A dynamic compositional equilibrium governs mRNA recognition by eIF3.

Eukaryotic translation initiation factor (eIF) 3 is a multi-subunit protein complex that binds both ribosomes and messenger RNAs (mRNAs) in order to drive a diverse set of mechanistic steps during translation. Despite its importance, a unifying framework explaining how eIF3 performs these numerous activities is lacking. Using single-molecule light scattering microscopy, we demonstrate that Saccharomyces cerevisiae eIF3 is an equilibrium mixture of the full complex, subcomplexes, and subunits. By extending our microscopy approach to an in vitro reconstituted eIF3 and complementing it with biochemical assays, we define the subspecies comprising this equilibrium and show that, rather than being driven by the full complex, mRNA binding by eIF3 is instead driven by the eIF3a subunit within eIF3a-containing subcomplexes. Our findings provide a mechanistic model for the role of eIF3 in the mRNA recruitment step of translation initiation and establish a mechanistic framework for explaining and investigating the other activities of eIF3.

The mechanism of mRNA activation.

During translation initiation, messenger RNA molecules must be identified and activated for loading into a ribosome. In this rate-limiting step, the heterotrimeric protein eukaryotic initiation factor eIF4F must recognize and productively interact with the 7-methylguanosine cap at the 5' end of the messenger RNA and subsequently activate the message. Despite its fundamental, regulatory role in gene expression, the molecular events underlying cap recognition and messenger RNA activation remain mysterious. Here, we generate a unique, single-molecule fluorescence imaging system to interrogate the dynamics with which eIF4F discriminates productive and non-productive locations on full-length, native messenger RNA molecules. At the single-molecule level, we observe stochastic sampling of eIF4F along the length of the messenger RNA and identify allosteric communication between the eIF4F subunits which ultimately drive cap-recognition and subsequent activation of the message. Our experiments uncover novel functions for each subunit of eIF4F and we conclude by presenting a model for messenger RNA activation which precisely defines the composition of the activated message. This model provides a general framework for understanding how messenger RNA molecules may be discriminated from one another, and how other RNA-binding proteins may control the efficiency of translation initiation.

Effects of Porting Essie Tokenization and Normalization to Solr.

Search for information is now an integral part of healthcare. Searches are enabled by search engines whose objective is to efficiently retrieve the relevant information for the user query. When it comes to retrieving biomedical text and literature, Essie search engine developed at the National Library of Medicine (NLM) performs exceptionally well. However, Essie is a software system developed for NLM that has ceased development and support. On the other hand, Solr is a popular opensource enterprise search engine used by many of the world's largest internet sites, offering continuous developments and improvements along with the state-of-the-art features. In this paper, we present our approach to porting the key features of Essie and developing custom components to be used in Solr. We demonstrate the effectiveness of the added components on three benchmark biomedical datasets. The custom components may aid the community in improving search methods for biomedical text retrieval.

Biogenesis of the Flagellar Switch Complex in Escherichia coli: Formation of Sub-Complexes Independently of the Basal-Body MS-Ring.

Direction switching in the flagellar motor of Escherichia coli is under the control of a complex on the rotor formed from the proteins FliG, FliM, and FliN. FliG lies at the top of the switch complex (i.e., nearest the membrane) and is arranged with its C-terminal domain (FliGC) resting on the middle domain (FliGM) of the neighboring subunit. This organization requires the protein to adopt an open conformation that exposes the surfaces engaging in intersubunit FliGC/FliGM contacts. In a recent study, Baker and coworkers [13] obtained evidence that FliG in the cytosol is monomeric and takes on a more compact conformation, with FliGC making intramolecular contact with FliGM of the same subunit. In the present work, we examine the conformational preferences and interactions of FliG through in vivo crosslinking experiments in cells that lack either all other flagellar proteins or just the MS-ring protein FliF. The results indicate that FliG has a significant tendency to form multimers independently of other flagellar components. The multimerization of FliG is promoted by FliF and also by FliM. FliM does not multimerize efficiently by itself but does so in the presence of FliG. Thus, pre-assemblies of the switch-complex proteins can form in the cytosol and might function as intermediates in assembly.

Architecture of the Flagellar Switch Complex of Escherichia coli: Conformational Plasticity of FliG and Implications for Adaptive Remodeling.

Structural models of the complex that regulates the direction of flagellar rotation assume either ~34 or ~25 copies of the protein FliG. Support for ~34 came from crosslinking experiments identifying an intersubunit contact most consistent with that number; support for ~25 came from the observation that flagella can assemble and rotate when FliG is genetically fused to FliF, for which the accepted number is ~25. Here, we have undertaken crosslinking and other experiments to address more fully the question of FliG number. The results indicate a copy number of ~25 for FliG. An interaction between the C-terminal and middle domains, which has been taken to support a model with ~34 copies, is also supported. To reconcile the interaction with a FliG number of ~25, we hypothesize conformational plasticity in an interdomain segment of FliG that allows some subunits to bridge gaps created by the number mismatch. This proposal is supported by mutant phenotypes and other results indicating that the normally helical segment adopts a more extended conformation in some subunits. The FliG amino-terminal domain is organized in a regular array with dimensions matching a ring in the upper part of the complex. The model predicts that FliG copy number should be tied to that of FliF, whereas FliM copy number can increase or decrease according to the number of FliG subunits that adopt the extended conformation. This has implications for the phenomenon of adaptive switch remodeling, in which the FliM copy number varies to adjust the bias of the switch.

The ClinicalTrials.gov results database--update and key issues.

BACKGROUND: The ClinicalTrials.gov trial registry was expanded in 2008 to include a database for reporting summary results. We summarize the structure and contents of the results database, provide an update of relevant policies, and show how the data can be used to gain insight into the state of clinical research. METHODS: We analyzed ClinicalTrials.gov data that were publicly available between September 2009 and September 2010. RESULTS: As of September 27, 2010, ClinicalTrials.gov received approximately 330 new and 2000 revised registrations each week, along with 30 new and 80 revised results submissions. We characterized the 79,413 registry and 2178 results of trial records available as of September 2010. From a sample cohort of results records, 78 of 150 (52%) had associated publications within 2 years after posting. Of results records available publicly, 20% reported more than two primary outcome measures and 5% reported more than five. Of a sample of 100 registry record outcome measures, 61% lacked specificity in describing the metric used in the planned analysis. In a sample of 700 results records, the mean number of different analysis populations per study group was 2.5 (median, 1; range, 1 to 25). Of these trials, 24% reported results for 90% or less of their participants. CONCLUSIONS: ClinicalTrials.gov provides access to study results not otherwise available to the public. Although the database allows examination of various aspects of ongoing and completed clinical trials, its ultimate usefulness depends on the research community to submit accurate, informative data.

Knowledge-based methods to help clinicians find answers in MEDLINE.

OBJECTIVES: Large databases of published medical research can support clinical decision making by providing physicians with the best available evidence. The time required to obtain optimal results from these databases using traditional systems often makes accessing the databases impractical for clinicians. This article explores whether a hybrid approach of augmenting traditional information retrieval with knowledge-based methods facilitates finding practical clinical advice in the research literature. DESIGN: Three experimental systems were evaluated for their ability to find MEDLINE citations providing answers to clinical questions of different complexity. The systems (SemRep, Essie, and CQA-1.0), which rely on domain knowledge and semantic processing to varying extents, were evaluated separately and in combination. Fifteen therapy and prevention questions in three categories (general, intermediate, and specific questions) were searched. The first 10 citations retrieved by each system were randomized, anonymized, and evaluated on a three-point scale. The reasons for ratings were documented. MEASUREMENTS: Metrics evaluating the overall performance of a system (mean average precision, binary preference) and metrics evaluating the number of relevant documents in the first several presented to a physician were used. RESULTS: Scores (mean average precision = 0.57, binary preference = 0.71) for fusion of the retrieval results of the three systems are significantly (p < 0.01) better than those for any individual system. All three systems present three to four relevant citations in the first five for any question type. CONCLUSION: The improvements in finding relevant MEDLINE citations due to knowledge-based processing show promise in assisting physicians to answer questions in clinical practice.

Issues in the registration of clinical trials.

Public concerns about the perils associated with incomplete or delayed reporting of results from clinical trials has heightened interest in trial registries and results databases. Here we review the current status of trial registration efforts and the challenges in developing a comprehensive system of trial registration and reporting of results. ClinicalTrials.gov, the largest trial registry with 36 249 trials from approximately 140 countries, has procedures in place to help ensure that records are valid and informative. Key challenges include the need to minimize inadvertent duplicate registrations, to ensure that interventions have unambiguous names, and to have a search engine that identifies all trials that meet a user's specifications. Recent policy initiatives have called for the development of a database of trial results. Several issues confound the implementation of such a database, including the lack of an accepted format or process for providing summaries of trial results to the public and concerns about disseminating data in the absence of independent scientific review.

Essie: a concept-based search engine for structured biomedical text.

This article describes the algorithms implemented in the Essie search engine that is currently serving several Web sites at the National Library of Medicine. Essie is a phrase-based search engine with term and concept query expansion and probabilistic relevancy ranking. Essie's design is motivated by an observation that query terms are often conceptually related to terms in a document, without actually occurring in the document text. Essie's performance was evaluated using data and standard evaluation methods from the 2003 and 2006 Text REtrieval Conference (TREC) Genomics track. Essie was the best-performing search engine in the 2003 TREC Genomics track and achieved results comparable to those of the highest-ranking systems on the 2006 TREC Genomics track task. Essie shows that a judicious combination of exploiting document structure, phrase searching, and concept based query expansion is a useful approach for information retrieval in the biomedical domain.

Trial Registration at ClinicalTrials.gov between May and October 2005.

BACKGROUND: Clinical trial registration allows interested parties to obtain information about ongoing and completed trials, but there are few data indicating the quality of the information provided during the registration process. We used information in the publicly available ClinicalTrials.gov database to describe patterns of trial registration before and after the implementation by journal editors of a new policy requiring registration as a prerequisite for publication. METHODS: We reviewed ClinicalTrials.gov records to determine patterns of completion of the "Intervention Name" and "Primary Outcome Measure" data fields for trials registered on May 20 and October 11, 2005, and for trials registered during the interval between these two dates, inclusively. RESULTS: During the interval studied, the number of registrations in ClinicalTrials.gov increased by 73 percent from 13,153 to 22,714. The percentage of interventional trials registered by industry with nonspecific Intervention Name entries (attributable to four drug companies) decreased from 10 percent to 2 percent; all other industry and nonindustry records contained specific entries in this field. Of the 2670 studies registered by industry between the two dates, 76 percent provided information in the Primary Outcome Measure field, although these entries varied markedly in their degree of specificity. In the remaining 24 percent of the records, this field was blank. CONCLUSIONS: During the summer of 2005, there were large increases in the number of clinical trial registrations. Overall, the data contained in records were more complete in October than they were in May, but there still is room for substantial improvement.

Strategies for supporting consumer health information seeking.

Despite a growing number of available Web-based health information resources, consumers continue to face a variety of barriers as they attempt to access these resources. Developing a system that appropriately responds to user queries poses several challenges. Guided by an earlier study that analyzed a large number of queries submitted to ClinicalTrials.gov, we developed a variety of techniques to assist user information seeking. We tested the efficacy of these techniques by submitting the original user queries to our new search engine to determine if these techniques would result in better system performance. Overall, the number of query failures was reduced, but the largest improvement was found in the system's query suggestion capability. For a subset of query failures, the current system was able to cut the earlier failure rate almost in half, in most cases providing a suggestion rather than directly finding records. The techniques described here provide a new approach for responding to user queries. The techniques are tolerant of certain types of errors and provide feedback to assist users in reformulating their queries.

Design, implementation and management of a web-based data entry system for ClinicalTrials.gov.

We describe the development and deployment of a web-based authoring capability, the first implementation of which is used for data entry and management in support of the ClinicalTrials.gov web site. The system facilitates efficient collection of summary protocol information from multiple geographically-dispersed organizations. We explain the motivation for developing this capability, and cite critical design goals. We then describe system design, implementation and operation, focusing on essential aspects of each. We conclude with a summary of the extent to which we met our stated objectives.