29 research outputs found
Effect of Correlated tRNA Abundances on Translation Errors and Evolution of Codon Usage Bias
Despite the fact that tRNA abundances are thought to play a major role in determining translation error rates, their distribution across the genetic code and the resulting implications have received little attention. In general, studies of codon usage bias (CUB) assume that codons with higher tRNA abundance have lower missense error rates. Using a model of protein translation based on tRNA competition and intra-ribosomal kinetics, we show that this assumption can be violated when tRNA abundances are positively correlated across the genetic code. Examining the distribution of tRNA abundances across 73 bacterial genomes from 20 different genera, we find a consistent positive correlation between tRNA abundances across the genetic code. This work challenges one of the fundamental assumptions made in over 30 years of research on CUB that codons with higher tRNA abundances have lower missense error rates and that missense errors are the primary selective force responsible for CUB
A kinetic study of in vitro lysis of \u3ci\u3eMycobacterium smegmatis \u3c/i\u3e
The traditional diagnostic tests for tuberculosis consist of an acid fast stain and a culture test from a sputum sample. With the emergence of drug resistant strains of tuberculosis, nucleic acid amplification has become the diagnostic test of choice. The nucleic acid amplification test consists of four steps: sputum sample collection, lysis of bacilli to release DNA, DNA amplification by PCR and detection of PCR products. The DNA extraction step has been largely overlooked and this study describes a systematic approach to measure the kinetics of cell lysis in a TrisāEDTA buffer. Mycobacterium smegmatis is a saphorytic, fast-growing mycobacterium that is often used as a surrogate of Mycobacterium tuberculosis in laboratory studies. M. smegmatis cells have been transformed with green fluorescent protein (GFP) genes. Transformed cells are lysed in a temperature-controlled cuvette that is equipped with optical input/output. The fluorescence signal increases when the GFP is released from lysed cells, and the extent of lysis of the loaded cells can be followed in real time. The experimental results are complemented by two theoretical models. The first model is based on a Monte Carlo simulation of the lysis process and the accompanying probability density function, as described by the FokkerāPlanck equation. The second model follows a chemical reaction engineering approach: the cell wall is modeled as layers, where each layer is made up of āblocks.ā Blocks can only be removed if they are exposed to the lysis solution and the model describes the rate of block exposure and removal. Both models are consistent with the experimental results. The main findings are: (1) the activation energy for M. smegmatis lysis in TrisāEDTA buffer is 22.1 kcal/mol, (2) cells lyse on the average after 14ā17% loss in cell wall thickness locally, (3) with the help of the models, the initial distribution in cell wall thickness of the population can be resolved and (4) near complete lysis of the cells is accomplished in 200 s at 80 Ā°C (90 s at 90 Ā°C). The results can be used to design an optimal lysis protocol that compromises between shorter processing times at higher temperature and reduced thermal damage to DNA at lower temperature. Supplementary data tables A-F are attached below as Related files
Transimulation - protein biosynthesis web service
Although translation is the key step during gene expression, it remains poorly characterized at the level of individual genes. For this reason, we developed Transimulation - a web service measuring translational activity of genes in three model organisms: Escherichia coli, Saccharomyces cerevisiae and Homo sapiens. The calculations are based on our previous computational model of translation and experimental data sets. Transimulation quantifies mean translation initiation and elongation time (expressed in SI units), and the number of proteins produced per transcript. It also approximates the number of ribosomes that typically occupy a transcript during translation, and simulates their propagation. The simulation of ribosomes' movement is interactive and allows modifying the coding sequence on the fly. It also enables uploading any coding sequence and simulating its translation in one of three model organisms. In such a case, ribosomes propagate according to mean codon elongation times of the host organism, which may prove useful for heterologous expression. Transimulation was used to examine evolutionary conservation of translational parameters of orthologous genes. Transimulation may be accessed at http://nexus.ibb.waw.pl/Transimulation (requires Java version 1.7 or higher). Its manual and source code, distributed under the GPL-2.0 license, is freely available at the website
Stochastic analysis of amino acid substitution in protein synthesis
We present a formal analysis of amino acid replacement during mRNA translation. Building on an abstract stochastic model of arrival of tRNAs and their processing at the ribosome, we compute probabilities of the insertion of amino acids into the nascent polypeptide chain. To this end, we integrate the probabilistic model checker Prism in the Matlab environment. We construct the substitution matrix containing the probabilities of an amino acid replacing another. The resulting matrix depends on various parameters, including availability and concentration of tRNA species, as well as their assignment to individual codons. We draw a parallel with the standard mutation matrices like Dayhoff and PET91, and analyze the mutual replacement of biologically similar amino acids.
Partially supported by EU FP6-project ESIGNET
In Silico Modelling and Analysis of Ribosome Kinetics and aa-tRNA Competition
We present a formal analysis of ribosome kinetics using probabilistic model checking and the tool Prism. We compute different parameters of the model, like probabilities of translation errors and average insertion times per codon. The model predicts strong correlation to the quotient of the concentrations of the so-called cognate and near-cognate tRNAs, in accord with experimental findings and other studies. Using piecewise analysis of the model, we are able to give an analytical explanation of this observation
In silico modelling and analysis of ribosome kinetics and aa-tRNA competition
We present a formal analysis of ribosome kinetics using probabilistic model checking and the tool Prism. We compute different parameters of the model, like probabilities of translation errors and average insertion times per codon. The model predicts strong correlation to the quotient of the concentrations of the socalled cognate and near-cognate tRNAs, in accord with experimental findings and other studies. Using piecewise analysis of the model, we are able to give an analytical
explanation of this observation
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Estrogen receptor Ī± protects pancreatic Ī²-cells from apoptosis by preserving mitochondrial function and suppressing endoplasmic reticulum stress.
Estrogen receptor Ī± (ERĪ±) action plays an important role in pancreatic Ī²-cell function and survival; thus, it is considered a potential therapeutic target for the treatment of type 2 diabetes in women. However, the mechanisms underlying the protective effects of ERĪ± remain unclear. Because ERĪ± regulates mitochondrial metabolism in other cell types, we hypothesized that ERĪ± may act to preserve insulin secretion and promote Ī²-cell survival by regulating mitochondrial-endoplasmic reticulum (EndoRetic) function. We tested this hypothesis using pancreatic islet-specific ERĪ± knockout (PERĪ±KO) mice and Min6 Ī²-cells in culture with Esr1 knockdown (KD). We found that Esr1-KD promoted reactive oxygen species production that associated with reduced fission/fusion dynamics and impaired mitophagy. Electron microscopy showed mitochondrial enlargement and a pro-fusion phenotype. Mitochondrial cristae and endoplasmic reticulum were dilated in Esr1-KD compared with ERĪ± replete Min6 Ī²-cells. Increased expression of Oma1 and Chop was paralleled by increased oxygen consumption and apoptosis susceptibility in ERĪ±-KD cells. In contrast, ERĪ± overexpression and ligand activation reduced both Chop and Oma1 expression, likely by ERĪ± binding to consensus estrogen-response element sites in the Oma1 and Chop promoters. Together, our findings suggest that ERĪ± promotes Ī²-cell survival and insulin secretion through maintenance of mitochondrial fission/fusion-mitophagy dynamics and EndoRetic function, in part by Oma1 and Chop repression