Autoinhibition by an internal nuclear localization signal revealed by the crystal structure of mammalian importin alpha

Importin alpha is the nuclear import receptor that recognizes classical monopartite and bipartite nuclear localization signals (NLSs). The structure of mouse importin alpha has been determined at 2.5 Angstrom resolution. The structure shows a large C-terminal domain containing armadillo repeats, and a less structured N-terminal importin beta-binding domain containing an internal NLS bound to the NLS-binding site. The structure explains the regulatory switch between the cytoplasmic, high-affinity form, and the nuclear, low-affinity form for NLS binding of the nuclear import receptor predicted by the current models of nuclear import. Importin beta conceivably converts the low- to high-affinity form by binding to a site overlapping the autoinhibitory sequence. The structure also has implications for understanding NLS recognition, and the structures of armadillo and HEAT repeats.

Analysis of the substrate specificity of human sulfotransferases SULT1A1 and SULT1A3: Site-directed mutagenesis and kinetic studies

Sulfonation is an important metabolic process involved in the excretion and in some cases activation of various endogenous compounds and xenobiotics. This reaction is catalyzed by a family of enzymes named sulfotransferases. The cytosolic human sulfotransferases SULT1A1 and SULT1A3 have overlapping yet distinct substrate specificities. SULT1A1 favors simple phenolic substrates such as p-nitrophenol, whereas SULT1A3 prefers monoamine substrates such as dopamine. In this study we have used a variety of phenolic substrates to functionally characterize the role of the amino acid at position 146 in SULT1A1 and SULT1A3. First, the mutation A146E in SULT1A1 yielded a SULT1A3-like protein with respect to the Michaelis constant for simple phenols. The mutation E146A in SULT1A3 resulted in a SULT1A1-like protein with respect to the Michaelis constant for both simple phenols and monoamine compounds. When comparing the specificity of SULT1A3 toward tyramine with that for p-ethylphenol (which differs from tyramine in having no amine group on the carbon side chain), we saw a 200-fold preference for tyramine. The kinetic data obtained with the E146A mutant of SULT1A3 for these two substrates clearly showed that this protein preferred substrates without an amine group attached. Second, changing the glutamic acid at position 146 of SULT1A3 to a glutamine, thereby neutralizing the negative charge at this position, resulted in a 360-fold decrease in the specificity constant for dopamine. The results provide strong evidence that residue 146 is crucial in determining the substrate specificity of both SULT1A1 and SULT1A3 and suggest that there is a direct interaction between glutamic acid 146 in SULT1A3 and monoamine substrates.

Miscibility and specific interactions in blends of poly(4-vinylphenol) and poly(2-ethoxyethyl methacrylate)

The properties of the hydrogen-bonded polymer blends of poly(4-vinylphenol) and poly(2-ethoxyethyl methacrylate) are presented. Spectroscopic techniques such as C-13 solid-state NMR and FT-IR are used to probe specific interactions of the blends at various compositions. Spectral features from both techniques revealed that site-specific interactions are present, consistent with a significant degree of mixing of the blend components. Changes in chemical shift and line shape of the phenolic carbon and carbonyl resonances in the C-13 CPMAS spectra of the blends as a function of composition are interpreted as resulting from changes in the relative intensities of two closely overlapped signals. A quantitative measure of hydrogen-bonded carbonyl groups using C-13 NMR has been obtained which agreed well with the results from FT-IR analyses. It is also shown that C-13 NMR can be used to measure the fraction of hydroxyl groups associated with carbonyl groups, which was not possible previously using FT-IR due to extensive overlapping of bands in the hydroxyl stretching region. The results of measurements of H-1 T-1 and 1H T-1 rho indicate that PVPh and PEEMA are intimately mixed on a scale less than 2-3 nm.

A model for assembly and activation of the GM-CSF, IL-3 and IL-5 receptors: Insights from activated mutants of the common beta subunit

Granulocyte-macrophage colony stimulating factor (GM-CSF), Interleukin-3 (IL-3) and Interleukin-5 (IL-5) have overlapping, pleiotropic effects on hematopoietic cells, including neutrophils, eosinophils, monocytes and early progenitor cells. The high-affinity receptors for human GM-CSF, IL-3, and IL-5 share a common beta-subunit (h beta(c)), which is essential for signalling and plays a major role in recruiting intracellular signalling molecules. While activation of the cytoplasmic tyrosine kinase JAK2 appears to be the initiating event for signalling, the immediate events that trigger this are still unclear. We have isolated a number of activated mutants of h beta(c), which can be grouped into classes defined by their state of receptor phosphorylation, their requirement for alpha subunit as a cofactor, and their activities in primary cells and cell lines. We discuss these findings with regard to the stoichiometry, activation, and signalling of the normal GM-CSF/IL-3/IL-5 receptor complexes. Specifically, this work has implications for the role of the ligand-specific alpha-subunits in initiating the signalling through the beta-subunit, the role of beta subunit dimerization as a receptor trigger, and the function of receptor tyrosine phosphorylation in generating growth and survival signals. Based on the properties of the activated mutants and the recent structures of erythropoietin receptor (Epo-R) complexes, we propose a model in which (1) activation of h beta(c) can occur via alternative states that differ with respect to stoichiometry and subunit assembly, but which all mediate proliferative responses, and (2) each of the different classes of activated mutants mimics one of these alternative states. (C) 2000 International Society for Experimental Hematology. Published by Elsevier Science Inc.

The Las Campanas/AAT rich cluster survey - I. Precision and reliability of the photometric catalogue

The Las Campanas Observatory and Anglo-Australian Telescope Rich Cluster Survey (LARCS) is a panoramic imaging and spectroscopic survey of an X-ray luminosity-selected sample of 21 clusters of galaxies at 0.07 < z < 0.16. Charge-coupled device (CCD) imaging was obtained in B and R of typically 2 degrees wide regions centred on the 21 clusters, and the galaxy sample selected from the imaging is being used for an on-going spectroscopic survey of the clusters with the 2dF spectrograph on the Anglo-Australian Telescope. This paper presents the reduction of the imaging data and the photometric analysis used in the survey. Based on an overlapping area of 12.3 deg(2) we compare the CCD-based LARCS catalogue with the photographic-based galaxy catalogue used for the input to the 2dF Galaxy Redshift Survey (2dFGRS) from the APM, to the completeness of the GRS/APM catalogue, b(J) = 19.45. This comparison confirms the reliability of the photometry across our mosaics and between the clusters in our survey. This comparison also provides useful information concerning the properties of the GRS/APM. The stellar contamination in the GRS/APM galaxy catalogue is confirmed as around 5-10 per cent, as originally estimated. However, using the superior sensitivity and spatial resolution in the LARCS survey evidence is found for four distinct populations of galaxies that are systematically omitted from the GRS/APM catalogue. The characteristics of the 'missing' galaxy populations are described, reasons for their absence examined and the impact they will have on the conclusions drawn from the 2dF Galaxy Redshift Survey are discussed.

Community structure of Quaternary coral reefs compared with Recent life and death assemblages

This paper assesses the reliability with which fossil reefs record the diversity and community structure of adjacent Recent reefs. The diversity and taxonomic composition of Holocene raised fossil reefs was compared with those of modern reef coral life and death assemblages in adjacent moderate and low-energy shallow reef habitats Of Madang Lagoon, Papua New Guinea. Species richness per sample area and Shannon-Wiener diversity (H') were highest in the fossil reefs, intermediate in the life assemblages, and lowest in the death assemblages. The taxonomic composition of the fossil reefs was most similar to the combination of the life and death assemblages from the modern reefs adjacent to the two fossil reefs. Depth zonation was recorded accurately in the fossil reefs. The Madang fossil reefs represent time-averaged composites of the combined life and death assemblages as they existed at the time the reef was uplifted. Because fossil reefs include overlapping cohorts from the life and death assemblages, lagoonal facies of fossil reefs are dominated by the dominant sediment-producing taxa, which are not necessarily the most abundant in the life assemblage. Rare or slow-growing taxa accumulate more slowly than the encasing sediments and are underrepresented in fossil reef lagoons. Time-averaging dilutes the contribution of rare taxa, rather than concentrating their contribution. Consequently, fidelity indices developed for mollusks in sediments yield low values in coral reef death and fossil assemblages. Branching corals dominate lagoonal facies of fossil reefs because they are abundant, they grow and produce sediment rapidly, and most of the sediment they produce is not exported. Fossil reefs distinguished kilometer-scale variations in community structure more clearly than did the modern life assemblages. This difference implies that fossil,reefs may provide a better long-term record of community structure than modern reefs. This difference also suggests that modern kilometer-scale variation in coral reef community structure may have been reduced by anthropogenic degradation, even in the relatively unimpacted reefs of Madang Lagoon. Holocene and Pleistocene fossil reefs provide a time-integrated historical record of community composition and may be used as long-term benchmarks for comparison with modern, degraded, nearshore reefs. Comparisons between fossil reefs and degraded modern reefs display gross changes in community structure more effectively than they demonstrate local extinction of rare taxa.

Methods for prediction of peptide binding to MHC molecules: A comparative study

Background: A variety of methods for prediction of peptide binding to major histocompatibility complex (MHC) have been proposed. These methods are based on binding motifs, binding matrices, hidden Markov models (HMM), or artificial neural networks (ANN). There has been little prior work on the comparative analysis of these methods. Materials and Methods: We performed a comparison of the performance of six methods applied to the prediction of two human MHC class I molecules, including binding matrices and motifs, ANNs, and HMMs. Results: The selection of the optimal prediction method depends on the amount of available data (the number of peptides of known binding affinity to the MHC molecule of interest), the biases in the data set and the intended purpose of the prediction (screening of a single protein versus mass screening). When little or no peptide data are available, binding motifs are the most useful alternative to random guessing or use of a complete overlapping set of peptides for selection of candidate binders. As the number of known peptide binders increases, binding matrices and HMM become more useful predictors. ANN and HMM are the predictive methods of choice for MHC alleles with more than 100 known binding peptides. Conclusion: The ability of bioinformatic methods to reliably predict MHC binding peptides, and thereby potential T-cell epitopes, has major implications for clinical immunology, particularly in the area of vaccine design.

The incidence of schizophrenia: A systematic review

There has been a debate on whether or not the incidence of schizophrenia varies across time and place. In order to optimise the evidence upon which this debate is based, we have undertaken a systematicsystematic review of the literature. In this paper we provide an overview of the methods of the review and a preliminary analysis of the studies identified to date. Electronic databases (Medline, Psychlnfo, Embase, LILAC) were systematically searched for articles published between January 1965 and December 2001. The search terms were: (schizo* OR psycho*)AND (incidence OR prevalence). References were also identified from review articles, reference list and by writing to authors. To date we have identified 137 papers drawn from 33 nations. 37 papers in language other than English await translation. The currently included papers have generated 1413 different items of rate information data. In order to analyze these data we have undertaken several sequential filters in order to identify (a) non-overlapping data, (b) birth cohort study versus noncohort studies, (c) overall and sex-specific rates, (d) diagnostic criteria, (e) age ranges, (f) epoch of study, and (g) data on migrant or other special interest groups. In addition, we will examine the impact of urbanicity of site, age and/or sex standardization, and quality score on the incidence rates. The various discrete incidence rates will be presented graphically and the impact of various filters on these rates will be inspected using meta-analytic techniques. The use of meta-analysis may help elucidate the epidemiological landscape with respect to the incidence of schizophrenia and aid in the generation of new hypothesis. Acknowledgements: The Stanley Medical Research Institute supported project

Two distinct regions in 2q24.2-q24.3 associated with idiopathic epilepsy

P>Approximately 50% of all carriers of 2q21-q31 deletions present epileptic seizures. The band 2q24 constitutes the smallest commonly deleted segment in these patients, and contains the voltage-gated sodium channel genes SCN1A and SCN2A, associated with Dravet syndrome and benign familial neonatal-infantile seizures, respectively. A further putative locus involving epilepsy in the region was previously identified through disruption of the SLC4A10 gene by translocation. In the course of performing high-resolution DNA copy number analyses on syndromic mentally impaired individuals, we encountered three patients with overlapping deletions in chromosome region 2q24. Two of these patients exhibited epileptic seizures in addition to mental deficiency. The deletion in one of the epileptic patients did not include the SCN cluster, demonstrating that a less severe form of epilepsy maps to an adjacent genomic region. This second region comprises about 3 Mb and contains the candidate gene SLC4A10, providing further support for the potential role of this gene in epilepsy.

A peptide-binding motif for I-A(g7), the class II major histocompatibility complex (MHC) molecule of NOD and Biozzi AB/H mice

The class II major histocompatibility complex molecule I-A(g7) is strongly linked to the development of spontaneous insulin-dependent diabetes mellitus (IDDM) in non obese diabetic mice and to the induction of experimental allergic encephalomyelitis in Biozzi AB/H mice. Structurally, it resembles the HLA-DQ molecules associated with human IDDM, in having a non-Asp residue at position 57 in its beta chain. To identify the requirements for peptide binding to I-A(g7) and thereby potentially pathogenic T cell epitopes, we analyzed a known I-A(g7)-restricted T cell epitope, hen egg white lysozyme (HEL) amino acids 9-27. NH2- and COOH-terminal truncations demonstrated that the minimal epitope for activation of the T cell hybridoma 2D12.1 was M12-R21 and the minimum sequence for direct binding to purified I-A(g7) M12-Y20/K13-R21. Alanine (A) scanning revealed two primary anchors for binding at relative positions (p) 6 (L) and 9 (Y) in the HEL epitope. The critical role of both anchors was demonstrated by incorporating L and Y in poly(A) backbones at the same relative positions as in the HEL epitope. Well-tolerated, weakly tolerated, and nontolerated residues were identified by analyzing the binding of peptides containing multiple substitutions at individual positions. Optimally, p6 was a large, hydrophobic residue (L, I, V, M), whereas p9 was aromatic and hydrophobic (Y or F) or positively charged (K, R). Specific residues were not tolerated at these and some other positions. A motif for binding to I-A(g7) deduced from analysis of the model HEL epitope was present in 27/30 (90%) of peptides reported to be I-A(g7)-restricted T cell epitopes or eluted from I-A(g7). Scanning a set of overlapping peptides encompassing human proinsulin revealed the motif in 6/6 good binders (sensitivity = 100%) and 4/13 weak or non-binders (specificity = 70%). This motif should facilitate identification of autoantigenic epitopes relevant to the pathogenesis and immunotherapy of IDDM.

Construction of a 1-Mb restriction-mapped cosmid contig containing the candidate region for the familial mediterranean fever locus (MEFV) on chromosome 16p13.3

In this paper we describe the assembly and restriction map of a 1.05-Mb cosmid contig spanning the candidate region for familial Mediterranean fever (FMF), a recessively inherited disorder of inflammation localized to 16p13.3. Using a combination of cosmid walking and screening for P1, PAC, BAG, and YAC clones, we have generated a contig of genomic clones spanning similar to 1050 kb that contains the FMF critical region. The map consists of 179 cosmid, 15 P1, 10 PAC, 3 BAG, and 17 YAC clones, anchored by 27 STS markers. Eight additional STSs have been developed from the similar to 700 kb immediately centromeric to this genomic region. Five of the 35 STSs are microsatellites that have not been previously reported. NotI and EcoRI mapping of the overlapping cosmids, hybridization of restriction fragments from cosmids to one another, and STS analyses have been used to validate the assembly of the contig. Our contig totally subsumes the 250-kb interval recently reported, by founder haplotype analysis, to contain the FMF gene. Thus, our high-resolution clone map provides an ideal resource for transcriptional mapping toward the eventual identification of this disease gene. (C) 1997 Academic Press.

Pathologic similarities and differences between asthma and chronic obstructive pulmonary disease

Recognizing the differences and similarities at pathological level in both diseases may lead to a better understanding of the overlapping clinical and physiological phenotypes, thereby helping to better plan specific treatment and long-term management.

Retinal neurons: cell types and coupled networks

Retinal neurons with distinct dendritic morphologies are likely to comprise different cell types, subject to three important caveats. First, it is necessary to avoid creating “artificial” cell types based on arbitrary criteria—for example, the presence of two or three primary dendrites. Second, it is essential to take into account changes in morphology with retinal eccentricity and cell density. Third, the retina contains imperfections like any natural system and a significant number of retinal neurons display aberrant morphologies or make aberrant connections that are not typical of the population as a whole. Many types of retinal ganglion cells show diverse patterns of tracer coupling, with the simplest pattern represented by the homologous coupling shown by On-Off direction-selective (DS) ganglion cells in the rabbit retina. Neighboring DS ganglion cells with a common preferred direction have regularly spaced somata and territorial dendritic fields, whereas DS ganglion cells with different preferred directions may have closely spaced somata and overlapping dendritic fields.

Autoinflammatory diseases: Mimics of Autoimmunity or part of its spectrum? Case presentation

Introduction: Autoinflammatory diseases are very rare diseases presenting within a wide clinical spectrum. Recognition of the main clinical features are challenging due to overlapping or mimicking with autoimmune diseases. Discussion: A case series is reviewed to illustrate typical and atypical features and the difficulties of these diagnoses in the low prevalence areas-a typical unrecognized case of familial Mediterranean fever (FMF) in a youngster, an atypical adult case with overlapping of IMF with Behcet disease, and an early presentation of FMF in infant presenting with inflammatory colitis, as well as the overlapping features within the cryopirin diseases spectrum in an 8-year-old boy who presented with systemic onset arthritis. Conclusion: These cases may represent examples of a very puzzling relationship among disorders of innate and adaptive immune systems and inflammation.

Electromyographic standardized indices in healthy Brazilian young adults and data reproducibility

P>The determination of normal parameters is an important procedure in the evaluation of the stomatognathic system. We used the surface electromyography standardization protocol described by Ferrario et al. (J Oral Rehabil. 2000;27:33-40, 2006;33:341) to determine reference values of the electromyographic standardized indices for the assessment of muscular symmetry (left and right side, percentage overlapping coefficient, POC), potential lateral displacing components (unbalanced contractile activities of contralateral masseter and temporalis muscles, TC), relative activity (most prevalent pair of masticatory muscles, ATTIV) and total activity (integrated areas of the electromyographic potentials over time, IMPACT) in healthy Brazilian young adults, and the relevant data reproducibility. Electromyography of the right and left masseter and temporalis muscles was performed during maximum teeth clenching in 20 healthy subjects (10 women and 10 men, mean age 23 years, s.d. 3), free from periodontal problems, temporomandibular disorders, oro-facial myofunctional disorder, and with full permanent dentition (28 teeth at least). Data reproducibility was computed for 75% of the sample. The values obtained were POC Temporal (88 center dot 11 +/- 1 center dot 45%), POC masseter (87 center dot 11 +/- 1 center dot 60%), TC (8 center dot 79 +/- 1 center dot 20%), ATTIV (-0 center dot 33 +/- 9 center dot 65%) and IMPACT (110 center dot 40 +/- 23 center dot 69 mu V/mu V center dot s %). There were no statistical differences between test and retest values (P > 0 center dot 05). The Technical Errors of Measurement (TEM) for 50% of subjects assessed during the same session were 1 center dot 5, 1 center dot 39, 1 center dot 06, 3 center dot 83 and 10 center dot 04. For 25% of the subjects assessed after a 6-month interval, the TEM were 0 center dot 80, 1 center dot 03, 0 center dot 73, 12 center dot 70 and 19 center dot 10. For all indices, there was good reproducibility. These electromyographic indices could be used in the assessment of patients with stomatognathic dysfunction.