Detection of changes in gene regulatory patterns, elicited by perturbations of the Hsp90 molecular chaperone complex, by visualizing multiple experiments with an animation

<p>Abstract</p> <p>Background</p> <p>To make sense out of gene expression profiles, such analyses must be pushed beyond the mere listing of affected genes. For example, if a group of genes persistently display similar changes in expression levels under particular experimental conditions, and the proteins encoded by these genes interact and function in the same cellular compartments, this could be taken as very strong indicators for co-regulated protein complexes. One of the key requirements is having appropriate tools to detect such regulatory patterns.</p> <p>Results</p> <p>We have analyzed the global adaptations in gene expression patterns in the budding yeast when the Hsp90 molecular chaperone complex is perturbed either pharmacologically or genetically. We integrated these results with publicly accessible expression, protein-protein interaction and intracellular localization data. But most importantly, all experimental conditions were simultaneously and dynamically visualized with an animation. This critically facilitated the detection of patterns of gene expression changes that suggested underlying regulatory networks that a standard analysis by pairwise comparison and clustering could not have revealed.</p> <p>Conclusions</p> <p>The results of the animation-assisted detection of changes in gene regulatory patterns make predictions about the potential roles of Hsp90 and its co-chaperone p23 in regulating whole sets of genes. The simultaneous dynamic visualization of microarray experiments, represented in networks built by integrating one's own experimental with publicly accessible data, represents a powerful discovery tool that allows the generation of new interpretations and hypotheses.</p

p23/Sba1p protects against Hsp90 inhibitors

La chaperonne moléculaire Hsp90 est essentielle chez les eukaryotes. Hsp90 assiste différentes protéines cellulaires, agissant en complexe avec des co-chaperonnes spécifiques. Les fonctions in vivo de la co-chaperonne p23 et ses relations structure-fonction restent peu connues. Le gene SBA1, codant l'orthologue de p23 chez Saccharomyces cerevisiae, n'est pas essentiel. Nous avons démontré qu'en absence de p23/Sba1p, la levure et les cellules de mammifère sont hypersensibles aux inhibituers de Hsp90. Cette fonction protectrice de Sba1p dépend de sa capacité à lier Hsp90, à bloquer son activité ATPase et sa liaison aux inhibiteurs, mais ne met pas en jeu son activité de chaperonne moléculaire indépendante de Hsp90. L'analyse structure-fonction suggère que p23/Sba1 subit d'importants réarrangements structuraux après liaison à Hsp90. L'importance de la surface d'interaction entre p23/Sba1p et Hsp90 pourrait contribuer à préserver une fontion protectrice dans un environnement où les composés inhibiteurs de Hsp90 peuvent être produits par divers microorganismes

Deciphering interactions used by the influenza virus NS1 protein to silence the host antiviral sensor protein RIG-I using a bacterial reverse two-hybrid system

The majority of biological processes are controlled and regulated by an intricate network of thousands of interacting proteins. Identifying and understanding the key components of these protein networks, especially those that play a critical role in disease, is a challenge that promises to dramatically alter our current approach to healthcare. To facilitate this process, we have developed a method for the rapid construction of a chromosomally integrated, bacterial reverse two-hybrid system (RTHS) that enables the identification of interacting protein partners. Chromosomal integration of the RTHS enables stable protein expression, free of plasmid copy-number effects, as well as eliminating false positives arising from plasmid ejection. We have utilized this approach to identify the interactions used by the influenza virus NS1 protein to silence the host's antiviral defences

p23/Sba1p protects against Hsp90 inhibitors independently of its intrinsic chaperone activity

The molecular chaperone Hsp90 assists a subset of cellular proteins and is essential in eukaryotes. A cohort of cochaperones contributes to and regulates the multicomponent Hsp90 machine. Unlike the biochemical activities of the cochaperone p23, its in vivo functions and the structure-function relationship remain poorly understood, even in the genetically tractable model organism Saccharomyces cerevisiae. The SBA1 gene that encodes the p23 ortholog in this species is not an essential gene. We found that in the absence of p23/Sba1p, yeast and mammalian cells are hypersensitive to Hsp90 inhibitors. This protective function of Sba1p depends on its abilities to bind Hsp90 and to block the Hsp90 ATPase and inhibitor binding. In contrast, the protective function of Sba1p does not require the Hsp90-independent molecular chaperone activity of Sba1p. The structure-function analysis suggests that Sba1p undergoes considerable structural rearrangements upon binding Hsp90 and that the large size of the p23/Sba1p-Hsp90 interaction surface facilitates maintenance of high affinity despite sequence divergence during evolution. The large interface may also contribute to preserving a protective function in an environment in which Hsp90 inhibitory compounds can be produced by various microorganisms

Targeting Bacillus anthracis toxicity with a genetically selected inhibitor of the PA/CMG2 protein-protein interaction.

Raw data for &#39;Targeting Bacillus anthracis toxicity with a genetically selected inhibitor of the PA/CMG2 protein-protein interaction.&#39; The dataset contains the raw data used to generate the figures in the above manuscript. Each data file is labelled with the corresponding figure number used in the manuscript. The files were generated with Graphpad Prism 7. The folder contains the following files: * data used in Figure 1B * data used in Figure 2A-2C * data used in Figure 2D * data used in Figure 2E * data used in Figure 3 * data used in Figure 4A-4K * data used in Figure 5C If you require additional information please contact the corresponding author of the manuscript.</span

Targeting Bacillus anthracis toxicity with a genetically selected inhibitor of the PA/CMG2 protein-protein interaction

The protein-protein interaction between the human CMG2 receptor and the Bacillus anthracis protective antigen (PA) is essential for the transport of anthrax lethal and edema toxins into human cells. We used a genetically encoded high throughput screening platform to screen a SICLOPPS library of 3.2 million cyclic hexapeptides for inhibitors of this protein-protein interaction. Unusually, the top 3 hits all contained stop codons in the randomized region of the library, resulting in linear rather than cyclic peptides. These peptides disrupted the targeted interaction in vitro; two act by binding to CMG2 while one binds PA. The efficacy of the most potent CMG2-binding inhibitor was improved through the incorporation of non-natural phenylalanine analogues. Cell based assays demonstrated that the optimized inhibitor protects macrophages from the toxicity of lethal factor