Computational Prediction and Experimental Verification of New MAP Kinase Docking Sites and Substrates Including Gli Transcription Factors

In order to fully understand protein kinase networks, new methods are needed to identify regulators and substrates of kinases, especially for weakly expressed proteins. Here we have developed a hybrid computational search algorithm that combines machine learning and expert knowledge to identify kinase docking sites, and used this algorithm to search the human genome for novel MAP kinase substrates and regulators focused on the JNK family of MAP kinases. Predictions were tested by peptide array followed by rigorous biochemical verification with in vitro binding and kinase assays on wild-type and mutant proteins. Using this procedure, we found new ‘D-site’ class docking sites in previously known JNK substrates (hnRNP-K, PPM1J/PP2Czeta), as well as new JNK-interacting proteins (MLL4, NEIL1). Finally, we identified new D-site-dependent MAPK substrates, including the hedgehog-regulated transcription factors Gli1 and Gli3, suggesting that a direct connection between MAP kinase and hedgehog signaling may occur at the level of these key regulators. These results demonstrate that a genome-wide search for MAP kinase docking sites can be used to find new docking sites and substrates

Robust metrics for assessing the performance of different verbal autopsy cause assignment methods in validation studies

<p>Abstract</p> <p>Background</p> <p>Verbal autopsy (VA) is an important method for obtaining cause of death information in settings without vital registration and medical certification of causes of death. An array of methods, including physician review and computer-automated methods, have been proposed and used. Choosing the best method for VA requires the appropriate metrics for assessing performance. Currently used metrics such as sensitivity, specificity, and cause-specific mortality fraction (CSMF) errors do not provide a robust basis for comparison.</p> <p>Methods</p> <p>We use simple simulations of populations with three causes of death to demonstrate that most metrics used in VA validation studies are extremely sensitive to the CSMF composition of the test dataset. Simulations also demonstrate that an inferior method can appear to have better performance than an alternative due strictly to the CSMF composition of the test set.</p> <p>Results</p> <p>VA methods need to be evaluated across a set of test datasets with widely varying CSMF compositions. We propose two metrics for assessing the performance of a proposed VA method. For assessing how well a method does at individual cause of death assignment, we recommend the average chance-corrected concordance across causes. This metric is insensitive to the CSMF composition of the test sets and corrects for the degree to which a method will get the cause correct due strictly to chance. For the evaluation of CSMF estimation, we propose CSMF accuracy. CSMF accuracy is defined as one minus the sum of all absolute CSMF errors across causes divided by the maximum total error. It is scaled from zero to one and can generalize a method's CSMF estimation capability regardless of the number of causes. Performance of a VA method for CSMF estimation by cause can be assessed by examining the relationship across test datasets between the estimated CSMF and the true CSMF.</p> <p>Conclusions</p> <p>With an increasing range of VA methods available, it will be critical to objectively assess their performance in assigning cause of death. Chance-corrected concordance and CSMF accuracy assessed across a large number of test datasets with widely varying CSMF composition provide a robust strategy for this assessment.</p

Predicting DNA-Binding Specificities of Eukaryotic Transcription Factors

Today, annotated amino acid sequences of more and more transcription factors (TFs) are readily available. Quantitative information about their DNA-binding specificities, however, are hard to obtain. Position frequency matrices (PFMs), the most widely used models to represent binding specificities, are experimentally characterized only for a small fraction of all TFs. Even for some of the most intensively studied eukaryotic organisms (i.e., human, rat and mouse), roughly one-sixth of all proteins with annotated DNA-binding domain have been characterized experimentally. Here, we present a new method based on support vector regression for predicting quantitative DNA-binding specificities of TFs in different eukaryotic species. This approach estimates a quantitative measure for the PFM similarity of two proteins, based on various features derived from their protein sequences. The method is trained and tested on a dataset containing 1 239 TFs with known DNA-binding specificity, and used to predict specific DNA target motifs for 645 TFs with high accuracy

Accuracy of five electronic foramen locators with different operating systems: an ex vivo study

OBJECTIVE: The aim of this study was to evaluate, ex vivo, the precision of five electronic root canal length measurement devices (ERCLMDs) with different operating systems: the Root ZX, Mini Apex Locator, Propex II, iPex, and RomiApex A-15, and the possible influence of the positioning of the instrument tips short of the apical foramen. MATERIAL AND METHODS: Forty-two mandibular bicuspids had their real canal lengths (RL) previously determined. Electronic measurements were performed 1.0 mm short of the apical foramen (-1.0), followed by measurements at the apical foramen (0.0). The data resulting from the comparison of the ERCLMD measurements and the RL were evaluated by the Wilcoxon and Friedman tests at a significance level of 5%. RESULTS: Considering the measurements performed at 0.0 and -1.0, the precision rates for the ERCLMDs were: 73.5% and 47.1% (Root ZX), 73.5% and 55.9% (Mini Apex Locator), 67.6% and 41.1% (Propex II), 61.7% and 44.1% (iPex), and 79.4% and 44.1% (RomiApex A-15), respectively, considering ±0.5 mm of tolerance. Regarding the mean discrepancies, no differences were observed at 0.0; however, in the measurements at -1.0, the iPex, a multi-frequency ERCLMD, had significantly more discrepant readings short of the apical foramen than the other devices, except for the Propex II, which had intermediate results. When the ERCLMDs measurements at -1.0 were compared with those at 0.0, the Propex II, iPex and RomiApex A-15 presented significantly higher discrepancies in their readings. CONCLUSIONS: Under the conditions of the present study, all the ERCLMDs provided acceptable measurements at the 0.0 position. However, at the -1.0 position, the ERCLMDs had a lower precision, with statistically significant differences for the Propex II, iPex, and RomiApex A-15

Large expert-curated database for benchmarking document similarity detection in biomedical literature search

Document recommendation systems for locating relevant literature have mostly relied on methods developed a decade ago. This is largely due to the lack of a large offline gold-standard benchmark of relevant documents that cover a variety of research fields such that newly developed literature search techniques can be compared, improved and translated into practice. To overcome this bottleneck, we have established the RElevant LIterature SearcH consortium consisting of more than 1500 scientists from 84 countries, who have collectively annotated the relevance of over 180 000 PubMed-listed articles with regard to their respective seed (input) article/s. The majority of annotations were contributed by highly experienced, original authors of the seed articles. The collected data cover 76% of all unique PubMed Medical Subject Headings descriptors. No systematic biases were observed across different experience levels, research fields or time spent on annotations. More importantly, annotations of the same document pairs contributed by different scientists were highly concordant. We further show that the three representative baseline methods used to generate recommended articles for evaluation (Okapi Best Matching 25, Term Frequency–Inverse Document Frequency and PubMed Related Articles) had similar overall performances. Additionally, we found that these methods each tend to produce distinct collections of recommended articles, suggesting that a hybrid method may be required to completely capture all relevant articles. The established database server located at https://relishdb.ict.griffith.edu.au is freely available for the downloading of annotation data and the blind testing of new methods. We expect that this benchmark will be useful for stimulating the development of new powerful techniques for title and title/abstract-based search engines for relevant articles in biomedical research

Biomaterials for pulp-capping: calcium-ions pulpflux through dentin in indirect pulp-capping.

Indirect pulp-cap is a procedure following caries removal in which a material is placed on remaining (infected/carious) dentin thickness (RDT). Ca(OH)2-&-CaO-based materials and calcium-silicate MTA (Mineral Trioxide Aggregate) cements are conventionally used for indirect pulp-capping. Ca ions induce the proliferation/differentiation of human dental pulp cells into odontoblasts, the formation of a tertiary/reparative dentin bridge, the mineralization of dentin. OH ions (alkaline pH) create an antibacterial/bacteriostatic environment, cause an inflammatory reaction with the formation of reparative dentin and promote the formation of hydroxyapatite. Objective: To verify the ability of four commercial Ca-containing materials for pulp-capping to generate a Ca ions outflux through dentin in indirect pulp-capping in vitro. Method: Pulpdent Paste Kit (Pulpdent Corp.-USA; lot.n.110411), Dycal (Dentsply-USA; lot.n.110630), Tech Biosealer capping (Isasan-Italy; lot.n.E10046) and ProRoot MTA (Dentsply-USA; lot.n.09001920) were tested. Standardized cavities were prepared (by a high-speed inverted-cone flat-end diamond bur, diam. 0.2mm) in erupted sound human molars. Pulp was removed. EDTA17% was used to remove the smear layer. The outer-surface of samples was covered by nail varnish to hamper Ca-releasing from the dental tissues. Indirect pulp-capping was performed on RDT (1±0.2mm thick) by layering one test material, a glass ionomer cement, a composite, and nail varnish. Samples were maintained deionized water (10ml, 37°C). After 3,24-hours and 3,7,14-days the leached Ca-&-OH ions were measured using ion-selective electrodes. Result: Ca-releasing was Tech Biosealer capping > ProRoot MTA > Pulpdent > Dycal. Only Tech Biosealer capping released significant amount of calcium. Alkaline pH of soaking water was observed after 24-hours. Conclusion: Only Tech Biosealer capping was able to create a calcium ions outflux through dentin. In clinical situation, the wet surgical site owing to the presence of biological fluids (blood, exudates, dentinal fluid) may supply calcium ions, with beneficial effects on pulp cells, tissue regeneration (reparative dentin formation) and clinical healing

On the multiwavelength variability of Mrk 110: two components acting at different time-scales

We present the first intensive continuum reverberation mapping study of the high accretion-rate Seyfert galaxy Mrk 110. The source was monitored almost daily for more than 200 d with the Swift X-ray and ultraviolet (UV)/optical telescopes, supported by ground-based observations from Las Cumbres Observatory, the Liverpool Telescope, and the Zowada Observatory, thus extending the wavelength coverage to 9100 Å. Mrk 110 was found to be significantly variable at all wavebands. Analysis of the intraband lags reveals two different behaviours, depending on the time-scale. On time-scales shorter than 10 d the lags, relative to the shortest UV waveband (∼1928 Å), increase with increasing wavelength up to a maximum of ∼2 d lag for the longest waveband (∼9100 Å), consistent with the expectation from disc reverberation. On longer time-scales, however, the g-band lags the Swift BAT hard X-rays by ∼10 d, with the z-band lagging the g-band by a similar amount, which cannot be explained in terms of simple reprocessing from the accretion disc. We interpret this result as an interplay between the emission from the accretion disc and diffuse continuum radiation from the broad-line region