Phenotypic plasticity in breeding plumage signals in both sexes of a migratory bird: responses to breeding conditions

Adaptive phenotypic plasticity may respond to present ambient conditions. Sexual and social signals in both sexes may express phenotype performance. Plumage signals that change discontinuously allow relating discrete variation to previous performance. Both sexes of the pied flycatcher Ficedula hypoleuca present white patches on the wings and on the forehead, which constitute sexual and social signals. Forehead patches are moulted together with body plumage in Africa, while wing patches are partly moulted in Africa and partly in the breeding area soon after breeding. We studied individual inter‐year changes (corrected for regression to the mean) in the size of forehead and wing patches of both sexes in seven years for females or six years for males in two nearby study areas in central Spain. We found that initial signal extent strongly delimits the possible subsequent changes negatively. There is a negative association of male age with forehead patch changes. Cold and rainy springs are associated in females with decreases in both patch areas and vice versa, while no association with climate is observed in male wing patch changes. Cold pre‐breeding conditions predict positive changes in female wing and male forehead patches. Breeding success is positively associated with forehead patch changes in females. Late‐breeding males experience more positive changes in forehead patch size than early‐breeding males. Some of these trends can be explained by variable costs of breeding in certain conditions for subsequent signal production and/or maintenance, while absence of trends in some cases may be explained by sex differences in costs of breeding and interactions with phenotypic quality of breeders.     

Brain proteomics identifies potential simvastatin targets in acute phase of stroke in a rat embolic model

Finding an efficient neuroprotectant is of urgent need in the field of stroke research. The goal of this study was to test the effect of acute simvastatin administration after stroke in a rat embolic model and to explore its mechanism of action through brain proteomics. To that end, male Wistar rats were subjected to a Middle Cerebral Arteria Occlusion and simvastatin (20 mg/kg s.c) (n = 11) or vehicle (n = 9) were administered 15 min after. To evaluate the neuroprotective mechanisms of simvastatin, brain homogenates after 48 h were analyzed by two‐dimensional fluorescence Difference in Gel Electrophoresis (DIGE) technology. We confirmed that simvastatin reduced the infarct volume and improved neurological impairment at 48 h after the stroke in this model. Considering our proteomics analysis, 66 spots, which revealed significant differences between groups, were analyzed by matrix‐assisted laser desorption/ionization‐time of flight mass spectrometry allowing the identification of 27 proteins. From these results, we suggest that simvastatin protective effect can be partly explained by the attenuation of the oxidative and stress response at blood–brain barrier level after cerebral ischemia. Interestingly, analyzing one of the proteins (HSP75) in plasma from stroke patients who had received simvastatin during the acute phase, we confirmed the results found in the pre‐clinical model.

     

Microrheology of human lung epithelial cells measured by atomic force microscopy

Lung epithelial cells are subjected to large cyclic forces from breathing. However, their response to dynamic stresses is poorly defined. We measured the complex shear modulus (G(*)(omega)) of human alveolar (A549) and bronchial (BEAS-2B) epithelial cells over three frequency decades (0.1-100 Hz) and at different loading forces (0.1-0.9 nN) with atomic force microscopy. G(*)(omega) was computed by correcting force-indentation oscillatory data for the tip-cell contact geometry and for the hydrodynamic viscous drag. Both cell types displayed similar viscoelastic properties. The storage modulus G'(omega) increased with frequency following a power law with exponent approximately 0.2. The loss modulus G"(omega) was approximately 2/3 lower and increased similarly to G'(omega) up to approximately 10 Hz, but exhibited a steeper rise at higher frequencies. The cells showed a weak force dependence of G'(omega) and G"(omega). G(*)(omega) conformed to the power-law model with a structural damping coefficient of approximately 0.3, indicating a coupling of elastic and dissipative processes within the cell. Power-law behavior implies a continuum distribution of stress relaxation time constants. This complex dynamics is consistent with the rheology of soft glassy materials close to a glass transition, thereby suggesting that structural disorder and metastability may be fundamental features of cell architecture.     

Twin seasonality in a rural Catalonian population

The seasonality of twinning in the Spanish populations has not been studied until now. Differences between seasonal distribution of the twin conceptions and those of the single births have been observed in other populations. The aim of this work is to explore the frequency of twinning in a rural population from Catalonia during the nineteenth century, as well as the seasonality patterns characterizing each of the twinning types. Data corresponding to all births recorded at Tortosa (South Catalonia) from 1801 to 1900 have been analyzed in order to study the twinning distribution. The distribution of the moving averages of the monthly rates of twins shows a peak in autumn. Twinning distribution differs from the total births' distribution in Tortosa. This fact is very clear in the case of unlike-sexed twins that have their greater incidence in the last quarter of the year, while the total maternities have their peak in the first one.     

Thrombin and histamine induce stiffening of alveolar epithelial cells.

The mechanical properties of alveolar epithelial cells play a central role in maintaining the physical integrity of the alveolar epithelium. We studied the viscoelastic properties of alveolar epithelial cells (A549) in response to thrombin and histamine with optical magnetic twisting cytometry. Ferrimagnetic beads coated with Arg-Gly-Asp (RGD)-peptide or acetylated low-density lipoprotein were bound to cell surface receptors and subsequently twisted in an oscillatory magnetic field (0.1-100 Hz). The cell storage (G') and loss (G') moduli were computed from twisting torque and bead displacement. In measurements with RGD-coated beads, thrombin (0.5 U/ml) induced a rapid and sustained threefold increase in G' and G' at approximately 100 s after challenge. Histamine (100 microM) induced a rapid but transient twofold increase in G' and G' with maximum values 60 s after challenge. Posttreatment with cytochalasin D abolished thrombin-induced cell stiffening. G' increased with frequency following a power law with exponent 0.214. G' increased proportionally to G' up to 10 Hz but showed a steeper rise at higher frequencies. Thrombin caused a fall in the power-law exponent (0.164). In measurements with acetylated low-density lipoprotein-coated beads, minor changes (<20%) were observed in G' and G' after the addition of thrombin and histamine. F-actin staining revealed that thrombin and histamine induced a profound reorganization of the actin cytoskeleton at the cell periphery and formation of actin bundles. In the mechanically dynamic environment of the lung, cell stiffening induced by thrombin and histamine increases centripetal tension, which could contribute to alveolar barrier dysfunction.     

Mechanism of p38 MAPK–induced EGFR endocytosis and its crosstalk with ligand-induced pathways

Ligand binding triggers clathrin-mediated and, at high ligand concentrations, clathrin-independent endocytosis of EGFR. Clathrin-mediated endocytosis (CME) of EGFR is also induced by stimuli activating p38 MAPK. Mechanisms of both ligand- and p38-induced endocytosis are not fully understood, and how these pathways intermingle when concurrently activated remains unknown. Here we dissect the mechanisms of p38-induced endocytosis using a pH-sensitive model of endogenous EGFR, which is extracellularly tagged with a fluorogen-activating protein, and propose a unifying model of the crosstalk between multiple EGFR endocytosis pathways. We found that a new locus of p38-dependent phosphorylation in EGFR is essential for the receptor dileucine motif interaction with the σ2 subunit of clathrin adaptor AP2 and concomitant receptor internalization. p38-dependent endocytosis of EGFR induced by cytokines was additive to CME induced by picomolar EGF concentrations but constrained to internalizing ligand-free EGFRs due to Grb2 recruitment by ligand-activated EGFRs. Nanomolar EGF concentrations rerouted EGFR from CME to clathrin-independent endocytosis, primarily by diminishing p38-dependent endocytosis.     

Cell Envelope Components Influencing Filament Length in the Heterocyst-Forming Cyanobacterium Anabaena sp. Strain PCC 7120

Heterocyst-forming cyanobacteria grow as chains of cells (known as trichomes or filaments) that can be hundreds of cells long. The filament consists of individual cells surrounded by a cytoplasmic membrane and peptidoglycan layers. The cells, however, share a continuous outer membrane, and septal proteins, such as SepJ, are important for cell-cell contact and filament formation. Here, we addressed a possible role of cell envelope components in filamentation, the process of producing and maintaining filaments, in the model cyanobacterium Anabaena sp. strain PCC 7120. We studied filament length and the response of the filaments to mechanical fragmentation in a number of strains with mutations in genes encoding cell envelope components. Previously published peptidoglycan- and outer membrane-related gene mutants and strains with mutations in two genes (all5045 and alr0718) encoding class B penicillin-binding proteins isolated in this work were used. Our results show that filament length is affected in most cell envelope mutants, but the filaments of alr5045 and alr2270 gene mutants were particularly fragmented. All5045 is a dd-transpeptidase involved in peptidoglycan elongation during cell growth, and Alr2270 is an enzyme involved in the biosynthesis of lipid A, a key component of lipopolysaccharide. These results indicate that both components of the cell envelope, the murein sacculus and the outer membrane, influence filamentation. As deduced from the filament fragmentation phenotypes of their mutants, however, none of these elements is as important for filamentation as the septal protein SepJ.     

p53 Gene repair with zinc finger nucleases optimised by yeast 1-hybrid and validated by Solexa sequencing

The tumor suppressor gene p53 is mutated or deleted in over 50% of human tumors. As functional p53 plays a pivotal role in protecting against cancer development, several strategies for restoring wild-type (wt) p53 function have been investigated. In this study, we applied an approach using gene repair with zinc finger nucleases (ZFNs). We adapted a commercially-available yeast one-hybrid (Y1H) selection kit to allow rapid building and optimization of 4-finger constructs from randomized PCR libraries. We thus generated novel functional zinc finger nucleases against two DNA sites in the human p53 gene, near cancer mutation 'hotspots'. The ZFNs were first validated using in vitro cleavage assays and in vivo episomal gene repair assays in HEK293T cells. Subsequently, the ZFNs were used to restore wt-p53 status in the SF268 human cancer cell line, via ZFN-induced homologous recombination. The frequency of gene repair and mutation by non-homologous end-joining was then ascertained in several cancer cell lines, using a deep sequencing strategy. Our Y1H system facilitates the generation and optimisation of novel, sequence-specific four- to six-finger peptides, and the p53-specific ZFN described here can be used to mutate or repair p53 in genomic loci.     

Functional dissection of the <Emphasis Type="Italic">ash2</Emphasis> and <Emphasis Type="Italic">ash1</Emphasis> transcriptomes provides insights into the transcriptional basis of wing phenotypes and reveals conserved protein interactions

Background  

The trithorax group (trxG) genes absent, small or homeotic discs 1 (ash1) and 2 (ash2) were isolated in a screen for mutants with abnormal imaginal discs. Mutations in either gene cause homeotic transformations but Hox genes are not their only targets. Although analysis of double mutants revealed that ash2 and ash1 mutations enhance each other's phenotypes, suggesting they are functionally related, it was shown that these proteins are subunits of distinct complexes.     

pH-responsive polysaccharide-based polyelectrolyte complexes as nanocarriers for lysosomal delivery of therapeutic proteins

Nanopharmaceutics composed of a carrier and a protein have the potential to improve the activity of therapeutical proteins. Therapy for lysosomal diseases is limited by the lack of effective protein delivery systems that allow the controlled release of specific proteins to the lysosomes. Here we address this problem by developing functional polyelectrolyte-based nanoparticles able to promote acidic pH-triggered release of the loaded protein. Trimethyl chitosan (TMC) was synthesized and allowed to form polyelectrolyte complexes (PECs) with the lysosomal enzyme α-GAL through self-assembly and ionotropic gelation, with average particle size <200 nm, polydispersity index (PDI) <0.2, ζ potential of ～ 20 mV, and a protein loading efficiency close to 65%. These polyelectrolyte nanoparticles were stable and active under physiological conditions and able to release the enzyme at acidic pH, as demonstrated by in situ atomic force microscopy (AFM). These nanoparticles were further functionalized with Atto 647N for single-particle characterization and tracking their cellular uptake and fate using high-resolution fluorescence microscopy. In contrast with their precursor, TMC, PECs were efficiently internalized by human endothelial cells and mostly accumulated in lysosomal compartments. The superior physicochemical characteristics of the TMC/α-GAL PECs together with their excellent cellular uptake properties indicate their enormous potential as advanced protein delivery systems for the treatment of lysosomal storage diseases.