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Molecular Medicine Reports Apr 2015Tendinopathy is a critical clinical problem as it is often asymptomatic at onset and during development, and is only recognized upon rupture of the tendon. It is common...
Tendinopathy is a critical clinical problem as it is often asymptomatic at onset and during development, and is only recognized upon rupture of the tendon. It is common among recreational and competitive athletes. The present study sought to examine the molecular mechanism of the progression of tendinopathy by screening out differentially expressed genes (DEGs) and investigating their functions. In addition, the present study aimed to identify the small molecules, which exhibit potential effects, which could be utilized for the treatment of tendinopathy. The gene expression profile of tendinopathy, GSE26051 was downloaded from the Gene Expression Omnibus database, which included 23 control samples and 18 samples of tendinopathy. The DEGs were identified using the Limma package in the R programming language, and gene ontology and pathway enrichment analysis were performed. In addition, the potential regulatory microRNAs and the target sites of the transcription factors were screened out based on the molecular signature database. In addition, the DEGs were mapped to the connectivity map database to identify the potential small molecule drugs. A total of 318 genes were filtered as DEGs between diseased samples and normal control tendons. Additionally, genes, including laminin, α4, platelet‑derived growth factor α, laminin γ1 and Src homology 2 transforming protein 1 may induce tendinopathy through the focal adhesion pathway. Furthermore, the transcription factor, lymphoid enhancer‑binding factor 1 and its target genes, pantothenate kinase 2 and G protein‑coupled receptor kinase 5 were identified. The most significant microRNA, miR‑499, was screened and was found to regulate specific genes, including CUGBP2 and MYB. Additionally, the small molecules, Prestwick‑1082 and viomycin were identified to have the potential to repair disordered metabolic pathways and furthermore to remedy tendinopathy. The results of the present study assessed the mechanism of tendinopathy and screened small molecule drugs as potential treatments for this condition. In addition, the present findings have the potential for use in a clinical setting for the treatment of tendinopathy in the future.
Topics: Binding Sites; Computational Biology; Databases, Nucleic Acid; Datasets as Topic; Drug Discovery; Drug Evaluation, Preclinical; Gene Expression Profiling; Gene Expression Regulation; Gene Regulatory Networks; Humans; MicroRNAs; Protein Binding; Signal Transduction; Small Molecule Libraries; Tendinopathy; Transcription Factors
PubMed: 25502513
DOI: 10.3892/mmr.2014.3081 -
Journal of Molecular Biology Jan 2015Previous structural studies suggested that ribosomal translocation is accompanied by large interdomain rearrangements of elongation factor G (EF-G). Here, we follow the...
Previous structural studies suggested that ribosomal translocation is accompanied by large interdomain rearrangements of elongation factor G (EF-G). Here, we follow the movement of domain IV of EF-G relative to domain II of EF-G using ensemble and single-molecule Förster resonance energy transfer. Our results indicate that ribosome-free EF-G predominantly adopts a compact conformation that can also, albeit infrequently, transition into a more extended conformation in which domain IV moves away from domain II. By contrast, ribosome-bound EF-G predominantly adopts an extended conformation regardless of whether it is interacting with pretranslocation ribosomes or with posttranslocation ribosomes. Our data suggest that ribosome-bound EF-G may also occasionally sample at least one more compact conformation. GTP hydrolysis catalyzed by EF-G does not affect the relative stability of the observed conformations in ribosome-free and ribosome-bound EF-G. Our data support a model suggesting that, upon binding to a pretranslocation ribosome, EF-G moves from a compact to a more extended conformation. This transition is not coupled to but likely precedes both GTP hydrolysis and mRNA/tRNA translocation.
Topics: Escherichia coli; Gene Rearrangement; Peptide Elongation Factor G; Protein Conformation; RNA, Messenger; RNA, Transfer; Ribosomes; Sequence Analysis, DNA; Translocation, Genetic
PubMed: 25463439
DOI: 10.1016/j.jmb.2014.11.010 -
Proceedings of the National Academy of... Oct 2014Translocation of mRNA and tRNAs through the ribosome is catalyzed by a universally conserved elongation factor (EF-G in prokaryotes and EF-2 in eukaryotes). Previous...
Translocation of mRNA and tRNAs through the ribosome is catalyzed by a universally conserved elongation factor (EF-G in prokaryotes and EF-2 in eukaryotes). Previous studies have suggested that ribosome-bound EF-G undergoes significant structural rearrangements. Here, we follow the movement of domain IV of EF-G, which is critical for the catalysis of translocation, relative to protein S12 of the small ribosomal subunit using single-molecule FRET. We show that ribosome-bound EF-G adopts distinct conformations corresponding to the pre- and posttranslocation states of the ribosome. Our results suggest that, upon ribosomal translocation, domain IV of EF-G moves toward the A site of the small ribosomal subunit and facilitates the movement of peptidyl-tRNA from the A to the P site. We found no evidence of direct coupling between the observed movement of domain IV of EF-G and GTP hydrolysis. In addition, our results suggest that the pretranslocation conformation of the EF-G-ribosome complex is significantly less stable than the posttranslocation conformation. Hence, the structural rearrangement of EF-G makes a considerable energetic contribution to promoting tRNA translocation.
Topics: Biological Transport; Catalysis; Fluorescence Resonance Energy Transfer; Guanosine Triphosphate; Microscopy; Peptide Elongation Factor G; Protein Binding; Protein Structure, Tertiary; Protein Synthesis Inhibitors; Protein Transport; RNA, Messenger; RNA, Transfer; Ribosomes; Viomycin
PubMed: 25288752
DOI: 10.1073/pnas.1410873111 -
Nature Communications Sep 2014The antibiotic streptomycin is widely used in the treatment of microbial infections. The primary mechanism of action is inhibition of translation by binding to the...
The antibiotic streptomycin is widely used in the treatment of microbial infections. The primary mechanism of action is inhibition of translation by binding to the ribosome, but how it enters the bacterial cell is unclear. Early in the study of this antibiotic, a mysterious streptomycin-induced potassium efflux preceding any decrease in viability was observed; it was speculated that this changed the electrochemical gradient such that streptomycin better accessed the cytoplasm. Here we use a high-throughput screen to search for compounds targeting the mechanosensitive channel of large conductance (MscL) and find dihydrostreptomycin among the 'hits'. Furthermore, we find that MscL is not only necessary for the previously described streptomycin-induced potassium efflux, but also directly increases MscL activity in electrophysiological studies. The data suggest that gating MscL is a novel mode of action of dihydrostreptomycin, and that MscL's large pore may provide a mechanism for cell entry.
Topics: Anti-Bacterial Agents; Dihydrostreptomycin Sulfate; Escherichia coli; Escherichia coli Proteins; High-Throughput Screening Assays; Ion Channels; Patch-Clamp Techniques; Potassium; Spectinomycin; Streptomycin; Viomycin
PubMed: 25205267
DOI: 10.1038/ncomms5891