Low incidence of polyploids and high uniformity of karyotypes displayed by Delphinium (Ranunculaceae) in the Hengduan Mountains region of south‐west China

The chromosome numbers and morphology in 92 populations belonging to 49 species and three varieties in the genus Delphinium L. (Ranunculaceae), mostly from the Hengduan Mountains region of south‐west China, were studied. Forty seven species and three varieties were diploid, with 2n = 16, one species was tetraploid, with 2n = 32, and one species had diploid and tetraploid cytotypes. Three species had B chromosomes, representing the first time the occurrence of B chromosomes has been reported in the genus. The karyotypes of all the diploid species were quite uniform, commonly bimodal, and usually consisted of one pair of large median‐centromeric (m), one pair of large submedian‐centromeric (sm), five pairs of medium‐sized subterminal‐centromeric (st), and one pair of smaller sm (rarely st) chromosomes. The low incidence of polyploids in Delphinium from the Hengduan Mountains region indicates that polyploidy has played a minor role in the speciation of this highly diversified genus in the region. © 2008 The Linnean Society of London, Botanical Journal of the Linnean Society, 2008, 158 , 172–188.     

Myosin from pancreatic acinar carcinoma cells. Isolation, characterization and demonstration of heavy- and light-chain phosphorylation.

Myosin has been identified in a variety of non-muscle cells, and is believed to play a role in maintenance of cell shape, locomotion, cytokinesis, exocytosis and other cellular functions. In this paper we describe the purification of myosin from a pancreatic acinar-cell carcinoma of the rat which forms solid tumours, but retains many differentiated functions. The purified myosin was composed of a 200,000 Da heavy chain and two or three classes of light chains. Electron-microscopic examination of rotary-shadowed preparations revealed that individual molecules had two globular heads and a long tail measuring approx. 149 nm. The myosin was soluble in high-salt buffers and became sedimentable as the ionic strength was lowered. Examination of negative-stained preparations showed that this sedimentable myosin consisted of short, bipolar, thick filaments which had a strong tendency to aggregate in a head-to-head manner. The ATPase activity of the purified myosin was stimulated by EDTA or Ca2+, but not by Mg2+. In low ionic strength the Mg2+-dependent ATPase activity was activated by muscle f-actin. The pancreatic myosin bound to actin and could be dissociated by the addition of MgATP. Myosin purified from cells cultured in media containing [32P]Pi was phosphorylated on one of the light chains as well as the heavy chain. Thus pancreatic acinar cells contain a typical non-muscle myosin, and the subunits of this molecule are subject to post-translational modification by phosphorylation.     

Dictyostelium discoideum myosin: isolation and characterization of cDNAs encoding the regulatory light chain.

Phosphorylation of the regulatory light chains (RMLC) of nonmuscle myosin can increase the actin-activated ATPase activity and filament formation. Little is known about these regulatory mechanisms and how the RMLC are involved in ATP hydrolysis. To better characterize the nonmuscle RMLC, we isolated cDNAs encoding the Dictyostelium RMLC. Using an antibody specific for the RMLC, we screened a lambda gt11 expression library and obtained a 200-base-pair clone that encoded a portion of the RMLC. The remainder of the sequence was obtained from two clones identified by DNA hybridization, using the 200-base-pair cDNA. The composite RMLC cDNA was 645 nucleotides long. It contained 60 base pairs of 5' untranslated, 483 bases of coding, and 102 base pairs of 3' untranslated sequence. The amino acid sequence predicted an 18,300-dalton protein that shares 42% amino acid identity with Dictyostelium calmodulin and 30% identity with the chicken skeletal myosin RMLC. This sequence contained three regions that were similar to the E-F hand calcium-binding domains found in calmodulin, troponin C, and other myosin light chains. A sequence similar to the phosphorylation sequence found in chicken gizzard and skeletal myosin light chains was found at the amino terminus. Genomic Southern blot analysis suggested that the Dictyostelium genome contains a single gene encoding the RMLC. Analysis of RMLC expression patterns during Dictyostelium development indicated that accumulation of this mRNA increases just before aggregation and again during culmination. This pattern is similar to that obtained for the Dictyostelium essential myosin light chain and suggests that expression of the two light chains is coordinated during development.     

Studies on the organization and localization of actin and myosin in neurons

The organization of actin in mouse neuroblastoma and chicken dorsal root ganglion (DRG) nerve cells was investigated by means of a variety of electron microscope techniques. Microspikes of neuroblastoma cells contained bundles of 7- to 8-nm actin filaments which originated in the interior of the neurite. In the presence of high concentrations of Mg++ ion, filaments in these bundles became highly ordered to form paracrystals. Actin filaments, but not bundles, were observed in growth cones of DRG cells. Actin was localized in the cell body, neurites, and microspikes of both DRG and neuroblastoma nerve cells by fluorescein-labeled S1. Myosin was localized primarily in the neurites of chick DRG nerve cells with fluorescein-labeled anti-brain myosin antibody. This antibody also stained stress fibers in fibroblasts and myoblasts but did not stain muscle myofibrils.     

MannDB – A microbial database of automated protein sequence analyses and evidence integration for protein characterization

Background  

MannDB was created to meet a need for rapid, comprehensive automated protein sequence analyses to support selection of proteins suitable as targets for driving the development of reagents for pathogen or protein toxin detection. Because a large number of open-source tools were needed, it was necessary to produce a software system to scale the computations for whole-proteome analysis. Thus, we built a fully automated system for executing software tools and for storage, integration, and display of automated protein sequence analysis and annotation data.     

Ladder webs in orb‐web spiders: ontogenetic and evolutionary patterns in Nephilidae

Spider web research bridges ethology, ecology, functional morphology, material science, development, genetics, and evolution. Recent work proposes the aerial orb web as a one‐time key evolutionary innovation that has freed spider‐web architecture from substrate constraints. However, the orb has repeatedly been modified or lost within araneoid spiders. Modifications include not only sheet‐ and cobwebs, but also ladder webs, which secondarily utilize the substrate. A recent nephilid species level phylogeny suggests that the ancestral nephilid web architecture was an arboricolous ladder and that round aerial webs were derived. Because the web biology of the basalmost Clitaetra and the derived Nephila are well understood, the present study focuses on the webs of the two phylogenetically intervening genera, Herennia and Nephilengys, to establish ontogenetic and macroevolutionary patterns across the nephilid tree. We compared juvenile and adult webs of 95 Herennia multipuncta and 143 Nephilengys malabarensis for two measures of ontogenetic allometric web changes: web asymmetry quantified by the ladder index, and hub asymmetry quantified by the hub displacement index. We define a ‘ladder web’ as a vertically elongated orb exceeding twice the length over width (ladder index ≥ 2) and possessing (sub)parallel rather than round side frames. Webs in both genera allometrically grew from orbs to ladders, more so in Herennia. Such allometric web growth enables the spider to maintain its arboricolous web site. Unexpectedly, hub asymmetry only increased significantly in heavy‐bodied Nephilengys females, and not in Herennia, challenging the commonly invoked gravity hypothesis. The findings obtained in the present study support the intrageneric uniformness of nephilid webs, with Herennia etruscilla webs being identical to H. multipuncta. The nephilid web evolution suggests that the ancestor of Nephila reinvented the aerial orb web because the orb arises at a much more inclusive phylogenetic level, and all intervening nephilids retained the secondarily acquired substrate‐dependent ladder web. © 2010 The Linnean Society of London, Biological Journal of the Linnean Society, 2010, 99 , 849–866.     

In adult onset myositis, the presence of interstitial lung disease and myositis specific/associated antibodies are governed by HLA class II haplotype, rather than by myositis subtype

The Fcγ receptors play important roles in the initiation and regulation of many immunological and inflammatory processes, and genetic variants (FCGR) have been associated with numerous autoimmune and infectious diseases. The data in rheumatoid arthritis (RA) are conflicting and we previously demonstrated an association between FCGR3A and RA. In view of the close molecular proximity with FCGR2A, FCGR2B and FCGR3B, additional polymorphisms within these genes and FCGR haplotypes were examined to refine the extent of association with RA. Biallelic polymorphisms in FCGR2A, FCGR2B and FCGR3B were examined for association with RA in two well characterized UK Caucasian and North Indian/Pakistani cohorts, in which FCGR3A genotyping had previously been undertaken. Haplotype frequencies and linkage disequilibrium were estimated across the FCGR locus and a model-free analysis was performed to determine association with RA. This was followed by regression analysis, allowing for phase uncertainty, to identify the particular haplotype(s) that influences disease risk. Our results reveal that FCGR2A, FCGR2B and FCGR3B were not associated with RA. The haplotype with the strongest association with RA susceptibility was the FCGR3A–FCGR3B 158V-NA2 haplotype (odds ratio 3.18, 95% confidence interval 1.13–8.92 [P = 0.03] for homozygotes compared with all genotypes). The association was stronger in the presence of nodules (odds ratio 5.03, 95% confidence interval 1.44–17.56; P = 0.01). This haplotype was also more common in North Indian/Pakistani RA patients than in control individuals, but not significantly so. Logistic regression analyses suggested that FCGR3A remained the most significant gene at this locus. The increased association with an FCGR3A–FCGR3B haplotype suggests that other polymorphic variants within FCGR3A or FCGR3B, or in linkage disequilibrium with this haplotype, may additionally contribute to disease pathogenesis.     

Pulse Decomposition Analysis of the digital arterial pulse during hemorrhage simulation

Background

Markers of temporal changes in central blood volume are required to non-invasively detect hemorrhage and the onset of hemorrhagic shock. Recent work suggests that pulse pressure may be such a marker. A new approach to tracking blood pressure, and pulse pressure specifically is presented that is based on a new form of pulse pressure wave analysis called Pulse Decomposition Analysis (PDA). The premise of the PDA model is that the peripheral arterial pressure pulse is a superposition of five individual component pressure pulses, the first of which is due to the left ventricular ejection from the heart while the remaining component pressure pulses are reflections and re-reflections that originate from only two reflection sites within the central arteries. The hypothesis examined here is that the PDA parameter T13, the timing delay between the first and third component pulses, correlates with pulse pressure. T13 was monitored along with blood pressure, as determined by an automatic cuff and another continuous blood pressure monitor, during the course of lower body negative pressure (LBNP) sessions involving four stages, -15 mmHg, -30 mmHg, -45 mmHg, and -60 mmHg, in fifteen subjects (average age: 24.4 years, SD: 3.0 years; average height: 168.6 cm, SD: 8.0 cm; average weight: 64.0 kg, SD: 9.1 kg).

Results

Statistically significant correlations between T13 and pulse pressure as well as the ability of T13 to resolve the effects of different LBNP stages were established. Experimental T13 values were compared with predictions of the PDA model. These interventions resulted in pulse pressure changes of up to 7.8 mmHg (SE = 3.49 mmHg) as determined by the automatic cuff. Corresponding changes in T13 were a shortening by -72 milliseconds (SE = 4.17 milliseconds). In contrast to the other two methodologies, T13 was able to resolve the effects of the two least negative pressure stages with significance set at p < 0.01.

Conclusions

The agreement of observations and measurements provides a preliminary validation of the PDA model regarding the origin of the arterial pressure pulse reflections. The proposed physical picture of the PDA model is attractive because it identifies the contributions of distinct reflecting arterial tree components to the peripheral pressure pulse envelope. Since the importance of arterial pressure reflections to cardiovascular health is well known, the PDA pulse analysis could provide, beyond the tracking of blood pressure, an assessment tool of those reflections as well as the health of the sites that give rise to them.     

Stopped-flow measurement of cytoskeletal contraction: Dictyostelium myosin II is specifically required for contraction of amoeba cytoskeletons

Cytoskeletons provide valuable information on the composition and organization of the cell's contractile machinery, and in many cases these cell models retain the ability to contract. To quantitate contraction rates, we developed a novel stopped-flow assay permitting simultaneous analysis of thousands of Dictyostelium cytoskeletons within milliseconds of mixing with Mg-ATP. Cytoskeletons were placed in one syringe of the stopped flow apparatus and the appropriate buffer was placed in the second syringe. Mixing with Mg-ATP caused an immediate increase in the absorbance at 310 nm. Rapid fixation of the cytoskeletons during the reaction confirmed that this change in absorbance was highly correlated with contraction of the cytoskeletons. This spectroscopic change was used to measure the effects of temperature, pH, ionic strength, and nucleotides on contraction rate. Treatment with high salt and ATP removed most of the myosin, some actin, and small amounts of minor proteins. These extracted cytoskeletons lost the ability to contract, but after the addition of purified Dictyostelium myosin they regained full function. In contrast, rabbit skeletal muscle myosin was unable to restore contractility, even though it bound to the extracted cytoskeletons. Cytoskeletons prepared from a myosin-null mutant did not contract. Upon the addition of purified ameba myosin, however, they became contractile. These results suggest that filamentous Dictyostelium myosin II is essential for contraction, and that the actin cytoskeleton and associated proteins retain their functional organization in the absence of myosin.