An ACAP1-containing clathrin coat complex for endocytic recycling

Whether coat proteins play a widespread role in endocytic recycling remains unclear. We find that ACAP1, a GTPase-activating protein (GAP) for ADP-ribosylation factor (ARF) 6, is part of a novel clathrin coat complex that is regulated by ARF6 for endocytic recycling in two key physiological settings, stimulation-dependent recycling of integrin that is critical for cell migration and insulin-stimulated recycling of glucose transporter type 4 (Glut4), which is required for glucose homeostasis. These findings not only advance a basic understanding of an early mechanistic step in endocytic recycling but also shed key mechanistic insights into major physiological events for which this transport plays a critical role.     

Epitope mapping of the hemagglutinin molecule of a highly pathogenic H5N1 influenza virus by using monoclonal antibodies

We mapped the hemagglutinin (HA) antigenic epitopes of a highly pathogenic H5N1 influenza virus on the three-dimensional HA structure by characterizing escape mutants of a recombinant virus containing A/Vietnam/1203/04 (H5N1) ΔHA and neuraminidase genes in the genetic background of A/Puerto Rico/8/34 (H1N1) virus. The mutants were selected with a panel of eight anti-HA monoclonal antibodies (MAbs), seven to A/Vietnam/1203/04 (H5N1) virus and one to A/Chicken/Pennsylvania/8125/83 (H5N2) virus, and the mutants’ HA genes were sequenced. The amino acid changes suggested three MAb groups: four MAbs reacted with the complex epitope comprising parts of the antigenic site B of H3 HA and site Sa of H1 HA, two MAbs reacted with the epitope corresponding to the antigenic site A in H3 HA, and two MAbs displayed unusual behavior: each recognized amino acid changes at two widely separate antigenic sites. Five changes were detected in amino acid residues not previously reported as changed in H5 escape mutants, and four others had substitutions not previously described. The HA antigenic structure differs substantially between A/Vietnam/1203/04 (H5N1) virus and the low-pathogenic A/Mallard/Pennsylvania/10218/84 (H5N2) virus we previously characterized (N. V. Kaverin et al., J. Gen. Virol. 83:2497-2505, 2002). The hemagglutination inhibition reactions of the MAbs with recent highly pathogenic H5N1 viruses were consistent with the antigenic-site amino acid changes but not with clades and subclades based on H5 phylogenetic analysis. These results provide information on the recognition sites of the MAbs widely used to study H5N1 viruses and demonstrate the involvement of the HA antigenic sites in the evolution of highly pathogenic H5N1 viruses, findings that can be critical for characterizing pathogenesis and vaccine design.     

Stimulation of fibroblast proliferation by neokyotorphin requires Ca influx and activation of PKA, CaMK II and MAPK/ERK

Neokyotorphin [TSKYR, hemoglobin alpha-chain fragment (137-141)] has previously been shown to enhance fibroblast proliferation, its effect depending on cell density and serum level. Here we show the dependence of the effect of neokyotorphin on cell type and its correlation with the effect of protein kinase A (PKA) activator 8-Br-cAMP, but not the PKC activator 4beta-phorbol 12-myristate, 13-acetate (PMA). In L929 fibroblasts, the proliferative effect of neokyotorphin was suppressed by the Ca2+ L-type channel inhibitors verapamil or nifedipine, the intracellular Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid acetoxymethyl ester, kinase inhibitors H-89 (PKA), KN-62 (Ca2+/calmodulin-dependent kinase II) and PD98059 (mitogen-activated protein kinase). The proliferative effect of 8-Br-cAMP was also suppressed by KN-62 and PD98059. PKC suppression (downregulation with PMA or inhibition with bisindolylmaleimide XI) did not affect neokyotorphin action. The results obtained point to a cAMP-like action for neokyotorphin.     

A first-principles model of early evolution: emergence of gene families, species, and preferred protein folds

In this work we develop a microscopic physical model of early evolution where phenotype—organism life expectancy—is directly related to genotype—the stability of its proteins in their native conformations—which can be determined exactly in the model. Simulating the model on a computer, we consistently observe the “Big Bang” scenario whereby exponential population growth ensues as soon as favorable sequence–structure combinations (precursors of stable proteins) are discovered. Upon that, random diversity of the structural space abruptly collapses into a small set of preferred proteins. We observe that protein folds remain stable and abundant in the population at timescales much greater than mutation or organism lifetime, and the distribution of the lifetimes of dominant folds in a population approximately follows a power law. The separation of evolutionary timescales between discovery of new folds and generation of new sequences gives rise to emergence of protein families and superfamilies whose sizes are power-law distributed, closely matching the same distributions for real proteins. On the population level we observe emergence of species—subpopulations that carry similar genomes. Further, we present a simple theory that relates stability of evolving proteins to the sizes of emerging genomes. Together, these results provide a microscopic first-principles picture of how first-gene families developed in the course of early evolution.     

Evolution of interdisciplinarity in biodiversity science

The study of biodiversity has grown exponentially in the last thirty years in response to demands for greater understanding of the function and importance of Earth's biodiversity and finding solutions to conserve it. Here, we test the hypothesis that biodiversity science has become more interdisciplinary over time. To do so, we analyze 97,945 peer‐reviewed articles over a twenty‐two‐year time period (1990–2012) with a continuous time dynamic model, which classifies articles into concepts (i.e., topics and ideas) based on word co‐occurrences. Using the model output, we then quantify different aspects of interdisciplinarity: concept diversity, that is, the diversity of topics and ideas across subdisciplines in biodiversity science, subdiscipline diversity, that is, the diversity of subdisciplines across concepts, and network structure, which captures interactions between concepts and subdisciplines. We found that, on average, concept and subdiscipline diversity in biodiversity science were either stable or declining, patterns which were driven by the persistence of rare concepts and subdisciplines and a decline in the diversity of common concepts and subdisciplines, respectively. Moreover, our results provide evidence that conceptual homogenization, that is, decreases in temporal β concept diversity, underlies the observed trends in interdisciplinarity. Together, our results reveal that biodiversity science is undergoing a dynamic phase as a scientific discipline that is consolidating around a core set of concepts. Our results suggest that progress toward addressing the biodiversity crisis via greater interdisciplinarity during the study period may have been slowed by extrinsic factors, such as the failure to invest in research spanning across concepts and disciplines. However, recent initiatives such as the Intergovernmental Science‐Policy Platform on Biodiversity and Ecosystem Services (IPBES) may attract broader support for biodiversity‐related issues and hence interdisciplinary approaches to address scientific, political, and societal challenges in the coming years.     

GMDP: unusual physico‐chemical and biological properties of the anomeriс forms

Disaccharide containing unit of peptidoglycan from bacterial cell wall, N‐acetyl‐d ‐glucosaminyl‐N‐acetylmuramyl‐l ‐alanyl‐d ‐glutaminamide (gluсosaminyl‐muramyl‐dipeptide) registered in Russia as an immunomodulatory drug, is shown to participate in slow equilibrium of α and β anomeric forms. Data of NMR spectra and molecular dynamics indicate that the α‐anomer predominantly acquires a folded conformation stabilized by intramolecular hydrogen bond between the alanyl carbonyl and muramyl NH proton. The β‐form displays a considerable fraction of extended, non‐hydrogen bonded structures. In the standard immunoadjuvant test system, the α‐form is practically inactive, and the activity of the equilibrium mixture with α : β = 68 : 32 ratio is due to the presence of β‐anomer. Such unique α–β selectivity of biological action must be considered at the design of related immunoactive glycopeptides. Copyright © 2015 European Peptide Society and John Wiley & Sons, Ltd.     

Effects of Melatonin‐Aided Therapy on the Glutathione Antioxidant System Activity and Liver Protection

Acute hepatitis results from oxidative stress triggered by hepatotoxic drugs causing liver injury and the activation of caspases cascade. The glutathione antioxidant system protects against reactive oxygen species and mitigates development of these processes. The effectiveness of silymarin, a polyphenolic flavonoid, essenthiale, composed of phosphatidyl choline, and melaxen, a melatonin‐correcting drug, as hepatoprotectors has been investigated. The variation of 6‐sulfatoxymelatonin (aMT6s), resulting from the biotransformation of melatonin, and GSH has been measured. The activities of caspase‐1 and caspase‐3, glutathione antioxidant system, and NADPH‐generating enzymes were determined. The aMT6s decreases in patients with drug hepatitis and recovers with administration of mexalen. GSH increased in the presence of the studied hepatoprotectors. Pathologically activated caspase‐1 and caspase‐3 decreased their activities in the presence of hepatoprotectors with melaxen showing the highest effect. The positive effect of melatonin appears to be related to the suppression of decompensation of the glutathione antioxidant system functions, recovery of liver redox status, and the attenuation of inhibition of the NADPH supply.     

Systematic analysis of Type I‐E Escherichia coli CRISPR‐Cas PAM sequences ability to promote interference and primed adaptation

CRISPR interference occurs when a protospacer recognized by the CRISPR RNA is destroyed by Cas effectors. In Type I CRISPR‐Cas systems, protospacer recognition can lead to «primed adaptation» – acquisition of new spacers from in cis located sequences. Type I CRISPR‐Cas systems require the presence of a trinucleotide protospacer adjacent motif (PAM) for efficient interference. Here, we investigated the ability of each of 64 possible trinucleotides located at the PAM position to induce CRISPR interference and primed adaptation by the Escherichia coli Type I‐E CRISPR‐Cas system. We observed clear separation of PAM variants into three groups: those unable to cause interference, those that support rapid interference and those that lead to reduced interference that occurs over extended periods of time. PAM variants unable to support interference also did not support primed adaptation; those that supported rapid interference led to no or low levels of adaptation, while those that caused attenuated levels of interference consistently led to highest levels of adaptation. The results suggest that primed adaptation is fueled by the products of CRISPR interference. Extended over time interference with targets containing «attenuated» PAM variants provides a continuous source of new spacers leading to high overall level of spacer acquisition.     

Interferon gamma-dependent transactivation of epidermal growth factor receptor

The present report provides evidence that, in A431 cells, interferon gamma (IFNgamma) induces the rapid (within 5 min), and reversible, tyrosine phosphorylation of the epidermal growth factor receptor (EGFR). IFNgamma-induced EGFR transactivation requires EGFR kinase activity, as well as activity of the Src-family tyrosine kinases and JAK2. Here, we show that IFNgamma-induced STAT1 activation in A431 and HeLa cells partially depends on the kinase activity of both EGFR and Src. Furthermore, in these cells, EGFR kinase activity is essential for IFNgamma-induced ERK1,2 activation. This study is the first to demonstrate that EGFR is implicated in IFNgamma-dependent signaling pathways.