant roles in a wide range of disciplines, including environmental toxicology, genetics, developmental biology, and physiology, among others. Much still remains to be learned from these animals, and there is a growing need for new marine models. Models for expression of marine animal genes have been limited to heterologous expression systems. While there is still a great deal to gain from heterologous expression systems, the interactions of genes with one another can best be determined in homologous expression systems where appropriate interactions are possible. This has become particularly important with the development of functional genomics in marine models. These homologous gene expression systems will be key to the use of functional genomics for marine animal molecular physiology and toxicology.

Animals↗
PubMed2006

Switching the flip: protein phosphatase roles in signaling pathways.

Protein phosphatases are the obligate partners of protein kinases in cellular control circuitry. Several specific roles of phosphatases have been revealed in plant systems through recent work capitalizing on functional genomics, interaction screens and activation-tagging approaches. Historically, the redundancy of genes that encode phosphatase functions has impeded genetic analysis in plant systems, and relatively few elucidated signaling pathways include protein phosphatases that have clearly defined substrates. Functional genomics, interaction and suppression screens, activation tagging and phosphoproteomics now hold great promise for identifying phosphatases that are active in signaling and regulation, and for revealing their in vivo substrates. This level of analysis will be essential for a full understanding of the effects of reversible protein phosphorylation in modulating and integrating cellular activities.

Abscisic Acid↗
PubMed2004

Whole-genome annotation by using evidence integration in functional-linkage networks.

The advent of high-throughput biology has catalyzed a remarkable improvement in our ability to identify new genes. A large fraction of newly discovered genes have an unknown functional role, particularly when they are specific to a particular lineage or organism. These genes, currently labeled "hypothetical," might support important biological cell functions and could potentially serve as targets for medical, diagnostic, or pharmacogenomic studies. An important challenge to the scientific community is to associate these newly predicted genes with a biological function that can be validated by experimental screens. In the absence of sequence or structural homology to known genes, we must rely on advanced biotechnological methods, such as DNA chips and protein-protein interaction screens as well as computational techniques to assign putative functions to these genes. In this article, we propose an effective methodology for combining biological evidence obtained in several high-throughput experimental screens and integrating this evidence in a way that provides consistent functional assignments to hypothetical genes. We use the visualization method of propagation diagrams to illustrate the flow of functional evidence that supports the functional assignments produced by the algorithm. Our results contain a number of predictions and furnish strong evidence that integration of functional information is indeed a promising direction for improving the accuracy and robustness of functional genomics.

Chromosome Mapping↗
PubMed2002

Relationship of the glyoxylate pathway to the pathogenesis of Cryptococcus neoformans.

Functional genomics has become a major focus in the study of microbial pathogenesis. This study used a functional genomic tool, differential display reverse transcription-PCR, to identify a transcriptional profile of Cryptococcus neoformans cells as they produced meningitis in an immunosuppressed host. This serial global gene expression during infection allowed for the identification of up- and down-regulated genes during infection. During this profiling, a single gene for the enzyme isocitrate lyase (ICL1) was found to be up regulated at 1 week of infection in a rabbit meningitis model and during a time of maximum host cellular response. The finding suggested that this enzyme and the glyoxylate shunt pathway are important to this yeast's energy production during infection. However, site-directed icl1 mutants had no apparent virulence defect in two animal models and no growth defect within macrophages. These observations suggest that although the yeast responded to a certain environmental cue(s) by an increase in ICL1 expression during infection, this gene was not necessary for progression of a C. neoformans infection. Compounds that specifically target only ICL1 are unlikely to cripple C. neoformans growth in vivo.

Amino Acid Sequence↗
PubMed1999

Sequence and structural elements at the 3' terminus of bovine viral diarrhea virus genomic RNA: functional role during RNA replication.

Bovine viral diarrhea virus (BVDV), a member of the genus Pestivirus in the family Flaviviridae, has a positive-stranded RNA genome consisting of a single open reading frame and untranslated regions (UTRs) at the 5' and 3' ends. Computer modeling suggested the 3' UTR comprised single-stranded regions as well as stem-loop structures-features that were suspected of being essentially implicated in the viral RNA replication pathway. Employing a subgenomic BVDV RNA (DI9c) that was shown to function as an autonomous RNA replicon (S.-E. Behrens, C. W. Grassmann, H. J. Thiel, G. Meyers, and N. Tautz, J. Virol. 72:2364-2372, 1998) the goal of this study was to determine the RNA secondary structure of the 3' UTR by experimental means and to investigate the significance of defined RNA motifs for the RNA replication pathway. Enzymatic and chemical structure probing revealed mainly the conserved terminal part (termed 3'C) of the DI9c 3' UTR containing distinctive RNA motifs, i.e., a stable stem-loop, SL I, near the RNA 3' terminus and a considerably less stable stem-loop, SL II, that forms the 5' portion of 3'C. SL I and SL II are separated by a long single-stranded intervening sequence, denoted SS. The 3'-terminal four C residues of the viral RNA were confirmed to be single stranded as well. Other intramolecular interactions, e.g., with upstream DI9c RNA sequences, were not detected under the experimental conditions used. Mutagenesis of the DI9c RNA demonstrated that the SL I and SS motifs do indeed play essential roles during RNA replication. Abolition of RNA stems, which ought to maintain the overall folding of SL I, as well as substitution of certain single-stranded nucleotides located in the SS region or SL I loop region, gave rise to DI9c derivatives unable to replicate. Conversely, SL I stems comprising compensatory base exchanges turned out to support replication, but mostly to a lower degree than the original structure. Surprisingly, replacement of a number of residues, although they were previously defined as constituents of a highly conserved stretch of sequence of the SS motif, had little effect on the replication ability of DI9c. In summary, these results indicate that RNA structure as well as sequence elements harbored within the 3'C region of the BVDV 3' UTR create a common cis-acting element of the replication process. The data further point at possible interaction sites of host and/or viral proteins and thus provide valuable information for future experiments intended to identify and characterize these factors.

3' Untranslated Regions↗
PubMed2002

Microarray deacetylation maps determine genome-wide functions for yeast histone deacetylases.

Yeast contains a family of five related histone deacetylases (HDACs) whose functions are known at few genes. Therefore, we used chromatin immunoprecipitation and intergenic microarrays to generate genome-wide HDAC enzyme activity maps. Rpd3 and Hda1 deacetylate mainly distinct promoters and gene classes where they are recruited largely by novel mechanisms. Hda1 also deacetylates subtelomeric domains containing normally repressed genes that are used instead for gluconeogenesis, growth on carbon sources other than glucose, and adverse growth conditions. These domains have certain features of heterochromatin but are distinct from subtelomeric heterochromatin repressed by the deacetylase Sir2. Finally, Hos1/Hos3 and Hos2 preferentially affect ribosomal DNA and ribosomal protein genes, respectively. Thus, acetylation microarrays uncover the "division of labor" for yeast histone deacetylases.

Carrier Proteins↗
PubMed2003

Genome-wide analysis of epigenetics in cancer.

Human cancers are caused by multiple mechanisms. Research in the last 30 years has firmly established the roles of a group of genes including oncogenes, tumor suppressor genes, and DNA repair genes in human cancers. The activation and inactivation of these cancer genes can be caused by genetic mutations or epigenetic alterations. The epigenetic changes in cancers include methylation of CpG islands, loss of imprinting, and chromatin modification. The completion of the genome sequences of many organisms including the human has transformed the traditional approach to molecular biology research into an era of functional genome research. Traditional research usually involves the study of one or a few genes (proteins) in a particular biological process in normal physiology or disease. Functional genome research takes advantage of newly available genome sequences and high-throughput genome technologies to study genes and/or proteins to inform the perspective of entire biological processes. I will focus on recent progress in the identification of imprinted genes and methylation of CpG islands through genome-wide analysis.

Computational Biology↗
PubMed1998

[Trends of genomic researches for drug discoveries].

In drug discoveries it becomes more important to search and validate target genes that control diseases. The development of novel strategies for functional genomics is indispensable for large scale analysis of human genes. (1) Recently, several research projects sequencing 5'-end of full-length enriched cDNA libraries were started because most of expressed sequence tags(ESTs) lack the 5'-end information of mRNA that is necessary for identifying the correct coding region. (2) New key technologies to quantify numerous gene expression levels simultaneously are now in progress. (3) Toward the functional analysis of unpredictable human genes, new strategies by using model animals (mouse, yeast, nematode) are under construction. Advances in molecular genetics and functional genomics will create new challenges for both drug discoveries and clinical evaluations.

Animals↗
PubMed2004

Exploring interactions between rat hepatocytes and nonparenchymal cells using gene expression profiling.

Cocultivation of primary hepatocytes with a plethora of nonparenchymal cells (from within and outside the liver) has been shown to support hepatic functions in vitro. Despite significant investigation into this phenomenon, the molecular mechanism underlying epithelial-nonparenchymal interactions in hepatocyte cocultures remains poorly understood. In this study, we present a functional genomic approach utilizing gene expression profiling to isolate molecular mediators potentially involved in induction of liver-specific functions by nonparenchymal cells. Specifically, primary rat hepatocytes were cocultivated with closely related murine fibroblast cell types (3T3-J2, NIH-3T3, mouse embryonic fibroblasts) to allow their classification as "high," "medium," or "low" inducers of hepatic functions. These functional responses were correlated with fibroblast gene expression profiles obtained using Affymetrix GeneChips. Microarray data analysis provided us with 17 functionally characterized candidate genes in the cell communication category (cell surface, extracellular matrix, secreted factors) that may be involved in induction of hepatic functions. Further analysis using various databases (i.e., PubMed, GenBank) facilitated prioritization of the candidates for functional characterization. We experimentally validated the potential role of two candidates in our coculture model. The cell surface protein, neural cadherin (N-cadherin), was localized to hepatocyte-fibroblast junctions, while adsorbed decorin up-regulated hepatic functions in pure cultures as well as cocultures with low-inducing fibroblasts. In the future, identifying mediators of hepatocyte differentiation may have implications for both fundamental hepatology and cell-based therapies (e.g., bioartificial liver devices). In conclusion, the functional genomic approach presented in this study may be utilized to investigate mechanisms of cell-cell interaction in a variety of tissues and disease states.

Animals↗
PubMed2000

Inducible expression of double-stranded RNA directs specific genetic interference in Drosophila.

BACKGROUND: The introduction of double-stranded RNA (dsRNA) can selectively interfere with gene expression in a wide variety of organisms, providing an ideal approach for functional genomics. Although this method has been used in Drosophila, it has been limited to studies of embryonic gene function. Only inefficient effects have been seen at later stages of development. RESULTS: When expressed under the control of a heat-inducible promoter, dsRNA interfered efficiently and specifically with gene expression during larval and prepupal development in Drosophila. Expression of dsRNA corresponding to the EcR ecdysone receptor gene generated defects in larval molting and metamorphosis, resulting in animals that failed to pupariate or prepupae that died with defects in larval tissue cell death and adult leg formation. In contrast, expression of dsRNA corresponding to the coding region of the betaFTZ-F1 orphan nuclear receptor had no effect on puparium formation, but led to an arrest of prepupal development, generating more severe lethal phenotypes than those seen with a weak betaFTZ-F1 loss-of-function allele. Animals that expressed either EcR or betaFTZ-F1 dsRNA showed defects in the expression of corresponding target genes, indicating that the observed developmental defects are caused by disruption of the genetic cascades that control the onset of metamorphosis. CONCLUSIONS: These results confirm and extend our understanding of EcR and betaFTZ-F1 function. They also demonstrate that dsRNA expression can inactivate Drosophila gene function at later stages of development, providing a new tool for functional genomic studies in Drosophila.

Animals↗
PubMed2005

Design and analysis in genetic studies of human ageing and longevity.

With the success of the Human Genome Project and taking advantage of the recent developments in high-throughput genotyping techniques as well as in functional genomics, it is now feasible to collect vast quantities of genetic data with the aim of deciphering the genetics of human complex traits. As a result, the amount of research on human ageing and longevity has been growing rapidly in recent years. The situation raises questions concerning efficient choice of study population, sampling schemes, and methods of data analysis. In this article, we summarize the key issues in genetic studies of human ageing and longevity ranging from research design to statistical analyses. We discuss the virtues and drawbacks of the multidisciplinary approaches including the population-based cross-sectional and cohort studies, family-based linkage analysis, and functional genomics studies. Different analytical approaches are illustrated with their performances compared. In addition, important research topics are highlighted together with experiment design and data analyzing issues to serve as references for future studies.

Aging↗
PubMed2026

An ATP-Driven N Protein-DDX21 Molecular Switch Dynamically Controls SARS-CoV-2 RNA G-Quadruplex Heterogeneity.

The SARS-CoV-2 RNA genome functions as a highly structured regulatory scaffold. Although bioinformatic analyses predict widespread RNA G-quadruplexes (G4s) across the viral genome, their structural diversity and regulatory mechanisms remain poorly understood. Here, we report a diverse landscape of viral G4s encompassing parallel and non-canonical topologies with remarkable thermostability. Unlike typical eukaryotic G4s, these two-tetrad viral G4s exhibit a hierarchical ion-dependent mechanism, in which K+ establishes the core fold, and Mg2 + acts as a secondary regulator promoting conformational compaction. Single-molecule FRET analysis further distinguishes rigid, long-lived G4 folds from highly dynamic, metastable species, defining a continuum of conformational states along the viral genome. Functionally, we identify a synergistic yet competitive interplay between the viral nucleocapsid (N) protein and host helicase DDX21. While the N protein acts as a molecular chaperone to promote G4 folding, DDX21 selectively resolves these structures in an ATP-dependent manner. Strikingly, N and DDX21 jointly constitute a finely tuned, ATP-driven molecular switch, where ATP availability dictates the equilibrium between G4-stabilized and resolved states. Our findings establish a mechanistic framework for the active regulation of SARS-CoV-2 RNA architecture and reveal a multilayered host-virus regulatory axis that modulates viral genome heterogeneity.

DEAD‐box helicases↗
PubMed2003

The Drosophila phenotype gap - and how to close it.

Functional genomics (the elucidation of gene function in the context of a sequenced genome) depends critically on functional biology. Genetic model organisms have hitherto not attracted much physiological input, however. This skills mismatch, termed the phenotype gap, can be quantified by analysis of the annotations of sequenced genomes. This is illustrated in the context of Drosophila. In this case, it seems as if a shift from developmental biology to transport physiology and metabolism will be required to provide a more balanced skills base for post-genomics.

Animals↗
PubMed2025

Quantitative essentiality in a reduced genome: a functional, regulatory and structural fitness map.

Essentiality studies have traditionally focused on coding regions, often overlooking other small genetic regulatory elements. To address this, we combined transposon libraries containing promoter or terminator sequences to obtain a high-resolution essentiality map of a genome-reduced bacterium, at near-single-nucleotide precision when considering non-essential genes. By integrating temporal transposon-sequencing data by k-means unsupervised clustering, we present a novel essentiality assessment approach, providing dynamic and quantitative information on the fitness contribution of different genomic regions. We compared the insertion tolerance and persistence of the two engineered libraries, assessing the local impact of transcription and termination on cell fitness. Essentiality assessment at the local base-level revealed essential protein domains and small genomic regions that are either essential or inaccessible to transposon insertion. We also identified structural regions within essential genes that tolerate transposon disruptions, resulting in functionally split proteins. Overall, this study presents a nuanced view of gene essentiality, shifting from static and binary models to a more accurate perspective. Additionally, it provides valuable insights for genome engineering and enhances our understanding of the biology of genome-reduced cells.

DNA Transposable Elements↗
PubMed2025

Microfluidics to Follow Spatiotemporal Dynamics at the Nucleo-Cytoplasmic Interface During Plant Root Growth.

Nuclear dynamics refers to global/local changes in the molecular and spatial organization of genomic DNA that can occur during development or in response to environmental stress signals and eventually impact genomic functions. In plants, nuclear dynamics relies notably on the connection of the nucleus with the cytoskeleton during development. It orchestrates genomic functions in response to developmental and environmental cues. This is particularly true in the plant root system, which is constantly exposed to a wide range of internal and external stimuli. Currently, studying nuclear dynamics in a growing root is challenging due to limitations regarding real-time imaging for quantitative analyses under controlled conditions. Microfluidic systems for plant cell studies are valuable analytical tools that provide precise control of culture conditions together with live-imaging capabilities at high temporal and spatial resolutions. Herein, we describe a microfluidic platform to unravel dynamically and noninvasively nuclear organization in the seedling root system exposed to various treatments. As exemplified here, our microfluidic platform can be conveniently used for real-time microscopy imaging and quantitative analysis of fine nuclear morphological changes upon modifying cytoskeleton dynamics. Importantly, our system can be applied to a wide variety of microscopic means including high-resolution microscopy to investigate diverse subcellular compartments or nuclear domains in Arabidopsis thaliana roots.

Plant Roots↗