N-Glycosylation regulates endothelial lipase-mediated phospholipid hydrolysis in apoE- and apoA-I-containing high density lipoproteins

Endothelial lipase (EL) is a member of the triglyceride lipase gene family with high phospholipase and low triacylglycerol lipase activities and a distinct preference for hydrolyzing phospholipids in HDL. EL has five potential N-glycosylation sites, four of which are glycosylated. The aim of this study was to determine how glycosylation affects the phospholipase activity of EL in physiologically relevant substrates. Site-directed mutants of EL were generated by replacing asparagine (N) 62, 118, 375, and 473 with alanine (A). These glycan-deficient mutants were used to investigate the kinetics of phospholipid hydrolysis in fully characterized preparations of spherical reconstituted high density lipoprotein (rHDL) containing apolipoprotein E2 (apoE2) [(E2)rHDL], apoE3 [(E3)rHDL], apoE4 [(E4)rHDL], or apoA-I [(A-I)rHDL] as the sole apolipoprotein. Wild-type EL hydrolyzed the phospholipids in (A-I)rHDL, (E2)rHDL, (E3)rHDL, and (E4)rHDL to similar extents. The phospholipase activities of EL N118A, EL N375A, and EL N473A were significantly diminished relative to that of wild-type EL, with the greatest reduction being apparent for (E3)rHDL. The phospholipase activity of EL N62A was increased up to 6-fold relative to that of wild-type EL, with the greatest enhancement of activity being observed for (E2)rHDL. These data show that individual N-linked glycans have unique and important effects on the phospholipase activity and substrate specificity of EL.     

Tryptophan Biosynthesis in Stramenopiles: Eukaryotic Winners in the Diatom Complex Chloroplast

Tryptophan is an essential amino acid that, in eukaryotes, is synthesized either in the plastids of photoautotrophs or in the cytosol of fungi and oomycetes. Here we present an in silico analysis of the tryptophan biosynthetic pathway in stramenopiles, based on analysis of the genomes of the oomycetes Phytophthora sojae and P. ramorum and the diatoms Thalassiosira pseudonana and Phaeodactylum tricornutum. Although the complete pathway is putatively located in the complex chloroplast of diatoms, only one of the involved enzymes, indole-3-glycerol phosphate synthase (InGPS), displays a possible cyanobacterial origin. On the other hand, in P. tricornutum this gene is fused with the cyanobacteria-derived hypothetical protein COG4398. Anthranilate synthase is also fused in diatoms. This fusion gene is almost certainly of bacterial origin, although the particular source of the gene cannot be resolved. All other diatom enzymes originate from the nucleus of the primary host (red alga) or secondary host (ancestor of chromalveolates). The entire pathway is of eukaryotic origin and cytosolic localization in oomycetes; however, one of the enzymes, anthranilate phosphoribosyl transferase, was likely transferred to the oomycete nucleus from the red algal nucleus during secondary endosymbiosis. This suggests possible retention of the complex plastid in the ancestor of stramenopiles and later loss of this organelle in oomycetes. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Quantitative DNA hybridization in solution using magnetic/luminescent core-shell nanoparticles

Nanoscale magnetic/luminescent core-shell particles were used for DNA quantification in a hybridization-in-solution approach. We demonstrated a rapid, simple, and non-polymerase chain reaction-based DNA hybridization-in-solution assay for quantifying bacteria capable of biodegrading methyl tertiary-butyl ether. Fe3O4/Eu:Gd2O3 core-shell nanoparticles synthesized by spray pyrolysis were biofunctionalized with NeutrAvidin. Following immobilization of a biotinylated probe DNA on the particles' surfaces via passive adsorption, target DNA labeled with fluorescein isothiocyanate was hybridized with probe DNA. The hybridized DNA complex was separated from solution with a magnet, while nonhybridized DNA remained in solution. The normalized fluorescence (fluorescein isothiocyanate/nanoparticles) measured with a spectrofluorometer indicated a linear quantification (R(2)=0.98) of the target bacterial 16 S rDNA. The rate of hybridization increased concurrently with the target DNA concentration. In addition, this approach differentiated between the signal outputs from perfectly complementary target and two-base mismatched target DNA in a range of concentrations, showing the specificity of the assay and the possibility for environmental applications.     

Involving undergraduates in the annotation and analysis of global gene expression studies: creation of a maize shoot apical meristem expression database

Through a multi-university and interdisciplinary project we have involved undergraduate biology and computer science research students in the functional annotation of maize genes and the analysis of their microarray expression patterns. We have created a database to house the results of our functional annotation of >4400 genes identified as being differentially regulated in the maize shoot apical meristem (SAM). This database is located at http://sam.truman.edu and is now available for public use. The undergraduate students involved in constructing this unique SAM database received hands-on training in an intellectually challenging environment, which has prepared them for graduate and professional careers in biological sciences. We describe our experiences with this project as a model for effective research-based teaching of undergraduate biology and computer science students, as well as for a rich professional development experience for faculty at predominantly undergraduate institutions.     

Survivorship,growth and reproduction of the non-native <Emphasis Type="Italic">Caprella mutica</Emphasis> Schurin, 1935 (Crustacea: Amphipoda)

Caprella mutica Schurin is an epifaunal amphipod crustacean which originates in north-east Asia and has spread throughout the world, yet very little is known about fundamental aspects of this species biology. This paper examined the survivorship of C. mutica reared under laboratory conditions at 13–14 °C, 14 h light: 10 h dark photoperiod and fed commercial salmon feed, the diatom Cylindrotheca fusiformis Reumann and Lewin, the macroalgae, Fucus vesiculosus L. and given no additional feed. In addition, growth, maturation and reproduction of C. mutica fed C. fusiformis were assessed. No significant difference in survivorship of C. mutica was observed for the diet types over the experimental period. C. mutica was able to survive for upto 20 days without additional food. Average survival time of males and females fed the diatom, C. fusiformis was 68.8 d (range = 62–73 d) and 82.0 d (range = 76–92 d). Juvenile C. mutica emerged from the brood pouch at a body length of 1.33 mm and moulted at 5.0–11.0 day intervals. Males exhibited faster growth rates than females after Instar VII. Females produced their first brood at Instar VII, 24–26 days post-hatching and with an average body length of 8.5 mm. Each female had an average of two broods sequentially and these were released at 20.2 day intervals. Brood size for a single female increased from 11.3 (±9.9) hatchlings at Instar VII to 25.5 (±11.5) at Instar IX and the maximum number of hatchlings produced by a single female was 82. The results suggest that C. mutica exhibits a number of life-history traits that would potentially enable it to withstand global transportation and to rapidly become established in an introduced region, if environmental conditions are suitable.     

Structural basis of integrin activation by talin

Regulation of integrin affinity (activation) is essential for metazoan development and for many pathological processes. Binding of the talin phosphotyrosine-binding (PTB) domain to integrin beta subunit cytoplasmic domains (tails) causes activation, whereas numerous other PTB-domain-containing proteins bind integrins without activating them. Here we define the structure of a complex between talin and the membrane-proximal integrin beta3 cytoplasmic domain and identify specific contacts between talin and the integrin tail required for activation. We used structure-based mutagenesis to engineer talin and beta3 variants that interact with comparable affinity to the wild-type proteins but inhibit integrin activation by competing with endogenous talin. These results reveal the structural basis of talin's unique ability to activate integrins, identify an interaction that could aid in the design of therapeutics to block integrin activation, and enable engineering of cells with defects in the activation of multiple classes of integrins.     

Naricolax hoi n. sp. (Cyclopoida: Bomolochidae) from Arius maculatus (Siluriformes: Ariidae) off Taiwan and a redescription of N. chrysophryenus (Roubal, Armitage & Rohde, 1983) from a new host, Seriola lalandi (Perciformes: Carangidae), in Australian waters

We propose that Naricolax stocki (Roubal, 1981) (Cyclopoida: Bomolochidae) of Ho & Lin (2005), reported from the spotted catfish Arius maculatus (Thunburg) off Taiwan, represents a new species, N. hoi n. sp. N. hoi can be distinguished from six known congeners by the shape of the rostral area, the maxillary armature and the structural details of legs 3 and 4. N. chrysophryenus (Roubal, Armitage & Rohde, 1983) is redescribed on the basis of recently collected material from wild and farmed yellowtail kingfish Seriola lalandi Valenciennes in southern and eastern Australian waters, providing the first record of Naricolax Ho, Do & Kasahara, 1983 from a carangid host. A key to the species of Naricolax is provided.     

Intravenous CCK-8, but not GLP-1, suppresses ghrelin and stimulates PYY release in healthy men

We have investigated the effects of exogenous CCK-8 and GLP-1, alone and in combination, on ghrelin and PYY secretion. Nine healthy males were studied on four occasions. Plasma ghrelin and PYY concentrations were measured during 150 min intravenous infusions of: (i) isotonic saline, (ii) CCK-8 at 1.8 pmol/kg/min, (iii) GLP-1 at 0.9 pmol/kg/min or (iv) CCK-8 and GLP-1 combined. CCK-8 markedly suppressed ghrelin and stimulated PYY when compared with control between t=0-120 min (P<0.001 for both). GLP-1 had no effect on ghrelin, but decreased PYY slightly at 120 min (P<0.05). During infusion of CCK-8+GLP-1, there was comparable suppression of ghrelin (P<0.001), but the stimulation of PYY was less (P<0.001), than that induced by CCK-8, between t=20-120 min. In conclusion, in healthy subjects, in the doses evaluated, exogenous CCK-8 suppresses ghrelin and stimulates PYY, and exogenous GLP-1 has no effect on ghrelin and attenuates the effect of CCK-8 on PYY.