Plasmodium falciparum RuvB1 is an active DNA helicase and translocates in the 5′–3′ direction

RuvB family of protein contains two similar kinds of proteins i.e. RuvB1 and RuvB2 from yeast to human. These proteins belong to the AAA + class of proteins and are critical components of several multiprotein complexes involved in diverse cellular activities. There are two RuvB proteins annotated in the Plasmodium database but the identification of the third protein recently by our lab has raised the question why Plasmodium falciparum contains three RuvB proteins instead of two. Hence the biochemical characterizations of these proteins have become essential to understand the role of these proteins in the malaria parasite. Recently we have reported the characterization of the recombinant PfRuvB3, which contains ATPase activity but lacks DNA helicase activity. In the present study we report the phylogenetic analysis and detailed biochemical characterization of one of the other RuvB homologue RuvB1 from P. falciparum. PfRuvB1 shows considerable homology with human as well as yeast RuvB1 and contains Walker motif A and Walker motif B. The activity analysis of this protein revealed that PfRuvB1 is an ATPase and this activity increased significantly in the presence of ss-DNA. PfRuvB1 also contains DNA helicase activity and translocates preferentially in 5′ to 3′ direction. In vivo investigation of PfRuvB1 revealed that it is constitutively expressed during all the stages of intraerythrocytic cycle of P. falciparum and localizes mainly to the nucleus. These studies will make important contribution in understanding the role of RuvB protein in P. falciparum.     

Unique monoclonal antibodies specifically bind surface structures on human fetal erythroid blood cells

Background

Continuing efforts in development of non-invasive prenatal genetic tests have focused on the isolation of fetal nucleated red blood cells (NRBCs) from maternal blood for decades. Because no fetal cell-specific antibody has been described so far, the present study focused on the development of monoclonal antibodies (mAbs) to antigens that are expressed exclusively on fetal NRBCs.Methods: Mice were immunized with fetal erythroid cell membranes and hybridomas screened for Abs using a multi-parameter fluorescence-activated cell sorting (FACS). Selected mAbs were evaluated by comparative FACS analysis involving Abs known to bind erythroid cell surface markers (CD71, CD36, CD34), antigen-i, galactose, or glycophorin-A (GPA). Specificity was further confirmed by extensive immunohistological and immunocytological analyses of NRBCs from umbilical cord blood and fetal and adult cells from liver, bone marrow, peripheral blood, and lymphoid tissues.Results: Screening of 690 hybridomas yielded three clones of which Abs from 4B8 and 4B9 clones demonstrated the desired specificity for a novel antigenic structure expressed on fetal erythroblast cell membranes. The antigenic structure identified is different from known surface markers (CD36, CD71, GPA, antigen-i, and galactose), and is not present on circulating adult erythroid cells, except for occasional detectability in adult bone marrow cells.Conclusions:The new mAbs specifically bind the same or highly overlapping epitopes of a surface antigen that is almost exclusively expressed on fetal erythroid cells. The high specificity of the mAbs should facilitate development of simple methods for reliable isolation of fetal NRBCs and their use in non-invasive prenatal diagnosis of fetal genetic status.     

Phenylacetaldehyde attracts male and female apple clearwing moths,Synanthedon myopaeformis,to inflorescences of showy milkweed,Asclepias speciosa

Synanthedon myopaeformis Borkhausen (Lepidoptera: Sesiidae) is a diurnal clearwing moth native to Eurasia that was recently introduced into British Columbia (BC) and Ontario, Canada, where it has become a serious pest in apple orchards. In BC, these moths commonly feed on nectar of inflorescences, particularly that of showy milkweed, Asclepias speciosa Torrey (Apocynaceae). We investigated the relative importance of visual and olfactory cues, and the key floral semiochemical(s) mediating attraction of S. myopaeformis to A. speciosa. In field experiments, inflorescences left exposed or enclosed in cheesecloth bags dyed green induced similar visitation rates by moths, indicating that olfactory cues are attractive. Among the >10 floral odourants that elicited responses from moth antennae in coupled gas chromatographic‐electroantennographic detection analyses, phenylacetaldehyde induced the most frequent proboscis extension reflexes of male and female moths. Among eight floral odourants that were field‐tested singly, phenylacetaldehyde attracted 35 times more male and female moths than any other candidate semiochemical. Attractiveness of phenylacetaldehyde could not be enhanced by admixture with other floral odourants at the ratios or concentrations tested indicating that it alone may mediate attraction of S. myopaeformis to the inflorescences of A. speciosa. The potential use of phenylacetaldehyde as bait to monitor or mass‐trap populations of male and female S. myopaeformis should be investigated.     

Late‐glacial recolonization and phylogeography of European red deer (Cervus elaphus L.)

The Pleistocene was an epoch of extreme climatic and environmental changes. How individual species responded to the repeated cycles of warm and cold stages is a major topic of debate. For the European fauna and flora, an expansion–contraction model has been suggested, whereby temperate species were restricted to southern refugia during glacial times and expanded northwards during interglacials, including the present interglacial (Holocene). Here, we test this model on the red deer (Cervus elaphus) a large and highly mobile herbivore, using both modern and ancient mitochondrial DNA from the entire European range of the species over the last c. 40 000 years. Our results indicate that this species was sensitive to the effects of climate change. Prior to the Last Glacial Maximum (LGM) haplogroups restricted today to South‐East Europe and Western Asia reached as far west as the UK. During the LGM, red deer was mainly restricted to southern refugia, in Iberia, the Balkans and possibly in Italy and South‐Western Asia. At the end of the LGM, red deer expanded from the Iberian refugium, to Central and Northern Europe, including the UK, Belgium, Scandinavia, Germany, Poland and Belarus. Ancient DNA data cannot rule out refugial survival of red deer in North‐West Europe through the LGM. Had such deer survived, though, they were replaced by deer migrating from Iberia at the end of the glacial. The Balkans served as a separate LGM refugium and were probably connected to Western Asia with genetic exchange between the two areas.     

The reproductive phenology of nine species of Rhodophyta in the subtidal region of the Isle of Man

Monthly samples of about 40 separate plants of each species were collected from 1 to 3 m below lowest astronomical tide on Port Erin breakwater, Isle of Man, Irish Sea. In three species growing on rock, Plocamium cartilagineum, Cryptopleura ramosa and Callophyllis laciniata, about 90% of the plants were fertile in late summer but less than 10% in spring although some fertile plants were always present. Delesseria sanguinea and Odonthalia dentata, also epilithic, had a winter sporing season, Odonthalia extending into late spring, and all plants were sterile in summer. Three species growing epiphytically, Palmaria palmata, Membranoptera alata and Phycodrys rubens, reproduced maximally in the first half of the year at the time when the stipes of the host species, Laminaria hyperborea, grow fastest. Only Palmaria had a sterile season, late summer. The encrusting Cruoria pellita showed little seasonality. The first three species, which reproduce mainly when the sea temperature is above average, are in the northern part of their geographical range. The remaining species (apart from Cruoria) reproduce mainly at low temperatures and are in the southern half of their ranges. Male plants appear to be in a minority in all species, presumably because they were manifest for a shorter period than carposporic plants. They appeared first after sterile periods and were absent as sporing declined. Plocamium and to a lesser extent Cryptopleura show an extremely high preponderance of tetrasporophytes in the population. This is attributed to perennation and some factor disallowing the survival of most of the tetraspores.     

Light is a crucial signal for zoosporogenesis and gametogenesis in some green microalgae

Patterns of reproduction were investigated in some microalgal species of Chlorophyceae (Botryosphaerella sudetica, Neochloris aquatica, Neochloris vigensis, Bracteacoccus minor). Under continuous light, the microalgae reproduced asexually producing autospores. However, appropriate manipulation of external conditions led to a change in the reproduction pattern towards production of zoospores or gametes. Production of zoospores and gametes was inhibited by light; motile cells emerged when microalgae were cultivated in darkness. The period of dark treatment necessary for zoosporogenesis or gametogenesis differed substantially among species that were tested. Sexual reproduction was observed in Neochloris vigensis and Bracteacoccus minor, whose generative life cycle had not been previously reported. The morphology of motile cells, the mode of sexual reproduction, and the efficiency of both the production of motile stages and mating events, were investigated. In order to gain detailed insights into patterns of reproduction, Botryosphaerella sudetica was selected for investigation under different light treatments. Non-actinic red light applied in the early phase of dark cultivation (up to 2 h) suppressed both zoosporogenesis and gametogenesis. However, after a 3-h dark pre-treatment, red light treatment had no effect on zoosporogenesis or gametogenesis. In contrast, non-actinic blue light did not block zoosporogenesis or gametogenesis, regardless of the time of treatment. The possible role of a red-light photoreceptor in zoosporogenesis and gametogenesis is discussed.     

Formation and Some Properties of the Acid Phosphatase in Aspergillus terreus

Substrate specificity of purified preparations of phytase from Asp, terreus was examined. The enzyme showed broad specificity. It was found that Asp, terreus produced only one kind of acid phosphatase and it had phytase activity.

Effective materials for the enzyme formation were examined. The formation of the enzyme occurred only during times that mycelia was in contact with inositol.

By differential centrifugation and electron-microscopic autoradiography, it was determined that inositol was incorporated into the mycelia and that it was located at almost the same point as where the active enzyme was located.     

Production of Lysine by Pyruvate Dehydrogenase Mutants of Brevibacterium flavum

Mutants with low pyruvate dehydrogenase (PD) activities were derived from a pyruvate kinase-deficient lysine-producing mutant of Brevibacterium flavum, No. 22. They were selected as prototrophic revertants of the acetate auxotrophs of strain No. 22. Among them strain KD-11 produced 55g/liter of lysine as its HCI salt when cultured for 72 hr in a medium containing lOOg/liter of glucose, soybean-meal hydrolysate and methionine. The lysine yield of strain KD-11 was the highest ever reported (55%). The mutant required a higher concentration of methionine for maximum production and gave a smaller amount of cell mass in cultivation than its parent. PD activity of strain No. 22 was stimulated by cysteine, stabilized by glycerol, and gave apparent Kms of 89, 22, 380, 83 μM for pyruvate, coenzyme A, 3-acetylpyridine adenine dinucleotide, and NAD, respectively, under standard conditions. The apparent Km for NAD of PD from strain KD-11 was 10-times higher than that from No. 22. When the concentration of NAD was low, the cell extracts of strain KD-11 showed low PD activity. The specific activity of phosphoenolpyruvate carboxylase of strain KD-11 was slightly higher than that of strain No. 22, while the inhibition by aspartate of the former enzyme was weaker than that of the latter.     

Effects of Korean Red Ginseng extract on hepatic lipid accumulation in HepG2 cells

In this study, we investigated the effects of Korean red ginseng water extract (KRGE) on hepatic lipid accumulation in HepG2 cells. KRGE decreased hepatic triglyceride and cholesterol levels. Further, KRGE suppressed expression of fatty acid synthase (FAS) and 3-hydroxy-3-methyl glutaryl coenzyme A (HMG-CoA) reductase. These results suggest that KRGE may reduce hepatic lipid accumulation by inhibition of FAS and HMG-CoA reductase expression in HepG2 cells.