Binding sites for tachykinin peptides in the brain and stomach of the dogfish, Scyliorhinus canicula.

In membranes of dogfish brain and stomach, two binding sites for tachykinins were identified. One site specifically bound [125I]-Bolton-Hunter substance P (BH-SP) and the rank potency of tachykinins to compete for BH-SP binding revealed similarities with the rank potency of an NK1 receptor. The pharmacology of the other site, which specifically bound [125I]-Bolton-Hunter scyliorhinin II (BH-Scy II), did not resemble any of the mammalian tachykinin receptors. The rank potency to inhibit BH-Scy II binding to this second site was: scyliorhinin II approximately scyliorhinin I greater than eledoisin approximately substance P approximately neurokinin A greater than phyllomedusin approximately physalaemin greater than [Sar9Met(O2)11]substance P. Neurokinin B and senktide did not displace BH-Scy II binding. In addition, nucleotide analogues inhibited BH-SP binding but not BH-Scy II binding. Our binding data suggest the existence of a mammalian-like NK1 receptor and of a nonmammalian tachykinin receptor in the dogfish.     

Regulatory peptide and serotonin content and brush-border enzyme activity in the rat gastrointestinal tract following neonatal treatment with capsaicin; lack of effect on epithelial markers.

The possible trophic influence of the capsaicin-sensitive extrinsic innervation of the gastrointestinal mucosa was investigated. Rats were treated neonatally with capsaicin. The gastrointestinal content of serotonin and glucagon-like immunoreactivity were used as a measure of the effect on the endocrine gut mucosa and gastrointestinal aminopeptidase and alkaline phosphatase activities were used as a measure of the effect on the gut brush-border. The gastrointestinal content of the neuropeptides substance P, VIP and CGRP were used to monitor effects on the innervation of the gut. The depletion of substance P-immunoreactivity(-IR) and calcitonin gene-related peptide(CGRP)-IR in extracts of urinary bladder and lung from the capsaicin-treated rats is evidence of the efficacy of capsaicin treatment in affecting a loss of C-fibre sensory nerves. The significant depletion of CGRP-IR measured in the stomach and duodenum of capsaicin-treated rats indicated the loss of the C-fibre sensory innervation to the gastrointestinal tract. The gastrointestinal content of VIP and substance P, which are predominantly within intrinsic gut neurones, were unaffected by capsaicin treatment. In all regions of the gastrointestinal tract of capsaicin-treated rats, the serotonin and glucagon-IR levels were not significantly different from those in controls. Similarly the levels of activity of the brush-border enzymes were not significantly effected by capsaicin treatment. This suggest the absence of any major trophic influence of capsaicin-sensitive sensory nerves on the gut endocrine mucosa and the brush border.     

Neuropeptide K-(1-24)-peptide: storage and release by carcinoid tumors

An antiserum directed against the COOH-terminal region of neuropeptide K-(1-24)-peptide that shows only 0.5% reactivity with neuropeptide K has been used in radioimmunoassay to study the posttranslation processing of human beta-preprotachykinin. A primary midgut carcinoid tumor contained high concentration of substance P (2970 pmol/g), neurokinin A (3660 pmol/g) and neuropeptide K-(1-24)-peptide (3430 pmol/g) but only a very low concentration (less than 5 pmol/g) of intact neuropeptide K. Neuropeptide K-(1-24)-peptide was also detected in extracts of metastatic tumor tissue from four patients with midgut carcinoid tumors. The amino acid sequence of tumor neuropeptide K-(1-24)-peptide was identical to that predicted from the nucleotide sequence of a human beta-preprotachykinin cDNA. The fasting plasma concentration of neuropeptide K-(1-24)-peptide was elevated in a patient with the carcinoid syndrome (821 fmol/ml compared with less than 18 fmol/ml in healthy subjects) and rose approximately 2-fold after intravenous pentagastrin. The study has demonstrated that the Lys25-Arg26 bond in neuropeptide K (corresponding to Lys96-Arg97 in the precursor) is an important processing site in human beta-preprotachykinin.     

A peptide from the eel pancreas with structural similarity to human pancreatic secretory trypsin inhibitor

The primary structure of a 61-amino-acid residue peptide from the pancreas of the European eel (Anguilla anguilla) has been established as E E K S G(5)L Y R K P(10)S C G E M(15)S A M H A(20)C P M N F(25)A P V C G(30)T D G N T(35)Y P N E C(40)S L C F Q(45)R Q N T K(50)T D I L I(55)T K D D R(60)C. There was no indication of microheterogeneity. This peptide shows structural similarity to pancreatic secretory trypsin inhibitors from several mammalian species and to a cholecystokinin-releasing peptide isolated from rat pancreatic juice. A comparison of the amino acid sequences of the peptides has identified a domain in the central region of the molecules that has been strongly conserved during evolution. In contrast, the amino acid sequence in the region corresponding to the reactive centre of the mammalian trypsin inhibitors is very poorly conserved in the eel peptide. The P1-P1' reactive site lysine-isoleucine (or arginine-isoleucine) bond in the mammalian trypsin inhibitors is replaced by a methionine-asparagine bond. This region does, however, show limited homology to the reactive centre of human alpha 1-protease inhibitor suggesting that the eel peptide may function as an inhibitor of other proteolytic enzymes in the pancreas.     

Purification of herpesvirus nucleocapsids by fluorocarbon extraction.

Purified nucleocapsids were prepared from herpes simplex virus type-1 and herpes simplex virus type-2 infected cells, using a fluorocarbon extraction technique with Freon 113. Fifty to 85% of the total nucleocapsid population remained 'nucleated' or full in terms of the nucleic acid core making them useful as a source of DNA. CsCl banded DNA represented 30--40% of the theoretical yield of the total nucleocapsid population.     

Structural and positional studies of the antimicrobial peptide brevinin‐1BYa in membrane‐mimetic environments

Brevinin‐1BYa (FLPILASLAAKFGPKLFCLVTKKC), first isolated from skin secretions of the foothill yellow‐legged frog Rana boylii, shows broad‐spectrum activity, being particularly effective against opportunistic yeast pathogens. The structure of brevinin‐1BYa was investigated in various solution and membrane‐mimicking environments by proton nuclear magnetic resonance (¹H‐NMR) spectroscopy and molecular modelling. The peptide does not possess a secondary structure in aqueous solution. In a 33% 2,2,2‐trifluoroethanol (TFE‐d₃)‐H₂O solvent mixture, as well as in membrane‐mimicking sodium dodecyl sulfate and dodecylphosphocholine micelles, the peptide's structure is characterised by a flexible helix‐hinge‐helix motif, with the hinge located at the Gly¹³/Pro¹⁴ residues, and the two α‐helices extending from Pro³ to Phe¹² and from Pro¹⁴ to Thr²¹. Positional studies involving the peptide in sodium dodecyl sulfate and dodecylphosphocholine micelles using 5‐doxyl‐labelled stearic acid and manganese chloride paramagnetic probes show that the peptide's helical segments lie parallel to the micellar surface, with the residues on the hydrophobic face of the amphipathic helices facing towards the micelle core and the hydrophilic residues pointing outwards, suggesting that the peptide exerts its biological activity by a non–pore‐forming mechanism.     

Bayesian meta-analysis models for microarray data: a comparative study

Background

With the growing abundance of microarray data, statistical methods are increasingly needed to integrate results across studies. Two common approaches for meta-analysis of microarrays include either combining gene expression measures across studies or combining summaries such as p-values, probabilities or ranks. Here, we compare two Bayesian meta-analysis models that are analogous to these methods.

Results

Two Bayesian meta-analysis models for microarray data have recently been introduced. The first model combines standardized gene expression measures across studies into an overall mean, accounting for inter-study variability, while the second combines probabilities of differential expression without combining expression values. Both models produce the gene-specific posterior probability of differential expression, which is the basis for inference. Since the standardized expression integration model includes inter-study variability, it may improve accuracy of results versus the probability integration model. However, due to the small number of studies typical in microarray meta-analyses, the variability between studies is challenging to estimate. The probability integration model eliminates the need to model variability between studies, and thus its implementation is more straightforward. We found in simulations of two and five studies that combining probabilities outperformed combining standardized gene expression measures for three comparison values: the percent of true discovered genes in meta-analysis versus individual studies; the percent of true genes omitted in meta-analysis versus separate studies, and the number of true discovered genes for fixed levels of Bayesian false discovery. We identified similar results when pooling two independent studies of Bacillus subtilis. We assumed that each study was produced from the same microarray platform with only two conditions: a treatment and control, and that the data sets were pre-scaled.

Conclusion

The Bayesian meta-analysis model that combines probabilities across studies does not aggregate gene expression measures, thus an inter-study variability parameter is not included in the model. This results in a simpler modeling approach than aggregating expression measures, which accounts for variability across studies. The probability integration model identified more true discovered genes and fewer true omitted genes than combining expression measures, for our data sets.     

Decoding development in Xenopus tropicalis

Xenopus tropicalis is rapidly being adopted as a model organism for developmental biology research and has enormous potential for increasing our understanding of how embryonic development is controlled. In recent years there has been a well-organized initiative within the Xenopus community, funded largely through the support of the National Institutes of Health in the US, to develop X. tropicalis as a new genetic model system with the potential to impact diverse fields of research. Concerted efforts have been made both to adapt established methodologies for use in X. tropicalis and to develop new techniques. A key resource to come out of these efforts is the genome sequence, produced by the US Department of Energy's Joint Genome Institute and made freely available to the community in draft form for the past three years. In this review, we focus on how advances in X. tropicalis genetics coupled with the sequencing of its genome are likely to form a foundation from which we can build a better understanding of the genetic control of vertebrate development and why, when we already have other vertebrate genetic models, we should want to develop genetic analysis in the frog.     

Small heat shock protein Hsp27 is required for proper heart tube formation

The small heat shock protein Hsp27 has been shown to be involved in a diverse array of cellular processes, including cellular stress response, protein chaperone activity, regulation of cellular glutathione levels, apoptotic signaling, and regulation of actin polymerization and stability. Furthermore, mutation within Hsp27 has been associated with the human congenital neuropathy Charcot-Marie Tooth (CMT) disease. Hsp27 is known to be expressed in developing embryonic tissues; however, little has been done to determine the endogenous requirement for Hsp27 in developing embryos. In this study, we show that depletion of XHSP27 protein results in a failure of cardiac progenitor fusion resulting in cardia bifida. Furthermore, we demonstrate a concomitant disorganization of actin filament organization and defects in myofibril assembly. Moreover, these defects are not associated with alterations in specification or differentiation. We have thus demonstrated a critical requirement for XHSP27 in developing cardiac and skeletal muscle tissues.