Arg-Gly-Asp (RGD) sequence conjugated in a synthetic copolymer bearing a sugar moiety for improved culture of parenchymal cells (hepatocytes)

A copolymer, including a Gly-Arg-Gly-Asp-Ser (GRGDS) sequence and sugar moieties, was synthesized for the culturing of parenchymal cells (hepatocytes). Hepatocyte cells attached to poly[N-p-vinylbenzyl-d-maltonamide-co-6-(p-vinylbenzamido)-hexanoic acid-GRGDS] [poly(VMA-co-VBRGD)]-coated dishes grew approximately 60% better than on other polymer-coated surface for 12 h. Also, about 80% greater albumin secretion (0.38 pg ml^–1) and about 70% greater urea synthesis (0.495 pg ml^–1) from hepatocytes were produced in this matrix as compared with unstimulated cells. The behaviour of hepatocytes on poly(VMA-co-VBGRGDS)-coated dishes was not distinct from those attached to a collagen. The conjugation of the adhesion molecules of the RGD peptide in the poly(VMA-co-VBGRGDS) copolymer therefore specifically interacts with integrin families on the hepatocyte cell membrane.     

Two oligosaccharides from Marsdenia roylei

Phytochemical analysis of dried twigs of Marsdenia roylei (family Asclepiadaceae) has resulted in the isolation of a trisaccharide, maryal, and a diglycoside, rolinose. Their structures were determined as O-beta-D-oleandropyranosyl-(1-->4)-O-beta-D-digitoxopyranosyl++ +-(1-->4)-D- cymaral and ethyl O-beta-D-oleandropyranosyl-(1-->4)-O-3-O-methyl-6-deoxy-beta-D- allopyranoside, respectively, by chemical degradation and spectroscopic methods.     

Sugar‐feeding status alters biting midge photoattraction

The biting midge Culicoides sonorensis Wirth and Jones (Diptera: Ceratopogonidae) transmits pathogens to both livestock and wildlife. Biting midge surveillance relies heavily on light traps for collection; however, little is known about the light spectra preferences of C. sonorensis midges. A light assay arena was constructed and light‐emitting diodes (LEDs) of various light spectra were used as light sources to evaluate midge photoattraction. A comparison of responses to light spectra indicated the highest proportions of C. sonorensis were attracted to ultraviolet (UV) light and that midges differentiated 10‐nm differences in wavelength. Stronger intensities of UV light resulted in greater attraction. Midges exhibited both sugar‐seeking and escape behaviours under different conditions of sugar supplementation before and during the experiment. These behaviours occurred with lights of 355 nm and 365 nm in wavelength. Based on the results of this study, the attraction of C. sonorensis to light traps can be improved through the use of bright LEDs at 355 nm or 365 nm.     

Significant impact of divalent metal ions on the fidelity,sugar selectivity,and drug incorporation efficiency of human PrimPol

Human PrimPol is a recently discovered bifunctional enzyme that displays DNA template-directed primase and polymerase activities. PrimPol has been implicated in nuclear and mitochondrial DNA replication fork progression and restart as well as DNA lesion bypass. Published evidence suggests that PrimPol is a Mn²⁺-dependent enzyme as it shows significantly improved primase and polymerase activities when binding Mn²⁺, rather than Mg²⁺, as a divalent metal ion cofactor. Consistently, our fluorescence anisotropy assays determined that PrimPol binds to a primer/template DNA substrate with affinities of 29 and 979 nM in the presence of Mn²⁺ and Mg²⁺, respectively. Our pre-steady-state kinetic analysis revealed that PrimPol incorporates correct dNTPs with 100-fold higher efficiency with Mn²⁺ than with Mg²⁺. Notably, the substitution fidelity of PrimPol in the presence of Mn²⁺ was determined to be in the range of 3.4 × 10⁻² to 3.8 × 10⁻¹, indicating that PrimPol is an error-prone polymerase. Furthermore, we kinetically determined the sugar selectivity of PrimPol to be 57–1800 with Mn²⁺ and 150–4500 with Mg²⁺, and found that PrimPol was able to incorporate the triphosphates of two anticancer drugs (cytarabine and gemcitabine), but not two antiviral drugs (emtricitabine and lamivudine).     

Relationship between the concentration of sugars in the roots and VA mycorrhizal infection

Summary The VA-infected wheat varieties showed an increase of total (Lozano var.) and reducing (Lozano and Pane vars.) sugars in their root extracts. However, no clear relationship between sugar concentration in the root and VA mycorrhizal infection level could be established.In addition, the VA mycorrhizal hosts sorghum, alfalfa, sunflower and maize, and non-host radish and cabbage plants were tested for sugar content in their root extracts after fifteen days of growth. Sugars present in the root extracts of these plants did not seem to be a decisive factor in plant susceptibility to VA infection.     

Lysine Succinylation of VBS Contributes to Sclerotia Development and Aflatoxin Biosynthesis in Aspergillus flavus

Aspergillus flavus is a common saprophytic and pathogenic fungus, and its secondary metabolic pathways are one of the most highly characterized owing to its aflatoxin (AF) metabolite affecting global economic crops and human health. Different natural environments can cause significant variations in AF synthesis. Succinylation was recently identified as one of the most critical regulatory post-translational modifications affecting metabolic pathways. It is primarily reported in human cells and bacteria with few studies on fungi. Proteomic quantification of lysine succinylation (Ksuc) exploring its potential involvement in secondary metabolism regulation (including AF production) has not been performed under natural conditions in A. flavus. In this study, a quantification method was performed based on tandem mass tag labeling and antibody-based affinity enrichment of succinylated peptides via high accuracy nano-liquid chromatography with tandem mass spectrometry to explore the succinylation mechanism affecting the pathogenicity of naturally isolated A. flavus strains with varying toxin production. Altogether, 1240 Ksuc sites in 768 proteins were identified with 1103 sites in 685 proteins quantified. Comparing succinylated protein levels between high and low AF-producing A. flavus strains, bioinformatics analysis indicated that most succinylated proteins located in the AF biosynthetic pathway were downregulated, which directly affected AF synthesis. Versicolorin B synthase is a key catalytic enzyme for heterochrome B synthesis during AF synthesis. Site-directed mutagenesis and biochemical studies revealed that versicolorin B synthase succinylation is an important regulatory mechanism affecting sclerotia development and AF biosynthesis in A. flavus. In summary, our quantitative study of the lysine succinylome in high/low AF-producing strains revealed the role of Ksuc in regulating AF biosynthesis. We revealed novel insights into the metabolism of AF biosynthesis using naturally isolated A. flavus strains and identified a rich source of metabolism-related enzymes regulated by succinylation.     

6-Shogaol induces apoptosis in acute lymphoblastic leukaemia cells by targeting p53 signalling pathway and generation of reactive oxygen species

Combination therapies, using medicinal herbs, are broadly recommended to attenuate the chemotherapy adverse effects. Based on our previous findings considering the anti-leukaemic effects of ginger extract on acute lymphoblastic leukaemia (ALL) cells, the present study was aimed to investigate the anti-cancer role of this pharmaceutical plant on ALL mice models. Moreover, we worked towards identifying the most anti-leukaemic derivative of ginger and the mechanism through which it may exert its cytotoxic impact. In vivo experiments were performed using five groups of six C57BL/6 nude mice, and the anti-leukaemic activity of ginger extract alone or in combination with methotrexate (MTX) was examined. Results showed increased survival rate and reduced damages in mice brain and liver tissues. Subsequently, MTT assay demonstrated synergistic growth inhibitory effect of 6-shogaol (6Sh) and MTX on ALL cell lines and patients primary cells. Eventually, the molecular anti-neoplastic mechanism of 6Sh was evaluated using Bioinformatics. Flow cytometry illustrated 6Sh-mediated apoptosis in Nalm-6 cells confirmed by Western blotting and RT-PCR assays. Further analyses exhibited the generation of reactive oxygen species (ROS) through 6Sh. The current study revealed the in vivo novel anti-leukaemic role of ginger extract, promoted by MTX. Moreover, 6-shogaol was introduced as the major player of ginger cytotoxicity through inducing p53 activity and ROS generation.     

In vivo control of Mycosphaerella pinodes on pea leaves by a crude bulb extract of Eucomis autumnalis

We previously reported on the in vitro antifungal activity of a crude whole plant extract from Eucomis autumnalis against seven economically important plant pathogenic fungi. A crude extract of the bulb showed similar in vitro mycelial growth inhibition of the same plant pathogenic fungi as well as that of an eighth fungus, Mycosphaerella pinodes, the cause of black spot or Ascochyta blight, in peas. Subsequently, fourth internode leaves were removed from 4 wk old pea plants, placed on moist filter paper in Petri dishes and inoculated with an M. pinodes spore suspension before and after treatment with the extract. The control of Ascochyta blight by different concentrations of the crude E. autumnalis extract was followed in vivo by leaf symptoms over a 6 day period at 20°C in a growth cabinet. The crude extract prevented M. pinodes spore infection of the leaves when the leaves were inoculated with spores both before or after treatment with the extract, confirming complete inhibition of spore germination. The crude E. autumnalis extract showed no phytotoxic reaction on the leaves even at the highest concentration applied.     

Transport of sugars across the plasma membrane of beetroot protoplasts

Protoplasts isolated from beetroot tissue took up glucose preferentially whereas sucrose was transported more slowly. The ¹⁴C-label from [¹⁴C]glucose and [¹⁴C]sucrose taken up by the cells could be detected rapidly in phosphate esters and, after feeding of [¹⁴C]glucose was found also in sucrose. The temperature-dependent uptake process (activation energy E_A about 50 kJ · mol^–1) seems to be carrier mediated as indicated by its substrate saturation and, for glucose, by competition experiments which revealed positions C₁, C₅ and C₆ of the D-glucose molecule as important for effective uptake. The apparent K_m(20° C) for glucose (3-O-methylglucose) was about 1 mM whereas for sucrose a significantly lower apparent affinity was determined (K_m about 10 mM). When higher concentrations of glucose (5 mM) or sucrose (20 mM) were administered, the uptake process followed first-order kinetics. Carrier-mediated transport was inhibited by N,N-dicyclohexylcarbodiimide, Na-orthovanadate, p–chloromercuribenzenesulfonic acid, and by uncouplers and ionophores. The uptake system exhibited a distinct pH optimum at pH 5.0. The results indicate that generation of a proton gradient is a prerequisite for sugar uptake across the plasma membrane. Protoplasts from the bundle regions in the hypocotyl take up glucose at higher rates than those derived from bundle-free regions. The results favour the idea that apoplastic transport of assimilates en route of unloading might be restricted to distinct areas within the storage organ (i.e. the bundle region) whereas distribution in the storage parenchyma is symplastic.Abbreviations CCCP Carbonylcyanide m–chlorophenylhydrazone - DCCD N,N-dicyclohexylcarbodiimide - DOG deoxyglucose - Mes 2-(N-morpholino)ethanesulfonic acid - 3-OMG 3-O-methylglucose - PCMBS p–chloromercuribenzenesulfonic acid - SDS Sodium dodecyl sulfate - Tris 2-amino-2-(hydroxymethyl)-1,3-propanediol