Lichen secondary metabolites from the cultured lichen mycobionts of Teloschistes chrysophthalmus and Ramalina celastri and their antiviral activities

Lichens and spore-derived cultured mycobionts of Teloschistes chrysophthalmus and Ramalina celastri were studied chemically, and results indicated that they produced, respectively, parietin and usnic acid as major secondary metabolites, which were purified and identified. Identification of the compounds was performed by high performance liquid chromatography and structural elucidation by nuclear magnetic resonance (1H) and electron impact mass spectrometry. Usnic acid exhibited antiviral activity whereas parietin had a virucidal effect against the arenaviruses Junin and Tacaribe.     

Monomeric and dimeric dibenzofurans from cultured mycobionts of Lecanora iseana

Spore-derived mycobionts of the lichen Lecanora iseana were cultivated on a malt-yeast extract medium supplemented with 10% sucrose and their metabolites were investigated. Four 3,7-dihydroxy-1,9-dimethyldibenzofuran derivatives along with the known 3,7-dihydroxy-1,9-dimethyldibenzofuran and five norlichexanthone derivatives were isolated. Their structures were determined by spectroscopic methods.     

Biosynthetic origin of graphenone in cultured lichen mycobionts of Graphis handelii

The biosynthetic origins of the carbon skeleton in graphenone were verified by feeding the culture of spore-derived mycobionts of the lichen Graphis handelii with sodium [1-13C]-acetate, sodium [1,2-13C2]-acetate, sodium [2-13C]-pyruvate, [1,2,3-13C]-glycerol, [13CH3]-methionine and sodium [1,4-13C2]-succinate.     

Alterations in growth and crassulacean acid metabolism (CAM) activity of in vitro cultured cactus

Unlike C-3 plants, cacti possess a crassulacean acid metabolism (CAM) physiology that can alter the pattern of carbon uptake and affect plant growth under artificial environmental conditions, especially in tissue culture. In vitro-derived plantlets of Coryphantha minima grew 7-fold larger than plants cultured under similar ex vitro conditions. Growth regulators incorporated into the culture media during shoot proliferation stage of micropropagation had a strong influence on this increased growth. Other important factors that contributed to increased growth under in vitro conditions were high relative humidity and sugar in the culture medium. An analysis of gas exchange and daily fluctuations of malic acid levels revealed an increase in net photosynthetic rate, in terms of carbon assimilation, by in vitro plants compared with that of ex vitro plants. This stimulated photosynthesis in the presence of an external carbon source was unexpected but apparently true for cacti exhibiting CAM physiology. Unlike CAM plants grown in ex vitro conditions, net CO₂ uptake by in vitro-cultured cacti occurred continuously in the light as well as the dark. Once regenerated, cacti were transferred to ex vitro conditions where the normal CAM pathway resumed with a concomitant reduction in growth and CO₂ uptake. These results showed that growth of cacti can be considerably accelerated by in vitro culture. This revised version was published online in June 2006 with corrections to the Cover Date.     

Phytic acid-mediated regulation of secondary metabolism in Streptomyces thermoviolaceus grown in simple and complex media

Primary and secondary metabolism in the thermophilic actinomycete Streptomycesthermoviolaceus were found to be strongly regulated by phosphate in complexand defined media. Increasing phosphate levels in glutamate minimal salts media led to peakproduction of granaticin at 5 mmol phosphate, a concentration that was growth-limiting, beforetotal inhibition of antibiotic production at 50 mmol. Product formation in particulate rapeseedmeal-based media was found to be less affected by the initial phosphate concentration. Theaddition of 5 mmol phytic acid to proline minimal salts media led to an increase in theconcentration of phosphate optimal for antibiotic production from 5·7 mmol to 15 mmol andreduced inhibition at higher concentrations. Phytic acid was shown to bind phosphate fromminimal salts media and inhibit the growth of the organism at high concentrations. Differences inthe production of granaticin by S. thermoviolaceus in two rapeseed meal-derived mediawere shown to be phosphate and phytic acid-related. In particulate rapeseed, the additionalphosphate from minimal salts media was predominantly bound in an organic-soluble complex,while in extracted rapemeal media, phosphate was present predominantly in the free form.Overall, the work suggests that reduction in growth rate,which can be brought about by a varietyof factors including low phosphate concentrations,is the critical factor for the onset of secondarymetabolism in S.thermoviolaceus.     

Three isocoumarins and a benzofuran from the cultured lichen mycobionts of Pyrenula sp

The spore-derived mycobionts of the lichen Pyrenula sp. were cultivated on a malt-yeast extract medium supplemented with 10% sucrose. The investigation of their metabolites resulted in isolation of four compounds, three isocoumarins and a biogenetically related benzofuran; their structures were determined by spectroscopic methods.     

Parietin,a photoprotective secondary product of the lichen Xanthoria parietina

Secondary lichen products can be extracted from air-dry thalli of Xanthoria parietina, Xanthoparmelia conspersa and Parmelina tiliacea with 100% acetone without affecting either short-or long-term viability. In Xanthoria parientina damage by acetone started to occur as water content reached the critical lower limit for photosystem II (PSII) activity. Extraction of the blue-light absorbing cortical pigment parietin increased the susceptibility of both air-dry and hydrated thalli to high light. Damage by high light levels caused a permanent reduction in F _v/F_m, quantum yield for photosynthetic O₂ production and photosynthetic capacity measured after a 2-day recovery period at low light levels (20 mol photons m^-2 s^-1). Parietin therefore protects the photosynthetic apparatus of Xanthoria parietina against damage by high light levels. Extraction of UV-absorbing pigments from Xanthoparmelia conspersa and Parmelina tiliacea did not increase photoinhibition after 24 h exposure to high light.     

山西省太岳山区有害元素大气污染的地衣(丽石黄衣)监测

山西太岳山区大气元素沉降研究较少。为摸清该区域大气污染水平及其受盆地的可能影响,以ICP-MS法测定采自15个样点的丽石黄衣(Xanthoria elegans)地衣体内7种常见有害元素(As、Cd、Cu、Pb、S、Sb和Zn)的含量,并进行相关性分析、方差分析和聚类分析。结果显示:该区域元素含量一般高于"清洁区",但低于"严重污染区"的相应数据,表明太岳山区具有一定程度的大气污染。聚类分析将15个样点分为两类,两类样点在距盆地距离(distance from the nearest basin, DFNB)和多数元素的含量方面均差异显著,近盆地区(DFNB=13.7±7.987 km)中的6种元素(As、Cd、Cu、Pb、Sb和Zn)含量均显著高于远盆地区(DFNB=32.5±6.062 km), 5种元素(As、Cu、Pb、Sb和Zn)的含量与DFNB之间的负相关关系显著(r≤-0.59; P≤0.02),表明山区内部的大气质量受盆地地区的大气污染物传输的影响。S含量的空间异质性低(变异系数CV10%),和DFNB的相关性及在近盆地区和远盆地区之间的差异均不显著(P0.05),但仍随DFNB增加呈下降趋势,表明盆地人类活动所释放的S可能主要是以更易于扩散的气态形式向山区传输的。本研究结果表明丽石黄衣是太岳山区大气有害元素污染的良好监测生物。     

Interaction between exogenous brassinolide, IAA and BAP in secondary metabolism of cultured Onosma paniculatum cells

Brassinolide (BL) together with IAA (indoleacetic acid) and BAP (6-benzylaminopurine) have been reported to enhance shikonin formation in cultured Onosma paniculatum cells . In this paper, we show that BL interacted significantly with both IAA and BAP to influence cell growth. In a BL/IAA interaction experiment, the optimal BL concentration for cell growth increased with IAA concentration. Thus, with IAA concentrations of 0.05, 0.1, 1.0, and 10 mg/L in the growth medium, the optimal BL concentrations for cell growth were 10, 10³, 10⁵, and 10⁷pg/L, respectively. In a BL/BAP interaction experiment, cell growth decreased with increasing concentration of BL at any given concentration of BAP. The optimal concentrations of BL and IAA for cell growth were 10 pg/L and 0.05 mg/L, respectively, among all BL/IAA combinations, and concentrations of 10 pg/L and 0.5 mg/L for BL and BAP were optimal among all BL/BAP combinations. Shikonin formation was affected significantly by both BL/IAA and BL/BAP combinations. Shikonin content was enhanced by increasing BL concentrations with IAA concentrations in the range of 0.05–10 mg/L and with BAP concentrations in the range of 0.5–5 mg/L in BL/IAA and BL/BAP experiments, respectively. The optimal combination of BL and IAA for enhanced shikonin formation was 10⁷pg/L and 0.05 mg/L, and BL and BAP concentrations of 10⁵pg/L and 0.5 mg/L optimal for shikonin formation. These results indicate that BL-stimulated cell growth occurs at lower concentration (10 pg/L) and enhanced shikonin formation at higher concentration (10⁵–10⁷pg/L), in combination with IAA or BAP at appropriate concentrations. Furthermore, BL increased phenylalanine ammonia-lyase (PAL) and p-hydroxybenzoic acid -geranyltransferase (PHB-geranyltransferase) activities, but decreased the activity of PHB–O–glucosyltransferase. These results suggest that enhanced shikonin formation induced by BL involves regulation of these key enzyme activities.     

Effects of abscisic acid on the secondary metabolism of cultured Onosma paniculatum cells

ABA addition to B5 or M9 medium at the concentrations from 0.1 to 5.0 mg/l suppressed growth of Onosma paniculatum cells. The addition of these ABA concentrations to M9 medium also significantly decreased the formation of shikonin and its derivatives in the cultured cells during the entire course of culturing. The enzyme activity assay showed that, on the 4th day after inoculation, 0.1 mg/l ABA significantly decreased the activities of phenylalanine ammonia-lyase, the first enzyme, and p-hydroxybenzoic acid-geranyltransferase, a key enzyme involved in shikonin biosynthesis. However, no significant change in these two enzyme activities was found during the following days for testing (8, 12, and 16 days). Interestingly, RT-PCR analysis of the PAL1 and LePGT1 gene expression showed no significant changes on the 4th day after inoculation. Furthermore, we found that the inhibitory effect of ABA on the secondary metabolism could be alleviated significantly by the addition of 2-aminoethyl diphenyl borate (an inhibitor of inositol triphosphate-receptor) or nicotinamide (an inhibitor of ADP-ribose cyclase), which functions by decreasing the intracellular calcium concentration. Published in Russian in Fiziologiya Rastenii, 2007, Vol. 54, No. 4, pp. 597–603. The text was submitted by the authors in English.     

Response to copper stress in aposymbiotically grown lichen mycobiont Cladonia cristatella: uptake,viability, ergosterol and production of non-protein thiols

The mycobiont of lichens usually determines the morphology of the symbiotic organism and is also dominates in terms of biomass. However, its role for sensitivity or tolerance of lichens to heavy metals is almost unknown. In the present study, the influence of copper (Cu) on the aposymbiotically-grown mycobiont of Cladonia cristatella was assessed. Intracellular Cu uptake was correlated with increasing Cu concentrations over a 24-h exposure time. Viability, measured as the degree of reduction of triphenyltetrazolium chloride to triphenyl formazan, as well as to ergosterol levels, decreased with growing Cu concentrations tested. Reduced glutathione (GSH) was found to be the most abundant low-molecular-weight thiol in the hyphae of C. cristatella and its intracellular content increased at concentrations of 10 μm Cu. Higher Cu concentrations caused a significant decrease in GSH, possibly due to heavy metal-induced oxidation of GSH to glutathione disulphide (GSSG). Free cysteine levels were relatively constant. As expected, we did not observe the production of phytochelatins in the mycobiont, contrary to what is found in intact lichens and axenic cultures of their photobionts.     

Variation in the ribosomal ITS-sequence of the lichens Buellia frigida and Xanthoria elegans from the Vestfold Hills,eastern Antarctica

Abstract:Thalli of the lichens Buellia frigida and Xanthoria elegans were collected from five different locations each 5–15 km apart in the Vestfold Hills, Princess Elizabeth Land, eastern Antarctica. A further collection was made from Mawson Station, Mac Robertson Land, eastern Antarctica, 660 km away. DNA was extracted from whole thalli and the ribosomal ITS region amplified by PCR using fungal specific primers. Resulting products were sequenced to gain an indication of whether or not variation was present within populations of lichen-forming fungi from continental Antarctica, and therefore of the availability of genetic resources to react to pressures such as climate change. Three genotypes ofB. frigida and two of X. elegans were detected in the Vestfold Hill collections. However, these differed by only one nucleotide position suggesting the presence of relatively little genetic variation, if the ITS region is indicative of the overall genome. Buellia frigida collected from Mawson Station had an identical ITS region sequence to the most common Vestfold Hills genotype, indicating that this species may have a low level of genetic variation across much of eastern Antarctica. In contrast, X. elegans collected from Mawson showed considerable genetic variation from the Vestfolds thalli, differing at 14·2% of nucleotide positions and had an identical ITS region sequence to an isolate from maritime Antarctica 4960 km away. Samples from the Vestfold Hills formed a distinct cluster in a phylogenetic analysis of ITS sequences from a worldwide collection of X. elegans isolates.     

Habitat selection and ecology of Xanthoria elegans (Link) Th. Fr. in glacier forefields: implications for lichenometry

Habitats occupied by the largest Xanthoria elegans (Link) Th. Fr. thalli at seven glacier forefields in the Canadian Rockies were studied to investigate the lichenometric assumption that large thalli occupy ideal sites for growth. The largest thalli were found on steep or overhanging facets at the base of grey limestone clasts that were embedded in moraines. These thalli were unfragmented, had nearly circular outlines, were bordered by barren rock and had SSE to S orientations. This is consistent with the general expectation that south-facing sites offer high solar input and a long snow-free season. Orientations other than south could result by chance or may reflect the importance of microscale factors (e.g. reflected rather than direct solar input). Closure of X. elegans communities and coalescence of thalli was only found at sites that were naturally fertilized with dung. It is concluded that all clasts do not afford homogeneous or ideal environments for lichen growth and do not have an equal chance of being colonized. This raises doubts concerning the validity of statistical normality assumptions in lichenometry and the use of grids to assess closure in lichen communities.     

Mechanisms of aging and energy metabolism in Caenorhabditis elegans

Aging studies on diverse species ranging from yeast to man have culminated in the delineation of several signaling pathways that influence the process of senescent decline and aging. While understanding these interlinked signal-transduction cascades is becoming even more detailed and comprehensive, the cellular and biochemical processes they impinge upon to modulate the rate of senescent decline and aging have lagged considerably behind. This fundamental question is one of the most important challenges of modern aging research and has been the focus of recent research efforts. Emerging findings provide insight into the facets of cellular metabolism which can be fine-tuned by upstream signaling events to ultimately promote longevity. Here, we survey the mechanisms regulating aging in the simple nematode worm Caenorhabditis elegans, aiming to highlight recent discoveries that shed light into the interface between aging signaling pathways and cellular energy metabolism. Our objective is to review the current understanding of the processes involved and discuss mechanisms that are likely conserved in higher organisms.     

Alterations of inosinate branchpoint enzymes in cultured human lymphoblasts

The specific activities of the three enzymes of the inosinate branchpoint are independently regulated when lymphoblasts are grown under various tissue culture conditions. In comparison to rapidly dividing cells, lymphoblasts at high cell density with no cellular division have decreased activity of the enzymes which commit inosinate to adenylate or guanylate, while cytoplasmic 5'-nucleotidase is relatively preserved. A linear relationship between inosinate dehydrogenase activity and growth rate (r = 0.92) exists in lymphoblasts with slowed growth rates. In contrast, in dividing cells adenylosuccinate synthetase and 5'-nucleotidase do not vary with growth rate. Adenylosuccinate synthetase and inosinate dehydrogenase activities appear to be related to the presence or rate of cellular division, as opposed to the presence or degree of neoplastic transformation. Lymphoblast lines with alterations of specific purine metabolic enzymes have characteristic alteration of the inosinate utilizing enzymes. Deficiencies of purine nucleoside phosphorylase or hypoxanthine phosphoribosyltransferase, abnormalities which render the cell unable to salvage purine effectively, are associated with depressed inosinate dehydrogenase activity. Insertion of the hypoxanthine phosphoribosyltransferase gene into hypoxanthine phosphoribosyltransferase-deficient cells normalizes inosinate dehydrogenase activity, while a hypoxanthine phosphoribosyltransferase-deficient mutant selected from a hypoxanthine phosphoribosyltransferase-containing line has depressed inosinate dehydrogenase activity. In contrast, overactivity of phosphoribosylpyrophosphate synthetase, with enhanced excretion of purines due to excessive production, is associated with elevated inosinate dehydrogenase activity. Inosinate dehydrogenase appears to be regulated according to the availability of purine nucleotides. Patients who overproduce uric acid and potentially have undescribed purine metabolic defects are now being screened for abnormalities in the inosinate branchpoint enzymes.