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At least 1,423 records · Page 79Linked to original sources
PubMed2005

Emerging issues of the expression profiling technologies for the study of gynecologic cancer.

Evaluation of the prognostic parameters of gynecologic cancer has shown their failure for classification according to the clinical behavior or the prediction of its outcome. This weakness has important implications on prognosis and treatment. The increasing understanding of the complexity of the human genome, coupled with the development of high throughput analysis techniques and bioinformatics tools, has changed our concepts on cancer biology, by shifting our targets to a global analysis of the transcriptome and the proteome, linking genes and their products into functional pathways. These approaches permit the documentation of expression patterns of thousands of genes within a cell. With the use of DNA microarray technology, it is feasible to identify signature patterns of expression in tumor samples that faithfully correlate with its biology, providing accurate prognosis for each cancer patient and thus a rational customized treatment. At this stage, there is a need for systematic studies for the validation of these novel approaches. In this review, we provide a basic background of the concept of the technology, highlight several emerging issues from their applications on gynecologic cancer, discuss a series of important themes and problems regarding their interpretation and relevance for the clinicians, and comment on future areas of research.

Computational Biology↗
PubMed2004

The thioredoxin h system of higher plants.

In plants, thioredoxins h are encoded by a multigenic family of genes (eight in Arabidopsis thaliana, at least five in Populus sp.). The multiplicity of these isoforms raises the question of their specificity. This review focuses on thioredoxins h in two plant models: Arabidopsis and poplar. Thioredoxins h can be divided into three different subgroups according to the analysis of their primary structure. This paper describes the biochemical properties of each subgroup. Recent data in the field indicate that subgroup members differ by their subcellular localization as well as their reduction pathways suggesting specific functions for each subgroup. The development of proteomic tools has also increased considerably the number of potential thioredoxin targets, showing the importance of thioredoxins h in plants.

Amino Acid Sequence↗
PubMed2006

Progress in in silico functional genomics: the malaria Metabolic Pathways database.

Genomic, transcriptomic and proteomic data can be turned into biologically meaningful information if they are synthesized into processes. Such amalgamation has been done for the most virulent malaria parasite Plasmodium falciparum in the Metabolic Pathways database. The dialectics of construction of metabolic pathways and other biological processes using this database is presented here. Additional features, such as links to other databases and the incorporation of transcriptomic clocks, are elucidated. Comparison of Metabolic Pathways to other similar databases is analyzed.

Animals↗
PubMed2002

Can we infer peptide recognition specificity mediated by SH3 domains?

Protein interaction domain families that modulate the formation of macromolecular complexes recognize specific sequence or structural motifs. For instance SH3 and WW domains bind to polyproline peptides while SH2 and FHA domains bind to peptides phosphorylated in Tyr and Thr respectively. Within each family, variations in the chemical characteristics of the domain binding pocket modulate a finer peptide recognition specificity and, as a consequence, determine the selection of functional protein partners in vivo. In the proteomic era there is the need for reliable inference methods to help restricting the sequence space of the putative targets to be confirmed experimentally by more laborious experimental approaches. Here we will review the published data about the peptide recognition specificity of the SH3 domain family and we will propose a classification of SH3 domains into eight classes. Finally, we will discuss whether the available information is sufficient to infer the recognition specificity of any uncharacterized SH3 domain.

Amino Acid Sequence↗
PubMed2003

Call volume and insulin signaling.

Perturbations of cell hydration as provoked by changes in ambient osmolarity or under isoosmotic conditions by hormones, second messengers, intracellular substrate accumulation, or reactive oxygen intermediates critically contribute to the physiological regulation of cell function. In general an increase in cell hydration stimulates anabolic metabolism and proliferation and provides cytoprotection, whereas cellular dehydration leads to a catabolic situation and sensitizes cells to apoptotic stimuli. Insulin produces cell swelling by inducing a net K+ and Na+ accumulation inside the cell, which results from a concerted activation of Na+/H+ exchange, Na+/K+/2Cl- symport, and the Na+/K(+)-ATPase. In the liver, insulin-induced cell swelling is critical for stimulation of glycogen and protein synthesis as well as inhibition of autophagic proteolysis. These insulin effects can largely be mimicked by hypoosmotic cell swelling, pointing to a role of cell swelling as a trigger of signal transduction. This article discusses insulin-induced signal transduction upstream of swelling and introduces the hypothesis that cell swelling as a signal amplifyer represents an essential component in insulin signaling, which contributes to the full response to insulin at the level of signal transduction and function. Cellular dehydration impairs insulin signaling and may be a major cause of insulin resistance, which develops in systemic hyperosmolarity, nutrient deprivation, uremia, oxidative challenges, and unbalanced production of insulin-counteracting hormones. Hydration changes affect cell functions at multiple levels (such as transcriptom, proteom, phosphoproteom, and the metabolom) and a system biological approach may allow us to develop a more holistic view on the hydration dependence of insulin signaling in the future.

Animals↗
PubMed2006

A tagging-via-substrate technology for genome-wide detection and identification of farnesylated proteins.

Protein farnesylation is one of the most common lipid modifications and has an important role in the regulation of various cellular functions. We have recently developed a novel proteomics strategy, designated the tagging-via-substrate (TAS) approach, for the detection and proteomic analysis of farnesylated proteins. This chapter describes the principle of TAS technology and details the method for detection and enrichment of farnesylated proteins.

Animals↗
PubMed2002

A single-chain antibody/epitope system for functional analysis of protein-protein interactions.

Protein-protein interactions play a critical role in cellular processes such as signal transduction. Although many methods for identifying the binding partners of a protein of interest are available, it is currently difficult or impossible to assess the functional consequences of a specific interaction in vivo. To address this issue, we propose to modify proteins by addition of an artificial protein binding interface, thereby forcing them to interact in the cell in a pairwise fashion and allowing the functional consequences to be determined. For this purpose, we have developed an artificial binding interface consisting of a anti-Myc single-chain antibody (ScFv) and its peptide epitope. We found that the binding of an ScFv derived from anti-Myc monoclonal antibody 9E10 was relatively weak in vivo, so we selected an improved clone, 3DX, by in vitro mutagenesis and phage display. 3DX bound well to its epitope in a yeast two-hybrid system, and GST-fused 3DX also bound to several Myc-tagged proteins in mammalian cells. In vivo binding was relatively insensitive to the position of the ScFv in a fusion protein, but was improved by including multiple tandem copies of the Myc epitope in the binding partner. To test the system, we successfully replaced the SH3 domain-mediated interaction between the Abl tyrosine kinase and adaptor proteins Crk and Nck with an engineered interaction between 3DX and multiple Myc tags. We expect that this approach, which we term a functional interaction trap, will be a powerful proteomic tool for investigating protein-protein interactions.

Adaptor Proteins, Signal Transducing↗
PubMedDate not supplied

Affinity ligand selection from a library of small molecules: assay development, screening, and application.

A facile and cost-effective process for screening synthetic libraries for an affinity ligand is described. A high throughput 96-well plate filtration method was designed to screen both discrete compounds and mixtures of compounds attached to a solid support. Human serum albumin (HSA) was used as a target protein to demonstrate the proof of concept. Detection and quantitation by fluorescence was accomplished with the use of fluorescamine to conjugate the protein in the filtrate. It is found that mixtures demonstrating low average binding reflect an overall lower hit rate of the components, whereas deconvolution of mixtures with high protein binding consistently provides a high hit rate. This differs from many of the previous experiences screening solid-phase mixtures in which high false positive rates are noted to occur. A total of 100K compounds were tested: 25K as discrete samples and 75K as mixtures. An overall hit rate of 8% was observed. Secondary screening of compounds measured specificity, recovery, and dynamic binding capacity. The effectiveness of the method is illustrated using an affinity column made with a representative lead compound. A similar purity was achieved in a single-step purification of HSA from serum as compared to that obtained by two steps of ion-exchange chromatography. The process for primary screening of a large number of compounds is simple, inexpensive, and applicable to any soluble target protein of known or unknown function from crude mixtures and may have additional utility as a generic chemical affinity tool for the functional characterization of novel proteins emerging from proteomics work.

Chromatography, Affinity↗
PubMed2002

DNA microarrays for functional plant genomics.

DNA microarray technology is a key element in today's functional genomics toolbox. The power of the method lies in miniaturization, automation and parallelism permitting large-scale and genome-wide acquisition of quantitative biological information from multiple samples. DNA microarrays are currently fabricated and assayed by two main approaches involving either in situ synthesis of oligonucleotides ('oligonucleotide microarrays') or deposition of pre-synthesized DNA fragments ('cDNA microarrays') on solid surfaces. To date, the main applications of microarrays are in comprehensive, simultaneous gene expression monitoring and in DNA variation analyses for the identification and genotyping of mutations and polymorphisms. Already at a relatively early stage of its application in plant science, microarrays are being utilized to examine a range of biological issues including the circadian clock, plant defence, environmental stress responses, fruit ripening, phytochrome A signalling, seed development and nitrate assimilation. Novel insights are obtained into the molecular mechanisms co-ordinating metabolic pathways, regulatory and signalling networks. Exciting new information will be gained in the years to come not only from genome-wide expression analyses on a few model plant species, but also from extensive studies of less thoroughly studied species on a more limited scale. The value of microarray technology to our understanding of living processes will depend both on the amount of data to be generated and on its clever exploration and integration with other biological knowledge arising from complementary functional genomics tools for 'profiling' the genome, proteome, metabolome and phenome.

Gene Expression Profiling↗
PubMed2003

ER-mediated phagocytosis: a new membrane for new functions.

Genomics and other high-throughput approaches, such as proteomics, are changing the way we study complex biological systems. In the past few years, these approaches have contributed markedly to improving our understanding of phagocytosis. Indeed, the ability to study biological systems by monitoring hundreds of proteins provides a level of resolution that is not attainable by the usual 'one protein at a time' approach. In this article, I discuss how proteomic approaches have revealed surprising findings that enable us to revisit established concepts, such as the origin of the phagosome membrane, and to propose new models of cell organization and the link between innate and adaptive immunity.

Animals↗
PubMed2024

Kick-starting the zygotic genome: licensors, specifiers, and beyond.

Zygotic genome activation (ZGA), the first transcription event following fertilization, kickstarts the embryonic program that takes over the control of early development from the maternal products. How ZGA occurs, especially in mammals, is poorly understood due to the limited amount of research materials. With the rapid development of single-cell and low-input technologies, remarkable progress made in the past decade has unveiled dramatic transitions of the epigenomes, transcriptomes, proteomes, and metabolomes associated with ZGA. Moreover, functional investigations are yielding insights into the key regulators of ZGA, among which two major classes of players are emerging: licensors and specifiers. Licensors would control the permission of transcription and its timing during ZGA. Accumulating evidence suggests that such licensors of ZGA include regulators of the transcription apparatus and nuclear gatekeepers. Specifiers would instruct the activation of specific genes during ZGA. These specifiers include key transcription factors present at this stage, often facilitated by epigenetic regulators. Based on data primarily from mammals but also results from other species, we discuss in this review how recent research sheds light on the molecular regulation of ZGA and its executors, including the licensors and specifiers.

Animals↗
PubMed2003

DNA vaccines: a review.

The DNA vaccines are simple rings of DNA containing a gene encoding an antigen, and a promoter/terminator to make the gene express in mammalian cells. They are a promising new approach for generating all types of desired immunity: cytolytic T lymphocytes (CTL), T helper cells and antibodies, whilst being a technology that has the potential for global usage in terms of manufacturing ease, broad population administration and safety. This review gives an overview of the mechanisms, preclinical and clinical efficacy of DNA vaccines, and point out the limitations of the first generation of such vaccines, and some of the promising second-generation developments. This technology is also being utilized in the field of proteomics as a tool to elucidate the function of genes. The breadth of applications for DNA vaccines thus ranges from prophylactic vaccines to immunotherapy for infectious diseases, cancer, and autoimmune and allergic diseases.

Antigens↗
PubMed2004

Global analyses of sumoylated proteins in Saccharomyces cerevisiae. Induction of protein sumoylation by cellular stresses.

We have undertaken a global analysis of sumoylated proteins in Saccharomyces cerevisiae by tandem mass spectrometry. Exposure of cells to oxidative and ethanol stresses caused large increases in protein sumoylation. A large number of new sumoylated proteins were identified in untreated, hydrogen peroxide-treated, and ethanol-treated cells. These proteins are known to be involved in diverse cellular processes, including gene transcription, protein translation, DNA replication, chromosome segregation, metabolic processes, and stress responses. Additionally, the known enzymes, including E1, E2, and E3 of the sumoylation cascade were found to be auto-sumoylated. Taken together, these results show that protein sumoylation is broadly involved in many cellular functions and this mass spectrometry-based proteomic approach is useful in studying the regulation of protein sumoylation in the cells.

Blotting, Western↗
PubMed2025

Knowledge-enhanced protein subcellular localization prediction from 3D fluorescence microscope images.

MOTIVATION: Pinpointing the subcellular location of proteins is essential for studying protein function and related diseases. Advances in spatial proteomics have shown that automatic recognition of protein subcellular localization from images could highly facilitate protein translocation analysis and biomarker discovery, but existing machine-learning works have been mostly limited to processing 2D images. By contrast, 3D images have higher spatial resolution and allow researchers to observe cellular structures in their natural context, but currently, there are only a few studies of 3D image processing for protein distribution analysis due to the lack of data and complexity of modeling. RESULTS: We developed a knowledge-enhanced protein subcellular localization model, KE3DLoc, which could recognize distribution patterns in 3D fluorescence microscope images using deep learning methods. The model designs an image feature extraction module that incorporates information from 3D and 2D projected cells and implements asymmetric loss and confidence weights to address data imbalance and weak cell annotation issues. Besides, considering that the biological knowledge in the Gene Ontology (GO) database can provide valuable support for protein location understanding, the KE3DLoc model incorporates a novel knowledge enhancement module that optimizes the protein representation by related knowledge graphs derived from the GO. Since the image module and the knowledge module calculate features from different levels, KE3DLoc designs protein ID aggregation to enhance the consistency of protein features across different cells. Experimental results on three public datasets have demonstrated that the KE3DLoc significantly outperforms existing methods and provides valuable insights for spatial proteomics research. AVAILABILITY AND IMPLEMENTATION: All datasets and codes used in this study are available at GitHub: https://github.com/PRBioimages/KE3DLoc.

Microscopy, Fluorescence↗
PubMed2003

Control of protein life-span by N-terminal methionine excision.

Peptide deformylases (PDFs) have been discovered recently in eukaryotic genomes, and it appears that N-terminal methionine excision (NME) is a conserved pathway in all compartments where protein synthesis occurs. This work aimed at uncovering the function(s) of NME in a whole proteome, using the chloroplast-encoded proteins of both Arabidopsis thaliana and Chlamydomonas reinhardtii as model systems. Disruption of PDF1B in A.thaliana led to an albino phenotype, and an extreme sensitivity to the PDF- specific inhibitor actinonin. In contrast, a knockout line for PDF1A exhibited no apparent phenotype. Photosystem II activity in C.reinhardtii cells was substantially reduced by the presence of actinonin. Pulse-chase experiments revealed that PDF inhibition leads to destabilization of a crucial subset of chloroplast-encoded photosystem II components in C. reinhardtii. The same proteins were destabilized in pdf1b. Site-directed substitutions altering NME of the most sensitive target, subunit D2, resulted in similar effects. Thus, plastid NME is a critical mechanism specifically influencing the life-span of photosystem II polypeptides. A general role of NME in modulating the half-life of key subsets of proteins is suggested.

Amidohydrolases↗
PubMed2006

Peroxisome proliferator-activated receptor alpha plays a vital role in inducing a detoxification system against plant compounds with crosstalk with other xenobiotic nuclear receptors.

Peroxisome proliferator-activated receptor alpha (PPARalpha) is thought to play an important role in lipid metabolism in the liver. To clarify the extra-hepatic and/or unknown function of PPARalpha, we previously performed a proteome analysis of the intestinal proteins and identified 17beta-hydroxysteroid dehydrogenase type 11 as a mostly induced protein by a PPARalpha ligand [Motojima, K. (2004) Eur. J. Biochem. 271, 4141-4146]. Because of its supposed wide substrate specificity, we examined the possibility that PPARalpha plays an important role in inducing detoxification systems for some natural foods by feeding mice with various plant seeds and grains. Feeding with sesame but not others often killed PPARalpha knockout mice but not wild-type mice. A microarray analysis of the sesame-induced mRNAs in the intestine revealed that PPARalpha plays a vital role in inducing various xenobiotic metabolizing enzymes in the mouse intestine and liver. A PPARalpha ligand alone could not induce most of these enzymes, suggesting that there is an essential crosstalk among PPARalpha and other xenobiotic nuclear receptors to induce a detoxification system for plant compounds.

Amino Acid Sequence↗
PubMed2002

A role for the protease falcipain 1 in host cell invasion by the human malaria parasite.

Cysteine proteases of Plasmodium falciparum are required for survival of the malaria parasite, yet their specific cellular functions remain unclear. We used a chemical proteomic screen with a small-molecule probe to characterize the predominant cysteine proteases throughout the parasite life cycle. Only one protease, falcipain 1, was active during the invasive merozoite stage. Falcipain 1-specific inhibitors, identified by screening of chemical libraries, blocked parasite invasion of host erythrocytes, yet had no effect on normal parasite processes such as hemoglobin degradation. These results demonstrate a specific role for falcipain 1 in host cell invasion and establish a potential new target for antimalarial therapeutics.

Animals↗
PubMed2005

The unique stacked rings in the nucleocapsid of the white spot syndrome virus virion are formed by the major structural protein VP664, the largest viral structural protein ever found.

One unique feature of the shrimp white spot syndrome virus (WSSV) genome is the presence of a giant open reading frame (ORF) of 18,234 nucleotides that encodes a long polypeptide of 6,077 amino acids with a hitherto unknown function. In the present study, by applying proteomic methodology to analyze the sodium dodecyl sulfate-polyacrylamide gel electrophoresis profile of purified WSSV virions by liquid chromatography-mass spectrometry (LC-MS/MS), we found that this giant polypeptide, designated VP664, is one of the viral structural proteins. The existence of the corresponding 18-kb transcript was confirmed by sequencing analysis of reverse transcription-PCR products, which also showed that vp664 was intron-less. A time course analysis showed that this transcript was actively transcribed at the late stage, suggesting that this gene product should contribute primarily to the assembly and morphogenesis of the virion. Several polyclonal antisera against this giant protein were prepared, and one of them was successfully used for immunoelectron microscopy analysis to localize the protein in the virion. Immunoelectron microscopy with a gold-labeled secondary antibody showed that the gold particles were regularly distributed around the periphery of the nucleocapsid with a periodicity that matched the characteristic stacked ring subunits that appear as striations. From this and other evidence, we argue that this giant ORF in fact encodes the major WSSV nucleocapsid protein.

Amino Acid Sequence↗