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1.  BioArchitecture 
Bioarchitecture  2012;2(6):200-203.
BioArchitecture is a term used to describe the organization and regulation of biological space. It applies to the principles which govern the structure of molecules, polymers and mutiprotein complexes, organelles, membranes and their organization in the cytoplasm and the nucleus. It also covers the integration of cells into their three dimensional environment at the level of cell-matrix, cell-cell interactions, integration into tissue/organ structure and function and finally into the structure of the organism. This review will highlight studies at all these levels which are providing a new way to think about the relationship between the organization of biological space and the function of biological systems.
PMCID: PMC3527313  PMID: 23267413
actin; cytoskeleton; microtubules; intermediate filaments; nuclear structure; protein folding; isoform sorting
2.  Welcome to BioArchitecture 2013 
Bioarchitecture  2013;3(1):1.
The first major hurdle for any new journal is to achieve acceptance into Medline/PubMed. I am pleased to report that BioArchitecture was accepted into Medline/PubMed in August 2012. This means that accepted manuscripts are immediately visible through their listing on PubMed. This is very welcome news to contributors to the journal and makes the journal a more attractive destination for publication of new research findings.
PMCID: PMC3639238
3.  Shaping muscle bioarchitecture for the fin to limb transition 
Bioarchitecture  2012;2(3):98-103.
Our recent paper examined how pelvic fins and their musculature form developmentally and how these mechanisms have evolved within the vertebrate lineage, a process fundamental to the tetrapod transition. The transition from the water onto the land is among one of the most well studied steps in the evolutionary history of vertebrates, yet the genetic basis of this evolutionary transition is little studied and ill-defined. The advent of these terrestrial species resulted in a shift in locomotor strategies from the rhythmic undulating muscles of the fish body to a reliance upon powerful weight bearing muscles of the limbs to generate movement. We demonstrated that the pelvic fin muscles of bony fish are generated by a mechanism that has features of both of limb/fin muscle formation in tetrapods and primitive cartilaginous fish. We hypothesize that the adoption of the fully derived mode of hindlimb muscle formation, was a further modification of the mode of development deployed to generate pelvic fin muscles, a shift in overall muscle bioarchitecture we believe was critical to the success of the tetrapod transition.
PMCID: PMC3414388  PMID: 22880150
muscle; evolution; fin; limb; zebrafish; tetrapod
4.  GNU polyxmass: a software framework for mass spectrometric simulations of linear (bio-)polymeric analytes 
BMC Bioinformatics  2006;7:226.
Nowadays, a variety of (bio-)polymers can be analyzed by mass spectrometry. The detailed interpretation of the spectra requires a huge number of "hypothesis cycles", comprising the following three actions 1) put forth a structural hypothesis, 2) test it, 3) (in)validate it. This time-consuming and painstaking data scrutiny is alleviated by using specialized software tools. However, all the software tools available to date are polymer chemistry-specific. This imposes a heavy overhead to researchers who do mass spectrometry on a variety of (bio-)polymers, as each polymer type will require a different software tool to perform data simulations and analyses. We developed a software to address the lack of an integrated software framework able to deal with different polymer chemistries.
The GNU polyxmass software framework performs common (bio-)chemical simulations–along with simultaneous mass spectrometric calculations–for any kind of linear bio-polymeric analyte (DNA, RNA, saccharides or proteins). The framework is organized into three modules, all accessible from one single binary program. The modules let the user to 1) define brand new polymer chemistries, 2) perform quick mass calculations using a desktop calculator paradigm, 3) graphically edit polymer sequences and perform (bio-)chemical/mass spectrometric simulations. Any aspect of the mass calculations, polymer chemistry reactions or graphical polymer sequence editing is configurable.
The scientist who uses mass spectrometry to characterize (bio-)polymeric analytes of different chemistries is provided with a single software framework for his data prediction/analysis needs, whatever the polymer chemistry being involved.
PMCID: PMC1524818  PMID: 16643644
5.  Soluble Polymer Carriers for the Treatment of Cancer: The Importance of Molecular Architecture 
Accounts of chemical research  2009;42(8):1141-1151.
Chemotherapy can destroy tumors and arrest cancer progress. Unfortunately, severe side effects—treatment is usually a series of injections of highly toxic drugs—often restrict the frequency and size of dosages, much to the detriment of tumor inhibition. Most chemotherapeutic drugs have pharmacokinetic profiles with tremendous potential for improvement.
Water-soluble polymers offer the potential to increase drug circulation time, improve drug solubility, prolong drug residence time in a tumor, and reduce toxicity. Cytotoxic drugs that are covalently attached to water-soluble polymers via reversible linkages more effectively target tumor tissue than the drugs alone. Macromolecules passively target solid tumor tissue through a combination of reduced renal clearance and exploitation of the enhanced permeation and retention (EPR) effect, which prevails for fast-growing tumors.
Effective drug delivery involves a balance between (i) elimination of the polymeric drug conjugate from the bloodstream by the kidneys, liver, and other organs and (ii) movement of the drug out of the blood vasculature and into the tumor (that is, extravasation). Polymers are eliminated in the kidney by filtration through pores with a size comparable to the hydrodynamic diameter of the polymer; in contrast, the openings in the blood vessel structures that traverse tumors are an order of magnitude greater than the diameter of the polymer. Thus, features that may broadly be grouped as the “molecular architecture” of the polymer—such as its hydrodynamic volume (or molecular weight), molecular conformation, chain flexibility, branching, and location of the attached drug—can greatly impact elimination of the polymer from the body through the kidney but have a much smaller effect on the extravasation of the polymer into the tumor. Molecular architecture can in theory be adjusted to assert essentially independent control over elimination and extravasation. Understanding how molecular architecture affects passage of a polymer through a pore is therefore essential for designing polymer drug carriers that are effective in passively delivering a drug payload while conforming to the requirement that the polymers must eventually be eliminated from the body.
In this Account, we discuss examples from in vivo studies that demonstrate how polymer architectural features impact the renal filtration of a polymer as well as tumor penetration and tumor accumulation. In brief, features that inhibit passage of a polymer through a pore—such as higher molecular weight, decreased flexibility, and an increased number of polymer chain ends—help prevent elimination of the polymer by the kidneys and can improve blood circulation times and tumor accumulation, thus improving therapeutic effectiveness.
PMCID: PMC2759385  PMID: 19555070
6.  Biosurveillance Adaptable Framework for Teaming, Exploration and Reuse (BioAFTER) 
Next-generation software environments for disease surveillance will need to have several important characteristics, among which are collaboration and search and discovery features, access to various data sets, and a variety of analytic methods. However, perhaps the most important feature is the least often mentioned - the ability to have the system adapt over time without high reengineering cost. The public health community cannot afford software redesigns every few years as data sets expand, analysis needs evolve, and software deficiencies are exposed. In addition to the need to adapt an environment over longer time periods, epidemiologists have high variability in their day-to-day needs that require adaptability over short time periods as well. Each outbreak or health situation has unique aspects, and analysts need to be able to bring in data and methods unique to that situation that may not be easily anticipated a priori.
The most common approach to increasing reusability and decreasing upgrade costs are open architecture software frameworks such as Service-Oriented Architectures (SOAs). If well implemented, SOAs can significantly reduce software upgrade costs by allowing services (a software module) to be easily swapped out for improvements or supplemented with additional services. SOAs can help with long-term adaptability, but are not useful in short-term adaptability, since the software development team must be engaged in each cycle. Another approach is to include an App Store. Unfortunately, App Stores for government use have often been disappointing. Apps can tend to be quite simple, and even slight changes from what is programmed - a predictable situation with the variability seen in disease surveillance realm - will result in an epidemiologist having to get a software developer to make them a new App.
Instead of the power for adaptability remaining solely in the control of software developers, that power needs to also be in the hands of the users themselves. The BioAFTER project builds upon SOA and App Store concepts by allowing Apps to be strung together in unique combinations, according to the problem of the day. As examples, these Apps can be data access programs, data quality editing, algorithms of various complexity, or reporting and visualization modules. The App Store feature allows software developers, including the public health academic community, to add new methods to the environment, while the composability feature allows ad-hoc combinations of Apps to suit particular situations.
The user composability feature would be of limited value without collaboration features in the environment. We expect that only a subset of users will make the effort to do composition. The rest will want to learn from those “super users”. The BioAFTER environment allows data, Apps (including new Apps created by compositions of other Apps), and analysis results to be shared with the entire epidemiological community, or with a set of “friends” or people with common interests. Analysts who use various Apps can rate their value. The environment is intended to allow search and discovery of Apps, expertise, or even people who may have needed expertise or experience with similar situations (these features are in development).
The motivations and features of the BioAFTER environment will be described at the ISDS Conference, and a brief demonstration of the capabilities on an example use case will be given.
PMCID: PMC3692856
biosurveillance; collaboration; discovery; composition; adaptability
7.  An ex situ evaluation of TBA- and MTBE-baited bio-traps 
Water research  2012;46(12):3879-3888.
Aquifer microbial communities can be investigated using Bio-traps® (“bio-traps”), passive samplers containing Bio-Sep® beads (“bio-beads”) that are deployed in monitoring wells to be colonized by bacteria delivered via groundwater flow through the well. When bio-beads are “baited” with organic contaminants enriched in 13C, stable isotope probing allows assessment of the composition and activity of the microbial community. This study used an ex situ system fed by groundwater continuously extracted from an adjacent monitoring well within an experimentally-created aerobic zone treating a tert-butyl alcohol (TBA) plume. The goal was to evaluate aspects of bio-trap performance that cannot be studied quantitatively in situ. The measured groundwater flow through a bio-trap housing suggests that such traps might typically “sample” about 1.8 L per month. The desorption of TBA or methyl tert-butyl ether (MTBE) bait from bio-traps during a typical deployment duration of 6 weeks was approximately 90% and 45%, respectively, of the total initial bait load, with initially high rate of mass loss that decreased markedly after a few days. The concentration of TBA in groundwater flowing by the TBA-baited bio-beads was estimated to be as high as 3400 mg/L during the first few days, which would be expected to inhibit growth of TBA-degrading microbes. Initial inhibition was also implied for the MTBE-baited bio-trap, but at lower concentrations and for a shorter time. After a few days, concentrations in groundwater flowing through the bio-traps dropped below inhibitory concentrations but remained 4–5 orders of magnitude higher than TBA or MTBE concentrations within the aquifer at the experimental site. Desorption from the bio-beads during ex situ deployment occurred at first as predicted by prior sorption analyses of bio-beads but with apparent hysteresis thereafter, possibly due to mass transfer limitations caused by colonizing microbes. These results suggest that TBA- or MTBE-baited bio-traps could be baited at lower initial total mass loading with no detriment to trapping ability. The bio-traps were able to collect detectable amounts of microbial DNA and thus allow some insight into the sparse microbial community present in the aquifer during remediation of the low concentration plume.
PMCID: PMC3582699  PMID: 22621895
MTBE; TBA; Bio-trap; In situ microcosm; Biodegradation; Stable isotope probing
8.  Molecular cloning of a novel bioH gene from an environmental metagenome encoding a carboxylesterase with exceptional tolerance to organic solvents 
BMC Biotechnology  2013;13:13.
BioH is one of the key enzymes to produce the precursor pimeloyl-ACP to initiate biotin biosynthesis de novo in bacteria. To date, very few bioH genes have been characterized. In this study, we cloned and identified a novel bioH gene, bioHx, from an environmental metagenome by a functional metagenomic approach. The bioHx gene, encoding an enzyme that is capable of hydrolysis of p-nitrophenyl esters of fatty acids, was expressed in Escherichia coli BL21 using the pET expression system. The biochemical property of the purified BioHx protein was also investigated.
Screening of an unamplified metagenomic library with a tributyrin-containing medium led to the isolation of a clone exhibiting lipolytic activity. This clone carried a 4,570-bp DNA fragment encoding for six genes, designated bioF, bioHx, fabG, bioC, orf5 and sdh, four of which were implicated in the de novo biotin biosynthesis. The bioHx gene encodes a protein of 259 aa with a calculated molecular mass of 28.60 kDa, displaying 24-39% amino acid sequence identity to a few characterized bacterial BioH enzymes. It contains a pentapeptide motif (Gly76-Trp77-Ser78-Met79-Gly80) and a catalytic triad (Ser78-His230-Asp202), both of which are characteristic for lipolytic enzymes. BioHx was expressed as a recombinant protein and characterized. The purified BioHx protein displayed carboxylesterase activity, and it was most active on p-nitrophenyl esters of fatty acids substrate with a short acyl chain (C4). Comparing BioHx with other known BioH proteins revealed interesting diversity in their sensitivity to ionic and nonionic detergents and organic solvents, and BioHx exhibited exceptional resistance to organic solvents, being the most tolerant one amongst all known BioH enzymes. This ascribed BioHx as a novel carboxylesterase with a strong potential in industrial applications.
This study constituted the first investigation of a novel bioHx gene in a biotin biosynthetic gene cluster cloned from an environmental metagenome. The bioHx gene was successfully cloned, expressed and characterized. The results demonstrated that BioHx is a novel carboxylesterase, displaying distinct biochemical properties with strong application potential in industry. Our results also provided the evidence for the effectiveness of functional metagenomic approach for identifying novel bioH genes from complex ecosystem.
PMCID: PMC3583802  PMID: 23413993
BioH; Biotin biosynthetic pathway; Carboxylesterase; Metagenome; Aqueous environment
9.  Negated bio-events: analysis and identification 
BMC Bioinformatics  2013;14:14.
Negation occurs frequently in scientific literature, especially in biomedical literature. It has previously been reported that around 13% of sentences found in biomedical research articles contain negation. Historically, the main motivation for identifying negated events has been to ensure their exclusion from lists of extracted interactions. However, recently, there has been a growing interest in negative results, which has resulted in negation detection being identified as a key challenge in biomedical relation extraction. In this article, we focus on the problem of identifying negated bio-events, given gold standard event annotations.
We have conducted a detailed analysis of three open access bio-event corpora containing negation information (i.e., GENIA Event, BioInfer and BioNLP’09 ST), and have identified the main types of negated bio-events. We have analysed the key aspects of a machine learning solution to the problem of detecting negated events, including selection of negation cues, feature engineering and the choice of learning algorithm. Combining the best solutions for each aspect of the problem, we propose a novel framework for the identification of negated bio-events. We have evaluated our system on each of the three open access corpora mentioned above. The performance of the system significantly surpasses the best results previously reported on the BioNLP’09 ST corpus, and achieves even better results on the GENIA Event and BioInfer corpora, both of which contain more varied and complex events.
Recently, in the field of biomedical text mining, the development and enhancement of event-based systems has received significant interest. The ability to identify negated events is a key performance element for these systems. We have conducted the first detailed study on the analysis and identification of negated bio-events. Our proposed framework can be integrated with state-of-the-art event extraction systems. The resulting systems will be able to extract bio-events with attached polarities from textual documents, which can serve as the foundation for more elaborate systems that are able to detect mutually contradicting bio-events.
PMCID: PMC3561152  PMID: 23323936
10.  DNA Sequences at a Glance 
PLoS ONE  2013;8(11):e79922.
Data summarization and triage is one of the current top challenges in visual analytics. The goal is to let users visually inspect large data sets and examine or request data with particular characteristics. The need for summarization and visual analytics is also felt when dealing with digital representations of DNA sequences. Genomic data sets are growing rapidly, making their analysis increasingly more difficult, and raising the need for new, scalable tools. For example, being able to look at very large DNA sequences while immediately identifying potentially interesting regions would provide the biologist with a flexible exploratory and analytical tool. In this paper we present a new concept, the “information profile”, which provides a quantitative measure of the local complexity of a DNA sequence, independently of the direction of processing. The computation of the information profiles is computationally tractable: we show that it can be done in time proportional to the length of the sequence. We also describe a tool to compute the information profiles of a given DNA sequence, and use the genome of the fission yeast Schizosaccharomyces pombe strain 972 h− and five human chromosomes 22 for illustration. We show that information profiles are useful for detecting large-scale genomic regularities by visual inspection. Several discovery strategies are possible, including the standalone analysis of single sequences, the comparative analysis of sequences from individuals from the same species, and the comparative analysis of sequences from different organisms. The comparison scale can be varied, allowing the users to zoom-in on specific details, or obtain a broad overview of a long segment. Software applications have been made available for non-commercial use at
PMCID: PMC3836782  PMID: 24278218
11.  Engineering BioBrick vectors from BioBrick parts 
The underlying goal of synthetic biology is to make the process of engineering biological systems easier. Recent work has focused on defining and developing standard biological parts. The technical standard that has gained the most traction in the synthetic biology community is the BioBrick standard for physical composition of genetic parts. Parts that conform to the BioBrick assembly standard are BioBrick standard biological parts. To date, over 2,000 BioBrick parts have been contributed to, and are available from, the Registry of Standard Biological Parts.
Here we extended the same advantages of BioBrick standard biological parts to the plasmid-based vectors that are used to provide and propagate BioBrick parts. We developed a process for engineering BioBrick vectors from BioBrick parts. We designed a new set of BioBrick parts that encode many useful vector functions. We combined the new parts to make a BioBrick base vector that facilitates BioBrick vector construction. We demonstrated the utility of the process by constructing seven new BioBrick vectors. We also successfully used the resulting vectors to assemble and propagate other BioBrick standard biological parts.
We extended the principles of part reuse and standardization to BioBrick vectors. As a result, myriad new BioBrick vectors can be readily produced from all existing and newly designed BioBrick parts. We invite the synthetic biology community to (1) use the process to make and share new BioBrick vectors; (2) expand the current collection of BioBrick vector parts; and (3) characterize and improve the available collection of BioBrick vector parts.
PMCID: PMC2373286  PMID: 18410688
Genetic association studies have rapidly become a major tool for identifying the genetic basis of common human diseases. The advent of cost-effective genotyping coupled with large collections of samples linked to clinical outcomes and quantitative traits now make it possible to systematically characterize genotype-phenotype relationships in diverse populations and extensive datasets. To capitalize on these advancements, the Epidemiologic Architecture for Genes Linked to Environment (EAGLE) project, as part of the collaborative Population Architecture using Genomics and Epidemiology (PAGE) study, accesses two collections: the National Health and Nutrition Examination Surveys (NHANES) and BioVU, Vanderbilt University’s biorepository linked to de-identified electronic medical records. We describe herein the workflows for accessing and using the epidemiologic (NHANES) and clinical (BioVU) collections, where each workflow has been customized to reflect the content and data access limitations of each respective source. We also describe the process by which these data are generated, standardized, and shared for meta-analysis among the PAGE study sites. As a specific example of the use of BioVU, we describe the data mining efforts to define cases and controls for genetic association studies of common cancers in PAGE. Collectively, the efforts described here are a generalized outline for many of the successful approaches that can be used in the era of high-throughput genotype-phenotype associations for moving biomedical discovery forward to new frontiers of data generation and analysis.
PMCID: PMC3579641  PMID: 23424142
13.  Interactions among the A and T Units of an ECF-Type Biotin Transporter Analyzed by Site-Specific Crosslinking 
PLoS ONE  2011;6(12):e29087.
Energy-coupling factor (ECF) transporters are a huge group of micronutrient importers in prokaryotes. They are composed of a substrate-specific transmembrane protein (S component) and a module consisting of a moderately conserved transmembrane protein (T component) and two ABC ATPase domains (A components). Modules of A and T units may be dedicated to a specific S component or shared by many different S units in an organism. The mode of subunit interactions in ECF transporters is largely unknown. BioMNY, the focus of the present study, is a biotin transporter with a dedicated AT module. It consists of the S unit BioY, the A unit BioM and the T unit BioN. Like all T units, BioN contains two three-amino-acid signatures with a central Arg residue in a cytoplasmic helical region. Our previous work had demonstrated a central role of the two motifs in T units for stability and function of BioMNY and other ECF transporters. Here we show by site-specific crosslinking of pairs of mono-cysteine variants that the Ala-Arg-Ser and Ala-Arg-Gly signatures in BioN are coupling sites to the BioM ATPases. Analysis of 64 BioN-BioM pairs uncovered interactions of both signatures predominantly with a segment of ∼13 amino acid residues C-terminal of the Q loop of BioM. Our results further demonstrate that portions of all BioN variants with single Cys residues in the two signatures are crosslinked to homodimers. This finding may point to a dimeric architecture of the T unit in BioMNY complexes.
PMCID: PMC3246461  PMID: 22216173
14.  Extracorporal hemodialysis with acute or decompensated chronical hepatic failure 
Conventional diagnostic procedures and therapy of acute liver failure (ALF) and acute-on-chronic liver failure (ACLF) focus on to identify triggering events of the acute deterioration of the liver function and to avoid them. Further objectives are to prevent the development respectively the progression of secondary organ dysfunctions or organ failure. Most of the times the endocrinological function of the liver can to a wide extent be compensated, but the removal of toxins can only marginally be substituted by conventional conservative therapy. To improve this component of the liver function is the main objective of extracorporal liver support systems. The following principles of liver support systems can be differentiated: Artificial systems, bioartifical systems and extracorporal liver perfusion systems. This HTA report focuses on artificial systems (e.g. BioLogic-DT/-DTPF, MARS, Prometheus), because only these approaches currently are relevant in the German health care system. In 2004 a category "Extracorporal liver assist device" was introduced in the list of "additional payments" in the German DRG-system, which makes reimbursement for hospitals using the technology in inpatient care possible, based on an hospital's individual contract with statutory sickness funds.
To report the present evidence and future research need on medical efficacy and economic effectiveness of extracorporal liver support devices for treatment of patients with ALF or ACLF based on published literature data. Are artificial liver support systems efficient and effective in the treatment of ALF or ACLF?
An extensive, systematic literature search in medical, economic, and HTA literature data bases was performed. Relevant data were extracted and synthesised.
Relevant controlled trials were detected for BioLogic-DT and MARS. No randomised controlled trial on Prometheus was found. None of the included studies on BioLogic-DT showed advantages of the technology compared with standard conventional therapy concerning survival, clinical scores or clinical surrogate parameter like laboratory tests of liver function. Some studies reported complications and side effects of BioLogic-DT. All studies were methodologically insufficient. Concerning the use of MARS overall five studies - three of them randomised - were identified. Two studies reported a significant higher 30d-survival after MARS compared to controls, one study showed a non-significant trend to a better survival probability after one year. The studies showed statistically significant advantages in severity of hepatic encephalopathy, routine lab tests and hemodynamic parameter of the MARS group. None of the studies reported relevant complications or side effects. Although the methodological quality of the studies is seen as slightly better than in the studies on BioLogic-DT, there are methodological limitations: The largest sample size of the randomised trials was twelve patients per group and the study population was highly selected. Because of the methodological limitations the results can hardly be generalised. Only two economic publications presenting analyses of MARS could be de-tected. One publication shows major methodological mistakes which make a further interpretation of the results impossible. The other publication presents an incremental cost-effectiveness of MARS of 29,719 EUR per life year gained after one year from a payer's perspective (German statutory sickness fund, neglecting the intervention costs because of lacking reimbursement at this time), respectively 79,075 EUR per life year gained from a societal perspective. Including health related quality of life aspects the incremental costs per QALY (Quality adjusted life years) gained were calculated to be 44,784 EUR from a payer's perspective respectively 119,162 EUR from a societal perspective. The authors state that prolonging the time horizon of the calculations would improve cost-effectiveness ratios. The limitations of the study design also limit the scientific evidence of the results.
The results of the detected publications do not give any evidence for a positive medical efficacy of BioLogic-DT. Concerning MARS there is some evidence for positive effects on 30d-survival, clinical parameter, and some lab tests, although the evidence is limited by the small number of studies and their methodological weakness. The currently strongly limited evidence shows a trend to an acceptable cost-effectiveness of MARS, although the results are based on only one non-randomised trial.
To give valid recommendations concerning the medical efficacy as well as the cost-effectiveness of artificial liver support systems further studies are necessary.
PMCID: PMC3011342  PMID: 21289959
extracorporal liver perfusion; hemodialysis; liver insufficiency; liver injury; hepatitic injury
15.  Systematic characterization of the conformation and dynamics of budding yeast chromosome XII 
The Journal of Cell Biology  2013;202(2):201-210.
Comprehensive analysis of the intranuclear territories and motion of budding yeast chromosome XII loci suggests that long-range chromosome architecture is mainly determined by the physical principles of polymers.
Chromosomes architecture is viewed as a key component of gene regulation, but principles of chromosomal folding remain elusive. Here we used high-throughput live cell microscopy to characterize the conformation and dynamics of the longest chromosome of Saccharomyces cerevisiae (XII). Chromosome XII carries the ribosomal DNA (rDNA) that defines the nucleolus, a major hallmark of nuclear organization. We determined intranuclear positions of 15 loci distributed every ∼100 kb along the chromosome, and investigated their motion over broad time scales (0.2–400 s). Loci positions and motions, except for the rDNA, were consistent with a computational model of chromosomes based on tethered polymers and with the Rouse model from polymer physics, respectively. Furthermore, rapamycin-dependent transcriptional reprogramming of the genome only marginally affected the chromosome XII internal large-scale organization. Our comprehensive investigation of chromosome XII is thus in agreement with recent studies and models in which long-range architecture is largely determined by the physical principles of tethered polymers and volume exclusion.
PMCID: PMC3718979  PMID: 23878273
16.  Introduction of customized inserts for streamlined assembly and optimization of BioBrick synthetic genetic circuits 
BioBrick standard biological parts are designed to make biological systems easier to engineer (e.g. assemble, manipulate, and modify). There are over 5,000 parts available in the Registry of Standard Biological Parts that can be easily assembled into genetic circuits using a standard assembly technique. The standardization of the assembly technique has allowed for wide distribution to a large number of users -- the parts are reusable and interchangeable during the assembly process. The standard assembly process, however, has some limitations. In particular it does not allow for modification of already assembled biological circuits, addition of protein tags to pre-existing BioBrick parts, or addition of non-BioBrick parts to assemblies.
In this paper we describe a simple technique for rapid generation of synthetic biological circuits using introduction of customized inserts. We demonstrate its use in Escherichia coli (E. coli) to express green fluorescent protein (GFP) at pre-calculated relative levels and to add an N-terminal tag to GFP. The technique uses a new BioBrick part (called a BioScaffold) that can be inserted into cloning vectors and excised from them to leave a gap into which other DNA elements can be placed. The removal of the BioScaffold is performed by a Type IIB restriction enzyme (REase) that recognizes the BioScaffold but cuts into the surrounding sequences; therefore, the placement and removal of the BioScaffold allows the creation of seamless connections between arbitrary DNA sequences in cloning vectors. The BioScaffold contains a built-in red fluorescent protein (RFP) reporter; successful insertion of the BioScaffold is, thus, accompanied by gain of red fluorescence and its removal is manifested by disappearance of the red fluorescence.
The ability to perform targeted modifications of existing BioBrick circuits with BioScaffolds (1) simplifies and speeds up the iterative design-build-test process through direct reuse of existing circuits, (2) allows incorporation of sequences incompatible with BioBrick assembly into BioBrick circuits (3) removes scar sequences between standard biological parts, and (4) provides a route to adapt synthetic biology innovations to BioBrick assembly through the creation of new parts rather than new assembly standards or parts collections.
PMCID: PMC3022552  PMID: 21172029
17.  Evolving BioAssay Ontology (BAO): modularization, integration and applications 
Journal of Biomedical Semantics  2014;5(Suppl 1):S5.
The lack of established standards to describe and annotate biological assays and screening outcomes in the domain of drug and chemical probe discovery is a severe limitation to utilize public and proprietary drug screening data to their maximum potential. We have created the BioAssay Ontology (BAO) project ( to develop common reference metadata terms and definitions required for describing relevant information of low-and high-throughput drug and probe screening assays and results. The main objectives of BAO are to enable effective integration, aggregation, retrieval, and analyses of drug screening data. Since we first released BAO on the BioPortal in 2010 we have considerably expanded and enhanced BAO and we have applied the ontology in several internal and external collaborative projects, for example the BioAssay Research Database (BARD). We describe the evolution of BAO with a design that enables modeling complex assays including profile and panel assays such as those in the Library of Integrated Network-based Cellular Signatures (LINCS). One of the critical questions in evolving BAO is the following: how can we provide a way to efficiently reuse and share among various research projects specific parts of our ontologies without violating the integrity of the ontology and without creating redundancies. This paper provides a comprehensive answer to this question with a description of a methodology for ontology modularization using a layered architecture. Our modularization approach defines several distinct BAO components and separates internal from external modules and domain-level from structural components. This approach facilitates the generation/extraction of derived ontologies (or perspectives) that can suit particular use cases or software applications. We describe the evolution of BAO related to its formal structures, engineering approaches, and content to enable modeling of complex assays and integration with other ontologies and datasets.
PMCID: PMC4108877  PMID: 25093074
18.  Linking Changes in Epithelial Morphogenesis to Cancer Mutations Using Computational Modeling 
PLoS Computational Biology  2010;6(8):e1000900.
Most tumors arise from epithelial tissues, such as mammary glands and lobules, and their initiation is associated with the disruption of a finely defined epithelial architecture. Progression from intraductal to invasive tumors is related to genetic mutations that occur at a subcellular level but manifest themselves as functional and morphological changes at the cellular and tissue scales, respectively. Elevated proliferation and loss of epithelial polarization are the two most noticeable changes in cell phenotypes during this process. As a result, many three-dimensional cultures of tumorigenic clones show highly aberrant morphologies when compared to regular epithelial monolayers enclosing the hollow lumen (acini). In order to shed light on phenotypic changes associated with tumor cells, we applied the bio-mechanical IBCell model of normal epithelial morphogenesis quantitatively matched to data acquired from the non-tumorigenic human mammary cell line, MCF10A. We then used a high-throughput simulation study to reveal how modifications in model parameters influence changes in the simulated architecture. Three parameters have been considered in our study, which define cell sensitivity to proliferative, apoptotic and cell-ECM adhesive cues. By mapping experimental morphologies of four MCF10A-derived cell lines carrying different oncogenic mutations onto the model parameter space, we identified changes in cellular processes potentially underlying structural modifications of these mutants. As a case study, we focused on MCF10A cells expressing an oncogenic mutant HER2-YVMA to quantitatively assess changes in cell doubling time, cell apoptotic rate, and cell sensitivity to ECM accumulation when compared to the parental non-tumorigenic cell line. By mapping in vitro mutant morphologies onto in silico ones we have generated a means of linking the morphological and molecular scales via computational modeling. Thus, IBCell in combination with 3D acini cultures can form a computational/experimental platform for suggesting the relationship between the histopathology of neoplastic lesions and their underlying molecular defects.
Author Summary
The majority of tumors arise in epithelial tissues that form monolayers of tightly packed cells enclosing the inner ductal or lobular cavities. Epithelial tumors (carcinomas) are associated with a disruption of epithelial architecture, such as filling of the inner lumen in the early stages of cancer, or the distortion of the ductal structure and spreading to the surrounding stroma in the subsequent invasive stages of tumor. Non-tumorigenic epithelial cells grown in 3D in vitro cultures form regular monolayered spheroids with hollow lumen (acini, Fig. 1a) resembling the architecture of normal epithelial cysts. In contrast, tumor cells taken from patients' biopsies and grown in 3D culture acquire various morphologies, often loosing the epithelial-like architecture. How these molecular defects produce such abnormal morphologies remains an open issue. We propose here to use the bio-mechanical model of epithelial morphogenesis, IBCell, to quantitatively investigate the phenotypical changes that the epithelial cells need to obtain in order to produce the aberrant morphologies observable experimentally and clinically. IBCell in combination with 3D acini cultures can form a computational/experimental platform for suggesting the link between histopathology of early tumors and underlying molecular defects.
PMCID: PMC2928778  PMID: 20865159
19.  Synthesis of a crosslinked branched polymer network in the interior of a protein cage 
A goal of biomimetic chemistry is to use the hierarchical architecture inherent in biological systems to guide the synthesis of functional three dimensional structures. Viruses and other highly symmetrical protein cage architectures provide defined scaffolds to initiate hierarchical structure assembly. Here we demonstrate that a crosslinked branched polymer can be initiated and synthesized within the interior cavity of a protein cage architecture. Creating this polymer network allows for the spatial control of pendant reactive sites and dramatically increases the stability of the cage architecture. This material was generated by the sequential coupling of multifunctional monomers using click chemistry to create a branched crosslinked polymer network. Analysis of polymer growth by mass spectrometry demonstrated that the polymer was initiated at the interior surface of the cage at genetically introduced cysteine reactive sites. The polymer grew as expected to generation 2.5 where it was limited by the size constraints of the cavity. The polymer network was fully crosslinked across protein subunits that make up the cage and extended the thermal stability for the cage to at least 120°C. The introduced reactive centers were shown to be active and their number density increased with increasing generation. This synthetic approach provides a new avenue for creating defined polymer networks, spatially constrained by a biological template.
PMCID: PMC2752147  PMID: 19317506
20.  Professional Uncertainty and Disempowerment Responding to Ethnic Diversity in Health Care: A Qualitative Study 
PLoS Medicine  2007;4(11):e323.
While ethnic disparities in health and health care are increasing, evidence on how to enhance quality of care and reduce inequalities remains limited. Despite growth in the scope and application of guidelines on “cultural competence,” remarkably little is known about how practising health professionals experience and perceive their work with patients from diverse ethnic communities. Using cancer care as a clinical context, we aimed to explore this with a range of health professionals to inform interventions to enhance quality of care.
Methods and Findings
We conducted a qualitative study involving 18 focus groups with a purposeful sample of 106 health professionals of differing disciplines, in primary and secondary care settings, working with patient populations of varying ethnic diversity in the Midlands of the UK. Data were analysed by constant comparison and we undertook processes for validation of analysis. We found that, as they sought to offer appropriate care, health professionals wrestled with considerable uncertainty and apprehension in responding to the needs of patients of ethnicities different from their own. They emphasised their perceived ignorance about cultural difference and were anxious about being culturally inappropriate, causing affront, or appearing discriminatory or racist. Professionals' ability to think and act flexibly or creatively faltered. Although trying to do their best, professionals' uncertainty was disempowering, creating a disabling hesitancy and inertia in their practice. Most professionals sought and applied a knowledge-based cultural expertise approach to patients, though some identified the risk of engendering stereotypical expectations of patients. Professionals' uncertainty and disempowerment had the potential to perpetuate each other, to the detriment of patient care.
This study suggests potential mechanisms by which health professionals may inadvertently contribute to ethnic disparities in health care. It identifies critical opportunities to empower health professionals to respond more effectively. Interventions should help professionals acknowledge their uncertainty and its potential to create inertia in their practice. A shift away from a cultural expertise model toward a greater focus on each patient as an individual may help.
From a qualitative study, Joe Kai and colleagues have identified opportunities to empower health professionals to respond more effectively to challenges in their work with patients from diverse ethnic communities.
Editors' Summary
Communities are increasingly diverse in terms of ethnicity (belonging to a group of people defined by social characteristics such as cultural tradition or national origin) and race (belonging to a group identified by inherited physical characteristics). Although health professionals and governments are striving to ensure that everybody has the same access to health care, there is increasing evidence of ethnic inequalities in health-care outcomes. Some of these inequalities reflect intrinsic differences between groups of people—Ashkenazi Jews, for example, often carry an altered gene that increases their chance of developing aggressive breast cancer. Often, however, these differences reflect inequalities in the health care received by different ethnic groups. To improve this situation, “cultural competence” has been promoted over recent years. Cultural competence is the development of skills by individuals and organizations that allow them to work effectively with people from different cultures. Health professionals are now taught about ethnic differences in health beliefs and practices, religion, and communication styles to help them provide the best service to all their patients.
Why Was This Study Done?
Numerous guidelines aim to improve cultural competency but little is known about how health professionals experience and perceive their work with patients from diverse ethnic groups. Is their behavior influenced by ethnicity in ways that might contribute to health care disparities? For example, do doctors sometimes avoid medical examinations for fear of causing offence because of cultural differences? If more were known about how health professionals handle ethnic diversity (a term used here to include both ethnicity and race) it might be possible to reduce ethnic inequalities in health care. In this qualitative study, the researchers have explored how health professionals involved in cancer care are affected by working with ethnically diverse patients. A qualitative study is one that collects nonquantitative data such as how doctors “feel” about treating people of different ethnic backgrounds; a quantitative study might compare clinical outcomes in different ethnic groups.
What Did the Researchers Do and Find?
The researchers enrolled 106 doctors, nurses, and other health-related professionals from different health-service settings in the Midlands, an ethnically diverse region of the UK. They organized 18 focus groups in which the health professionals described their experiences of caring for people from ethnic minority backgrounds. The participants were encouraged to recall actual cases and to identify what they saw as problems and strengths in their interactions with these patients. The researchers found that the health professionals wrestled with many challenges when providing health care for patients from diverse ethnic backgrounds. These challenges included problems with language and with general communication (for example, deciding when it was acceptable to touch a patient to show empathy). Health professionals also worried they did not know enough about cultural differences. As a result, they said they often felt uncertain of their ability to avoid causing affront or appearing racist. This uncertainty, the researchers report, disempowered the health professionals, sometimes making them hesitate or fail to do what was best for their patient.
What Do These Findings Mean?
These findings reveal that health professionals often experience considerable uncertainty when caring for ethnically diverse patients, even after training in cultural competency. They also show that this uncertainty can lead to hesitancy and inertia, which might contribute to ethnic health care inequalities. Because the study participants were probably already interested in ethnic diversity and health care, interviews with other health professionals (and investigations of patient experiences) are needed to confirm these findings. Nevertheless, the researchers suggest several interventions that might reduce health care inequalities caused by ethnic diversity. For example, health professionals should be encouraged to recognize their uncertainty and should have access to more information and training about ethnic differences. In addition, there should be a shift in emphasis away from relying on knowledge-based cultural information towards taking an “ethnographic” approach. In other words, health professionals should be helped to feel able to ask their patients about what matters most to them as individuals about their illness and treatment.
Additional Information.
Please access these Web sites via the online version of this summary at
Information on cultural competence and health care is available from the US National Center for Cultural Competence (in English and Spanish) and DiversityRx
PROCEED (Professionals Responding to Cancer in Ethnic Diversity) is a multimedia training tool for educators within the health and allied professions developed from the results of this study; a press release on PROCEED is available from the University of Nottingham
Transcultural Health Care Practice: An educational resource for nurses and health care practitioners is available on the web site of the UK Royal College of Nursing
PMCID: PMC2071935  PMID: 18001148
21.  Mapping the bacterial cell architecture into the chromosome. 
A genome is not a simple collection of genes. We propose here that it can be viewed as being organized as a 'celluloculus' similar to the homunculus of preformists, but pertaining to the category of programmes (or algorithms) rather than to that of architectures or structures: a significant correlation exists between the distribution of genes along the chromosome and the physical architecture of the cell. We review here data supporting this observation, stressing physical constraints operating on the cell's architecture and dynamics, and their consequences in terms of gene and genome structure. If such a correlation exists, it derives from some selection pressure: simple and general physical principles acting at the level of the cell structure are discussed. As a first case in point we see the piling up of planar modules as a stable, entropy-driven, architectural principle that could be at the root of the coupling between the architecture of the cell and the location of genes at specific places in the chromosome. We propose that the specific organization of certain genes whose products have a general tendency to form easily planar modules is a general motor for architectural organization in the bacterial cell. A second mechanism, operating at the transcription level, is described that could account for the efficient building up of complex structures. As an organizing principle we suggest that exploration by biological polymers of the vast space of possible conformation states is constrained by anchoring points. In particular, we suggest that transcription does not always allow the 5'-end of the transcript to go free and explore the many conformations available, but that, in many cases, it remains linked to the transcribing RNA polymerase complex in such a way that loops of RNA, rather than threads with a free end, explore the surrounding medium. In bacteria, extension of the loops throughout the cytoplasm would therefore be mediated by the de novo synthesis of ribosomes in growing cells. Termination of transcription and mRNA turnover would accordingly be expected to be controlled by sequence features at both the 3'- and 5'-ends of the molecule. These concepts are discussed taking into account in vitro analysis of genome sequences and experimental data about cell compartmentalization, mRNA folding and turnover, as well as known structural features of protein and membrane complexes.
PMCID: PMC1692725  PMID: 10724454
22.  Nanophotonic Filters and Integrated Networks in Flexible 2D Polymer Photonic Crystals 
Scientific Reports  2013;3:2145.
Polymers have appealing optical, biochemical, and mechanical qualities, including broadband transparency, ease of functionalization, and biocompatibility. However, their low refractive indices have precluded wavelength-scale optical confinement and nanophotonic applications in polymers. Here, we introduce a suspended polymer photonic crystal (SPPC) architecture that enables the implementation of nanophotonic structures typically limited to high-index materials. Using the SPPC platform, we demonstrate nanophotonic band-edge filters, waveguides, and nanocavities featuring quality (Q) factors exceeding 2, 300 and mode volumes (Vmode) below 1.7(λ/n)3. The unprecedentedly high Q/Vmode ratio results in a spectrally selective enhancement of radiative transitions of embedded emitters via the cavity Purcell effect with an enhancement factor exceeding 100. Moreover, the SPPC architecture allows straightforward integration of nanophotonic networks, shown here by a waveguide-coupled cavity drop filter with sub-nanometer spectral resolution. The nanoscale optical confinement in polymer promises new applications ranging from optical communications to organic opto-electronics, and nanophotonic polymer sensors.
PMCID: PMC3701891  PMID: 23828320
23.  Collenchyma: a versatile mechanical tissue with dynamic cell walls 
Annals of Botany  2012;110(6):1083-1098.
Collenchyma has remained in the shadow of commercially exploited mechanical tissues such as wood and fibres, and therefore has received little attention since it was first described. However, collenchyma is highly dynamic, especially compared with sclerenchyma. It is the main supporting tissue of growing organs with walls thickening during and after elongation. In older organs, collenchyma may become more rigid due to changes in cell wall composition or may undergo sclerification through lignification of newly deposited cell wall material. While much is known about the systematic and organographic distribution of collenchyma, there is rather less information regarding the molecular architecture and properties of its cell walls.
Scope and conclusions
This review summarizes several aspects that have not previously been extensively discussed including the origin of the term ‘collenchyma’ and the history of its typology. As the cell walls of collenchyma largely determine the dynamic characteristics of this tissue, I summarize the current state of knowledge regarding their structure and molecular composition. Unfortunately, to date, detailed studies specifically focusing on collenchyma cell walls have not been undertaken. However, generating a more detailed understanding of the structural and compositional modifications associated with the transition from plastic to elastic collenchyma cell wall properties is likely to provide significant insights into how specific configurations of cell wall polymers result in specific functional properties. This approach, focusing on architecture and functional properties, is likely to provide improved clarity on the controversial definition of collenchyma.
PMCID: PMC3478049  PMID: 22933416
Collenchyma; histology; plant anatomy; mechanical tissue; plant cell wall; primary and secondary cell walls; plant biomechanics
24.  Development of functionalised polyelectrolyte capsules using filamentous Escherichia coli cells 
Escherichia coli is one of the best studied microorganisms and finds multiple applications especially as tool in the heterologous production of interesting proteins of other organisms. The heterologous expression of special surface (S-) layer proteins caused the formation of extremely long E. coli cells which leave transparent tubes when they divide into single E. coli cells. Such natural structures are of high value as bio-templates for the development of bio-inorganic composites for many applications. In this study we used genetically modified filamentous Escherichia coli cells as template for the design of polyelectrolyte tubes that can be used as carrier for functional molecules or particles. Diversity of structures of biogenic materials has the potential to be used to construct inorganic or polymeric superior hybrid materials that reflect the form of the bio-template. Such bio-inspired materials are of great interest in diverse scientific fields like Biology, Chemistry and Material Science and can find application for the construction of functional materials or the bio-inspired synthesis of inorganic nanoparticles.
Genetically modified filamentous E. coli cells were fixed in 2% glutaraldehyde and coated with alternating six layers of the polyanion polyelectrolyte poly(sodium-4styrenesulfonate) (PSS) and polycation polyelectrolyte poly(allylamine-hydrochloride) (PAH). Afterwards we dissolved the E. coli cells with 1.2% sodium hypochlorite, thus obtaining hollow polyelectrolyte tubes of 0.7 μm in diameter and 5–50 μm in length. For functionalisation the polyelectrolyte tubes were coated with S-layer protein polymers followed by metallisation with Pd(0) particles. These assemblies were analysed with light microscopy, scanning electron microscopy, energy dispersive X-ray spectroscopy and transmission electron microscopy.
The thus constructed new material offers possibilities for diverse applications like novel catalysts or metal nanowires for electrical devices. The novelty of this work is the use of filamentous E. coli templates and the use of S-layer proteins in a new material construct.
PMCID: PMC3546914  PMID: 23259586
Escherichia coli; S-layer; Polyelectrolytes; Layer-by-layer (LbL); Palladium; SEM; TEM; Nanoparticle
25.  Brucella BioR Regulator Defines a Complex Regulatory Mechanism for Bacterial Biotin Metabolism 
Journal of Bacteriology  2013;195(15):3451-3467.
The enzyme cofactor biotin (vitamin H or B7) is an energetically expensive molecule whose de novo biosynthesis requires 20 ATP equivalents. It seems quite likely that diverse mechanisms have evolved to tightly regulate its biosynthesis. Unlike the model regulator BirA, a bifunctional biotin protein ligase with the capability of repressing the biotin biosynthetic pathway, BioR has been recently reported by us as an alternative machinery and a new type of GntR family transcriptional factor that can repress the expression of the bioBFDAZ operon in the plant pathogen Agrobacterium tumefaciens. However, quite unusually, a closely related human pathogen, Brucella melitensis, has four putative BioR-binding sites (both bioR and bioY possess one site in the promoter region, whereas the bioBFDAZ [bio] operon contains two tandem BioR boxes). This raised the question of whether BioR mediates the complex regulatory network of biotin metabolism. Here, we report that this is the case. The B. melitensis BioR ortholog was overexpressed and purified to homogeneity, and its solution structure was found to be dimeric. Functional complementation in a bioR isogenic mutant of A. tumefaciens elucidated that Brucella BioR is a functional repressor. Electrophoretic mobility shift assays demonstrated that the four predicted BioR sites of Brucella plus the BioR site of A. tumefaciens can all interact with the Brucella BioR protein. In a reporter strain that we developed on the basis of a double mutant of A. tumefaciens (the ΔbioR ΔbioBFDA mutant), the β-galactosidase (β-Gal) activity of three plasmid-borne transcriptional fusions (bioBbme-lacZ, bioYbme-lacZ, and bioRbme-lacZ) was dramatically decreased upon overexpression of Brucella bioR. Real-time quantitative PCR analyses showed that the expression of bioBFDA and bioY is significantly elevated upon removal of bioR from B. melitensis. Together, we conclude that Brucella BioR is not only a negative autoregulator but also a repressor of expression of bioY and bio operons that separately function in biotin transport and the biosynthesis pathway.
PMCID: PMC3719536  PMID: 23729648

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