2D differential in-gel electrophoresis for the identification of esophageal scans cell cancer-specific protein markers

The reproducibility of conventional two-dimensional (2D) gel electrophoresis can be improved using differential in-gel electrophoresis (DIGE), a new emerging technology for proteomic analysis. In DIGE, two pools of proteins are labeled with 1-(5-carboxypentyl)-1'-propylindocarbocyanine halide (Cy3) N-hydroxy-succinimidyl ester and 1-(5-carboxypentyl)-1'-methylindodi-carbocyanine halide (Cy5) N-hydroxysuccinimidyl ester fluorescent dyes, respectively. The labeled proteins are mixed and separated in the same 2D gel. 2D DIGE was applied to quantify the differences in protein expression between laser capture microdissection-procured esophageal carcinoma cells and normal epithelial cells and to define cancer-specific and normal-specific protein markers. Analysis of the 2D images from protein lysates of approximately 250,000 cancer cells and normal cells identified 1038 protein spots in cancer cell lysates and 1088 protein spots in normal cell lysates. Of the detected proteins, 58 spots were up-regulated by >3-fold and 107 were down-regulated by >3-fold in cancer cells. In addition to previously identified down-regulated protein annexin I, tumor rejection antigen (gp96) was found up-regulated in esophageal squamous cell cancer. Global quantification of protein expression between laser capture-microdissected patient-matched cancer cells and normal cells using 2D DIGE in combination with mass spectrometry is a powerful tool for the molecular characterization of cancer progression and identification of cancer-specific protein markers.     

Discordant protein and mRNA expression in lung adenocarcinomas

The relationship between gene expression measured at the mRNA level and the corresponding protein level is not well characterized in human cancer. In this study, we compared mRNA and protein expression for a cohort of genes in the same lung adenocarcinomas. The abundance of 165 protein spots representing 98 individual genes was analyzed in 76 lung adenocarcinomas and nine non-neoplastic lung tissues using two-dimensional polyacrylamide gel electrophoresis. Specific polypeptides were identified using matrix-assisted laser desorption/ionization mass spectrometry. For the same 85 samples, mRNA levels were determined using oligonucleotide microarrays, allowing a comparative analysis of mRNA and protein expression among the 165 protein spots. Twenty-eight of the 165 protein spots (17%) or 21 of 98 genes (21.4%) had a statistically significant correlation between protein and mRNA expression (r > 0.2445; p < 0.05); however, among all 165 proteins the correlation coefficient values (r) ranged from -0.467 to 0.442. Correlation coefficient values were not related to protein abundance. Further, no significant correlation between mRNA and protein expression was found (r = -0.025) if the average levels of mRNA or protein among all samples were applied across the 165 protein spots (98 genes). The mRNA/protein correlation coefficient also varied among proteins with multiple isoforms, indicating potentially separate isoform-specific mechanisms for the regulation of protein abundance. Among the 21 genes with a significant correlation between mRNA and protein, five genes differed significantly between stage I and stage III lung adenocarcinomas. Using a quantitative analysis of mRNA and protein expression within the same lung adenocarcinomas, we showed that only a subset of the proteins exhibited a significant correlation with mRNA abundance.     

Inter- and intra-individual variation in resting oxygen consumption in post-larvae of the giant freshwater prawn,Macrobrachium rosenbergii (De Man)

In a study of the factors that influence metabolic rate in the giant freshwater prawn Macrobrachium rosenbergii, resting oxygen consumption (ROC) was measured in 90 post-larvae ranging in size from 0.1 to 2.8 g. As in many other animal species, ROC was strongly negatively related to body weight. A stressful event (anaesthesia with or without tagging) caused a sharp increase in the ROC that disappeared over a time scale of hours. As has been demonstrated for other species of crustaceans, ROC was highest in prawns in the pre-moult stage. Individual differences in ROC among prawns handled in the same way and in the same moult stage persisted over a period of hours, but not over days. It remains unclear, therefore, whether early differences in resting metabolic rate can explain the conspicuous differences in growth rate that are found in this species during the first few weeks of life and that profoundly influence subsequent life history events.     

Osteoblastic glutamate receptor function regulates bone formation and resorption

Previous studies showed that a variety of bone cells express protein components necessary for neuronal-like glutamatergic signaling and implicated glutamate as having a role in mechanically induced bone remodeling. Initial functional studies concentrated on the role of glutamate signaling in bone resorption and provided compelling evidence to suggest that glutamate signaling through functional NMDA type ionotropic glutamate receptors (iGluRs) is a prerequisite for in vitro osteoclastogenesis. Originally, effects of iGluR antagonists seen in co-cultures were attributed to antagonists acting directly on osteoclast precursors. However, in the light of recent osteoblast studies it now seems likely that the observed effects on osteoclastogenesis are an indirect effect of modulating the function of pre-osteoblast present within these cultures. The presence of iGluRs in osteoblasts suggests a role for them in bone formation and this paper reviews and discusses the emerging data relating to the role of glutamate signaling in osteoblasts. A number of recently published studies have shown that osteoblasts not only express a wide number of 'pre-synaptic' glutamatergic proteins but also possess the ability to both regulate glutamate release and actively recycle extracellular glutamate. The functionality of osteoblastic 'post-synaptic' glutamatergic components has also been shown as both primary and clonal osteoblasts express electrophysiologically active iGluRs, metabotropic type glutamate receptors (mGluRs) along with a variety of glutamate receptor associated signaling proteins. There is, however, little published data regarding the actual role of glutamatergic signaling in osteoblastic bone formation. In vivo and in vitro studies performed provide evidence that glutamatergic signaling is a necessity for normal osteoblast function. In a number of different models of in vitro bone formation, the addition of non-competitive antagonists of iGluRs prevents the formation of mineralized bone, moreover antagonizing some sub-types of iGluR mediates the differentiation of pre-osteoblasts. iGluR antagonists modulate osteoblast function in a manner that correlates with the previously reported data regarding in vitro osteoclastogenesis. Interestingly iGluR mediated glutamate signaling appears to function differently in osteoblasts derived from flat and long bones. This implies the components of osteoblastic glutamatergic signaling may be adapted in vivo possibly to reflect the differential function of osteoblasts in those regions of the skeleton.     

Structure motif discovery and mining the PDB

MOTIVATION: Many of the most interesting functional and evolutionary relationships among proteins are so ancient that they cannot be reliably detected through sequence analysis and are apparent only through a comparison of the tertiary structures. The conserved features can often be described as structural motifs consisting of a few single residues or Secondary Structure (SS) elements. Confidence in such motifs is greatly boosted when they are found in more than a pair of proteins. RESULTS: We describe an algorithm for the automatic discovery of recurring patterns in protein structures. The patterns consist of individual residues having a defined order along the protein's backbone that come close together in the structure and whose spatial conformations are similar. The residues in a pattern need not be close in the protein's sequence. The work described in this paper builds on an earlier reported algorithm for motif discovery. This paper describes a significant improvement of the algorithm which makes it very efficient. The improved efficiency allows us to use it for doing unsupervised learning of patterns occurring in small subsets in a large set of structures, a non-redundant subset of the Protein Data Bank (PDB) database of all known protein structures.     

A comparative map of bovine chromosome 25 with human chromosomes 7 and 16

A comparative genome map is necessary for the implementation of comparative positional candidate gene cloning in cattle. We have developed a medium density comparative gene map of bovine chromosome 25 (BTA25). A radiation hybrid (RH) panel was used to map nine microsatellites and nine genes. Eight of the nine comparative loci were also mapped by FISH. These results were combined with data from published articles to create a comprehensive comparative map of BTA25 with human chromosomes 7 (HSA7) and 16 (HSA16). This map should facilitate the cloning of genes of interest on bovine chromosome 25.     

Validation of single nucleotide polymorphism quantification in pooled DNA samples with SNaPIT. A glycosylase-mediated methods for polymorphism detection method

Association studies using genome scans to identify quantitative trait loci for multifactorial disorders, with anything approaching reasonable power, have been compromised by the need for a very dense array of genetic markers and large numbers of affected individuals. These requirements impose enormous burdens on the genotyping capacity for most laboratories. DNA pooling has been proposed as a possible approach to reduce genotyping costs and effort. We report on the application of the SNaPIT™ technology to evaluate allele frequencies in pooled DNA samples and conclude that it offers a cost effective, efficient and accurate estimator and provides several advantages over competing technologies in this regard.     

The molecular genetics of schizophrenia: an emerging consensus

Schizophrenia is perhaps the most debilitating mental disease and determining the underlying cause has become a challenging area of psychiatric research. It is relatively well established that genes play a role in the aetiology of schizophrenia. In this article, a review of important findings related to schizophrenia as a genetic trait will be provided, including a discussion of family, twin and adoption studies. Molecular genetic studies of specific candidate genes are then reviewed. Some controversies within the literature are examined and possible directions for future research are discussed.