Levels of endogenous abscisic acid and indole-3-acetic acid influence shoot organogenesis in callus cultures of rice subjected to osmotic stress

Osmotic stress and endogenous hormone levels may have a role in shoot organogenesis, but a systematic study has not yet to investigate the links. We evaluated the changes of the endogenous indole-3-acetic acid (IAA) and abscisic acid (ABA) levels in rice (Oryza sativa L. cv. Tainan 5) callus during shoot organogenesis induced by exogenous plant growth regulator treatments or under osmotic stress. Non-regenerable callus showed low levels of endogenous ABA and IAA, with no fluctuation in level during the period evaluated. The addition of 100 μM ABA or 2 mM anthranilic acid (IAA precursor) into Murashige and Skoog basal induction medium containing 10 μM 2,4-D enhanced the regeneration frequency slightly, to 5 and 35%, respectively, and their total cellular ABA or IAA levels were increased significantly, correspondingly to the treatments. However, the regeneration frequency was greatly increased to 80% after treatment with 0.6 M sorbitol or 100 μM ABA and 2 mM anthranilic acid combined. Both treatments produced high levels of total cellular ABA and IAA at the callus stage, which was quickly decreased on the first day after transfer to regeneration medium. Thus, osmotic stress-induced simultaneous accumulation of endogenous ABA and IAA is involved in shoot regeneration in rice callus.     

Enzymatic biotransformation of ginsenoside Rb1 to 20(<Emphasis Type="Italic">S</Emphasis>)-Rg3 by recombinant β-glucosidase from <Emphasis Type="Italic">Microbacterium esteraromaticum</Emphasis>

Microbacterium esteraromaticum was isolated from ginseng field. The β-glucosidase gene (bgp1) from M. esteraromaticum was cloned and expressed in Escherichia coli BL21 (DE3). The bgp1 gene consists of 2,496 bp encoding 831 amino acids which have homology to the glycosyl hydrolase family 3 protein domain. The recombinant β-glucosidase enzyme (Bgp1) was purified and characterized. The molecular mass of purified Bgp1 was 87.5 kDa, as determined by SDS-PAGE. Using 0.1 mg ml⁻¹ enzyme in 20 mM sodium phosphate buffer at 37°C and pH 7.0, 1.0 mg ml⁻¹ ginsenoside Rb1 was transformed into 0.444 mg ml⁻¹ ginsenoside Rg3 within 6 h. The Bgp1 sequentially hydrolyzed the outer and inner glucose attached to the C-20 position of ginsenosides Rb1. Bgp1 hydrolyzed the ginsenoside Rb1 along the following pathway: Rb1 → Rd → 20(S)-Rg3. This is the first report of the biotransformation of ginsenoside Rb1 to ginsenoside 20(S)-Rg3 using the recombinant β-glucosidase.     

Dynamic 1H NMR-based extracellular metabonomic analysis of oligodendroglia cells infected with herpes simplex virus type 1

Herpes simplex virus type 1 (HSV-1) is a large, neurotropic, double-stranded DNA virus that establishes a lifelong latent infection in neurons and glial cells. Previous studies reveal that several metabolic perturbations are associated with HSV-1 infection. However, the extracellular metabolic alterations associated with HSV-1 infection have not been systematically profiled in human cells. Here, a proton nuclear magnetic resonance-based metabonomic approach was applied to differentiate the extracellular metabonomic profiles of HSV-1 infected human oligodendroglia cells (n = 18) and matched control cells (n = 18) at three time points (12, 24, and 36 h post-infection). Resulting spectra were analyzed by chemometric and statistical methods. Metabonomic profiling revealed perturbations in 21 extracellular metabolites. Partial least squares discriminant analysis demonstrated that the whole metabolic patterns enabled statistical discrimination between HSV-1 infected human oligodendroglia cells and control cells. Eight extracellular metabolites, seven of which were amino acids, were primarily responsible for score plot discrimination between HSV-1 infected human oligodendroglia cells and control cells at 36 h post-infection: alanine, glycine, isoleucine, leucine, glutamate, glutamine, histidine, and lactate. HSV-1 infection alters amino acid metabolism in human oligodendroglia cells cultured in vitro. HSV-1 infection may disturb these host cellular pathways to support viral replication. Through elucidating the extracellular metabolic changes incident to HSV-1 infection, this study also provides future directions for investigation into the pathogenic mechanism of HSV-1.     

Applying Imaging Genetics to ADHD: the Promises and the Challenges

Attention deficit/hyperactivity disorder (ADHD) is one of the most prevalent childhood psychiatric disorders. During the past decades, studies have focused on its genetic background and the abnormality of the brain structure and function. Recently, an advanced technique linking these two areas, namely, the imaging genetics emerged and kept growing. Imaging genetics primarily identifies genes that influence the brain variations. There are three main strategies of doing an imaging genetic study: using neuroimaging as endophenotypes to find the associated DNA variants, finding neuroimaging effects of risk genes, and hypothesis-free whole-brain voxelwise genome-wide association study. In this review, we begin with demonstrating the basic principles of imaging genetics precisely, with examples from other psychiatric conditions, and then go on to synthesize the existing imaging genetic studies in ADHD. Finally, we elaborate the challenges of applying imaging genetics to ADHD. We conclude that imaging genetics has somewhat showed its potential to provide a more precise understanding of how the genes shape the brain variations and further the clinical features of ADHD.     

A facile method for controlling the reaction equilibrium of sphingolipid ceramide N-deacylase for lyso-glycosphingolipid production

Lyso-glycosphingolipids (lyso-GSLs), the N-deacylated forms of glycosphingolipids (GSLs), are important synthetic intermediates for the preparation of GSL analogs. Although lyso-GSLs can be produced by hydrolyzing natural GSLs using sphingolipid ceramide N-deacylase (SCDase), the yield for this reaction is usually low because SCDase also catalyzes the reverse reaction, ultimately establishing an equilibrium between hydrolysis and synthesis. In the present study, we developed an efficient method for controlling the reaction equilibrium by introducing divalent metal cation and detergent in the enzymatic reaction system. In the presence of both Ca²⁺ and taurodeoxycholate hydrate, the generated fatty acids were precipitated by the formation of insoluble stearate salts and pushing the reaction equilibrium toward hydrolysis. The yield of GM1 hydrolysis can be achieved as high as 96%, with an improvement up to 45% compared with the nonoptimized condition. In preparative scale, 75 mg of lyso-GM1 was obtained from 100 mg of GM1 with a 90% yield, which is the highest reported yield to date. The method can also be used for the efficient hydrolysis of a variety of GSLs and sphingomyelin. Thus, this method should serve as a facile, easily scalable, and general tool for lyso-GSL production to facilitate further GSL research.     

Structural organization of an intact phycobilisome and its association with photosystem II

Phycobilisomes (PBSs) are light-harvesting antennae that transfer energy to photosynthetic reaction centers in cyanobacteria and red algae. PBSs are supermolecular complexes composed of phycobiliproteins (PBPs) that bear chromophores for energy absorption and linker proteins. Although the structures of some individual components have been determined using crystallography, the three-dimensional structure of an entire PBS complex, which is critical for understanding the energy transfer mechanism, remains unknown. Here, we report the structures of an intact PBS and a PBS in complex with photosystem II (PSII) from Anabaena sp. strain PCC 7120 using single-particle electron microscopy in combination with biochemical and molecular analyses. In the PBS structure, all PBP trimers and the conserved linker protein domains were unambiguously located, and the global distribution of all chromophores was determined. We provide evidence that ApcE and ApcF are critical for the formation of a protrusion at the bottom of PBS, which plays an important role in mediating PBS interaction with PSII. Our results provide insights into the molecular architecture of an intact PBS at different assembly levels and provide the basis for understanding how the light energy absorbed by PBS is transferred to PSII.