bluuebunny/crossref_metadata_2025 huggingface dataset:
Fishing with (Proto)Net—a principled approach to protein target selection
Authors: Linial Michal
Published: October 2003

Structural genomics strives to represent the entire protein space. The first step towards achieving this goal is by rationally selecting proteins whose structures have not been determined, but that represent an as yet unknown structural superfamily or fold. Once such a structure is solved, it can be used as a template for modelling homologous proteins. This will aid in unveiling the structural diversity of the protein space. Currently, no reliable method for accurate 3D structural prediction is available when a sequence or a structure homologue is not available. Here we present a systematic methodology for selecting target proteins whose structure is likely to adopt a new, as yet unknown superfamily or fold. Our method takes advantage of a global classification of the sequence space as presented by ProtoNet‐3D, which is a hierarchical agglomerative clustering of the proteins of interest (the proteins in Swiss‐Prot) along with all solved structures (taken from the PDB). By navigating in the scaffold of ProtoNet‐3D, we yield a prioritized list of proteins that are not yet structurally solved, along with the probability of each of the proteins belonging to a new superfamily or fold. The sorted list has been self‐validated against real structural data that was not available when the predictions were made. The practical application of using our computational–statistical method to determine novel superfamilies for structural genomics projects is also discussed. Copyright © 2003 John Wiley & Sons, Ltd.
Meeting Report: ESF Programme on Functional Genomics 1st European Conference: Functional Genomics and Disease 2003
Authors: Wixon Jo , Marsh Joan
Published: October 2003

In this report from the 1st European Conference of the European Science Foundation Programme on Functional Genomics, we provide coverage of the high‐profile plenary talks and a cross‐section of the many presentations in the disease analysis symposia and functional genomics technologies workshops. Copyright © 2003 John Wiley & Sons, Ltd.
Theoretical and numerical analyses of convective instability in porous media with temperature‐dependent viscosity
Authors: Lin Ge , Zhao Chongbin , Hobbs B. E. , Ord A. , Mühlhaus H. B.
Published: October 2003

Exact analytical solutions of the critical Rayleigh numbers have been obtained for a hydrothermal system consisting of a horizontal porous layer with temperature‐dependent viscosity. The boundary conditions considered are constant temperature and zero vertical Darcy velocity at both the top and bottom of the layer. Not only can the derived analytical solutions be readily used to examine the effect of the temperature‐dependent viscosity on the temperature‐gradient driven convective flow, but also they can be used to validate the numerical methods such as the finite‐element method and finite‐difference method for dealing with the same kind of problem. The related analytical and numerical results demonstrated that the temperature‐dependent viscosity destabilizes the temperature‐gradient driven convective flow and therefore, may affect the ore body formation and mineralization in the upper crust of the Earth. Copyright © 2003 John Wiley & Sons, Ltd.
An efficient and accurate algorithm for generating spatially‐correlated random fields
Authors: Fang Jiannong , Tacher Laurent
Published: October 2003

This paper presents a new computer algorithm for generating spatially‐correlated random fields. Such fields are often encountered in hydrology and hydrogeology and in the earth sciences and used as inputs for Monte Carlo simulations. The algorithm is designed by using a multilevel grid strategy and combining the matrix decomposition (MD) method and the screening sequential simulation (SSS) method. The idea originates from the facts: (i) the MD method accounts for all possible nodal correlation values and hence the accuracy of the method is high, but it can be extremely computationally intensive for fine meshes with large number of nodes, and (ii) the SSS method is more efficient because a small search neighbourhood for conditioning can be used due to the screening effect of measurements, however, for large separation distances, correlation values from the SSS method is significantly inferior to the values obtained by the MD method. Numerical examples are presented to demonstrate the new method. It is shown that the presented method is much more efficient than the MD method and more accurate than the SSS method. The new algorithm is also versatile: it can directly simulate fields of irregular geometry without additional efforts. Copyright © 2003 John Wiley & Sons, Ltd.
Mixed variational methods for finite element analysis of geometrically non‐linear, inelastic Bernoulli–Euler beams
Authors: Hjelmstad K. D. , Taciroglu E.
Published: October 2003

Bernoulli–Euler beam theory has long been the standard for the analysis of reticulated structures. The need to accurately compute the non‐linear (material and geometric) response of structures has renewed interest in the application of mixed variational approaches to this venerable beam theory. Recent contributions in the literature on mixed methods and the so‐called (but quite related) non‐linear flexibility methods have left open the question of what is the best approach to the analysis of beams. In this paper we present a consistent computational approach to one‐, two‐, and three‐field variational formulations of non‐linear Bernoulli–Euler beam theory, including the effects of non‐linear geometry and inelasticity. We examine the question of superiority of methods through a set of benchmark problems with features typical of those encountered in the structural analysis of frames. We conclude that there is no clear winner among the various approaches, even though each has predictable computational strengths. Copyright © 2003 John Wiley & Sons, Ltd.
Optimizing operating system performance for CC‐NUMA architectures
Authors: Chang Moon Seok
Published: October 2003

The CC‐NUMA (cache‐coherent non‐uniform memory access) architecture is an attractive solution to scalable servers. The performance of a CC‐NUMA system heavily depends on the number of accesses to remote memory through an interconnection network. To reduce the number of remote accesses, an operating system needs to exploit the potential locality of the architecture. This paper describes the design and implementation of a UNIX‐based operating system supporting the CC‐NUMA architecture. The operating system implements various enhancements by revising kernel algorithms and data structures. This paper also analyzes the performance of the enhanced operating system by running commercial benchmarks on a real CC‐NUMA system. The performance analysis shows that the operating system can achieve improved performance and scalability for CC‐NUMA by implementing kernel data striping, localization and load balancing. Copyright © 2003 John Wiley & Sons, Ltd.
Determining the appropriate amount of data for classifying consumers for direct marketing purposes
Authors: Heilman Carrie M. , Kaefer Frederick , Ramenofsky Samuel D.
Published: October 2003

This article examines the impact of using incremental amounts of purchasing data on the ability to classify consumers in consumer packaged goods categories for direct marketing purposes. Building on the work of Rossi, McCulloch, and Allenby (1996), who focused on the impact of three information sets—(a) demographics only, (b) demographics and one purchase made by a consumer, and (c) demographics plus an entire purchasing history of a consumer—we examine the impact of each additional purchase, starting with no purchasing information (i.e., demographics only) through 20 purchases. Using two different classification models, a Multinomial Logit model and an Artificial Neural Network model, we examine the sensitivity of classification accuracy to each additional purchase. We use these results in a profitability analysis of a hypothetical direct marketing campaign to determine the optimal number of purchases to use for classification in the category studied. The findings suggest an optimal number of purchasing observations exists for classification and targeting purposes and this optimal number falls between one purchase and a “history” of purchases as studied by Rossi et al. Our findings illustrate the importance of conducting a sensitivity analysis to identify the optimal amount of purchasing data to use when classifying consumers for the purpose of a direct marketing campaign.
Consumer search for information in the digital age: An empirical study of prepurchase search for automobiles
Authors: Klein Lisa R. , Ford Gary T.
Published: October 2003

This research explores whether earlier findings on the economics of consumer information search still hold in the digital age. Through an Internet-based survey conducted with automobile shoppers and purchasers, we examine how individuals differ in the use of the Internet and in the patterns of substitution they exhibit across sources and media. Survey results show that basic economics continues to drive information search in general, measured in terms of amount (time) and breadth (number of sources). However, analysis of consumers’ information search behavior suggests that an additional dimension of search—whether the source is offline or online—needs to be added to the traditional two dimensions of impersonal/personal and independent/seller-dominated sources. We also analyze trends in the use of the Internet as an information source based on recency of respondents’ shopping experiences.
Should a company have message boards on its Web sites?
Authors: Chiou Jyh-Shen , Cheng Cathy
Published: October 2003

This study examined the impact of messages on a discussion forum regarding a consumer's brand evaluation and attitude toward the Web owner. Four hypotheses were developed based on consumer behavior theories. An experiment using a 2×3×2 between participants factorial design was conducted to test the hypotheses. The results showed that message favorableness and message number on an Internet discussion forum can influence a consumer's brand evaluation and attitude toward the Web owner. However, the effects are different under different levels of preexisting brand image. Negative messages hurt mainly low-image brands, whereas message repetition helps only high-image brands.
An experimental study of the role of brand strength in the relationship between the medium of communication and perceived credibility of the message
Authors: Marshall Roger , WoonBong Na
Published: October 2003

Does a message on the Internet have less credibility than an equivalent message in conventional print media? What is the role of brand strength in the relation between the medium of communication and perceived credibility of the message? We address these questions through an experimental study. The results show that a message on the Internet has less credibility than the same message in print. For messages of strong brands, however, there is no significant difference in their credibility between the Internet and print media.
Genetic and genomic tools for zebrafish research: The NIH zebrafish initiative
Authors: Rasooly Rebekah S. , Henken Deborah , Freeman Nancy , Tompkins Laurie , Badman David , Briggs Josephine , Hewitt A. Tyl
Published: October 2003

The National Institutes of Health (NIH) has been a leading advocate of the zebrafish as a model organism for the study of vertebrate development, physiology, and disease. Genomic tools, developed with the support of NIH funding, have made zebrafish even more attractive as a genetic system and have stimulated research using this model. The NIH continues to provide support for new and existing community resources, such as the Zebrafish International Resource Center and the Zebrafish Information Network, research tool development and mutant screens, and a wide array of investigator‐initiated studies that are using zebrafish to elucidate many aspects of vertebrate biology. Developmental Dynamics, 2003. Published 2003 Wiley‐Liss, Inc.
Expression of zebrafish Lc3 synthase gene in embryonic lens requires hedgehog signaling
Authors: Cao Ying , Zhao Jue , Wang Ying , Meng Anming
Published: October 2003

Glycosyltransferases are involved in synthesis of various glycolipids and glycoproteins that play important roles in many biological processes. We have identified a zebrafish gene encoding a member of β1,3‐N‐acetylglucosaminyltransferase family, the Lc3 synthase/bGn‐T5. Whole‐mount in situ hybridization reveals that the Lc3 synthase gene is expressed in two distinct phases during the zebrafish embryogenesis. The early phase extends from late blastulation to the completion of epiboly, during which the expression occurs in the superficial layer of the embryos. The second phase of expression starts during mid‐segmentation and persists until day 3, during which the expression occurs prominently in the developing lens. The expression of the Lc3 synthase gene in the lens is inhibited in you‐too (yot) mutant embryos that are defective in Hedgehog signaling. The expression in the lens also decreases in cyclops (cyc) and one‐eyed‐pinhead (oep) mutant embryos and lefty1‐injected embryos, which are deficient in Nodal signaling and lack Hedgehog activity in the ventral brain. These results suggest that Hedgehog signaling is required for the Lc3 synthase expression in embryonic lens. Developmental Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
Expression and phylogenetic analyses of three zebrafish FoxI class genes
Authors: Solomon Keely S. , Logsdon John M. , Fritz Andreas
Published: October 2003

We have identified three novel members of the zebrafish forkhead class I gene family, which we have named foxi2, foxi3a, and foxi3b. We have reported previously the identification of zebrafish foxi1, which is required for otic placode and jaw development. Expression analysis shows that foxi2 is expressed within the chordamesoderm during early somitogenesis and the retina and pharyngeal arches during later stages. The foxi3a and foxi3b genes likely represent a recently duplicated pair, and they are similarly expressed in epidermal mucous cells throughout embryogenesis and early larval stages. None of these newly identified FoxI genes are expressed in otic precursor cells and, therefore, are unlikely to share functional overlap with foxi1 in the development of the inner ear. In addition to these zebrafish FoxI paralogs, we have identified 16 new FoxI sequences in species ranging from Ciona intestinalis to Homo sapiens. We present an extensive phylogenetic analysis of the FoxI class that includes these new sequences together with those previously reported. This analysis supports the existence of three subfamilies within the FoxI class, each containing at least one zebrafish member. Developmental Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
Insertional mutagenesis in zebrafish
Authors: Amsterdam Adam
Published: October 2003

Insertional mutagenesis is a method for identifying genes essential for a given biological process by using the integration of DNA as the mutagen, thereby facilitating the cloning of the mutated gene. The use of retrovirus‐mediated insertional mutagenesis in zebrafish has led to the mutation and rapid identification of hundreds of genes required for embryonic development and cell viability and growth, revealing the diversity of gene products required for the development of this vertebrate. Here, I will review the methodology of this approach and the results to date, as well as other potential ways to use insertional mutagenesis for genetic screens. Developmental Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
Integrity of the midbrain region is required to maintain the diencephalic–mesencephalic boundary in zebrafishno isthmus/pax2.1mutants
Authors: Scholpp Steffen , Brand Michael
Published: October 2003

Initial anterior–posterior patterning of the neural tube into forebrain, midbrain, and hindbrain primordia occurs already during gastrulation, in response to signals patterning the gastrula embryo. After the initial establishment, further development within each brain part is thought to proceed largely independently of the others. However, mechanisms should exist that ensure proper delineation of brain subdivisions also at later stages; such mechanisms are, however, poorly understood. In zebrafishno isthmusmutant embryos, inactivation of thepax2.1gene leads to a failure of the midbrain and isthmus primordium to develop normally from the gastrula stage onward (Lun and Brand [1998] Development 125:3049–3062). Here, we report that, after the initially correct establishment during gastrulation stages, the neighbouring forebrain primordium and, partially, the hindbrain primordium expand into the misspecified midbrain territory inno isthmusmutant embryos. The expansion is particularly evident for the posterior part of the diencephalon and less so for the first rhombomeric segment, the territories immediately abutting the midbrain/isthmus primordium. The nucleus of the posterior commissure is expanded in size, and marker genes of the forebrain and rhombomere 1 expand progressively into the misspecified midbrain primordium, eventually resulting in respecification of the midbrain primordium. We therefore suggest that the genetic program controlled by Pax2.1 is not only involved in initiating but also in maintaining the identity of midbrain and isthmus cells to prevent them from assuming a forebrain or hindbrain fate. Developmental Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
Postembryonic development of the cranial lateral line canals and neuromasts in zebrafish
Authors: Webb Jacqueline F. , Shirey Jonathan E.
Published: October 2003

The development of the cranial lateral line canals and neuromast organs are described in postembryonic zebrafish (0–80 days postfertilization). Cranial canal development commences several weeks after hatch, is initiated in the vicinity of individual neuromasts, and occurs in four discrete stages that are described histologically. Neuromasts remain in open canal grooves for several weeks during which they dramatically change shape and increase in size by adding hair cells at a rate one‐tenth that in the zebrafish inner ear. Scanning electron microscopy demonstrates that neuromasts elongate perpendicular to the canal axis and the axis of hair cell polarization and that they lack a prominent nonsensory cell population surrounding the hair cells—features that make zebrafish neuromasts unusual among fishes. These results demand a reassessment of neuromast and lateral line canal diversity among fishes and highlight the utility of the lateral line system of postembryonic zebrafish for experimental and genetic studies of the development and growth of hair cell epithelia. Developmental Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
Two divergent slit1 genes in zebrafish
Authors: Hutson Lara D. , Jurynec Michael J. , Yeo Sang‐Yeob , Okamoto Hitoshi , Chien Chi‐Bin
Published: October 2003

Members of the Slit family regulate axon guidance and cell migration. To date, three vertebrate slit1 genes have been identified in mammals and orthologs of two, slit2 and slit3, have been identified in zebrafish. Here, we describe the cloning of full‐length cDNAs for two zebrafish slit orthologs, slit1a and slit1b. Both predicted proteins contain the conserved motifs that characterize other vertebrate Slits. slit1a and slit1b are both expressed in the midline, hypochord, telencephalon, and hindbrain. Apart from these shared expression domains, however, their expression patterns largely differ. Whereas slit1a is expressed broadly in the central nervous system (CNS) and in the somites, pectoral fin buds, tail bud, and caudal fin folds, slit1b is expressed in the olfactory system throughout embryonic and larval development, and in the retina during larval stages. Their expression patterns, particularly that of slit1a, suggest that Slit proteins may have roles in tissue morphogenesis in addition to their established roles in axon guidance and cell migration. Developmental Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
Developmental morphology of the axial skeleton of the zebrafish,Danio rerio(Ostariophysi: Cyprinidae)
Authors: Bird Nathan C. , Mabee Paula M.
Published: October 2003

Before our rapidly increasing knowledge of gene interactions can be connected with the morphologic defects in mutant zebrafish, the normal course of skeletal development must be understood. Here, we describe the developmental morphology of the axial skeleton of zebrafish and review it in relation to the morphology of related species. The relative sequence of ossification in the skeleton is described. Two separate centers of development were found in the axial skeleton (Weberian apparatus and caudal fin) in contrast to tetrapods, which have a single anterior center. Slight variation was found in the overall relative timing of development. The extensive ichthyological literature on teleost anatomy and recent genetic data form the basis for our review and interpretation of homologies of various elements of the axial skeleton. Because homology forms the basis for all evolutionary comparisons, these data are critical for integration in evo–devo studies. Developmental Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
Runx3 is required for hematopoietic development in zebrafish
Authors: Kalev‐Zylinska Maggie L. , Horsfield Julia A. , Flores Maria Vega C. , Postlethwait John H. , Chau Jackie Y.M. , Cattin Peter M. , Vitas Maria R. , Crosier Philip S. , Crosier Kathryn E.
Published: October 2003

We cloned zebrafish runx3/aml2/cbfa3 and examined its expression and function during embryogenesis. In the developing embryo, runx3 is dynamically expressed in hematopoietic, neuronal, and cartilaginous tissues. Hematopoietic expression of runx3 commences late in embryogenesis in the ventral tail intermediate cell mass and later colocalizes with spi1 and lyz in circulating blood cells. In the cloche mutant, hematopoietic expression was absent, suggesting that Runx3 functions downstream of cloche in a hematopoietic pathway. Neuronal tissues expressing runx3 include the trigeminal ganglia and Rohon‐Beard neurons. Runx3 appears to contribute to normal development of primitive and definitive hematopoietic cells. When Runx3 function was compromised using morpholino oligonucleotides, a reduction in the number of mature blood cells was observed. Furthermore, Runx3 depletion decreased runx1 expression in the ventral wall of the dorsal aorta and reduced the number of spi1‐ and lyz‐containing blood cells. Conversely, ubiquitous overexpression of runx3 led to an increase in primitive blood cell numbers, together with an increase in runx1‐expressing cells in the ventral wall of the dorsal aorta. We propose a role for Runx3 in the regulation of blood cell numbers. Developmental Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
Functional genomic dissection of multimeric protein families in zebrafish
Authors: Cheng Keith C. , Levenson Robert , Robishaw Janet D.
Published: October 2003

The study of multimeric protein function in the postgenomicera has become complicated by the discovery of multiple isoforms for each subunit of those proteins. A correspondingly large number of potential isoform combinations offer the multicellular organism a constellation of protein assemblies from which to generate a variety of functions across different cells, tissues, and organs. At the same time, the multiplicity of potential subunit isoform combinations presents a significant challenge when attempting to dissect the functions of particular isoform combinations. Biochemical and cell culture methods have brought us to a significant state of understanding of multimeric proteins but are unable to answer questions of function within the context of the many tissues and developmental stages of the multicellular organism. Answering those questions can be greatly facilitated in model systems in which expression can be determined over time, in the context of the whole organism, and in which hypomorphic function of each subunit can be studied individually and in combination. Fortunately, the potential for high‐throughput in situ hybridization studies and antisense‐based reverse genetic knockdowns in zebrafish offers exciting opportunities to meet this challenge. Some of these opportunities, along with cautions of interpretation and gaps in the existing technologies, are discussed in the context of ongoing investigations of the dimeric Na,K‐ATPases and heterotrimeric G proteins. Development Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
Maternal factors in zebrafish development
Authors: Pelegri Francisco
Published: October 2003

All processes that occur before the activation of the zygotic genome at the midblastula transition are driven by maternal products, which are produced during oogenesis and stored in the mature oocyte. Upon egg activation and fertilization, these maternal factors initiate developmental cascades that carry out the embryonic developmental program. Even after the initiation of zygotic gene expression, perduring maternal products continue performing essential functions, either together with other maternal factors or through interactions with newly expressed zygotic products. Advances in zebrafish research have placed this organism in a unique position to contribute to a detailed understanding of the role of maternal factors in early vertebrate development. This review summarizes our knowledge on the processes involved in the production and redistribution of maternal factors during zebrafish oogenesis and early development, as well as our understanding of the function of these factors in axis formation, germ layer and germ cell specification, and other early embryonic processes. Developmental Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
Transgene driving GFP expression from the promoter of the zona pellucida gene zpc is expressed in oocytes and provides an early marker for gonad differentiation in zebrafish
Authors: Onichtchouk Daria , Aduroja Kristin , Belting Heinz‐Georg , Gnügge Lara , Driever Wolfgang
Published: October 2003

Although mechanisms of sex differentiation have been studied intensely in mammals, insects, and worms, little is known about this process in lower vertebrates. To establish a marker for female gonad differentiation in zebrafish, we generated a transgenic line in which 412 bp from the promoter and 5′ mRNA leader of the female‐specific zebrafish zona pellucida gene zpc are fused to the coding region of green fluorescent protein (GFP). The zpc0.5:GFP transgene is expressed exclusively in oocytes, starting from the onset of female‐specific differentiation, and closely resembles the expression pattern of the wild‐type zpc. Strong GFP expression persists throughout oogenesis and is visible through the body wall of females. We have also characterized a putative upstream factor of zpc, FIGalpha, and show that distribution of FIGalpha RNA is compatible with its postulated role in the regulation of zpc. The zpc0.5:GFP transgenic line described here will be useful for studying oocyte development and the mechanisms that determine sex‐specific gene expression in the zebrafish. It is also the first promoter characterized to date to drive stable and efficient expression specifically in the zebrafish female germline. Development Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
Modeling human hematopoietic and cardiovascular diseases in zebrafish
Authors: North Trista E. , Zon Leonard I.
Published: October 2003

Zebrafish have emerged as a useful vertebrate model system in which unbiased large‐scale screens have revealed hundreds of mutations affecting vertebrate development. Many zebrafish mutants closely resemble known human disorders, thus providing intriguing prospects for uncovering the genetic basis of human diseases and for the development of pharmacologic agents that inhibit or correct the progression of developmental disorders. The rapid pace of advances in genomic sequencing and map construction, in addition to morpholino targeting and transgenic techniques, have facilitated the identification and analysis of genes associated with zebrafish mutants, thus promoting the development of zebrafish as a model for human disorders. This review aims to illustrate how the zebrafish has been used to identify unknown genes, to assign function to known genes, and to delineate genetic pathways, all contributing valuable leads toward understanding human pathophysiology. Developmental Dynamics 2003. © 2003 Wiley‐Liss, Inc.
Targeted gene knockdown in zebrafish using negatively charged peptide nucleic acid mimics
Authors: Urtishak Karen A. , Choob Michael , Tian Xiaobing , Sternheim Nitzan , Talbot William S. , Wickstrom Eric , Farber Steven A.
Published: October 2003

Negatively charged homo‐oligomers of alternating trans‐4‐hydroxy‐L‐proline/phosphonate polyamides with DNA bases (HypNA‐pPNA) display excellent hybridization properties toward DNA and RNA, while preserving the mismatch discrimination, nuclease resistance, and protease resistance of peptide nucleic acids (PNAs). Similar properties are associated with morpholino phosphorodiamidate (MO) DNA mimics, which have been used in the model vertebrate zebrafish (Danio rerio) for genome‐wide, sequence‐based, reverse genetic screens during embryonic development. We evaluated mixed sequence HypNA‐pPNAs as an alternative to MOs, and found that even a single central DNA mismatch lowered the HypNA‐pPNA melting temperature by 16°C. We then observed that the melting temperatures of HypNA‐pPNA 18‐mers hybridized to RNA 25‐mers were comparable to the melting temperatures of MO 25‐mers, and that two HypNA‐pPNA mismatches lowered the melting temperature with RNA by 18°C. In zebrafish embryos we observed that HypNA‐pPNA 18‐mers displayed comparable potency to MO 25‐mers as knockdown agents against chordin, notail, and uroD, with greater mismatch stringency. Finally we observed that a specific HypNA‐pPNA 18‐mer elicited the dharma (bozozok)‐/‐ phenotype in zebrafish embryos, which MO 25‐mers do not. HypNA‐pPNAs designed to inhibit translation of specific zebrafish RNA targets thus demonstrated stringent hybridization properties, relative to DNA and MO oligomers, and present a valuable alternative for reverse genetic studies, enabling the targeting of previously inaccessible genes. Developmental Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
Imaging brain development and organogenesis in zebrafish using immobilized embryonic explants
Authors: Langenberg Tobias , Brand Michael , Cooper Mark S.
Published: October 2003

Owing to its optical clarity and rapid rate of development, the zebrafish embryo is an ideal model system for studying the cellular mechanics of organogenesis. Unfortunately, extended time‐lapse recordings of zebrafish embryos are often disrupted by the extension and straightening of the embryonic axis, as well as movement artifacts associated with developing musculature. In addition, the embryo's massive yolk cell often prevents optical access to tissues of interest. To circumvent these imaging problems, we have developed a procedure to deflate and mechanically remove the yolk cell. A “paralyzing” agent, AMP‐PNP (a membrane‐impermeant nonhydrolyzable analog of ATP), is first injected into the embryo's contractile yolk cell. The yolk cell is then removed using sharpened tungsten needles. Deyolked embryos, or organ rudiments explanted from them, are then immobilized on a microscope coverslip using a thin plasma clot. This plasma clot immobilization allows novel mountings of the explants so that ventral, lateral, and even cross‐sectional fields of views are possible using high numerical aperture objectives. We show that isolated head rudiments undergo normal morphogenesis and gene expression for at least 1 day after being explanted into organotypic culture. These procedures can be used to study the cellular mechanics of organogenesis in “deyolked” embryos, as well as in tissues explanted from green fluorescent protein transgenic animals. Developmental Dynamics 2003. © 2003 Wiley‐Liss, Inc.
A MAGE/NDN‐like gene in zebrafish
Authors: Bischof Jocelyn M. , Ekker Marc , Wevrick Rachel
Published: October 2003

The human necdin/MAGE gene family has over 50 members, but most of the proteins encoded by these genes are of unknown function. We have now identified a single locus in Danio rerio that encodes a putative protein with significant coding sequence similarity to the mammalian NDN/MAGE genes. Analysis of the complete Fugu ribripes genome sequence also suggests that there is only a single MAGE‐like gene in teleost fish. mage is expressed in the larval and adult brain, specifically the retina, the medial region of the telencephalon, periventricular gray zone of the optic tectum, and most highly in the cerebellar corpus. The discovery of a zebrafish NDN/MAGE gene expressed the developing brain facilitates studies of the MAGE homology domain in vertebrate development. Developmental Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
Histology‐based screen for zebrafish mutants with abnormal cell differentiation
Authors: Mohideen Manzoor‐Ali P.K. , Beckwith Lee G. , Tsao‐Wu Gladys S. , Moore Jessica L. , Wong Andrew C.C. , Chinoy Mala R. , Cheng Keith C.
Published: October 2003

The power of histology to define states of cell differentiation was used as the basis of a mutagenesis screen in zebrafish. In this screen, 7‐day‐old parthenogenetic half‐tetrad larvae from potential carrier females were screened for mutations affecting cell differentiation in hematoxylin and eosin‐stained tissue sections. Seven, noncomplementing, recessive mutations were found. Two mutations affect only the retina: segmented photoreceptors (spr) show a discontinuous photoreceptor cell layer; vestigial outer segments (vos) has fewer photoreceptor cells and degenerated outer segments within this cell layer. Three mutants have gut‐specific defects: the epithelial cells of kirby (kby) are replaced by ballooned cells; the intestines of stuffy (sfy) and stuffed (sfd) contain increased luminal mucus. Two mutations affect multiple organs: disordered neural retina (dnr) has disrupted retinal layering and mild nuclear abnormalities in the gut and liver; and in huli hutu (hht), the retinal cell layers are disorganized and multiple organs have mild to severe nuclear abnormalities that are reminiscent of the atypia of human neoplasia. Each mutation appears to be homozygous lethal. This screen is proof of principle for the feasibility of histologic screens to yield novel mutations, including potential models of human disease. The throughput for this type of screen may be enhanced by automation. Developmental Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
Direct binding of Lef1 to sites in the boz promoter may mediate pre–midblastula‐transition activation of boz expression
Authors: Leung Tinchung , Söll Iris , Arnold Sebastian J. , Kemler Rolf , Driever Wolfgang
Published: October 2003

The Nieuwkoop center provides signals essential for the establishment of the dorsal gastrula organizer in vertebrates. Activation of β‐catenin is one of the events in the Nieuwkoop center that lead to activation of dorsal‐specific genes during blastula and early gastrula stages. Zebrafish bozozok (boz) mutant embryos have severe defects in axial mesoderm and anterior neuroectoderm. The boz gene is activated in the organizer in response to β‐catenin signaling, and Boz protein has been demonstrated to contribute to organizer formation by repression of ventralizing genes, including bmp2b, vega1, and vega2. Here, we investigate the timing and molecular mechanism by which boz expression is activated in the organizer. We demonstrate that boz is already expressed before midblastula transition (MBT). We further identify high‐affinity binding sites for Tcf/Lef1 within the boz promoter region. These sites, together with the finding that β‐catenin induces boz expression, indicate that transcription of boz may be activated directly by β‐catenin/Lef1. We hypothesize that pre‐MBT activation of boz may be important to build up a sufficiently strong antagonizing activity against zygotic ventralizing genes activated immediately post‐MBT. Thus, the early onset of boz expression may be crucial for organizer establishment in the presence of ubiquitous maternal activators of ventralizing genes. Developmental Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
gbx2Homeobox gene is required for the maintenance of the isthmic region in the zebrafish embryonic brain
Authors: Kikuta Hiroshi , Kanai Maiko , Ito Yukiko , Yamasu Kyo
Published: October 2003

We isolated cDNA clones for the zebrafishgbx2gene, which is implicated in the establishment of the midbrain–hindbrain boundary (MHB) in other vertebrates. Spatially localized expression ofgbx2was observed at the MHB from 90% epiboly through to the hatching stage. Comparisons with the expression ofotx2,wnt1, andkrox20showed thatgbx2is expressed in the anterior hindbrain. Ectopic expression ofgbx2by mRNA injection caused cyclopia or truncation of the fore‐ and midbrain and severely affected isthmic and cerebellar structures, while hindbrain formation was not significantly affected. At the molecular level,gbx2suppressed the expression ofotx2in the fore/midbrain,six3in the anterior forebrain, and MHB‐specific genes such aseng2andwnt1. In contrast,gbx2did not down‐regulate the expression of the hindbrain marker genes. Therefore,gbx2specifically suppressed the formation of the entire fore/midbrain. Meanwhile, misexpression ofotx2suppressed the expression ofgbx2in the embryonic brain. Abrogation ofgbx2expression with an antisense morpholino oligonucleotide disrupted the midbrain/anterior hindbrain region, and these loss‐of‐function effects were rescued by activating the Gbx2 protein immediately after the end of gastrulation. Taken together, these results suggest that the zebrafishgbx2gene is essential for the maintenance of MHB and/or the formation of the isthmic structure during somitogenesis, rather than for the MHB establishment during gastrulation. We also suggest that other factors, includinggbx1, is required for the establishment of the MHB during gastrulation. Developmental Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
Genome duplication, subfunction partitioning, and lineage divergence:Sox9in stickleback and zebrafish
Authors: Cresko William A. , Yan Yi‐Lin , Baltrus David A. , Amores Angel , Singer Amy , Rodríguez‐Marí Adriana , Postlethwait John H.
Published: October 2003

Teleosts are the most species‐rich group of vertebrates, and a genome duplication (tetraploidization) event in ray‐fin fish appears to have preceded this remarkable explosion of biodiversity. What is the relationship of the ray‐fin genome duplication to the teleost radiation? Genome duplication may have facilitated lineage divergence by partitioning different ancestral gene subfunctions among co‐orthologs of tetrapod genes in different teleost lineages. To test this hypothesis, we investigated gene expression patterns forSox9gene duplicates in stickleback and zebrafish, teleosts whose lineages diverged early in Euteleost evolution. Most expression domains appear to have been partitioned betweenSox9aandSox9bbefore the divergence of stickleback and zebrafish lineages, but some ancestral expression domains were distributed differentially in each lineage. We conclude that some gene subfunctions, as represented by lineage‐specific expression domains, may have assorted differently in separate lineages and that these may have contributed to lineage diversification during teleost evolution. Developmental Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
Neurogenic phenotype of mind bomb mutants leads to severe patterning defects in the zebrafish hindbrain
Authors: Bingham Stephanie , Chaudhari Summer , Vanderlaan Gary , Itoh Motoyuki , Chitnis Ajay , Chandrasekhar Anand
Published: October 2003

Failure of Notch signaling in zebrafish mind bomb (mib) mutants results in a neurogenic phenotype where an overproduction of early differentiating neurons is accompanied by the loss of later‐differentiating cell types. We have characterized in detail the hindbrain phenotype of mib mutants. Hindbrain branchiomotor neurons (BMNs) are reduced in number but not missing in mib mutants. In addition, BMN clusters are frequently fused across the midline in mutants. Mosaic analysis indicates that the BMN patterning and fusion defects in the mib hindbrain arise non–cell autonomously. Ventral midline signaling is defective in the mutant hindbrain, in part due to the differentiation of some midline cells into neural cells. Interestingly, while early hindbrain patterning appears normal in mib mutants, subsequent rhombomere‐specific gene expression is completely lost. The defects in ventral midline signaling and rhombomere patterning are accompanied by an apparent loss of neuroepithelial cells in the mutant hindbrain. These observations suggest that, by regulating the differentiation of neuroepithelial cells into neurons, Notch signaling preserves a population of non‐neuronal cells that are essential for maintaining patterning mechanisms in the developing neural tube. Developmental Dynamics, 2003. © 2003 Wiley‐Liss, Inc.
Adults with learning disabilities: Differences between The Netherlands and Flanders
Authors: Ruijssenaars A.J.J.M. , Ghesquière Pol
Published: October 2003

In this paper, similarities and differences are outlined between adults with self reported learning disabilities (SRLD) drawn from the International Adult Literacy Survey (IALS) data on adult literacy in The Netherlands and Flanders. These results are discussed in terms of striking differences in the educational context within which the adults developed. The literacy skills of SRLD adults in The Netherlands are superior to SRLD adults in Flanders. This is interpreted in terms of societal acceptance of disability. Copyright © 2003 John Wiley & Sons, Ltd.
Adults with self‐reported learning disabilities in Slovenia: Findings from the international adult literacy survey on the incidence and correlates of learning disabilities in Slovenia
Authors: Magajna Lidija , Kavkler Marija , Ortar‐Križaj Martina
Published: October 2003

This study of adults with self‐reported learning disabilities (SRLD) in Slovenia is part of a larger secondary analysis of the data from the International Literacy Survey project (IALS). The purpose of the study was to examine the characteristics of 79 (2.68%) individuals who reported experiencing learning disabilities and compare them to the general population on a variety of indicators of educational background, employment status, and reading and writing activities at work and at home. The proficiency scores of the SRLD individuals were lower in all three literacy domains (prose, document and quantitative literacy). In prose literacy 77.9% of SRLD adults performed at Level 1 and only 7.8% reached the level necessary for a modern technological society. Experiencing learning disabilities was not related to gender or age, however, results showed significant differences between the levels achieved by older and younger people with SRLD. In SRLD groups aged 40 years and above, no one achieved more than the second level of literacy in any domain. Learning disabilities were reported more frequently in rural areas.

SRLD groups achieve significantly lower educational attainment, and lower employment status, with a preference for manual labour or craft. These findings are of critical importance. SRLD people report that poorer literacy skills are an obstacle to their progression in employment. In the Slovene sample, the SRLD group stands out for low scores in quantitative literacy. Results show that they are less active, pick up information only auditorily or in short written form. They need more frequent help from relatives in literacy activities.

Interpretation of the IALS data on SRLD presents many problems. These include amongst others, problems in terminology, different background factors, and the validity of self‐report measures. However, the study also raises many interesting challenges for future research and policy. Increasing the availability of support, assistance and counselling for adolescents and adults with learning disabilities remains a very important goal for dyslexia and LD policies in Slovenia. Copyright © 2003 John Wiley & Sons, Ltd.
A comparative study of adults with and without self‐reported learning disabilities in six English‐speaking populations: What have we learned?
Authors: Vogel Susan A. , Holt Janet K.
Published: October 2003

The purpose of this study was to compare adults with and without self‐reported learning disabilities (SRLD and NSRLD) from six English‐speaking populations including: English‐speaking Canada, Great Britain, The Republic of Ireland, New Zealand, Northern Ireland, and the United States. These six populations were selected because they were all English‐speaking populations, participated in the first administration of the International Adult Literacy Survey (IALS), and included the optional questions regarding the presence of a disability. In this study, we compared the groups on prevalence by population, percentage of each group by age and gender, awareness of learning disabilities and problems in school, document and quantitative literacy proficiency, educational attainment, reasons for dropping out of school, and employment, occupational and financial status. Findings were reported among these six populations within an historical perspective including differences in awareness and definition of learning disabilities, public policy, special education services, reading pedagogy, and teacher preparation. Recommendations are made for improving literacy and long‐term outcomes for those with learning disabilities in all nations as well as future research directions. Copyright © 2003 John Wiley & Sons, Ltd.
Endothelial cell cytoprotection induced in vitro by allo‐ or xenoreactive antibodies is mediated by signaling through adenosine A2 receptors
Authors: Delikouras Alexandros , Fairbanks Lynette D. , Simmonds Anne H. , Lechler Robert I. , Dorling Anthony
Published: October 2003

Endothelial cell (EC) expression of proteins such as hemoxygenase‐1 or Bcl‐xL is associated with enhanced survival of vascularized allo‐ or xenografts. These grafts are resistant to humoral rejection, a phenomenon called accommodation. In vitro, low concentrations of allo‐ and xenoreactive antibody (XNA) induce a cytoprotective phenotype in EC similar to that seen in accommodated grafts. In this study we examine whether adenosine plays a role in antibody‐induced cytoprotection. Porcine EC were incubated with human anti‐pig XNA or specific alloantibody. EC expressed Bcl‐xL and were protected from TNF‐mediated apoptosis. Bcl‐xL expression was inhibited by an adenosine A2 receptor antagonist. Human anti‐pig XNA were shown to bind and induce cyclic adenosine 3′,5′‐monophosphate (cAMP) generation through these receptors. This activity was abolished by depletion of anti‐galα(1–3)gal‐specific XNA. In contrast, alloantibody caused adenosine production. Protection from TNF‐mediated apoptosis was also mediated through A2 receptor but involved additional non‐cAMP‐dependent signaling. This study indicates a molecular mechanism common to both antibody‐mediated cytoprotection and ischemic preconditioning and suggests a potential therapeutic avenue based on adenosine for improving the outcome of transplanted grafts in those patients with pre‐existing anti‐graft antibody.