Acceptor specificity of the human leukocyte alpha3 fucosyltransferase: role of FucT-VII in the generation of selectin ligands

The alpha3 fucosyltransferase, FucT-VII, is one of the key glycosyltransferases involved in the biosynthesis of the sialyl Lewis X (sLex) antigen on human leukocytes. The sialyl Lewis X antigen (NeuAcalpha(2-3)Galbeta(1-4)[Fucalpha(1-3)]GlcNAc-R) is an essential component of the recruitment of leukocytes to sites of inflammation, mediating the primary interaction between circulating leukocytes and activated endothelium. In order to characterize the enzymatic properties of the leukocyte alpha3 fucosyltransferase FucT-VII, the enzyme has been expressed in Trichoplusia ni insect cells. The enzyme is capable of synthesizing both sLexand sialyl-dimeric-Lexstructures in vitro , from 3'-sialyl-lacNAc and VIM-2 structures, respectively, with only low levels of fucose transfer observed to neutral or 3'-sulfated acceptors. Studies using fucosylated NeuAcalpha(2-3)-(Galbeta(1- 4)GlcNAc)3-Me acceptors demonstrate that FucT-VII is able to synthesize both di-fucosylated and tri-fucosylated structures from mono- fucosylated precursors, but preferentially fucosylates the distal GlcNAc within a polylactosamine chain. Furthermore, the rate of fucosylation of the internal GlcNAc residues is reduced once fucose has been added to the distal GlcNAc. These results indicate that FucT-VII is capable of generating complex selectin ligands, in vitro , however the order of fucose addition to the lactosamine chain affects the rate of selectin ligand synthesis.      

Distinct cold tolerance traits independently vary across genotypes in Drosophila melanogaster

Cold tolerance, the ability to cope with low temperature stress, is a critical adaptation in thermally variable environments. An individual's cold tolerance comprises several traits including minimum temperatures for growth and activity, ability to survive severe cold, and ability to resume normal function after cold subsides. Across species, these traits are correlated, suggesting they were shaped by shared evolutionary processes or possibly share physiological mechanisms. However, the extent to which cold tolerance traits and their associated mechanisms covary within populations has not been assessed. We measured five cold tolerance traits—critical thermal minimum, chill coma recovery, short- and long-term cold tolerance, and cold-induced changes in locomotor behavior—along with cold-induced expression of two genes with possible roles in cold tolerance (heat shock protein 70 and frost)—across 12 lines of Drosophila melanogaster derived from a single population. We observed significant genetic variation in all traits, but few were correlated across genotypes, and these correlations were sex-specific. Further, cold-induced gene expression varied by genotype, but there was no evidence supporting our hypothesis that cold-hardy lines would have either higher baseline expression or induction of stress genes. These results suggest cold tolerance traits possess unique mechanisms and have the capacity to evolve independently.     

Avrainvillea rotumensis sp. nov. (Bryopsidales, Chlorophyta), a peltate species from the South Pacific

A new species of Avrainvillea was found on the South Pacific island of Rotuma, Fiji. Avrainvillea rotumensis sp. nov. occurs 1.5-3.0 m deep in a high energy current area of the Hoféa Passage, one of the few openings in the fringing reef that surrounds the island. The distinctive peltate growth habit of A. rotumensis is unique for the genus and facilitates quick and accurate field identification. The peltate blade (7–9 cm in diameter at maturity) is unusually thick (34 mm) tapering toward a short (up to 6 cm in length), thick (1.5-2.0 cm in diameter) stipe.     

Spatiotemporal Regulation of Chloride Intracellular Channel Protein CLIC4 by RhoA

Chloride intracellular channel (CLIC) 4 is a soluble protein structurally related to omega-type glutathione-S-transferases (GSTs) and implicated in various biological processes, ranging from chloride channel formation to vascular tubulogenesis. However, its function(s) and regulation remain unclear. Here, we show that cytosolic CLIC4 undergoes rapid but transient translocation to discrete domains at the plasma membrane upon stimulation of G₁₃-coupled, RhoA-activating receptors, such as those for lysophosphatidic acid, thrombin, and sphingosine-1-phosphate. CLIC4 recruitment is strictly dependent on Gα₁₃-mediated RhoA activation and F-actin integrity, but not on Rho kinase activity; it is constitutively induced upon enforced RhoA-GTP accumulation. Membrane-targeted CLIC4 does not seem to enter the plasma membrane or modulate transmembrane chloride currents. Mutational analysis reveals that CLIC4 translocation depends on at least six conserved residues, including reactive Cys35, whose equivalents are critical for the enzymatic function of GSTs. We conclude that CLIC4 is regulated by RhoA to be targeted to the plasma membrane, where it may function not as an inducible chloride channel but rather by displaying Cys-dependent transferase activity toward a yet unknown substrate.