Microsclerotial germination of Verticillium dahliae as affected by rape rhizosphere

Abstract The germination of nylon net-trapped microsclerotia of Verticillium dahliae pathogenic to rape ( Brassica napus ) was assessed in different systems by fluorescence microscopy using fluorescein diacetate. The influence of the culture's age and the size of the microsclerotia on germination percentages was assessed in water, mineral salts solution and mineral salts solution plus sucrose for 3 V. dahliae isolates. Large microsclerotia germinated better than smaller ones. The microsclerotia of 2 isolates showed decreased germination percentages with culture age over a 4–11-week period. Microsclerotial germination percentages were always higher in mineral salts solution plus sucrose than in mineral salts solution alone or water. In a sand culture system with the intact rape plant, microsclerotial germination percentages were high close to the root and decreased in a steep gradient to background levels within 5 mm from the root.     

Taste discrimination vs hedonic response to sucrose in coffee beverage. An interlaboratory study

Degree of liking, using a 17-point hedonic scale, and discriminationtaste thresholds, using a paired-comparison technique, weredetermined in a coffee beverage containing 0, 2.5, 5.0, 7.5,and 10.0% sucrose at laboratories in Brazil, Japan, Poland,Sweden and USA. Hedonic responses from the 122 subjects weresubdivided into four distinct sub-groups, according to differentpatterns as a function of sucrose concentration. Different frequenciesof these hedonic patterns resulted from the five laboratories.With few exceptions, repeated hedonic testing at the terminationof the experiment matched those from the beginning, indicatingstability of response during the lengthy study. No differenceswere observed in hedonic responses (nor in discrimination ability)between the male and female subjects at each laboratory. Discrimination thresholds at the five standard sucrose concentrations,and corresponding Weber ratios, were reported for the pooleddata within each laboratory. In general, the Weber ratios werehigher at the lower concentrations, indicating dependence ofdiscrimination upon the standard concentration. Notable differencesin discrimination ability were evident among the five laboratories,but were unrelated to degree of liking for sweetness in thecoffee. Subjects with low or with high degree of liking forall coffee samples, as well as those with increasing or decreasinghedonic responses as a function of sucrose concentration, discriminatedequally well among the concentration levels. The data from alllaboratories showed that ability to discriminate among sucroselevels and degree of liking for sucrose levels in coffee areindependent behavioral responses. ¹Present address: Department of Nutrition, Cornell University,Ithaca, New York, USA     

Effects of light and sound on parotid secretion and taste perception in response to sodium chloride

Salivary secretion in response to solutions of 1.6% NaCl wasslightly, but not significantly lower under red than under whiteillumination, and when wearing earphones which delivered intermittent"kitchen" noise, or the monotonous noise of a siren. Gustatoryresponses from the same subjects to a 15-sample concentrationseries of NaCl indicated that neither discrimination nor perceivedintensity of saltiness was significantly affected by the illuminationor by the kitchen noise. Fitting of a smooth, sigmoid line tothe group's discrimination data gave a value of 0.012% NaClfor the 75% correct level (50% above chance) and of 0.08% NaClfor 100% identification. The subjects showed no systematic relationshipamong salivary flow rate, discrimination, or perceived intensity. Contrary to previous findings, neither the light nor the noiseresulted in significantly lowered responses in 10 out of 12subjects in the salivation tests, nor in 45 out of 52 trials(13 subjects x 4 test conditions) in the taste tests. It appearsthat experienced subjects adapt well to systematic environmentaldistractions, which do not adversely affect their performanceof simple perceptual tasks. ¹ Based on research conducted while the senior author was avisiting professor at SIK in 1970–71.     

Increased vascular smooth muscle contractility in TRPC6-/- mice

Among the TRPC subfamily of TRP (classical transient receptor potential) channels, TRPC3, -6, and -7 are gated by signal transduction pathways that activate C-type phospholipases as well as by direct exposure to diacylglycerols. Since TRPC6 is highly expressed in pulmonary and vascular smooth muscle cells, it represents a likely molecular candidate for receptor-operated cation entry. To define the physiological role of TRPC6, we have developed a TRPC6-deficient mouse model. These mice showed an elevated blood pressure and enhanced agonist-induced contractility of isolated aortic rings as well as cerebral arteries. Smooth muscle cells of TRPC6-deficient mice have higher basal cation entry, increased TRPC-carried cation currents, and more depolarized membrane potentials. This higher basal cation entry, however, was completely abolished by the expression of a TRPC3-specific small interference RNA in primary TRPC6(-)(/)(-) smooth muscle cells. Along these lines, the expression of TRPC3 in wild-type cells resulted in increased basal activity, while TRPC6 expression in TRPC6(-/-) smooth muscle cells reduced basal cation influx. These findings imply that constitutively active TRPC3-type channels, which are up-regulated in TRPC6-deficient smooth muscle cells, are not able to functionally replace TRPC6. Thus, TRPC6 has distinct nonredundant roles in the control of vascular smooth muscle tone.     

A fast and flexible PEG-mediated transient expression system in plants for high level expression of secreted recombinant proteins

Plant expression systems offer a valuable alternative to traditional systems for the production of recombinant biopharmaceuticals. A highly efficient polyethyleneglycol (PEG)-mediated transient expression system for secreted recombinant proteins in plants has been developed. The human vascular endothelial growth factor 121 (rhVEGF) has been successfully expressed and efficiently secreted into the culture medium by transiently transformed moss protoplasts. In order to obtain secretion efficiency data, different expressed signal peptides were analysed and time course studies were performed with expression constructs containing different promoters. The transformation procedure was optimised for high level expression (up to 10 microg/ml) and successfully performed even with a transgenic glyco-engineered strain lacking plant-specific immunogenic sugar residues in N-glycans. The amount of rhVEGF was produced in such quantity that it allowed for the analysis of biological activity, silver-staining and Western blotting, revealing the correct formation and processing of the homodimer. This fast and flexible transient expression system enables feasibility studies and construct optimisation to be concluded within a few days, thus avoiding the time consuming step of having to generate stably transformed lines.     

Poly(3-hydroxyalkanoate) polymerase synthesis and in vitro activity in recombinant Escherichia coli and Pseudomonas putida

We tested the synthesis and in vitro activity of the poly(3-hydroxyalkanoate) (PHA) polymerase 1 from Pseudomonas putida GPo1 in both P. putida GPp104 and Escherichia coli JMU193. The polymerase encoding gene phaC1 was expressed using the inducible PalkB promoter. It was found that the production of polymerase could be modulated over a wide range of protein levels by varying inducer concentrations. The optimal inducer dicyclopropylketone concentrations for PHA production were at 0.03% (v/v) for P. putida and 0.005% (v/v) for E. coli. Under these concentrations the maximal polymerase level synthesized in the E. coli host (6% of total protein) was about three- to fourfold less than that in P. putida (20%), whereas the maximal level of PHA synthesized in the E. coli host (8% of total cell dry weight) was about fourfold less than that in P. putida (30%). In P. putida, the highest specific activity of polymerase was found in the mid-exponential growth phase with a maximum of 40 U/g polymerase, whereas in E. coli, the maximal specific polymerase activity was found in the early stationary growth phase (2 U/g polymerase). Our results suggest that optimal functioning of the PHA polymerase requires factors or a molecular environment that is available in P. putida but not in E. coli.     

Variation in structural location and amino acid conservation of functional sites in protein domain families

Background  

The functional sites of a protein present important information for determining its cellular function and are fundamental in drug design. Accordingly, accurate methods for the prediction of functional sites are of immense value. Most available methods are based on a set of homologous sequences and structural or evolutionary information, and assume that functional sites are more conserved than the average. In the analysis presented here, we have investigated the conservation of location and type of amino acids at functional sites, and compared the behaviour of functional sites between different protein domains.