bluuebunny/crossref_metadata_2025 huggingface dataset:
Hepatitis B immunity in a population of drug and alcohol users
Authors: POLIZZOTTO M. N. , WHELAN G.
Published: June 2007

The objective of this study was to determine the level of immunity from Hepatitis B infection in an Australian population of drug and alcohol users, and the validity of self‐reported immune status. A cross‐sectional survey was employed of drug and alcohol users presenting for treatment, who were clients of De Paul House, a community‐based drug and alcohol withdrawal service in Victoria. Outcome measures were: hepatitis B serology and personal reports of hepatitis B immunity. A total of 118 people were enrolled; 22% were injecting drug users and 48% reported past injecting drug use, while 55% were alcohol users; 51% had no general practitioner; 73% of participants were unaware of whether they were protected from hepatitis B infection, while 19.5% believed they were protected. However, serology demonstrated that 52.2% of those who believed they were protected were not immune. Only 21% of participants were immune. This is the first study in an Australian setting since the 1970s to examine the effectiveness of vaccination targeted to this population, and demonstrates low rates of immunity. New strategies are needed to deliver vaccination to this group. These will need to take account of the often chaotic nature of their lives, the poor validity of self‐reported immunity, and the lack of primary care links. [Polizzotto MN, Whelan G. Hepatitis B immunity in a population of drug and alcohol users. Drug Alcohol Rev 2007;26:417–419]
Photo-Induced Damages of Cytoplasmic and Mitochondrial Membranes by a [C60]Fullerene Malonic Acid Derivative
Authors: Yang Xinlin , Chen Lin , Qiao Xinge , Fan Cuihong
Published: June 2007

The biological activities of fullerene derivatives have attracted much attention in the last decade. In this paper, effects of dimalonic acid C60 (DMA C60) on cytoplasmic membrane, intracellular calcium concentration ([Ca2+]i), and mitochondrial membrane in HeLa cells were studied by using laser scanning confocal microscopy together with fluorescent probes propidium iodide (PI), fluo-3 acetoxymethyl ester (fluo-3 AM), and tetramethyl rhodamine methyl ester (TMRM). The data showed that under laser irradiation produced by a Kr/Ar laser source with a low power less than 1 mW, DMA C60 might induce damages against both cytoplasmic and mitochondrial membranes in a time- and dose-dependent manner. Prior to leakage of cytoplasmic membrane, a transient increase in [Ca2+]i occurred due to influx of calcium from the culture medium. These data provided some novel clues to explain the mechanisms involved in the photo-induced cytotoxicity of fullerene derivatives.
Toxicological Investigation of Drug-Related Cases in Greece: Interpretation of Analytical Findings
Authors: Stefanidou Maria , Maravelias Constantine , Dona Artemis , Athanaselis Sotiris , Spiliopoulou Hara
Published: June 2007

The process of toxicological analysis of postmortem specimens can reveal some special difficulty compared to the clinically derived specimens. Many drugs are not stable and the chemical changes that occur in the specimens, due to the hydrolysis processing, the time passed, the drug metabolism, and matrix effect, even when the postmortem interval is short, may affect the interpretation of the toxicological results. This interpretation may be critical, not only to the thorough investigation of different kind of forensic cases, but also to clinical or other cases as it provides very significant challenges to the scientists. This article reviews (a) particular toxicological issues associated with some toxic substances responsible for common lethal or nonlethal poisonings, such as opiates, cannabis, and cocaine and the vast number of factors that affect drug concentration; and (b) focuses on toxicological issues associated with the analytical findings of certain postmortem specimens. The toxic substances cited in the present paper are the most commonly found in forensic cases in Greece. The investigation of these drug-related deaths has revealed that heroin, alone or in combination with other psychoactive substances, such as cannabis and cocaine, is the main drug involved in these deaths.
Changes in Biochemical Processes in Cerebellar Granule Cells of Mice Exposed To Methylmercury
Authors: Bellum Sairam , Bawa Bhupinder , Thuett Kerry A. , Stoica Gheorghe , Abbott Louise C.
Published: June 2007

At postnatal day 34, male and female C57BL/6J mice were exposed orally once a day to a total of five doses totaling 1.0 or 5.0 mg/kg of methylmercuric chloride or sterile deionized water in moistened rodent chow. Eleven days after the last dose cerebellar granule cells were acutely isolated to measure reactive oxygen species (ROS) levels and mitochondrial membrane potential using CM-H2DCFDA and TMRM dyes, respectively. For visualizing intracellular calcium ion distribution using transmission electron microscopy, mice were perfused 11 days after the last dose of methylmercury (MeHg) using the oxalate-pyroantimonate method. Cytosolic and mitochondrial protein fractions from acutely isolated granule cells were analyzed for cytochrome c content using Western blot analysis. Histochemistry (Fluoro-Jade dye) and immunohistochemistry (activated caspase 3) was performed on frozen serial cerebellar sections to label granule cell death and activation of caspase 3, respectively. Granule cells isolated from MeHg-treated mice showed elevated ROS levels and decreased mitochondrial membrane potential when compared to granule cells from control mice. Electron photomicrographs of MeHg-treated granule cells showed altered intracellular calcium ion homeostasis ([Ca2+]i) when compared to control granule cells. However, in spite of these subcellular changes and moderate relocalization of cytochrome c into the cytosol, the concentrations of MeHg used in this study did not produce significant neuronal cell death/apoptosis at the time point examined, as evidenced by Fluoro-Jade and activated caspase 3 immunostaining, respectively. These results demonstrate that short-term in vivo exposure to total doses of 1.0 and 5.0 mg/kg MeHg through the most common exposure route (oral) can result in significant subcellular changes that are not accompanied by overt neuronal cell death.
The freshwater habitats, fishes, and fisheries of the Orinoco River basin
Authors: Rodríguez Marco A. , Winemiller Kirk O. , Lewis William M. , Taphorn Baechle Donald C.
Published: June 2007

The Orinoco River of Venezuela and Colombia is one of the great rivers of the world, ranking third by discharge after the Amazon and the Congo. In the Orinoco basin, riverine and floodplain habitats, including riparian forests, play key roles in the conservation of biodiversity and support commercial, sport, and subsistence fisheries. The basin's three major floodplains regulate the amplitude and duration of floods, maintain fertile agricultural terrain, provide habitat for numerous terrestrial and aquatic species, and support the fishery. The fish fauna, which includes some 1,000 species, encompasses a great deal of ecological diversity in terms of geographic distributions, habitat affinities, functional morphology, and reproductive and feeding strategies. The Orinoco fishery is still multispecific, with around 80 different species found in the fish markets at different times of year. Current estimates indicate that annual sustainable yield is 40,000 – 45,000 metric tons. Fish culture in the region is underdeveloped despite decades of research and promotion. There is no serious commercial trade for ornamental fishes. Large regions of the Orinoco basin are still in a relatively pristine state, but aquatic resources are increasingly threatened by habitat destruction, overharvesting, pollution, and hydrological perturbation. Scientific understanding of diversity hotspots, critical habitats, and conservation status of fishes in the basin is currently insufficient to satisfy management needs. Compliance to fishery regulations is low and fishing is drastically modifying the relative abundance, population structure, and distribution of fish stocks. However, many sectors of the Orinoco basin are unexploited or only lightly exploited, and fish stocks can recover quickly if given the opportunity. Stricter enforcement of current fishery regulations would reduce the likelihood of stock collapses and other, possibly irreversible, changes in the fishery.
Normal families of holomorphic functions with multiple zeros
Authors: Pang Xuecheng , Fang Mingliang , Zalcman Lawrence
Published: June 2007

Let

F

\mathcal F

be a family of functions holomorphic on a domain

D
D

in

C

,

\mathbb C,

all of whose zeros are multiple. Let

h
h

be a function meromorphic on

D
,

D,

h
≢
0
,
∞
.

h\not \equiv 0,\infty .

Suppose that for each

f
∈

F

,

f\in \mathcal F,

f
′

(
z
)
≠
h
(
z
)

f’(z)\ne h(z)

for

z
∈
D
.

z\in D.

Then

F

\mathcal F

is a normal family on

D
.

D.
Aurora B Phosphorylates Multiple Sites on Mitotic Centromere-associated Kinesin to Spatially and Temporally Regulate Its Function
Authors: Zhang Xin , Lan Weijie , Ems-McClung Stephanie C. , Stukenberg P. Todd , Walczak Claire E.
Published: June 2007

Chromosome congression and segregation require the proper attachment of microtubules to the two sister kinetochores. Disruption of either Aurora B kinase or the Kinesin-13 mitotic centromere-associated kinesin (MCAK) increases chromosome misalignment and missegregation due to improper kinetochore–microtubule attachments. MCAK localization and activity are regulated by Aurora B, but how Aurora B phosphorylation of MCAK affects spindle assembly is unclear. Here, we show that the binding of MCAK to chromosome arms is also regulated by Aurora B and that Aurora B-dependent chromosome arm and centromere localization is regulated by distinct two-site phosphoregulatory mechanisms. MCAK association with chromosome arms is promoted by phosphorylation of T95 on MCAK, whereas phosphorylation of S196 on MCAK promotes dissociation from the arms. Although targeting of MCAK to centromeres requires phosphorylation of S110 on MCAK, dephosphorylation of T95 on MCAK increases the binding of MCAK to centromeres. Our study reveals a new role for Aurora B, which is to prevent excess MCAK binding to chromatin to facilitate chromatin-nucleated spindle assembly. Our study also shows that the interplay between multiple phosphorylation sites of MCAK may be critical to temporally and spatially control MCAK function.
Cdc6 Stability Is Regulated by the Huwe1 Ubiquitin Ligase after DNA Damage
Authors: Hall Jonathan R. , Kow Evelyn , Nevis Kathleen R. , Lu Chiajung Karen , Luce K. Scott , Zhong Qing , Cook Jeanette Gowen
Published: June 2007

The Cdc6 protein is an essential component of pre-replication complexes (preRCs), which assemble at origins of DNA replication during the G1 phase of the cell cycle. Previous studies have demonstrated that, in response to ionizing radiation, Cdc6 is ubiquitinated by the anaphase promoting complex (APCCdh1) in a p53-dependent manner. We find, however, that DNA damage caused by UV irradiation or DNA alkylation by methyl methane sulfonate (MMS) induces Cdc6 degradation independently of p53. We further demonstrate that Cdc6 degradation after these forms of DNA damage is also independent of cell cycle phase, Cdc6 phosphorylation of the known Cdk target residues, or the Cul4/DDB1 and APCCdh1ubiquitin E3 ligases. Instead Cdc6 directly binds a HECT-family ubiquitin E3 ligase, Huwe1 (also known as Mule, UreB1, ARF-BP1, Lasu1, and HectH9), and Huwe1 polyubiquitinates Cdc6 in vitro. Degradation of Cdc6 in UV-irradiated cells or in cells treated with MMS requires Huwe1 and is associated with release of Cdc6 from chromatin. Furthermore, yeast cells lacking the Huwe1 ortholog, Tom1, have a similar defect in Cdc6 degradation. Together, these findings demonstrate an important and conserved role for Huwe1 in regulating Cdc6 abundance after DNA damage.
The Role of p58IPK in Protecting the Stressed Endoplasmic Reticulum
Authors: Rutkowski D. Thomas , Kang Sang-Wook , Goodman Alan G. , Garrison Jennifer L. , Taunton Jack , Katze Michael G. , Kaufman Randal J. , Hegde Ramanujan S.
Published: June 2007

The preemptive quality control (pQC) pathway protects cells from acute endoplasmic reticulum (ER) stress by attenuating translocation of nascent proteins despite their targeting to translocons at the ER membrane. Here, we investigate the hypothesis that the DnaJ protein p58IPK plays an essential role in this process via HSP70 recruitment to the cytosolic face of translocons for extraction of translocationally attenuated nascent chains. Our analyses revealed that the heightened stress sensitivity of p58−/− cells was not due to an impairment of the pQC pathway or elevated ER substrate burden during acute stress. Instead, the lesion was in the protein processing capacity of the ER lumen, where p58IPK was found to normally reside in association with BiP. ER lumenal p58IPK could be coimmunoprecipitated with a newly synthesized secretory protein in vitro and stimulated protein maturation upon overexpression in cells. These results identify a previously unanticipated location for p58IPK in the ER lumen where its putative function as a cochaperone explains the stress-sensitivity phenotype of knockout cells and mice.
Boltzmann probability of RNA structural neighbors and riboswitch detection
Authors: Freyhult Eva , Moulton Vincent , Clote Peter
Published: June 2007

Motivation: We describe algorithms implemented in a new software package, RNAbor, to investigate structures in a neighborhood of an input secondary structure S of an RNA sequence s. The input structure could be the minimum free energy structure, the secondary structure obtained by analysis of the X-ray structure or by comparative sequence analysis, or an arbitrary intermediate structure.

Results: A secondary structure T of s is called a δ-neighbor of S if T and S differ by exactly δ base pairs. RNAbor computes the number (Nδ), the Boltzmann partition function (Zδ) and the minimum free energy (MFEδ) and corresponding structure over the collection of all δ-neighbors of S. This computation is done simultaneously for all δ ≤ m, in run time O (mn3) and memory O(mn2), where n is the sequence length. We apply RNAbor for the detection of possible RNA conformational switches, and compare RNAbor with the switch detection method paRNAss. We also provide examples of how RNAbor can at times improve the accuracy of secondary structure prediction.

Availability:  http://bioinformatics.bc.edu/clotelab/RNAbor/

Contact:  clote@bc.edu

Supplementary information: Supplementary data are available at Bioinformatics online.
Julius Axelrod. 30 May 1912 — 29 December 2004
Authors: Iversen Leslie
Published: June 2007

Julie Axelrod was a laboratory technician until the age of 42 years, when he finally achieved his PhD and independence. He worked at the National Institutes of Health for most of his career. Among his early pioneering research achievements in applying chemical and biochemical approaches to neuroscience were the discoveries of the painkiller acetaminophen (Tylenol, Paracetamol) and the liver microsomal drug–metabolizing enzymes, and the establishment of catechol–
O
–methyltransferase as an important enzyme in catecholamine metabolism. He shared the Nobel Prize in Physiology or Medicine in 1970 for his discovery that the reuptake of noradrenaline (norepinephrine) into the nerve endings from which it was released represented a novel method of neurotransmitter inactivation. An important corollary was the finding that antidepressant drugs acted as inhibitors of this uptake process.
John Robert Stanley Fincham. 11 August 1926 — 9 February 2005
Authors: Holliday Robin , Flavell Richard B.
Published: June 2007

Professor John Fincham was one of the UK's leading geneticists, with a remarkably broad knowledge of the subject across all the biological kingdoms. He became an international leader through being at the forefront of microbial genetics as some of the founding principles of the relationships between gene structure, activity and enzyme functions were being uncovered. He spearheaded discoveries from the one gene–one enzyme concept, through genetic complementation, protein structure and recombination. Much of his experimental microbial research centred on the genetic and enzyme variants of glutamate dehydrogenase in the fungus
Neurospora
. He also brought his outstanding mind and comprehensive interest in genetics to the then obscure features of unstable genes and transposable elements in plants. His standing was recognized by holding prestigious chairs in Leeds, Edinburgh and Cambridge universities. He was a talented writer, producing several textbooks and especially the leading text
Fungal genetics
. He was also a practitioner and lover of sports and in his early career was politically active. His successes in life made him an extraordinarily talented man who achieved much as a leader in genetics in the UK and internationally.
Gordon Elliott Fogg CBE. 26 April 1919 — 30 January 2005
Authors: Walsby Anthony E.
Published: June 2007

Gordon Elliott (Tony) Fogg was a botanist who pioneered studies in cyanobacterial heterocysts, nitrogen fixation, extracellular products of algae and phytoplankton ecophysiology. He was a gifted lecturer and writer, and the author of books on algae, phytoplankton ecology and Antarctic science. After many years in Botany at University College and then Westfield College in the University of London, he completed his career in Marine Biology at the University College of North Wales. His natural diplomacy led to appointments on many scientific committees, where he was valued for his succinct observations and his ability to settle disputes behind the scenes. He made important contributions to many organizations, including the Institute of Biology, the Freshwater Biological Association, the British Antarctic Survey and the Royal Botanic Gardens, Kew. He was artistic, a fine water-colourist, and he enjoyed walking in Anglesey and Snowdonia.
Yasutomi Nishizuka. 12 July 1932 — 4 November 2004
Authors: Yamamura Hirohei , Nakamura Shun-Ichi
Published: June 2007

Yasutomi Nizhizuka was born in Seido Village, Muko–gun (now City of Ashiya), Hyogo–Prefecture, on 12 July 1932, as the second son of Yasunobu and Nobuko Nishizuka. His elder brother, Yasuaki (1922–95, pathologist, President of Aichi Cancer Center), was 10 years old, his eldest sister, Hiroko, was aged 8, his elder sister, Fumiko, was 5, and he was a heterozygotic twin with his sister Junko. His mother, Nobuko, was from the Ijichi family, a huge landlord, which traced its roots back to a powerful family of the Heian Era (eleventh century). When Yasutomi was an infant, he was so weak that his parents were not sure that he would survive to adulthood. He had intussusception (ileus) when he was six months old. He barely survived after emergency surgery at Daido Hospital in Osaka. Since then, he frequently suffered from diseases associated with stomach ache. He was always weak and ailing.
Sir Frederick William Page CBE FREng. 20 February 1917 — 29 May 2005
Authors: Yates I.R.
Published: June 2007

Sir Frederick Page was all round the most able member of the British aeronautical industry, a man of exceptional ability and integrity. Promoted rapidly in early life, he developed into a very progressive and innovative company manager, mastering all aspects as the industry went through a succession of painful mergers. Finally, as Chief Executive of the Aircraft Group of British Aerospace he developed very perceptive industrial strategies with excellent financial and risk management. The sum total of his contribution to the British aerospace community over a period of 45 years until his retirement in 1983 was greater than that of any other individual.
Autar Singh Paintal. 24 September 1925 — 21 December 2004
Authors: Iggo Ainsley
Published: June 2007

Autar Paintal began his scientific career at Lucknow University in India, enhanced it in David Whitteridge's Department of Physiology in Edinburgh and consolidated it in India, where he worked at the Technical Development Establishment, the All-India Institute of Medical Sciences and The Vallabhbhai Patel Chest Institute of Delhi University. He correctly identified atrial B receptors and by precise experimentation established the properties of atrial A and B fibres, of ventricular afferent fibres and the gastrointestinal sensory receptors. His enduring scientific contribution was to identify J receptors in the lung and to characterize their reflex and sensory actions, both at sea level and at Himalayan altitude. His distinguished contributions to medical science in India and abroad has been marked by the award of numerous honorary degrees and prizes.
Sir Alan Sterling Parkes. 10 September 1900 — 17 July 1990
Authors: Polge Christopher
Published: June 2007

Alan Parkes was one of the most influential figures in the field of reproductive biology in the twentieth century. He had a huge impact on its growth and development during that time, and the legacy of his work still remains.
Robert William Stewart. 21 August 1923 — 19 January 2005
Authors: Garrett Chris
Published: June 2007

Bob Stewart was a leading ocean physicist who made notable contributions to the study of turbulence, water waves, and air–sea interaction. He was also an influential educator, laboratory director, provincial deputy minister and an exemplary leader in international organizations involved in oceanographic and climate studies.
Michael Szwarc. 19 June 1909 — 4 August 2000
Authors: Jagur-Grodzinski Joseph , Penczek Stanislaw
Published: June 2007

Michael Szwarc was born in Będzin, an industrial town in the south of Poland, as an only son of a Jewish family. Michael received his pre–college and college education in Warsaw and in 1932 received his degree in chemical engineering from the Warsaw Polytechnic Institute. After graduation he married in 1933 Maria Frenkel (Marysia), whose brother married his younger sister Rala. From 1933 to 1935 he worked in Poland as a chemical engineer. In 1935 he emigrated to Palestine (today's Israel) and after a year was joined there by his sister and brother–in–law. Two of his three children were born during his stay in Jerusalem, where he was engaged in research at the Hebrew University of Jerusalem and from which he gained a PhD in organic chemistry. In late 1945 he went to England, where he joined the group under Professor Michael Polanyi FRS in Manchester.
Sylvia Agnes Sophia Tait. 8 January 1917 — 28 February 2003
Authors: Denton Derek A. , MacIntyre Iain
Published: June 2007

Sylvia Agnes Sophia Tait was born on 8 January 1917 in Tumen, Siberia, Russia. She was the daughter of James Wardropper, an agronomist and trader, working in Russia. It seems that James Wardropper worked there with his elder brother, Robert (Huntford 1997). The wife of James Wardropper, Ludmilla, was a Russian who had the rare distinction of graduating in mathematics from the University of Moscow in the time of the reign of the Tsar. James and Ludmilla Wardropper adopted a Russian girl, Pasha; she became part of the family and helped to look after Sylvia. During the revolution, in 1920 the whole family, including Pasha (but not including Robert) left Russia from Vladivostok for the UK, where James Wardropper eventually became a successful civil engineer. The fate of Robert Wardropper remains a mystery. The other Wardroppers first stayed in the UK in Ealing, London, where Sylvia attended the local secondary school, the Ealing County School for Girls. In her senior years there, she mainly studied languages, particularly German but also French and Latin. The Wardroppers had relatives in Germany and, before World War II, Sylvia spent some time in Germany, including Berlin, which improved her German. In addition, of course, at that time she spoke fairly fluent Russian with her mother and step-sister, Pasha. Sylvia had considerable trouble in establishing her citizenship because of her birthplace but eventually was officially declared British. Because of the nature of the father's history as a Scottish engineer in Russia and also the effects of the revolution, Sylvia never met her maternal grandparents and knew little about them.
Sir John Robert Vane. 29 March 1927 — 19 November 2004
Authors: Moncada Salvador
Published: June 2007

Sir John Robert Vane, who died on 19 November 2004, will be remembered as one of the most influential British pharmacologists. During his distinguished career he published more than 700 scientific papers and wrote or edited 20 books. His many awards include the Nobel Prize in Physiology or Medicine (1982) and a knighthood in 1984.
David John Wheeler. 9 February 1927 — 13 December 2004
Authors: Campbell-Kelly Martin
Published: June 2007

David Wheeler's scientific career was unusual. He was not the architect of a grand project, nor did he become a senior scientific administrator. Instead, his career had an unusually level trajectory. His individual style of working barely changed in half a century, from research student to professor emeritus. Apart from a few sabbatical leaves, his whole working life took place in the Mathematical (later Computer) Laboratory at Cambridge University.
Intracellular Targeting of CEA Results in Th1-Type Antibody Responses Following Intradermal Genetic Vaccination by a Needle-Free Jet Injection Device
Authors: Johansson Susanne , Ek Monica , Wahren Britta , Stout Richard , Liu Margaret A. , Hallermalm Kristian
Published: June 2007

The route and method of immunization, as well as the cellular localization of the antigen, can influence the generation of an immune response. In general, intramuscular immunization results in Th1 responses, whereas intradermal delivery of DNA by gene gun immunization often results in more Th2 responses. Here we investigate how altering the cellular localization of the tumor antigen CEA (carcinoembryonic antigen) affects the quality and amplitude of DNA vaccine-induced antibody responses in mice following intradermal delivery of DNA by a needle-free jet injection device (Biojector). CEA was expressed either in a membrane-bound form (wild-type CEA) or in two truncated forms (CEA6 and CEA66) with cytoplasmic localization, where CEA66 was fused to a promiscuous T-helper epitope from tetanus toxin. Repeated intradermal immunization of BALB/c mice with DNA encoding wild-type CEA produced high antibody titers of a mixed IgG1/IgG2a ratio. In contrast, utilizing the DNA construct that resulted in intracellular targeting of CEA led to a reduced capacity to induce CEA-specific antibodies, but instead induced a Th1-biased immune response.
Statistical analysis of the genomic distribution and correlation of regulatory elements in the ENCODE regions
Authors: Zhang Zhengdong D. , Paccanaro Alberto , Fu Yutao , Weissman Sherman , Weng Zhiping , Chang Joseph , Snyder Michael , Gerstein Mark B.
Published: June 2007

The comprehensive inventory of functional elements in 44 human genomic regions carried out by the ENCODE Project Consortium enables for the first time a global analysis of the genomic distribution of transcriptional regulatory elements. In this study we developed an intuitive and yet powerful approach to analyze the distribution of regulatory elements found in many different ChIP–chip experiments on a 10∼100-kb scale. First, we focus on the overall chromosomal distribution of regulatory elements in the ENCODE regions and show that it is highly nonuniform. We demonstrate, in fact, that regulatory elements are associated with the location of known genes. Further examination on a local, single-gene scale shows an enrichment of regulatory elements near both transcription start and end sites. Our results indicate that overall these elements are clustered into regulatory rich “islands” and poor “deserts.” Next, we examine how consistent the nonuniform distribution is between different transcription factors. We perform on all the factors a multivariate analysis in the framework of a biplot, which enhances biological signals in the experiments. This groups transcription factors into sequence-specific and sequence-nonspecific clusters. Moreover, with experimental variation carefully controlled, detailed correlations show that the distribution of sites was generally reproducible for a specific factor between different laboratories and microarray platforms. Data sets associated with histone modifications have particularly strong correlations. Finally, we show how the correlations between factors change when only regulatory elements far from the transcription start sites are considered.
A framework for collaborative analysis of ENCODE data: Making large-scale analyses biologist-friendly
Authors: Blankenberg Daniel , Taylor James , Schenck Ian , He Jianbin , Zhang Yi , Ghent Matthew , Veeraraghavan Narayanan , Albert Istvan , Miller Webb , Makova Kateryna D. , Hardison Ross C. , Nekrutenko Anton
Published: June 2007

The standardization and sharing of data and tools are the biggest challenges of large collaborative projects such as the Encyclopedia of DNA Elements (ENCODE). Here we describe a compact Web application, Galaxy2ENCODE, that effectively addresses these issues. It provides an intuitive interface for the deposition and access of data, and features a vast number of analysis tools including operations on genomic intervals, utilities for manipulation of multiple sequence alignments, and molecular evolution algorithms. By providing a direct link between data and analysis tools, Galaxy2ENCODE allows addressing biological questions that are beyond the reach of existing software. We use Galaxy2ENCODE to show that the ENCODE regions contain >2000 unannotated transcripts under strong purifying selection that are likely functional. We also show that the ENCODE regions are representative of the entire genome by estimating the rate of nucleotide substitution and comparing it to published data. Although each of these analyses is complex, none takes more than 15 min from beginning to end. Finally, we demonstrate how new tools can be added to Galaxy2ENCODE with almost no effort. Every section of the manuscript is supplemented with QuickTime screencasts. Galaxy2ENCODE and the screencasts can be accessed at http://g2.bx.psu.edu.
The ENCODEdb portal: Simplified access to ENCODE Consortium data
Authors: Elnitski Laura L. , Shah Prachi , Moreland R. Travis , Umayam Lowell , Wolfsberg Tyra G. , Baxevanis Andreas D.
Published: June 2007

The Encyclopedia of DNA Elements (ENCODE) project aims to identify and characterize all functional elements in a representative chromosomal sample comprising 1% of the human genome. Data generated by members of The ENCODE Project Consortium are housed in a number of public databases, such as the UCSC Genome Browser, NCBI’s Gene Expression Omnibus (GEO), and EBI’s ArrayExpress. As such, it is often difficult for biologists to gather all of the ENCODE data from a particular genomic region of interest and integrate them with relevant information found in other public databases. The ENCODEdb portal was developed to address this problem. ENCODEdb provides a unified, single point-of-access to data generated by the ENCODE Consortium, as well as to data from other source databases that lie within ENCODE regions; this provides the user a complete view of all known data in a particular region of interest. ENCODEdb Genomic Context searches allow for the retrieval of information on functional elements annotated within ENCODE regions, including mRNA, EST, and STS sequences; single nucleotide polymorphisms, and UniGene clusters. Information is also retrieved from GEO, OMIM, and major genome sequence browsers. ENCODEdb Consortium Data searches allow users to perform compound queries on array-based ENCODE data available both from GEO and from the UCSC Genome Browser. Results are retrieved from a specific genomic area of interest and can be further manipulated in a variety of contexts, including the UCSC Genome Browser and the Galaxy large-scale genome analysis platform. The ENCODEdb portal is freely accessible at http://research.nhgri.nih.gov/ENCODEdb.
Mapping of transcription factor binding regions in mammalian cells by ChIP: Comparison of array- and sequencing-based technologies
Authors: Euskirchen Ghia M. , Rozowsky Joel S. , Wei Chia-Lin , Lee Wah Heng , Zhang Zhengdong D. , Hartman Stephen , Emanuelsson Olof , Stolc Viktor , Weissman Sherman , Gerstein Mark B. , Ruan Yijun , Snyder Michael
Published: June 2007

Recent progress in mapping transcription factor (TF) binding regions can largely be credited to chromatin immunoprecipitation (ChIP) technologies. We compared strategies for mapping TF binding regions in mammalian cells using two different ChIP schemes: ChIP with DNA microarray analysis (ChIP-chip) and ChIP with DNA sequencing (ChIP-PET). We first investigated parameters central to obtaining robust ChIP-chip data sets by analyzing STAT1 targets in the ENCODE regions of the human genome, and then compared ChIP-chip to ChIP-PET. We devised methods for scoring and comparing results among various tiling arrays and examined parameters such as DNA microarray format, oligonucleotide length, hybridization conditions, and the use of competitor Cot-1 DNA. The best performance was achieved with high-density oligonucleotide arrays, oligonucleotides ≥50 bases (b), the presence of competitor Cot-1 DNA and hybridizations conducted in microfluidics stations. When target identification was evaluated as a function of array number, 80%–86% of targets were identified with three or more arrays. Comparison of ChIP-chip with ChIP-PET revealed strong agreement for the highest ranked targets with less overlap for the low ranked targets. With advantages and disadvantages unique to each approach, we found that ChIP-chip and ChIP-PET are frequently complementary in their relative abilities to detect STAT1 targets for the lower ranked targets; each method detected validated targets that were missed by the other method. The most comprehensive list of STAT1 binding regions is obtained by merging results from ChIP-chip and ChIP-sequencing. Overall, this study provides information for robust identification, scoring, and validation of TF targets using ChIP-based technologies.
Pseudogenes in the ENCODE regions: Consensus annotation, analysis of transcription, and evolution
Authors: Zheng Deyou , Frankish Adam , Baertsch Robert , Kapranov Philipp , Reymond Alexandre , Choo Siew Woh , Lu Yontao , Denoeud France , Antonarakis Stylianos E. , Snyder Michael , Ruan Yijun , Wei Chia-Lin , Gingeras Thomas R. , Guigó Roderic , Harrow Jennifer , Gerstein Mark B.
Published: June 2007

Arising from either retrotransposition or genomic duplication of functional genes, pseudogenes are “genomic fossils” valuable for exploring the dynamics and evolution of genes and genomes. Pseudogene identification is an important problem in computational genomics, and is also critical for obtaining an accurate picture of a genome’s structure and function. However, no consensus computational scheme for defining and detecting pseudogenes has been developed thus far. As part of the ENCyclopedia Of DNA Elements (ENCODE) project, we have compared several distinct pseudogene annotation strategies and found that different approaches and parameters often resulted in rather distinct sets of pseudogenes. We subsequently developed a consensus approach for annotating pseudogenes (derived from protein coding genes) in the ENCODE regions, resulting in 201 pseudogenes, two-thirds of which originated from retrotransposition. A survey of orthologs for these pseudogenes in 28 vertebrate genomes showed that a significant fraction (∼80%) of the processed pseudogenes are primate-specific sequences, highlighting the increasing retrotransposition activity in primates. Analysis of sequence conservation and variation also demonstrated that most pseudogenes evolve neutrally, and processed pseudogenes appear to have lost their coding potential immediately or soon after their emergence. In order to explore the functional implication of pseudogene prevalence, we have extensively examined the transcriptional activity of the ENCODE pseudogenes. We performed systematic series of pseudogene-specific RACE analyses. These, together with complementary evidence derived from tiling microarrays and high throughput sequencing, demonstrated that at least a fifth of the 201 pseudogenes are transcribed in one or more cell lines or tissues.
Detection of DNA structural motifs in functional genomic elements
Authors: Greenbaum Jason A. , Parker Stephen C.J. , Tullius Thomas D.
Published: June 2007

The completion of the human genome project has fueled the search for regulatory elements by a variety of different approaches. Many successful analyses have focused on examining primary DNA sequence and/or chromatin structure. However, it has been difficult to detect common sequence motifs within the feature of chromatin structure most closely associated with regulatory elements, DNase I hypersensitive sites (DHSs). Considering just the nucleotide sequence and/or the chromatin structure of regulatory elements may neglect a critical feature of what is recognized by the regulatory machinery—DNA structure. We introduce a new computational method to detect common DNA structural motifs in a large collection of DHSs that are found in the ENCODE regions of the human genome. We show that DHSs have common DNA structural motifs that show no apparent sequence consensus. One such structural motif is much more highly enriched in experimentally identified DHSs that are in CpG islands and near transcription start sites (TSSs), compared to DHSs not in CpG islands and farther from TSSs, suggesting that DNA structural motifs may participate in the formation of functional regulatory elements. We propose that studies of the conservation of DNA structure, independent of sequence conservation, will provide new information about the link between the nucleotide sequence of a DNA molecule and its experimentally demonstrated function.
Independent and complementary methods for large-scale structural analysis of mammalian chromatin
Authors: Dennis Jonathan H. , Fan Hua-Ying , Reynolds Sheila M. , Yuan Guocheng , Meldrim James C. , Richter Daniel J. , Peterson Daniel G. , Rando Oliver J. , Noble William S. , Kingston Robert E.
Published: June 2007

The fundamental building block of chromatin, the nucleosome, occupies 150 bp of DNA in a spaced arrangement that is a primary determinant in regulation of the genome. The nucleosomal organization of some regions of the human genome has been described, but mapping of these regions has been limited to a few kilobases. We have explored two independent and complementary methods for the high-throughput analysis of mammalian chromatin structure. Through adaptations to a protocol used to map yeast chromatin structure, we determined sites of nucleosomal protection over large regions of the mammalian genome using a tiling microarray. By modifying classical primer extension methods, we localized specific internucleosomally cleaved mammalian genomic sequences using a capillary electrophoresis sequencer in a manner that allows high-throughput nucleotide-resolution characterization of nucleosome protection patterns. We developed algorithms for the automated and unbiased analysis of the resulting data, a necessary step toward large-scale analysis. We validated these assays using the known positions of nucleosomes on the mouse mammary tumor virus LTR, and additionally, we characterized the previously unreported chromatin structure of the LCMT2 gene. These results demonstrate the effectiveness of the combined methods for reliable analysis of mammalian chromatin structure in a high-throughput manner.
Integrated analysis of experimental data sets reveals many novel promoters in 1% of the human genome
Authors: Trinklein Nathan D. , Karaöz Ulaş , Wu Jiaqian , Halees Anason , Force Aldred Shelley , Collins Patrick J. , Zheng Deyou , Zhang Zhengdong D. , Gerstein Mark B. , Snyder Michael , Myers Richard M. , Weng Zhiping
Published: June 2007

The regulation of transcriptional initiation in the human genome is a critical component of global gene regulation, but a complete catalog of human promoters currently does not exist. In order to identify regulatory regions, we developed four computational methods to integrate 129 sets of ENCODE-wide chromatin immunoprecipitation data. They collectively predicted 1393 regions. Roughly 47% of the regions were unique to one method, as each method makes different assumptions about the data. Overall, predicted regions tend to localize to highly conserved, DNase I hypersensitive, and actively transcribed regions in the genome. Interestingly, a significant portion of the regions overlaps with annotated 3′-UTRs, suggesting that some of them might regulate anti-sense transcription. The majority of the predicted regions are >2 kb away from the 5′-ends of previously annotated human cDNAs and hence are novel. These novel regions may regulate unannotated transcripts or may represent new alternative transcription start sites of known genes. We tested 163 such regions for promoter activity in four cell lines using transient transfection assays, and 25% of them showed transcriptional activity above background in at least one cell line. We also performed 5′-RACE experiments on 62 novel regions, and 76% of the regions were associated with the 5′-ends of at least two RACE products. Our results suggest that there are at least 35% more functional promoters in the human genome than currently annotated.
Analysis of overrepresented motifs in human core promoters reveals dual regulatory roles of YY1
Authors: Xi Hualin , Yu Yong , Fu Yutao , Foley Jonathan , Halees Anason , Weng Zhiping
Published: June 2007

A set of 723 high-quality human core promoter sequences were compiled and analyzed for overrepresented motifs. Beside the two well-characterized core promoter motifs (TATA and Inr), several known motifs (YY1, Sp1, NRF-1, NRF-2, CAAT, and CREB) and one potentially new motif (motif8) were found. Interestingly, YY1 and motif8 mostly reside immediately downstream from the TSS. In particular, the YY1 motif occurs primarily in genes with 5′-UTRs shorter than 40 base pairs (bp) and its locations coincide with the translation start site. We verified that the YY1 motif is bound by YY1 in vitro. We then performed detailed analysis on YY1 chromatin immunoprecipitation data with a whole-genome human promoter microarray (ChIP-chip) and revealed that the thus identified promoters in HeLa cells were highly enriched with the YY1 motif. Moreover, the motif overlapped with the translation start sites on the plus strand of a group of genes, many with short 5′-UTRs, and with the transcription start sites on the minus strand of another distinct group of genes; together, the two groups of genes accounted for the majority of the YY1-bound promoters in the ChIP-chip data. Furthermore, the first group of genes was highly enriched in the functional categories of ribosomal proteins and nuclear-encoded mitochondria proteins. We suggest that the YY1 motif plays a dual role in both transcription and translation initiation of these genes. We also discuss the evolutionary advantages of housing a transcriptional element inside the transcript in terms of the migration of these genes in the human genome.
Identification of an OCT4 and SRY regulatory module using integrated computational and experimental genomics approaches
Authors: Jin Victor X. , O’Geen Henriette , Iyengar Sushma , Green Roland , Farnham Peggy J.
Published: June 2007

ChIP-chip studies have revealed that many in vivo binding sites have a weak match to the consensus sequence for the transcription factor being analyzed. Possible explanations for these observations include (1) the in vitro-derived consensus site does not represent the in vivo binding site and/or (2) the factor is recruited to a weak binding site via interaction with another protein. To address these possibilities, we developed an approach (ChIPMotifs) that incorporates a bootstrap resampling method to statistically infer the optimal cutoff threshold for a position weight matrix (PWM) of a motif identified from ChIP-chip data by ab initio motif discovery programs. Using OCT4 ChIP-chip data and the ChIPMotifs approach, we first developed a refined OCT4 PWM. We then used the refined PWM and a ChIPModules approach to identify transcription factors colocalizing with OCT4 in Ntera2 testicular embryonal carcinoma cells. We found that the consensus binding site for SRY, a transcription factor critical for testis development, colocalizes with the OCT4 PWM. To further characterize the relationship between OCT4 and SRY, we performed ChIP-chip experiments with human promoter microarrays, and found that 49% of the top ∼1000 OCT4 target promoters were also bound by SRY. This analysis represents the first identification of SRY target promoters. Interestingly, we determined that promoters bound by OCT4 and SRY, but not those bound by SRY alone, were also bound by the transcriptional repressor KAP1. Our studies not only validate the ChIPMotifs and ChIPModules combinatorial approach but also identify a possible new regulatory partner of OCT4.
Construction of a genome-scale structural map at single-nucleotide resolution
Authors: Greenbaum Jason A. , Pang Bo , Tullius Thomas D.
Published: June 2007

Few methods are available for mapping the local structure of DNA throughout a genome. The hydroxyl radical cleavage pattern is a measure of the local variation in solvent-accessible surface area of duplex DNA, and thus provides information on the local shape and structure of DNA. We report the construction of a relational database, ORChID (OHRadical Cleavage Intensity Database), that contains extensive hydroxyl radical cleavage data produced from two DNA libraries. We have used the ORChID database to develop a set of algorithms that are capable of predicting the hydroxyl radical cleavage pattern of a DNA sequence of essentially any length, to high accuracy. We have used the prediction algorithm to produce a structural map of the 30 Mb of the ENCODE regions of the human genome.
Origin of phenotypes: Genes and transcripts
Authors: Gingeras Thomas R.
Published: June 2007

While the concept of a gene has been helpful in defining the relationship of a portion of a genome to a phenotype, this traditional term may not be as useful as it once was. Currently, “gene” has come to refer principally to a genomic region producing a polyadenylated mRNA that encodes a protein. However, the recent emergence of a large collection of unannotated transcripts with apparently little protein coding capacity, collectively called transcripts of unknown function (TUFs), has begun to blur the physical boundaries and genomic organization of genic regions with noncoding transcripts often overlapping protein-coding genes on the same (sense) and opposite strand (antisense). Moreover, they are often located in intergenic regions, making the genic portions of the human genome an interleaved network of both annotated polyadenylated and nonpolyadenylated transcripts, including splice variants with novel 5′ ends extending hundreds of kilobases. This complex transcriptional organization and other recently observed features of genomes argue for the reconsideration of the term “gene” and suggests that transcripts may be used to define the operational unit of a genome.
Threading a peptide through a peptide: Protein loops, rotaxanes, and knots
Authors: Blankenship John W. , Dawson Philip E.
Published: June 2007

Proteins adopt complex folds in nature that typically avoid conformations that are knotted or “threaded” through closed loops. Is this the result of fundamental barriers to folding, or have proteins simply evolved to avoid threaded conformations? Organic synthesis has been used in supramolecular chemistry to install topological links in small molecules. By following these principles, we now show that it is possible to assemble a topologically linked protein complex by threading a linear protein through a cyclic protein to form a [2]pseudo‐rotaxane. Subsequent ring closure using native chemical ligation cyclizes the linear protein, forming a [2]heterocatenane. Although the kinetics of protein threading are slower than the folding kinetics of the native protein, threading appears to be a highly efficient process.