Microheterogeneity of human galactose-1-Phosphate uridylyl transferase. Isoelectrofocusing results

Using thin-layer acrylamide gel isoelectrofocusing, several bands of galactose-1-Phosphate uridylyl transferase were found in various human tissues. Liver transferase, as well as that of some other tissues, was resolved into several bands with pHi between 5.30 and 5.80; red cell enzyme was resolved into five bands with pHi between 5.0 and 5.45. The comparison of erythrocytes with their precursors, reticulocytes and erythroblasts, showed a striking difference: the pHi of the erythroblast enzyme was between 5.55 and 5.90 and that of reticulocytes between 5.30 and 5.50. It is possible that molecular aging is the cause of the anodisation of the erythrocyte transferase and the microheterogeneity of the enzyme observed in other tissues.     

Enzymatic O-glycosylation of kappa-caseinomacropeptide by ovine mammary Golgi membranes

The results of subcellular fractionation of sheep mammary gland membranes indicate that N-acetylgalactosaminyl polypeptide transferase and galactosyl-N-acetylgalactosaminyl transferase, which are involved in the assembly of disaccharide units of kappa-casein, are localized chiefly in Golgi membranes. The glycosyltransferase activities incorporating N-acetyl [1-14C] galactosamine and [U-14C] galactose from uridine diphosphate N-acetyl [1-14C] galactosamine and uridine diphosphate [U-14C] galactose, respectively, were measured after membrane solubilization with Triton X-100 either with unglycosylated caseinomacropeptide, or with this polypeptide containing the N-acetylgalactosamine side chain residues (desialylated and degalactosylated caseinomacropeptide). Radioactive N-acetylgalactosamine was incorporated in the unglycosylated acceptor peptide, and the glycosidic bonds in the product were alkali labile, suggesting that they were linked to the hydroxyamino acid residues. In addition radioactive N-acetylgalactosamine was released after alpha N-acetyl-D-galactosaminidase treatment of labelled caseinomacropeptide. [U-14C] galactose was incorporated in the desialylated and degalactosylated acceptor peptide. Reductive alkaline treatment of [U-14C] galactose peptide resulted in the release of a major product, the chromatographic properties of which in TLC were identical with authentic galactosyl (1 leads to 3) N-acetylgalactosaminitol. The structure of the labelled disacchariditol determined after periodate oxidation (two equivalents) by gas liquid chromatography-mass spectrometry revealed that the [U-14C] galactose was linked to position C-3 on the N-acetylgalactosaminyl-residue. The anomery of the galactose, as determined by a chemical method, indicates unambiguously a beta configuration.     

The molecular size of glycans liberated by hydrazinolysis from semliki forest virus proteins

The glycans of well characterized, [6-³H]galactose-labelled glycopeptides, GC-4 from bovine IgG₁ as well as GP-V-2 and GP-V-5 from α₁-acid glycoprotein, were liberated by hydrazinolysis. Molecular weights close to the expected values were observed by gel filtration. Desialated glycans of Semliki Forest virus proteins were likewise liberated by hydrazinolysis and subjected to gel filtration. Metabolically labelled [1-³H]galactose-oligosaccharides of the mixed viral proteins revealed an apparent molecular weight of 1800. The bi-antennary glycan liberated from the reference glycopeptide GC-4 was of 1750 daltons. A mixture of [2-³H]mannose-labelled E₁-and E₂-proteins of the virus contained L-type glycans of 1800 daltons (formerly called A-type), and M-type glycans of 1200 daltons (formerly called B-type). A fraction of the E₃-glycans isolated by affinity chromatography on Concanavalin A-Sepharose showed an average molecular weight of 2150, a value intermediate between the three- and four-antennary glycans liberated from the reference glycopeptides GP-V-5 and GP-V-2. The rest of the E₃-glycans were of 1850 daltons, a value close to the bi-antennary GC-4 glycan. We suggest that the comparatively large size of the E₃-glycans and the exposed position of E₃-proteins on the viral surface may be interrelated.     

Ectopic expression of ecdysone oxidase impairs tissue degeneration in Bombyx mori

Metamorphosis in insects includes a series of programmed tissue histolysis and remolding processes that are controlled by two major classes of hormones, juvenile hormones and ecdysteroids. Precise pulses of ecdysteroids (the most active ecdysteroid is 20-hydroxyecdysone, 20E), are regulated by both biosynthesis and metabolism. In this study, we show that ecdysone oxidase (EO), a 20E inactivation enzyme, expresses predominantly in the midgut during the early pupal stage in the lepidopteran model insect, Bombyx mori. Depletion of BmEO using the transgenic CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/RNA-guided Cas9 nucleases) system extended the duration of the final instar larval stage. Ubiquitous transgenic overexpression of BmEO using the Gal4/UAS system induced lethality during the larval–pupal transition. When BmEO was specifically overexpressed in the middle silk gland (MSG), degeneration of MSG at the onset of metamorphosis was blocked. Transmission electron microscope and LysoTracker analyses showed that the autophagy pathway in MSG is inhibited by BmEO ectopic expression. Furthermore, RNA-seq analysis revealed that the genes involved in autophagic cell death and the mTOR signal pathway are affected by overexpression of BmEO. Taken together, BmEO functional studies reported here provide insights into ecdysone regulation of tissue degeneration during metamorphosis.     

Characterisation of Rubus niveus: a prerequisite to its biological control in oceanic islands

This study is intended to inform the search for a biocontrol solution to stem the spread of Rubus niveus, a morphologically-diverse and invasive Rubus species which has become an aggressive invader in areas of the world outside its native range. The fragile biodiversity of islands can be especially threatened by this plant – exemplars of this are the Galápagos archipelago and Hawaii. This report focuses on the morphological and molecular work carried out to establish the homogeneity or intra-specific variation of R. niveus populations within the Galápagos Islands, as well as their kinship with plants of the same name found in Asia. The premise of this study is that genetic closeness between these populations offers increased probability that pathogens of R. niveus in its native range will also provide effective control in the target environment. Increased knowledge of the genetic make-up of R. niveus in both its native and naturalised locations will also help to assess the feasibility of using organisms which have been found to be effective against other Rubus species, but not hitherto considered for R. niveus itself.     

酵母转录因子Gal4研究进展

胁迫应答基因的转录激活是细胞应答胁迫作用的关键步骤。转录激活因子与启动子顺式作用元件结合是胁迫应答基因转录激活的关键环节。进化保守的Gal4是半乳糖代谢相关基因的转录激活因子。酵母Gal4通过其N端的DNA结合结构域识别并结合启动子UAS,通过其C端的激活结构域与转录因子作用,起始RNA聚合酶Ⅱ复合体的组装和转录。该过程不仅受转录调控因子Gal80和Gal3的调节,还与Gal4二聚体的形成有关。概述了酵母半乳糖代谢相关基因转录激活因子Gal4的研究进展。     

DIATOMS (BACILLARIOPHYTA) OF ISOLATED ISLANDS: NEW TAXA IN THE GENUS NAVICULA SENSU STRICTO FROM THE GALáPAGOS ISLANDS1

The diatoms (Bacillariophyta) from a coastal lagoon from the Diablas wetlands (Isla Isabela, the Galápagos Islands) were studied in material from surface samples and a sediment core spanning the past 2,700 years in order to examine evidence of diatom evolution under geographic isolation. The total number of taxa found was ～100. Ultrastructural variation in valve morphology between members of Galápagos taxa was used to describe 10 species from the genus Navicula sensu stricto, which are new to science. Four taxa: N. isabelensis, N. isabelensoides, N. isabelensiformis, and N. isabelensiminor, shared several key characteristics that may be indicative of a common evolutionary heritage; these species therefore provide possible evidence for the in situ evolution of diatoms in the Galápagos coastal lagoons. Shared morphological characteristics include: (i) stria patterning in the central area, (ii) an elevated and thickened external raphe‐sternum, (iii) external central raphe endings that are slightly deflected toward the valve primary side, and (iv) an arched valve surface. To explain these findings, two models were proposed. The first suggested limited lateral diatomaceous transport of Navicula species between the Galápagos and continental South America. Alternatively, these new species may be ecological specialists arising from the unique environmental conditions of the Galápagos coastal lagoons, which restrict the colonization of common diatom taxa and enable the establishment of novel, rare species. The Diablas wetlands are an important site for diatom research, where local‐scale environmental changes have combined with global‐scale biogeographic processes resulting in unique diatom assemblages.