Ribonuclease P: the diversity of a ubiquitous RNA processing enzyme

Ribonuclease P is the endonuclease required for generating the mature tRNA 5'-end. The ribonucleoprotein character of this enzyme has now been proven in most organisms and organelles. Exceptions, however, are still the chloroplasts, plant nuclei and animal mitochondria where no associated RNAs have been detected to date. In contrast to the known RNA subunits, which are fairly well-conserved in size and structure among diverse phylogenetic groups, the protein contribution to the holoenzyme is highly variable in size and number of the individual components. The structure of the bacterial protein component has recently been solved. In contrast, the spatial arrangement of the multiple subunits in eukaryotic enzymes is still enigmatic. Substrate requirements of the enzymes or their catalytic RNA subunits are equally diverse, ranging from simple single domain mimics to an almost intact three-dimensional structure of the pre-tRNA substrate. As an example for an intermediate in the enzyme evolution, ribonuclease P from the Cyanophora paradoxa cyanelle will be discussed in more detail. This enzyme is unique, as it combines cyanobacterial and eukaryotic features in its function, subunit composition and holoenzyme topology.     

Pulmonary neuroendocrine cells sense succinate to stimulate myoepithelial cell contraction

1. Download : Download high-res image (185KB)
2. Download : Download full-size image

     

Biochemical Conservation and Evolution of Germacrene A Oxidase in Asteraceae

Sesquiterpene lactones are characteristic natural products in Asteraceae, which constitutes ∼8% of all plant species. Despite their physiological and pharmaceutical importance, the biochemistry and evolution of sesquiterpene lactones remain unexplored. Here we show that germacrene A oxidase (GAO), evolutionarily conserved in all major subfamilies of Asteraceae, catalyzes three consecutive oxidations of germacrene A to yield germacrene A acid. Furthermore, it is also capable of oxidizing non-natural substrate amorphadiene. Co-expression of lettuce GAO with germacrene synthase in engineered yeast synthesized aberrant products, costic acids and ilicic acid, in an acidic condition. However, cultivation in a neutral condition allowed the de novo synthesis of a single novel compound that was identified as germacrene A acid by gas and liquid chromatography and NMR analyses. To trace the evolutionary lineage of GAO in Asteraceae, homologous genes were further isolated from the representative species of three major subfamilies of Asteraceae (sunflower, chicory, and costus from Asteroideae, Cichorioideae, and Carduoideae, respectively) and also from the phylogenetically basal species, Barnadesia spinosa, from Barnadesioideae. The recombinant GAOs from these genes clearly showed germacrene A oxidase activities, suggesting that GAO activity is widely conserved in Asteraceae including the basal lineage. All GAOs could catalyze the three-step oxidation of non-natural substrate amorphadiene to artemisinic acid, whereas amorphadiene oxidase diverged from GAO displayed negligible activity for germacrene A oxidation. The observed amorphadiene oxidase activity in GAOs suggests that the catalytic plasticity is embedded in ancestral GAO enzymes that may contribute to the chemical and catalytic diversity in nature.     

Patterns and sources of variation in Daphnia phosphorus content in nature

It has recently been shown that Daphnia can vary in the phosphorus (P)-content of their body tissues, but the relative importance of genetic versus environmental causes for this variation is unexplored. We measured variation in P-content (as % body mass) of Daphnia from eight lakes and conducted experiments to contrast three sources of variation: interspecific variation, clonal variation and phenotypic plasticity. Daphnia P-content decreased with increasing seston C:P ratio across lakes. This relationship reflected both inter- and intraspecific variation. Daphnia parvula and D. dubia exhibited high P-content and were found in shallow lakes with low C:P seston, whereas D. pulicaria had low P-content and was found in deep, stratified lakes having high C:P seston. Populations of D. dentifera spanned this lake gradient and exhibited P-content that was negatively related to seston C:P. Evidence for phenotypic plasticity came from experiments with D. pulicaria and D. dentifera collected from a lake with P-deficient seston and fed a P-sufficient diet in the laboratory. In addition, populations of D. dentifera differed in P-content even after 7 d of feeding on P-sufficient resources, suggesting within-species clonal variation. However, mesocosm experiments revealed broad and surprisingly continuous variation in the P-content of individual clones of D. pulex (range 1.54–1.05%) and D. mendotae (1.51–1.07%) over a gradient in dietary C:P. The broad range in P-content exhibited by individual clones, acclimated for generations, suggests that variation in Daphnia P-content from laboratory experiments needs to be interpreted with caution. These results also show that phenotypic variation in response to environment can be a larger source of variation in P-content than genetic differences within or among species.     

Benzo(a)pyrene-induced genotoxicity: Attenuation by farnesol in a mouse model

In the present study we have evaluated the antigenotoxic effects of Farnesol (FL) a 15-carbon isoprenoid alcohol against benzo (a) pyrene [B(a)P] (125 mg kg^{? 1}.b.wt oral) induced toxicity. B(a)P administration lead to significant induction in Cytochrome P450 (CYP) content and aryl hydrocarbon hydrolase (AHH) activity (p < 0.001), DNA strand breaks and DNA adducts (p < 0.001) formation. FL was shown to suppress the activities of both CYP and AHH (p < 0.005) in modulator groups. FL pretreatment significantly (p < 0.001) restored depleted levels of reduced glutathione (GSH), quinone reductase (QR) and glutathione –S-transferase (GST). A simultaneous significant and at both the doses reduction was seen in DNA strand breaks and in in-vivo DNA adducts formation (p < 0.005), which gives some insight on restoration of DNA integrity. The results support the protective nature of FL. Hence present data supports FL as a future drug to preclude B (a) P induced toxicity.     

Influence of phosphatidylcholine/cholesterol molar ratios in liposomes on cholesterol reactivity with cholesterol:oxygen oxidoreductase]

The reactivity of sonicated phosphatidylcholine-cholesterol liposomes with cholesterol : oxygene oxydoreductase, an enzyme which catalyses the oxidation of the 3 beta hydroxyl group of cholesterol to a ketone group, is compared with that of ternary system phosphatidylcholine-cholesterol-Thesit. Regardless to the phosphatidylcholines nature and the phosphatidylcholine/cholesterol molar ratio (R), the enzymatic oxidation rate of liposomal cholesterol is slower than when the reaction is developed in the present of Thesit, a surfactif agent which destroyes the lamellar particles. This is true whether Thesit is added during preparation of dispersions or during incubation with cholesterol oxydase. The enzymatic oxydation rate of cholesterol of ternary systems phosphatidylcholine-cholesterol-Thesit is independent of the (R) value and the phosphatidylcholine fatty acid unsaturation, whereas that of phosphatidylcholine-cholesterol dispersions depends on these two parameters. The reaction rate increases in the order: dipalmitoylphosphatidylcholine to yolk egg phosphatidylcholines, and dioleylphosphatidylcholine. The optimal conditions for cholesterol oxidation were found to be R = 0.5. This result is not affected by the phosphatidylcholines nature. In order to explain these data, various hypotheses are considered. In particular, the weak liposomal cholesterol reactivity with cholesterol oxidase could result from an inhibitory effect on the enzyme-substrate combination due to the polar phosphorylcholine groups.     

Are Sulfurous Soil Amendments (S0, Fe(II)SO4, Fe(III)SO4) an Effective Tool in the Restoration of Heathland and Acidic Grassland after Four Decades of Rock Phosphate Fertilization?

This paper deals with the complex issue of reversing long‐term improvements of fertility in soils derived from heathlands and acidic grasslands using sulfur‐based amendments. The experiment was conducted on a former heathland and acid grassland in the U.K. that was heavily fertilized and limed with rock phosphate, chalk, and marl. The experimental work had three aims. First, to determine whether sulfurous soil amendments are able to lower pH to a level suitable for heathland and acidic grassland re‐creation (approximately 3 pH units). Second, to determine what effect the soil amendments have on the available pool of some basic cations and some potentially toxic acidic cations that may affect the plant community. Third, to determine whether the addition of Fe to the soil system would sequester PO₄^? ions that might be liberated from rock phosphate by the experimental treatments. The application of S⁰ and Fe^(II)SO₄^? to the soil was able to reduce pH. However, only the highest S⁰ treatment (2,000 kg/ha S) lowered pH sufficiently for heathland restoration purposes but effectively so. Where pH was lowered, basic cations were lost from the exchangeable pool and replaced by acidic cations. Where Fe was added to the soil, there was no evidence of PO₄^? sequestration from soil test data (Olsen P), but sequestration was apparent because of lower foliar P in the grass sward. The ability of the forb Rumex acetosella to apparently detoxify Al³⁺, prevalent in acidified soils, appeared to give it a competitive advantage over other less tolerant species. We would anticipate further changes in plant community structure through time, driven by Al³⁺ toxicity, leading to the competitive exclusion of less tolerant species. This, we suggest, is a key abiotic driver in the restoration of biotic (acidic plant) communities.