Hairy roots of Hypericum perforatum L.: a promising system for xanthone production

Hypericum perforatum L. is a common perennial plant with a reputed medicinal value. Investigations have been made to develop an efficient protocol for the identification and quantification of secondary metabolites in hairy roots (HR) of Hypericum perforatum L. HR were induced from root segments of in vitro grown seedlings from H. perforatum, after co-cultivation with Agrobacterium rhizogenes A4. Transgenic status of HR was confirmed by PCR analysis using rolB specific primers. HR had an altered phenolic profile with respect to phenolic acids, flavonol glycosides, flavan-3-ols, flavonoid aglycones and xanthones comparing to control roots. Phenolics in control and HR cultures were observed to be qualitatively and quantitatively distinct. Quinic acid was the only detectable phenolic acid in HR. Transgenic roots are capable of producing flavonol glycosides such as quercetin 6-C-glucoside, quercetin 3-O-rutinoside (rutin) and isorhamnetin O-hexoside. The HPLC analysis of flavonoid aglycones in HR resulted in the identification of kaempferol. Transformed roots yielded higher levels of catechin and epicatechin than untransformed roots. Among the twenty-eight detected xanthones, four of them were identified as 1,3,5,6-tetrahydroxyxanthone, 1,3,6,7-tetrahydroxyxanthone, γ-mangostin and garcinone C were de novo synthesized in HR. Altogether, these results indicated that H. perforatum HR represent a promising experimental system for enhanced production of xanthones.     

Cholecystokinin-producing (I) cells of intestinal mucosa in dexamethasone-treated rats

The aim of this study was to investigate the morphological, immunohistochemical and ultrastructural changes of cholecystokinin-producing (I) cells of gastrointestinal mucosa in dexamethasone-treated rats (D). After 12-daily intraperitoneal administration of 2mg/kg dexamethasone, rats developed diabetes similar to human diabetes mellitus type 2. The mean diameter of the duodenum was significantly decreased due to significant reduction of volume fraction and profile area of lamina propria. There was a decrease in volume fraction and number of cholecystokinin (CCK)-producing cells per mm(2) of mucosa, as well as their numerical density, but without statistical significance. Also, dexamethasone induced appearance of hyperactive duodenal I-cells with small number of granules and dilated endoplasmic reticulum. In conclusion, the present study showed that morphological changes in duodenum cholecystokinin-producing (I) cells occurred in diabetic rats, in a manner which, suggests compensatory effort of CCK cells in diabetic condition.     

Heterosis for biomass-related traits in Arabidopsis investigated by quantitative trait loci analysis of the triple testcross design with recombinant inbred lines

Arabidopsis thaliana has emerged as a leading model species in plant genetics and functional genomics including research on the genetic causes of heterosis. We applied a triple testcross (TTC) design and a novel biometrical approach to identify and characterize quantitative trait loci (QTL) for heterosis of five biomass-related traits by (i) estimating the number, genomic positions, and genetic effects of heterotic QTL, (ii) characterizing their mode of gene action, and (iii) testing for presence of epistatic effects by a genomewide scan and marker x marker interactions. In total, 234 recombinant inbred lines (RILs) of Arabidopsis hybrid C24 x Col-0 were crossed to both parental lines and their F1 and analyzed with 110 single-nucleotide polymorphism (SNP) markers. QTL analyses were conducted using linear transformations Z1, Z2, and Z3 calculated from the adjusted entry means of TTC progenies. With Z1, we detected 12 QTL displaying augmented additive effects. With Z2, we mapped six QTL for augmented dominance effects. A one-dimensional genome scan with Z3 revealed two genomic regions with significantly negative dominance x additive epistatic effects. Two-way analyses of variance between marker pairs revealed nine digenic epistatic interactions: six reflecting dominance x dominance effects with variable sign and three reflecting additive x additive effects with positive sign. We conclude that heterosis for biomass-related traits in Arabidopsis has a polygenic basis with overdominance and/or epistasis being presumably the main types of gene action.     

Genetic basis of heterosis for growth-related traits in Arabidopsis investigated by testcross progenies of near-isogenic lines reveals a significant role of epistasis

Epistasis seems to play a significant role in the manifestation of heterosis. However, the power of detecting epistatic interactions among quantitative trait loci (QTL) in segregating populations is low. We studied heterosis in Arabidopsis thaliana hybrid C24 x Col-0 by testing near-isogenic lines (NILs) and their triple testcross (TTC) progenies. Our objectives were to (i) provide the theoretical basis for estimating different types of genetic effects with this experimental design, (ii) determine the extent of heterosis for seven growth-related traits, (iii) map the underlying QTL, and (iv) determine their gene action. Two substitution libraries, each consisting of 28 NILs and covering approximately 61 and 39% of the Arabidopsis genome, were assayed by 110 single-nucleotide polymorphism (SNP) markers. With our novel generation means approach 38 QTL were detected, many of which confirmed heterotic QTL detected previously in the same cross with TTC progenies of recombinant inbred lines. Furthermore, many of the QTL were common for different traits and in common with the 58 QTL detected by a method that compares triplets consisting of a NIL, its recurrent parent, and their F(1) cross. While the latter approach revealed mostly (75%) overdominant QTL, the former approach allowed separation of dominance and epistasis by analyzing all materials simultaneously and yielded substantial positive additive x additive effects besides directional dominance. Positive epistatic effects reduced heterosis for growth-related traits in our materials.     

Long-term mineralocorticoid receptor blockade reduces fibrosis and improves cardiac performance and coronary hemodynamics in elderly SHR

Aldosterone has been implicated as one of the mediators of cardiovascular injury in various diseases. This study examines whether mineralocorticoid antagonism ameliorates or prevents the adverse cardiac effects of hypertension and aging. Male 22-wk-old spontaneously hypertensive rats (SHR) were divided into two groups, 15 rats in each. One group received no treatment; the other was given eplerenone ( approximately 100 mg.kg(-1).day(-1)). At the age of 54 wk, indexes of cardiovascular mass, systemic and regional hemodynamics, including coronary, left ventricular function, and myocardial collagen content, were determined in all rats. Hemodynamic studies were done in conscious rats. Arterial pressure was lowered only slightly in eplerenone-treated rats, and cardiac output and total peripheral resistance did not differ from control rats. Left and right ventricular and aortic mass indexes were unaffected by eplerenone; however, concentration of hydroxyproline in the right and left ventricle was decreased significantly (P < 0.05) by eplerenone. This was accompanied by an improvement in left ventricular diastolic function and coronary hemodynamics. In conclusion, long-term therapy with the mineralocorticoid receptor antagonist eplerenone ameliorated adverse cardiac effects of both hypertension and aging in SHR. Thus reduction in myocardial fibrosis, paralleled by improvements in left ventricular function and coronary hemodynamics, was observed in eplerenone-treated SHR.     

Ecologically Determined Variation in Leaf Anatomical Traits of Sesleria rigida (Poaceae) in Serbia – Multivariate Morphometric Evidence

This paper presents the results of a multivariate morphometric study of leaf anatomical characters in different populations of Sesleria rigida sensu lato in Serbia. Morphometric analyses were performed on cross-sections of 521 tiller leaves collected from 21 populations of S. rigida. Principal component analysis and canonical discriminant analysis were used to identify the structure of variability and to determine which variables discriminate between the groups. The UPGMA (unweighted pair-group average linkage) clustering analyses based on leaf anatomical characters and habitat climatic characteristics were employed to explore whether the observed anatomical differences are a result of adaptive responses. Regression analysis (linear regression) was performed to identify the level of correlation between leaf anatomical characters and basic orographic, geological, and bioclimatic habitat characteristics. DIVA-GIS software was used to extract 19 bioclimatic parameters from the WorldClim set of global climate layers. Our detailed analyses have shown that the studied populations are anatomically very well differentiated into the serpentine populations from western Serbia and carbonate populations from eastern Serbia. The regression analysis has shown that the geological substrate (serpentinites vs. carbonates) represents the most significant abiotic factor that is correlated with the anatomical differentiation of the populations in the studied area. Besides the geological substrate, bioclimatic parameters such as precipitation and habitat humidity are also highly correlated with the leaf anatomical characters, which was not the case with temperature.     

The rotation-coupled sliding of EcoRV

It has been proposed that certain type II restriction enzymes (REs), such as EcoRV, track the helical pitch of DNA as they diffuse along DNA, a so-called rotation-coupled sliding. As of yet, there is no direct experimental observation of this phenomenon, but mounting indirect evidence gained from single-molecule imaging of RE-DNA complexes support the hypothesis. We address this issue by conjugating fluorescent labels of varying size (organic dyes, proteins and quantum dots) to EcoRV, and by fusing it to the engineered Rop protein scRM6. Single-molecule imaging of these modified EcoRVs sliding along DNA provides us with their linear diffusion constant (D(1)), revealing a significant size dependency. To account for the dependence of D(1) on the size of the EcoRV label, we have developed four theoretical models describing different types of motion along DNA and find that our experimental results are best described by rotation-coupled sliding of the protein. The similarity of EcoRV to other type II REs and DNA binding proteins suggests that this type of motion could be widely preserved in other biological contexts.