PhyloFacts: an online structural phylogenomic encyclopedia for protein functional and structural classification

The Berkeley Phylogenomics Group presents PhyloFacts, a structural phylogenomic encyclopedia containing almost 10,000 'books' for protein families and domains, with pre-calculated structural, functional and evolutionary analyses. PhyloFacts enables biologists to avoid the systematic errors associated with function prediction by homology through the integration of a variety of experimental data and bioinformatics methods in an evolutionary framework. Users can submit sequences for classification to families and functional subfamilies. PhyloFacts is available as a worldwide web resource from .     

Entrapment of viral capsids in nuclear PML cages is an intrinsic antiviral host defense against varicella-zoster virus

The herpesviruses, like most other DNA viruses, replicate in the host cell nucleus. Subnuclear domains known as promyelocytic leukemia protein nuclear bodies (PML-NBs), or ND10 bodies, have been implicated in restricting early herpesviral gene expression. These viruses have evolved countermeasures to disperse PML-NBs, as shown in cells infected in vitro, but information about the fate of PML-NBs and their functions in herpesvirus infected cells in vivo is limited. Varicella-zoster virus (VZV) is an alphaherpesvirus with tropism for skin, lymphocytes and sensory ganglia, where it establishes latency. Here, we identify large PML-NBs that sequester newly assembled nucleocapsids (NC) in neurons and satellite cells of human dorsal root ganglia (DRG) and skin cells infected with VZV in vivo. Quantitative immuno-electron microscopy revealed that these distinctive nuclear bodies consisted of PML fibers forming spherical cages that enclosed mature and immature VZV NCs. Of six PML isoforms, only PML IV promoted the sequestration of NCs. PML IV significantly inhibited viral infection and interacted with the ORF23 capsid surface protein, which was identified as a target for PML-mediated NC sequestration. The unique PML IV C-terminal domain was required for both capsid entrapment and antiviral activity. Similar large PML-NBs, termed clastosomes, sequester aberrant polyglutamine (polyQ) proteins, such as Huntingtin (Htt), in several neurodegenerative disorders. We found that PML IV cages co-sequester HttQ72 and ORF23 protein in VZV infected cells. Our data show that PML cages contribute to the intrinsic antiviral defense by sensing and entrapping VZV nucleocapsids, thereby preventing their nuclear egress and inhibiting formation of infectious virus particles. The efficient sequestration of virion capsids in PML cages appears to be the outcome of a basic cytoprotective function of this distinctive category of PML-NBs in sensing and safely containing nuclear aggregates of aberrant proteins.     

C3G is required for c-Abl-induced filopodia and its overexpression promotes filopodia formation

The Rap1 guanine nucleotide exchange factor, C3G (also known as Rap1GEF-1) is involved in signaling from growth factors, cytokines and integrins and plays a role in cell adhesion and migration, but the mechanism by which C3G regulates various cellular functions is poorly understood. We, therefore, investigated the ability of C3G to affect actin cytoskeleton-dependent morphological changes in cells. Using RNA interference, we provide evidence that C3G is required for c-Abl-induced filopodia during cell spreading on fibronectin. C3G expression induces actin cytoskeletal reorganization and promotes filopodia formation independent of its catalytic activity. It showed enrichment at filopodia tips characteristic of molecules involved in filopodia dynamics. C3G-induced filopodia were not inhibited by dominant negative mutants of Rho, Rac and Cdc42, but required Abl catalytic activity. Coexpression of N-Wasp-Crib inhibited C3G induced as well as c-Abl-induced filopodia and wiskostatin, a pharmacological inhibitor of N-Wasp attenuates C3G-induced filopodia. Cellular C3G interacts with c-Abl and C3G expression results in enhanced localization of endogenous c-Abl in the cytoplasm. We suggest that C3G and c-Abl function in an interdependent manner, in linking external signals to remodeling the cytoskeleton to induce filopodia.     

Life table demography and population growth of Brachionus variabilis Hempel, 1896 in relation to Chlorella vulgaris densities

We studied the life history variables and population growth characteristics of Brachionus variabilis, which was recorded for the first time from Mexico. The animals were fed Chlorella, using five concentrations (0.25, 0.5, 1, 2 and 4 × 10⁶ cells ml^–1) at 25 °C. Food density was observed to have significant effect on life expectancy, average lifespan, gross reproductive rate, net reproductive rate, generation time and population growth rate. The average lifespan ranged from 3 to 6 days depending on the food density. The net reproductive rate ranged from 2 to 7 neonates female^–1 d^–1. The rate of population increase per day varied from 0.14 to 0.35. The highest net reproductive rate and average lifespan and life expectancy were recorded at Chlorella concentrations of 1 × 10⁶ and 2 × 10⁶ cells ml^–1.     

G-protein-dependent cell surface dynamics of the human serotonin1A receptor tagged to yellow fluorescent protein

Serotonergic signaling appears to play a key role in the generation and modulation of various cognitive, behavioral, and developmental processes. The serotonin(1A) receptor is an important member of the superfamily of seven transmembrane domain G-protein-coupled receptors and is the most extensively studied among the serotonin receptors. Several aspects of serotonin(1A) receptor biology such as cellular distribution and signal transduction characteristics are technically difficult to address in living cells on account of the inability to optically track these receptors with fluorescence-based techniques. We describe here the characterization of the serotonin(1A) receptor tagged to the enhanced yellow fluorescent protein (EYFP) stably expressed in Chinese hamster ovary (CHO) cells. These receptors were found to be essentially similar to the native receptor in pharmacological assays and can therefore be used to reliably explore aspects of receptor biology such as cellular distribution and dynamics on account of their intrinsic fluorescent properties. Analysis of the cell surface dynamics of these receptors by fluorescence recovery after photobleaching (FRAP) experiments has provided novel insight into the molecular mechanism of signal transduction of serotonin(1A) receptors in living cells. Interestingly, addition of pharmacologically well-characterized ligands or activators of G-proteins altered the diffusion characteristics of the receptor in a manner consistent with the G-protein activation model. These results demonstrate, for the first time, that membrane dynamics of this receptor is modulated in a G-protein-dependent manner.     

TGFbeta intercepts nuclear glycogen synthase kinase 3beta to inhibit PDGF-induced DNA synthesis in mesangial cells

Here, we demonstrate a mechanism of TGFbeta-mediated inhibition of PDGF-induced DNA synthesis in mesangial cells. TGFbeta significantly inhibited nuclear Akt phosphorylation without any effect on PDGF-stimulated phosphorylation of PDGFR at PI 3 kinase binding site (Tyr-751). Remarkably, TGFbeta inhibited cyclin D1 and cyclin E expression with concomitant decrease in CDK2 activity induced by PDGF. More importantly, we demonstrate that TGFbeta significantly abolished Akt-mediated serine-9 phosphorylation of glycogen synthase kinase 3beta (GSK3beta), thus prevented its inactivation. Expression of inactive GSK3betaK85R mutant increased cyclin D1 expression and DNA synthesis similar to PDGF. These results provide the first evidence that TGFbeta intercepts Akt kinase activity in the nucleus to block inactivation of GSK3beta, leading to attenuation of PDGF-induced CDK2 activity and DNA synthesis.     

Identification of prosaposin and transgelin as potential biomarkers for gallbladder cancer using quantitative proteomics

Gallbladder cancer is an uncommon but lethal malignancy with particularly high incidence in Chile, India, Japan and China. There is a paucity of unbiased large-scale studies investigating molecular basis of gallbladder cancer. To systematically identify differentially regulated proteins in gallbladder cancer, iTRAQ-based quantitative proteomics of gallbladder cancer was carried out using Fourier transform high resolution mass spectrometry. Of the 2575 proteins identified, proteins upregulated in gallbladder cancer included several lysosomal proteins such as prosaposin, cathepsin Z and cathepsin H. Downregulated proteins included serine protease HTRA1 and transgelin, which have been reported to be downregulated in several other cancers. Novel biomarker candidates including prosaposin and transgelin were validated to be upregulated and downregulated, respectively, in gallbladder cancer using tissue microarrays. Our study provides the first large scale proteomic characterization of gallbladder cancer which will serve as a resource for future discovery of biomarkers for gallbladder cancer.     

Interaction Among Copper Toxicity,Temperature and Salinity on the Population Dynamics of <Emphasis Type="Italic">Brachionus Rotundiformis</Emphasis> (Rotifera)

Heavy metals may interact with ecological factors such as temperature, food level and salinity, causing both mortality and reduced reproduction in organisms. Among different heavy metals, copper compounds are commonly used for eliminating algal blooms in aquaculture tanks. At certain concentrations, copper is toxic to rotifers. In the present work, we evaluated the combined effects of salt concentrations (2.5 and 5.0 g l⁻¹ NaCl), copper levels (0, 0.03125, 0.0625, 0.125 and 0.25 mg l⁻¹ as CuCl₂) and two temperatures (20 and 25 °C) on the population growth of B. rotundiformis using Chlorella as the algal food (at 0.5 × 10⁶ cells ml⁻¹ for every 24 h). Regardless of salinity and temperature, copper at concentrations as low as 0.03 mg l⁻¹ had an adverse effect on the population growth of rotifers and above 0.125 mg l⁻¹, the populations did not grow. The effect of the toxicant on B. rotundiformis was more severe at 25° than at 20 °C at lower salinity. In general, we observed peak densities of rotifers around day 12 at 20 °C but 6–8 days earlier at 25 °C. Peak population densities of B. rotundiformis in the controls at the salinity of 2.5 g l⁻¹ ranged from 90 to 180 ind. ml⁻¹, depending on temperature; at a salinity of 5.0 g l⁻¹, these were lower. The population growth rates, r, in our study varied from +0.31 to –0.12 depending on the test conditions. There was a significant impact of temperature, salinity and toxicity level on the population growth rate of B. rotundiformis. Our results suggested that even narrow changes in salinity could negatively influence the toxicity of heavy metal on the population growth rates of B. rotundiformis.