Different cell-wall components from Phytophthora megasperma f. sp. glycinea elicit phytoalexin production in soybean and parsley

Different components of a crude cell-wall preparation from the phytopathogenic fungus, Phytophthora megasperma f. sp. glycinea, act as elicitors of phytoalexin accumulation in parsley (Petroselinum crispum) and soybean (Glycine max). Treatments of cultured parsley cells and protoplasts or soybean cells and cotyledons with proteinase-digested or deglycosylated elicitor preparations identify proteinaceous constituents as active eliciting compounds in parsley, which are inactive in soybean. The proteinase-treated elicitor as well as a defined heptaglucan are active in soybean but do not stimulate phytoalexin synthesis in parsley. Soybean and parsley cells therefore not only perceive different signals from cell walls of Phytophthora megasperma f. sp. glycinea, but are unable to respond to the fungal compounds primarily recognized by the other plant.Abbreviations Pmg Phytophthora megasperma f. sp. glycinea     

Impaired ion channel function related to a common KCNQ1 mutation — Implications for risk stratification in long QT syndrome 1

Long QT syndrome (LQTS) 1 is the most common type of inherited LQTS and is linked to mutations in the KCNQ1 gene. We identified a KCNQ1 missense mutation, KCNQ1 G325R, in an asymptomatic patient presenting with significant QT prolongation (QTc, 448–600 ms). Prior clinical reports revealed phenotypic variability ranging from the absence of symptoms to syncope among KCNQ1 G325R mutation carriers. The present study was designed to determine the G325R ion channel phenotype and its association with the clinical LQTS presentation. Electrophysiological testing was performed using the Xenopus oocyte expression system. KCNQ1 G325R channels were non-functional and suppressed wild type (WT) currents by 71.1%. In the presence of the native cardiac regulatory ß-subunit, KCNE1, currents conducted by G325R and WT KCNQ1 were reduced by 52.9%. Co-expression of G325R and WT KCNQ1 with KCNE1 shifted the voltage-dependence of I_Ks activation by 12.0 mV, indicating co-assembly of mutant and WT subunits. The dysfunctional biophysical phenotype validates the pathogenicity of the KCNQ1 G325R mutation and corresponds well with the severe clinical presentation revealed in some reports. However, the index patient and other mutation carriers were asymptomatic, highlighting potential limitations of risk assessment schemes based on ion channel data.     

Long-term therapy using horse anti-dog lymphocyte globulin without sensitization against horse protein]

Eight mongrel dogs received a standard daily i.v. infusion of 20 mg/kg b.w. deaggregated horse-anti-dog-lymphocyte-globulin (ALG) and additional prednisolone (1 mg/kg b.w. daily i.v.) over a maximum period of 82 days following pretreatment with deaggregated normal horse IgG. No sensitization against horse protein was observed during therapy of afterwards as proved by lack of humoral antibodies against horse antigens, maintained lymphopenia, good compatibility, longterm prolongation of xenogeneic skin graft survival (85.6+/-20.6 days, n=8' untreated controls 12.5+/-1.3 days, n=4) and longterm suppression of cytotoxic antibodies against donor lymphocytes. The level of preformed agglutinating antibodies against horse erythrocytes was significantly reduced, while preformed antibodies against other species remained normal. The immune response to a challenge injection of anti-lymphocyte-serum (ALS) 6-11 weeks after termination of treatment was significantly lower in the ALG treated animals as compared to the control group. These results suggest the involvement of a specific mechanism of unresponsiveness against ALG other than immunosuppression only. It is concluded, that by the described method sensitization against ALG can be prevented during longterm treatment.     

The Multifunctional Enzyme CYP71B15 (PHYTOALEXIN DEFICIENT3) Converts Cysteine-Indole-3-Acetonitrile to Camalexin in the Indole-3-Acetonitrile Metabolic Network of Arabidopsis thaliana

Accumulation of camalexin, the characteristic phytoalexin of Arabidopsis thaliana, is induced by a great variety of plant pathogens. It is derived from Trp, which is converted to indole-3-acetonitrile (IAN) by successive action of the cytochrome P450 enzymes CYP79B2/B3 and CYP71A13. Extracts from wild-type plants and camalexin biosynthetic mutants, treated with silver nitrate or inoculated with Phytophthora infestans, were comprehensively analyzed by ultra-performance liquid chromatography electrospray ionization quadrupole time-of-flight mass spectrometry. This metabolomics approach was combined with precursor feeding experiments to characterize the IAN metabolic network and to identify novel biosynthetic intermediates and metabolites of camalexin. Indole-3-carbaldehyde and indole-3-carboxylic acid derivatives were shown to originate from IAN. IAN conjugates with glutathione, γ-glutamylcysteine, and cysteine [Cys(IAN)] accumulated in challenged phytoalexin deficient3 (pad3) mutants. Cys(IAN) rescued the camalexin-deficient phenotype of cyp79b2 cyp79b3 and was itself converted to dihydrocamalexic acid (DHCA), the known substrate of CYP71B15 (PAD3), by microsomes isolated from silver nitrate–treated Arabidopsis leaves. Surprisingly, yeast-expressed CYP71B15 also catalyzed thiazoline ring closure, DHCA formation, and cyanide release with Cys(IAN) as substrate. In conclusion, in the camalexin biosynthetic pathway, IAN is derivatized to the intermediate Cys(IAN), which serves as substrate of the multifunctional cytochrome P450 enzyme CYP71B15.     

Characterization of the ribonuclease activity on the skin surface

The rapid degradation of ribonucleic acids (RNA) by ubiquitous ribonucleases limits the efficacy of new therapies based on RNA molecules. Therefore, our aim was to characterize the natural ribonuclease activities on the skin and in blood plasma i.e. at sites where many drugs in development are applied. On the skin surfaces of Homo sapiens and Mus musculus we observed dominant pyrimidine-specific ribonuclease activity. This activity is not prevented by a cap structure at the 5'-end of messenger RNA (mRNA) and is not primarily of a 5'- or 3'-exonuclease type. Moreover, the ribonuclease activity on the skin or in blood plasma is not inhibited by chemical modifications introduced at the 2'OH group of cytidine or uridine residues. It is, however, inhibited by the ribonuclease inhibitor RNasin^® although not by the ribonuclease inhibitor SUPERase· In?. The application of our findings in the field of medical science may result in an improved efficiency of RNA-based therapies that are currently in development.