Accelerated search for biomolecular network models to interpret high-throughput experimental data
HoughFeature, a novel method for assessing drug effects in three-color cDNA microarrays
Three-color microarray experiments can be performed to assess drug effects on the genomic scale. The methodology may be useful in shortening the cycle, reducing the cost, and improving the efficiency in drug discovery and development compared with the commonly used dual-color technology. A visualization tool, the hexaMplot, is able to show the interrelations of gene expressions in normal-disease-drug samples in three-color microarray data. However, it is not enough to assess the complicated drug therapeutic effects based on the plot alone. It is important to explore more effective tools so that a deeper insight into gene.
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Tumor Drug Resistance via the same Signaling Pathway
Engelman et al.
A promising class of "smart" cancer drugs work by inhibiting specific tyrosine kinases linked to uncontrolled growth. Gefitinib and erlotinib, drugs that target the kinase activity of the epidermal growth factor receptor (EGFR), can be very effective when initially administered to lung cancer patients whose tumors contain activating mutations in the EGFR gene. Almost inevitably, however, these tumors develop resistance to the drugs and begin to regrow. Engelman et al. (p. 1039, published online 26 April) find that drug resistance in a subset of these tumors is caused by amplification of the MET oncogene, an event that in turn activates, via a different route, the same cellular signaling pathway originally activated by the mutant EGFR.
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Genome Expression Pathway Analysis Tool
Regulation of gene expression is relevant to many areas of biology and medicine, in the study of treatments, diseases, and developmental stages. Microarrays can be used to measure the expression level of thousands of mRNAs at the same time, allowing insight into or comparison of different cellular conditions. The data derived out of microarray experiments is highly dimensional and often noisy, and interpretation of the results can get intricate. Although programs for the statistical analysis of microarray data exist, most of them lack an integration of analysis results and biological interpretation. GEPAT is a modular, scalable and professional-grade software integrating analysis and interpretation of microarray gene expression data. An installation available for academic users can be found at http://gepat.bioapps.biozentrum.uni-wuerzburg.de.
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Combinatorial RNAi for quantitative protein network analysis
Özgür Sahin et al.
The elucidation of cross-talk events between intersecting signaling pathways is one main challenge in biological research. The complexity of protein networks, composed of different pathways, requires novel strategies and techniques to reveal relevant interrelations. Here, we established a combinatorial RNAi strategy for systematic single, double, and triple knockdown, and we measured the residual mRNAs and proteins quantitatively by quantitative real-time PCR and reverse-phase protein arrays, respectively, as a prerequisite for data analysis. Our results show that the parallel knockdown of at least three different genes is feasible while keeping both untargeted silencing and cytotoxicity low. The technique was validated by investigating the interplay of tyrosine kinase receptor ErbB2 and its downstream targets Akt-1 and MEK1 in cell invasion. This experimental approach combines multiple gene knockdown with a subsequent quantitative validation of reduced protein expression and is a major advancement toward the analysis of signaling pathways in systems biology.
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Plastid-to-nucleus retrograde signaling
Shai Koussevitzky et al.
Plastid-to-nucleus retrograde signaling coordinates nuclear gene expression with chloroplast function and is essential for the photoautotrophic life-style of plants. Three retrograde signals have been described, but little is known of their signaling pathways. We show here that GUN1, a chloroplast localized pentatricopeptide-repeat protein and ABI4, an AP2-type transcription factor, are common to all three pathways. ABI4 binds the promoter of a retrograde-regulated gene through a conserved motif found in close proximity to a light-regulatory element. We propose a model in which multiple indicators of aberrant plastid function are integrated upstream of GUN1 within plastids, leading to ABI4-mediated repression of nuclear-encoded genes.
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PageMan: An interactive ontology tool to generate, display, and annotate overview graphs for profiling experiments
Microarray technology has become a widely accepted and standardized tool in biology. The first microarray data analysis programs were developed to support pair-wise comparison. However, as microarray experiments have become more routine, large scale experiments have become more common, which investigate multiple time points or sets of mutants or transgenics. To extract biological information from such high-throughput expression data, it is necessary to develop efficient analytical platforms, which combine manually curated gene ontologies with efficient visualization and navigation tools. Currently, most tools focus on a few limited biological aspects, rather than offering a holistic, integrated analysis. PageMan allows multiple microarray experiments to be efficiently condensed into a single page graphical display. The flexible interface allows data to be quickly and easily visualized, facilitating comparisons within experiments and to published experiments, thus enabling researchers to gain a rapid overview of the biological responses in the experiments.
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The NeuARt II system: a viewing tool for neuroanatomical data based on published neuroanatomical atlases
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MathDAMP: a package for differential analysis of metabolite profiles
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MIMOX: a web tool for phage display based epitope mapping
Huang et al. 2006
A researcher who has identified a promising antibody, but who needs to discern the exact location of the epitope in the target antigen, faces a potentially time-consuming process. It is easy to see why the classical wet-lab approach of deriving a crystal structure of the antibody bound to its antigen would be unappealing. Fortunately, phage display has proven to be a powerful (and more convenient) approach. In this strategy, peptides that bind with high affinity to the antibody are analyzed and the consensus sequence is found. More often than not, the sequence is a mimotope, a sequence which mimics the true epitope's charge and shape rather than its exact sequence. Clearly, computational methods would be extremely useful in order to match the mimotope with the native epitope; however, no published software has hit the sweet spot of offering flexible features while remaining platform independent and freely available. Now, Huang et al. introduce MIMOX, a web-based tool for epitope mapping from phage display data. MIMOX begins with the mimotope sequences that have been determined via the phage display process; these are aligned using ClustalW and are processed in order to derive a consensus sequence. The software then combines this piece of information with a PDB file containing the antigen molecule's structure and provides the user with promising epitope candidates. Users can easily visualize mapping results in the context of the antigen's overall structure, and may peruse data regarding the surface accessibility of each candidate. MIMOX is implemented in Perl, is freely available for use by academic and commercial researchers alike, and can be accessed online at web.kuicr.kyoto-u.ac.jp/~hjian/mimox.
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Extended BiFC
Wolff et al.
Bimolecular fluorescence complementation (BiFC) is a powerful tool for detecting protein-protein interactions in living cells. Proteins of interest are tagged at the N (YN) or C terminus (YC) with complementary fragments of yellow fluorescence protein (YFP); when an interaction occurs, the complementary fragments come together, producing a yellow fluorescent signal. However, this approach does not provide information on expression or localization of the individual protein partners, which would require additional analysis (e.g., FRET). Wolff et al. have modified the BiFC assay to provide expression and localization data in addition to interaction data.
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Monitoring miRNAs
De Pietri Tonelli et al
MicroRNAs (miRNAs) are 20–25 nucleotide long stretches of noncoding RNAs that repress the translation of target mRNAs. They are usually tissue-specific and are often expressed in a developmentally regulated fashion. Thus, it would be very beneficial to be able to detect miRNA expression during cell fate changes in vivo. Unfortunately, the sensors used to detect miRNA expression often rely on negative data—the lack of translation of the sensor mRNA—so the presence of the miRNA must be confirmed. These sensors also require the generation of transgenic animals, which is laborious, expensive, and time-consuming. De Pietri Tonelli et al. have made a dual-fluorescence reporter/sensor plasmid that overcomes both of these limitations and allows the detection of miRNAs with cellular resolution in vivo. Their elegant system relies on a GFP reporter and an mRFP sensor, both under the control of identical constitutive promoters. Green fluorescence thus establishes the presence of the plasmid in a given cell, while red fluorescence indicates the lack of miRNA silencing. Using this technique, the group confirmed previous in situ data showing active miR-1 in zebrafish muscle fibers 33 h postfertilization. They also found that, contrary to currently held views, miR-124a is expressed not only in postmitotic neurons, but also in their progenitors, the neuroepithelial cells.
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Not Lost in Translation
PFINGSTEN ET AL.
The canonical mechanism for initiation of protein synthesis in eukaryotes involves a nucleotide cap on messenger RNA (mRNA) that is recognized by an initiation protein factor. However, a variety of pathogenic viruses and cellular mRNAs bypass the canonical mechanism by using structured RNA sequences, called internal ribosomal entry sites (IRESs), to initiate translation. Pfingsten et al. have determined the structure of the ribosome-binding domain of an IRES at 3.1 angstrom resolution. The RNA prefolds to create a specific ribosome-binding structure. By docking the structure onto cryoelectron microscopic reconstructions of an IRES-ribosome complex, contacts were identified that drive binding and induce conformational change in the ribosome.
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Chaperome: A Variety of Chaperone Proteins
Xiaodong Wang, John Venable, Paul LaPointe, Darren M. Hutt, Atanas V. Koulov, Judith Coppinger, Cemal Gurkan, Wendy Kellner, Jeanne Matteson, Helen Plutner,John R. Riordan, Jeffery W. Kelly,John R. Yates III and William E. Balch
The expression of misfolded or aberrant proteins on the cell surface could wreak havoc with the immune system. Cells have therefore developed an efficient quality-control system, which diverts misfolded membrane and secretory proteins from the secretory pathway by retaining and degrading them at the entry portal to the secretory pathway, the endoplasmic reticulum (ER). One well-studied example of quality control involves the cystic fibrosis transmembrane conductance regulator (CFTR), misfolding of which is responsible for disease in a large proportion of sufferers. However, sometimes quality control is too stringent, and functional, though mutant, proteins are retained. Wang et al. used a systematic approach to examine the folding pathway and protein interaction partners of CFTR and the common disease variant CFTR F508, which, even though functional, is retained in the ER. A variety of chaperone proteins, which help to promote protein folding, are present in the ER, and a chaperome of over 30 proteins involved in CFTR folding and transport was identified from among more than 200 interacting proteins. Interfering with CFTR-specific chaperone mechanisms may thus be a useful strategy to correct disease, and other protein misfolding diseases might be similarly amenable to equivalent interventions.
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3D Complex: A Structural Classification of Protein Complexes
The millions of genes sequenced over the past decade correspond to a much smaller set of protein structural domains, or folds—probably only a few thousand. Since structural data is being accumulated at a fast pace, classifications of domains such as SCOP help significantly in understanding the sequence–structure relationship. More recently, classifications of interacting domain pairs address the relationship between sequence divergence and domain–domain interaction. One level of description that has yet to be investigated is the protein complex level, which is the physiologically relevant state for most proteins within the cell. Here, Levy and colleagues propose a classification scheme for protein complexes, which will allow a better understanding of their structural properties and evolution.
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An Integrative Genomic Approach to Uncover Molecular Mechanisms of Prokaryotic Traits
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Computational Inference of Neural Information Flow Networks
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Modified cotton could help feed the world
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Scientists have genetically modified cotton to make its seeds — which are full of high-quality protein — fit for human consumption.
The research published this week in Proceedings of the National Academies of Science has the potential to feed half a billion people worldwide each year.
For every kilogram of fibre produced by cotton, the plant releases 1.65 kilograms of seed. These contain a toxin called gossypol that protects the crops from insects and pathogens and can only be digested by ruminant animals such as cattle.
The genetically modified cotton plants produce seeds with 98 per cent less gossypol, but have normal levels of the toxin in the rest of the plant, preserving its chemical defences.
Deborah Delmer, associate director of the Rockefeller Foundation in New York, United States, and an expert in agricultural food safety, says that the method — which turns off a gene process rather than introducing a new protein — raises less safety concerns than other genetic modification technologies.
But she adds that extensive field trials are still needed to test the plant's stability over many generations.
Link to full article in Nature
Link to abstract in Proceedings of the National Academies of Sciences
