Optimal analytical conditions for catalase in fresh water prawn,Macrobrachium malcolmsonii

The cytosol of hepatopancreas was prepared from the freshwater prawn Macrobrachium malcolmsonii, and optimal assay conditions, i.e., concentration of substrate, pH, and temperature, were determined to measure basal activities and kinetic constants of catalase activity. The properties of catalase were examined in M. macolmsonii, because quantitative data on catalase are limited in crustacean species. The optimal pH for catalase was 7.0. The activation energy was 3.55 Kcal/mol and energy inhibition value was 5.16 Kcal/mol. The value of energy inhibition is higher than that of energy activation. This may be due to inhibition of catalase by some substrate other than H2O2. A Km of 66.6?mM was also determined from various concentrations of substrate.     

Vitexin prevents Aβ proteotoxicity in transgenic Caenorhabditis elegans model of Alzheimer's disease by modulating unfolded protein response

Alzheimer's disease (AD) accounts for an estimated 60% to 80% of all dementia cases. The present study is aimed at evaluating the neuroprotective efficacy of vitexin, an apigenin flavone glycoside using transgenic Caenorhabditis elegans strain (CL2006) of AD. The neuroprotective effect of vitexin was determined using physiological assays, quantitative polymerase chain reaction, and Western blotting. The results of survival and paralysis assay indicate that vitexin (200 μM) significantly extended the lifespan of the nematodes. Vitexin‐treated nematodes showed a significant reduction in the expression of Aβ, ace‐1, and ace‐2 genes when compared to control. Further, vitexin significantly upregulated the expression of acr‐8 and dnj‐14, and increased the lifespan of the nematodes. Vitexin was also found to modulate the unfolded protein response genes (hsp‐4, pek‐1, ire‐1, and xbp‐1) and suppress the expression of Aβ. Overall, the results show that vitexin acts as a neuroprotective agent and protects transgenic C. elegans strains from Aβ proteotoxicity.     

Live cell imaging reveals structural associations between the actin and microtubule cytoskeleton in Arabidopsis

In eukaryotic cells, the actin and microtubule (MT) cytoskeletal networks are dynamic structures that organize intracellular processes and facilitate their rapid reorganization. In plant cells, actin filaments (AFs) and MTs are essential for cell growth and morphogenesis. However, dynamic interactions between these two essential components in live cells have not been explored. Here, we use spinning-disc confocal microscopy to dissect interaction and cooperation between cortical AFs and MTs in Arabidopsis thaliana, utilizing fluorescent reporter constructs for both components. Quantitative analyses revealed altered AF dynamics associated with the positions and orientations of cortical MTs. Reorganization and reassembly of the AF array was dependent on the MTs following drug-induced depolymerization, whereby short AFs initially appeared colocalized with MTs, and displayed motility along MTs. We also observed that light-induced reorganization of MTs occurred in concert with changes in AF behavior. Our results indicate dynamic interaction between the cortical actin and MT cytoskeletons in interphase plant cells.     

HIV-1 Gag Cytotoxic T Lymphocyte Epitopes Vary in Presentation Kinetics Relative to HLA Class I Downregulation

Although CD8⁺ cytotoxic T lymphocytes (CTLs) are protective in HIV-1 infection, the factors determining their antiviral efficiency are poorly defined. It is proposed that Gag targeting is superior because of very early Gag epitope presentation, allowing early killing of infected cells before Nef-mediated downregulation of human leukocyte antigen class I (HLA-I). To study Gag epitope presentation kinetics, three epitopes (SL9_77-85, KF11_162-172, and TW10_240-249) were genetically translocated from their endogenous location in the Rev-dependent (late) gag gene into the Rev-independent (early) nef gene with concomitant mutation of the corresponding endogenous epitopes to nonrecognized sequences. These viruses were compared to the index virus for CTL-mediated suppression of replication and the susceptibility of this antiviral activity to Nef-mediated HLA-I downregulation. SL9-specific CTLs gained activity after SL9 translocation to Nef, going from Nef sensitive to Nef insensitive, indicating that translocation accelerated infected cell recognition from after to before HLA-I downregulation. KF11-specific CTL antiviral activity was unchanged and insensitive to HLA-I downregulation before and after KF11 translocation, suggesting that already rapid recognition of infected cells was not accelerated. However, TW10-specific CTLs that were insensitive to Nef at the baseline became sensitive with reduced antiviral activity after translocation, indicating that translocation retarded epitope expression. Cytosolic peptide processing assays suggested that TW10 was inefficiently generated after translocation to Nef, compared to SL9 and KF11. As a whole, these data demonstrate that epitope presentation kinetics play an important role in CTL antiviral efficiency, that Gag epitopes are not uniformly presented early, and that the epitope context can play a major role in presentation kinetics.     

Dense fouling in acid transfer pipelines by an acidophilic rubber degrading fungus

An unique case of dense fouling by an acidophilic, hard rubber (polymerized rubber) degrading fungus in the acid transfer pipelines of a boron enrichment plant located at Kalpakkam, India is reported. In spite of a highly adverse environment for survival (pH 1.5, no dissolved nutrients), the fungus thrived and clogged the pipeline used for transferring 0.1N hydrochloric acid (HCl). Detailed investigations were carried out to isolate and identify the fungus and examine the nutrient source for such profuse growth inside the system. Microscopic observation showed the presence of a thick filamentous fungal biomass. Molecular characterization by 18S rRNA gene sequencing showed 98% similarity of the isolate with the acidophilic fungus Bispora sp. In laboratory studies the fungus showed luxuriant growth (specific growth rate of 13 mg day?1) when scrapings of the hard rubber were used as the sole source of carbon. Scanning electron microscopy revealed extensive incursion of the fungus into the hard rubber matrix. In the laboratory, fungal growth was completely inhibited by the antifungal agent sodium omadine. The study illustrates an interesting example of biofouling under extreme conditions and demonstrates that organisms can physiologically adapt to grow under unfavourable conditions, provided that a nutrient source is available and competition is low. The use of this fungal strain in biodegradation and in development of environmentally compatible processes for disposal of rubber wastes is envisaged.     

Potentiation of concurrent expression of lipogenic genes by novel strong promoters in the oleaginous microalga Phaeodactylum tricornutum

There has been growing interest in using microalgae as production hosts for a wide range of value-added compounds. However, microalgal genetic improvement is impeded by lack of genetic tools to concurrently control multiple genes. Here, we identified two novel strong promoters, designated Pt202 and Pt667, and delineated their potential role on simultaneously driving the expression of key lipogenic genes in Phaeodactylum tricornutum. In silico analyses of the identified promoter sequences predicted the presence of essential core cis elements such as TATA and CAAT boxes. Regulatory role of the promoters was preliminarily assessed by using GUS reporter which demonstrated strong GUS expression. Thereafter, two key lipogenic genes including malic enzyme (PtME) and 5-desaturase (PtD5b), were overexpressed by the two promoters Pt202 and Pt667, respectively, in P. tricornutum. Combinatorial gene overexpression did not impair general physiological performance, meanwhile neutral lipid content was remarkably increased by 2.4-fold. GC-MS analysis of fatty acid methyl esters revealed that eicosapentaenoic acid (EPA; C20:5) was increased significantly. The findings augment a crucial kit to microalgal genetic tools that could facilitate the multiple-gene expression driven by various promoters, and promote microalgae for industrial bioproduction.