The dissociation of chicken erythrocyte deoxyribonucleoprotein and some properties of its partial nucleoproteins

Histones were completely dissociated from their native complex with DNA in 2.0m-sodium chloride. Histone fractions IIb, V and I were dissociated in 1.2m-sodium chloride, fractions V and I in 0.7m-sodium chloride and fraction I in 0.45m-sodium chloride. Repeated extraction of partial dRNP (deoxyribonucleoprotein) preparations with sodium chloride of the same concentration as that from which they were prepared resulted in release of histones that previously had remained associated with the DNA of the complex. Gradual removal of histones from dRNP was paralleled by an improvement in solubility, a decrease in wavelength of the u.v.-absorption minimum, and a fall in sedimentation coefficient of the remaining partial dRNP. X-ray diffraction patterns of partial dRNP preparations showed that removal of histone fractions I and V from dRNP did not destroy the super-coil structure of the dRNP, but further removal of histones did. Infrared spectra of partial dRNP preparations showed that in native dRNP histone fraction I was present in the form of extended, isolated polypeptide chains, and that the other histone fractions probably contain a helical component that lies roughly parallel to the polynucleotide chains in the double helix and an extended polypeptide component that is more nearly parallel to the DNA helix axis. An analysis of the sedimentation of partial dRNP preparations on sucrose gradients showed that native dRNP consists of DNA molecules each complexed with histone fractions of all types.     

The Fine Structure of the Adipose Cell of the Leech Glossiphonia complanata

The two distinct types of cytoplasm seen with the light microscope in the adipose cell of the leech Glossiphonia complanata have been identified in the electron microscope image of this cell. One of these, the basophil cytoplasm, contains many well oriented, paired membranes which are much more clearly evident when calcium ions are added to the fixative. The membranes sometimes appear as concentric arrays of lamellae and are thought to represent sections through a phospholipide-containing body. The paired membranes and the concentric lamellae have granules attached to them and resemble in size and structure the membranes of the endoplasmic reticulum encountered in many mammalian cells. Small dense cytoplasmic particles are present throughout the cell; they may be ferritin molecules, derived from the breakdown of haemoglobin taken in as food. On the basis of a previous histochemical study and the present electron microscope investigation, it is suggested that these paired membranes are similar to the organized type of mammalian ER and the results seem to confirm the belief that these membranes are composed of layers of phospholipoprotein together with attached particles of ribonucleoprotein.     

Recruitment of TBK1 to cytosol‐invading Salmonella induces WIPI2‐dependent antibacterial autophagy

Mammalian cells deploy autophagy to defend their cytosol against bacterial invaders. Anti‐bacterial autophagy relies on the core autophagy machinery, cargo receptors, and “eat‐me” signals such as galectin‐8 and ubiquitin that label bacteria as autophagy cargo. Anti‐bacterial autophagy also requires the kinase TBK1, whose role in autophagy has remained enigmatic. Here we show that recruitment of WIPI2, itself essential for anti‐bacterial autophagy, is dependent on the localization of catalytically active TBK1 to the vicinity of cytosolic bacteria. Experimental manipulation of TBK1 recruitment revealed that engagement of TBK1 with any of a variety of Salmonella‐associated “eat‐me” signals, including host‐derived glycans and K48‐ and K63‐linked ubiquitin chains, suffices to restrict bacterial proliferation. Promiscuity in recruiting TBK1 via independent signals may buffer TBK1 functionality from potential bacterial antagonism and thus be of evolutionary advantage to the host.     

Neutral 5-substituted 4-indazolylaminoquinazolines as potent, orally active inhibitors of erbB2 receptor tyrosine kinase

We have identified a new series of C-5 substituted indazolylaminoquinazolines as potent erbB2 kinase inhibitors. The lead compound 22 showed excellent in vitro potency, good physical properties, acceptable oral pharmacokinetics in rat and dog, and low human in vitro clearance. It showed at least equivalent activity dose for dose compared to lapatinib in various erbB2- or EGFR-driven xenograft models after chronic oral administration.     

Estimating contemporary early life-history dispersal in an estuarine fish: integrating molecular and otolith elemental approaches

Dispersal during the early life history of the anadromous rainbow smelt, Osmerus mordax , was examined using assignment testing and mixture analysis of multilocus genotypes and otolith elemental composition. Six spawning areas and associated estuarine nurseries were sampled throughout southeastern Newfoundland. Samples of adults and juveniles isolated by > 25 km displayed moderate genetic differentiation ( F _ST ~ 0.05), whereas nearby (< 25 km) spawning and nursery samples displayed low differentiation ( F _ST < 0.01). Self-assignment and mixture analysis of adult spawning samples supported the hypothesis of independence of isolated spawning locations (> 80% self-assignment) with nearby runs self-assigning at rates between 50 % and 70%. Assignment and mixture analysis of juveniles using adult baselines indicated high local recruitment at several locations (70–90%). Nearby (< 25 km) estuaries at the head of St Mary's Bay showed mixtures of individuals (i.e. 20–40% assignment to adjacent spawning location). Laser ablation inductively coupled mass spectrometry transects across otoliths of spawning adults of unknown dispersal history were used to estimate dispersal among estuaries across the first year of life. Single-element trends and multivariate discriminant function analysis (Sr:Ca and Ba:Ca) classified the majority of samples as estuarine suggesting limited movement between estuaries (< 0.5%). The mixtures of juveniles evident in the genetic data at nearby sites and a lack of evidence of straying in the otolith data support a hypothesis of selective mortality of immigrants. If indeed selective mortality of immigrants reduces the survivorship of dispersers, estimates of dispersal in marine environments that neglect survival may significantly overestimate gene flow.     

Rab11b Regulates the Apical Recycling of the Cystic Fibrosis Transmembrane Conductance Regulator in Polarized Intestinal Epithelial Cells

The cystic fibrosis transmembrane conductance regulator (CFTR), a cAMP/PKA-activated anion channel, undergoes efficient apical recycling in polarized epithelia. The regulatory mechanisms underlying CFTR recycling are understood poorly, yet this process is required for proper channel copy number at the apical membrane, and it is defective in the common CFTR mutant, ΔF508. Herein, we investigated the function of Rab11 isoforms in regulating CFTR trafficking in T84 cells, a colonic epithelial line that expresses CFTR endogenously. Western blotting of immunoisolated Rab11a or Rab11b vesicles revealed localization of endogenous CFTR within both compartments. CFTR function assays performed on T84 cells expressing the Rab11a or Rab11b GDP-locked S25N mutants demonstrated that only the Rab11b mutant inhibited 80% of the cAMP-activated halide efflux and that only the constitutively active Rab11b-Q70L increased the rate constant for stimulated halide efflux. Similarly, RNAi knockdown of Rab11b, but not Rab11a, reduced by 50% the CFTR-mediated anion conductance response. In polarized T84 monolayers, adenoviral expression of Rab11b-S25N resulted in a 70% inhibition of forskolin-stimulated transepithelial anion secretion and a 50% decrease in apical membrane CFTR as assessed by cell surface biotinylation. Biotin protection assays revealed a robust inhibition of CFTR recycling in polarized T84 cells expressing Rab11b-S25N, demonstrating the selective requirement for the Rab11b isoform. This is the first report detailing apical CFTR recycling in a native expression system and to demonstrate that Rab11b regulates apical recycling in polarized epithelial cells.     

Structural and functional connectivity of marine fishes within a semi-enclosed Newfoundland fjord

The interplay between structural connectivity ( i.e. habitat continuity) and functional connectivity ( i.e. dispersal probability) in marine fishes was examined in a coastal fjord (Holyrood Pond, Newfoundland, Canada) that is completely isolated from the North Atlantic Ocean for most of the year. Genetic differentiation was described in three species (rainbow smelt Osmerus mordax , white hake Urophycis tenuis and Atlantic cod Gadus morhua ) with contrasting life histories using seven to 10 microsatellite loci and a protein-coding locus, Pan I ( G. morhua ). Analysis of microsatellite differentiation indicated clear genetic differences between the fjord and coastal regions; however, the magnitude of difference was no more elevated than adjacent bays and was not enhanced by the fjord's isolation. Osmerus mordax was characterized by the highest structure overall with moderate differentiation between the fjord and St Mary's Bay ( F _ST c. 0·047). In contrast, U. tenuis and G. morhua displayed weak differentiation ( F _ST < 0·01). Nonetheless, these populations did demonstrate high rates (< 75%) of Bayesian self-assignment. Furthermore, elevated differentiation was observed at the Pan I locus in G. morhua between the fjord and other coastal locations. Interestingly, locus-specific genetic differentiation and expected heterozygosity were negatively associated in O. mordax , in contrast to the positive associations observed in U. tenuis and G. morhua . Gene flow in these species is apparently unencumbered by limited structural connectivity, yet the observed differentiation suggests that population structuring exists over small scales despite high dispersal potential.     

Codon usage bias and the evolution of influenza A viruses. Codon Usage Biases of Influenza Virus

Background  

The influenza A virus is an important infectious cause of morbidity and mortality in humans and was responsible for 3 pandemics in the 20^th century. As the replication of the influenza virus is based on its host's machinery, codon usage of its viral genes might be subject to host selection pressures, especially after interspecies transmission. A better understanding of viral evolution and host adaptive responses might help control this disease.     

AKT1 polymorphisms are associated with risk for metabolic syndrome

Converging lines of evidence suggest that AKT1 is a major mediator of the responses to insulin,insulin-like growth factor 1 (IGF1), and glucose. AKT1 also plays a key role in the regulation of both muscle cell hypertrophy and atrophy. We hypothesized that AKT1 variants may play a role in the endophenotypes that makeup metabolic syndrome. We studied a 12-kb region including the ?rst exon of the AKT1 gene for association with metabolic syndrome-related phenotypes in four study populations [FAMUSS cohort (n = 574; age 23.7 ± 5.7 years), Strong Heart Study (SHS) (n = 2,134; age 55.5 ± 7.9 years), Dynamics of Health, Aging and Body Composition (Health ABC) (n = 3,075; age 73.6 ± 2.9 years), and Studies of a Targeted Risk Reduction Intervention through De?ned Exercise (STRRIDE)(n = 175; age 40–65 years)]. We identi?ed a three SNP haplotype that we call H1, which represents the ancestral alleles eles at the three loci and H2, which represents the derived alleles at the three loci. In young adult European Americans (FAMUSS), H1 was associated with higher fasting glucose levels in females. In middle age Native Americans (SHS), H1 carriers showed higher fasting insulin and HOMA in males, and higher BMI in females. Inolder African-American and European American subjects(Health ABC) H1 carriers showed a higher incidence of metabolic syndrome. Homozygotes for the H1 haplotype showed about twice the risk of metabolic syndrome in both males and females (p\0.001). In middle-aged European Americans with insulin resistance (STRRIDE) studied by intravenous glucose tolerance test (IVGTT), H1 carriers showed increased insulin resistance due to the Sg component (p = 0.021). The 12-kb haplotype is a risk factor for metabolic syndrome and insulin resistance that needs to be explored in further populations.