Non-native interactions play an effective role in protein folding dynamics

Systematic Monte Carlo simulations of simple lattice models show that the final stage of protein folding is an ordered process where native contacts get locked (i.e., the residues come into contact and remain in contact for the duration of the folding process) in a well‐defined order. The detailed study of the folding dynamics of protein‐like sequences designed as to exhibit different contact energy distributions, as well as different degrees of sequence optimization (i.e., participation of non‐native interactions in the folding process), reveals significant differences in the corresponding locking scenarios—the collection of native contacts and their average locking times, which are largely ascribable to the dynamics of non‐native contacts. Furthermore, strong evidence for a positive role played by non‐native contacts at an early folding stage was also found. Interestingly, for topologically simple target structures, a positive interplay between native and non‐native contacts is observed also toward the end of the folding process, suggesting that non‐native contacts may indeed affect the overall folding process. For target models exhibiting clear two‐state kinetics, the relation between the nucleation mechanism of folding and the locking scenario is investigated. Our results suggest that the stabilization of the folding transition state can be achieved through the establishment of a very small network of native contacts that are the first to lock during the folding process.     

Molecular parameters associated with insulinoma progression: chromosomal instability versus p53 and CK19 status

Insulinomas represent the predominant syndromic subtype of endocrine pancreatic tumors (EPTs). Their metastatic potential cannot be predicted reliably using histopathological criteria. In the past few years, several attempts have been made to identify prognostic markers, among them TP53 mutations and immunostaining of p53 and recently cytokeratin 19 (CK19). In a previous study using conventional comparative genomic hybridization (CGH) we have shown that chromosomal instability (CIN) is associated with metastatic disease in insulinomas. It was our aim to evaluate these potential parameters in a single study. For the determination of CIN, we applied CGH to microarrays because it allows a high-resolution detection of DNA copy number changes in comparison with conventional CGH as well as the analysis of chromosomal regions close to the centromeres and telomeres, and at 1pter-->p32, 16p, 19 and 22. These regions are usually excluded from conventional CGH analysis, because they may show DNA gains in negative control hybridizations. Array CGH analysis of 30 insulinomas (15 tumors of benign, eight tumors of uncertain and seven tumors of malignant behavior) revealed that >or=20 chromosomal alterations and >or=6 telomeric losses were the best predictors of malignant progression. A subset of 22 insulinomas was further investigated for TP53 exon 5-8 gene mutations, and p53 and CK19 expression. Only one malignant tumor was shown to harbor an arginine 273 serine mutation and immunopositivity for p53. CK19 immunopositivity was detected in three malignant tumors and one tumor with uncertain behavior. In conclusion, our results indicate that CIN as well as telomeric loss are very powerful indicators for malignant progression in sporadic insulinomas. Our data do not support a critical role for p53 and CK19 as molecular parameters for this purpose.     

Statistical modeling of biomedical corpora: mining the Caenorhabditis Genetic Center Bibliography for genes related to life span

Background  

The statistical modeling of biomedical corpora could yield integrated, coarse-to-fine views of biological phenomena that complement discoveries made from analysis of molecular sequence and profiling data. Here, the potential of such modeling is demonstrated by examining the 5,225 free-text items in the Caenorhabditis Genetic Center (CGC) Bibliography using techniques from statistical information retrieval. Items in the CGC biomedical text corpus were modeled using the Latent Dirichlet Allocation (LDA) model. LDA is a hierarchical Bayesian model which represents a document as a random mixture over latent topics; each topic is characterized by a distribution over words.     

The shape of human gene family phylogenies

Background  

The shape of phylogenetic trees has been used to make inferences about the evolutionary process by comparing the shapes of actual phylogenies with those expected under simple models of the speciation process. Previous studies have focused on speciation events, but gene duplication is another lineage splitting event, analogous to speciation, and gene loss or deletion is analogous to extinction. Measures of the shape of gene family phylogenies can thus be used to investigate the processes of gene duplication and loss. We make the first systematic attempt to use tree shape to study gene duplication using human gene phylogenies.     

Chemoattractant-stimulated calcium influx in Dictyostelium discoideum does not depend on cGMP

Chemoattractant stimulation of Dictyostelium cells leads to the opening of calcium channels in the plasma membrane, causing extracellular calcium to flux into the cell. The genetically uncharacterised mutants stmF and KI8 show strongly altered chemoattractant-stimulated cGMP responses. The aberrant calcium influx in these strains has provided evidence that the chemoattractant-stimulated calcium influx is potentiated by cGMP. We have tested this hypothesis in genetically defined mutants by measuring the calcium influx in a strain that lacks intracellular cGMP due to the disruption of two guanylyl cyclases, and in a strain with increased cGMP levels caused by the disruption of two cGMP-degrading phosphodiesterases. The results reveal that the calcium influx stimulated by cAMP or folic acid is essentially identical in these strains. We conclude that cGMP is not involved in chemoattractant-stimulated calcium influx.     

A Highlights from MBoC Selection: Forty-five years of cGMP research in Dictyostelium: understanding the regulation and function of the cGMP pathway for cell movement and chemotaxis

In Dictyostelium, chemoattractants induce a fast cGMP response that mediates myosin filament formation in the rear of the cell. The major cGMP signaling pathway consists of a soluble guanylyl cyclase sGC, a cGMP-stimulated cGMP-specific phosphodiesterase, and the cGMP-target protein GbpC. Here we combine published experiments with many unpublished experiments performed in the past 45 years on the regulation and function of the cGMP signaling pathway. The chemoattractants stimulate heterotrimeric Gαβγ and monomeric Ras proteins. A fraction of the soluble guanylyl cyclase sGC binds with high affinity to a limited number of membrane binding sites, which is essential for sGC to become activated by Ras and Gα proteins. sGC can also bind to F-actin; binding to branched F-actin in pseudopods enhances basal sGC activity, whereas binding to parallel F-actin in the cortex reduces sGC activity. The cGMP pathway mediates cell polarity by inhibiting the rear: in unstimulated cells by sGC activity in the branched F-actin of pseudopods, in a shallow gradient by stimulated cGMP formation in pseudopods at the leading edge, and during cAMP oscillation to erase the previous polarity and establish a new polarity axis that aligns with the direction of the passing cAMP wave.     

Activation of soluble guanylyl cyclase at the leading edge during Dictyostelium chemotaxis

Dictyostelium contains two guanylyl cyclases, GCA, a 12-transmembrane enzyme, and sGC, a homologue of mammalian soluble adenylyl cyclase. sGC provides nearly all chemoattractant-stimulated cGMP formation and is essential for efficient chemotaxis toward cAMP. We show that in resting cells the major fraction of the sGC-GFP fusion protein localizes to the cytosol, and a small fraction is associated to the cell cortex. With the artificial substrate Mn2+/GTP, sGC activity and protein exhibit a similar distribution between soluble and particulate fraction of cell lysates. However, with the physiological substrate Mg2+/GTP, sGC in the cytosol is nearly inactive, whereas the particulate enzyme shows high enzyme activity. Reconstitution experiments reveal that inactive cytosolic sGC acquires catalytic activity with Mg2+/GTP upon association to the membrane. Stimulation of cells with cAMP results in a twofold increase of membrane-localized sGC-GFP, which is accompanied by an increase of the membrane-associated guanylyl cyclase activity. In a cAMP gradient, sGC-GFP localizes to the anterior cell cortex, suggesting that in chemotacting cells, sGC is activated at the leading edge of the cell.     

Quality determination and the repair of poor quality spots in array experiments

Background  

A common feature of microarray experiments is the occurence of missing gene expression data. These missing values occur for a variety of reasons, in particular, because of the filtering of poor quality spots and the removal of undefined values when a logarithmic transformation is applied to negative background-corrected intensities. The efficiency and power of an analysis performed can be substantially reduced by having an incomplete matrix of gene intensities. Additionally, most statistical methods require a complete intensity matrix. Furthermore, biases may be introduced into analyses through missing information on some genes. Thus methods for appropriately replacing (imputing) missing data and/or weighting poor quality spots are required.     

A Taxonomic Search Engine: Federating taxonomic databases using web services

Background  

The taxonomic name of an organism is a key link between different databases that store information on that organism. However, in the absence of a single, comprehensive database of organism names, individual databases lack an easy means of checking the correctness of a name. Furthermore, the same organism may have more than one name, and the same name may apply to more than one organism.