Ion channels in smooth muscle: regulation by the sarcoplasmic reticulum and mitochondria

In smooth muscle, Ca(2+) regulates cell division, growth and cell death as well as providing the main trigger for contraction. Ion channels provide the major access route to elevate the cytoplasmic Ca(2+) concentration ([Ca(2+)](c)) in smooth muscle by permitting Ca(2+) entry across the plasma membrane and release of the ion from intracellular Ca(2+) stores. The control of [Ca(2+)](c) relies on feedback modulation of the entry and release channels by Ca(2+) itself. Local rises in [Ca(2+)](c) may promote or inhibit channel activity directly or indirectly. The latter may arise from Ca(2+) regulation of ionic conductances in the plasma membrane to provide control of cell excitability and so [Ca(2+)](c) entry. Organelles such as mitochondria may also contribute significantly to the feedback regulation of ion channel activity by the control of Ca(2+) or redox status of the cell. This brief review describes the feedback regulation of Ca(2+) release from the internal Ca(2+) store and of plasma membrane excitability in smooth muscle.     

Assessing the importance of genotype × environment interaction for root traits in rice using a mapping population II: conventional QTL analysis

Modifying plant root systems is considered a means of crop improvement targeted to low-resource environments, particularly low nutrient and drought-prone agriculture. The identification of quantitative trait loci (QTLs) for root traits has stimulated marker-assisted breeding to this end, but different QTLs have been detected in different populations of the same species, and importantly, in the same population when grown in different experimental environments. The presence of QTL × environment interaction is implicated, and this must be characterised if the utility of the target QTLs is to be realised. Previous attempts to do this suffer from a lack of control over replicate environments and inadequate statistical rigour. The Bala × Azucena mapping population was grown in two replicate experiments of four treatment environments, a control, a low light, a low soil nitrogen and a low soil water treatment. After a 4 weeks growth, maximum root length, maximum root thickness, root mass below 50 cm, total plant dry mass, % root mass and shoot length were measured. A summary of the overall results is presented in an accompanying paper. Here, QTL analysis by composite interval mapping is presented. A total of 145 QTLs were detected, mapping to 37 discrete loci on all chromosomes. Superficial evidence of QTL × E (great difference in LOD score) was tested by single-marker analysis which confirmed QTL × E for five loci representing only five individual trait-loci interactions. Some loci appeared to be stable across environments. Some QTLs were clearly more or less active under low light, low nitrogen or drought. A few notable loci on chromosomes 1, 2, 3, 5, 7 and 9 are briefly discussed. Also discussed are some remaining statistical shortcomings that will be addressed in another companion paper.     

Mgm101 is a Rad52-related protein required for mitochondrial DNA recombination

Homologous recombination is a conserved molecular process that has primarily evolved for the repair of double-stranded DNA breaks and stalled replication forks. However, the recombination machinery in mitochondria is poorly understood. Here, we show that the yeast mitochondrial nucleoid protein, Mgm101, is related to the Rad52-type recombination proteins that are widespread in organisms from bacteriophage to humans. Mgm101 is required for repeat-mediated recombination and suppression of mtDNA fragmentation in vivo. It preferentially binds to single-stranded DNA and catalyzes the annealing of ssDNA precomplexed with the mitochondrial ssDNA-binding protein, Rim1. Transmission electron microscopy showed that Mgm101 forms large oligomeric rings of ～14-fold symmetry and highly compressed helical filaments. Specific mutations affecting ring formation reduce protein stability in vitro. The data suggest that the ring structure may provide a scaffold for stabilization of Mgm101 by preventing the aggregation of the otherwise unstable monomeric conformation. Upon binding to ssDNA, Mgm101 is remobilized from the rings to form distinct nucleoprotein filaments. These studies reveal a recombination protein of likely bacteriophage origin in mitochondria and support the notion that recombination is indispensable for mtDNA integrity.     

Mapping Soil Properties of Africa at 250 m Resolution: Random Forests Significantly Improve Current Predictions

80% of arable land in Africa has low soil fertility and suffers from physical soil problems. Additionally, significant amounts of nutrients are lost every year due to unsustainable soil management practices. This is partially the result of insufficient use of soil management knowledge. To help bridge the soil information gap in Africa, the Africa Soil Information Service (AfSIS) project was established in 2008. Over the period 2008–2014, the AfSIS project compiled two point data sets: the Africa Soil Profiles (legacy) database and the AfSIS Sentinel Site database. These data sets contain over 28 thousand sampling locations and represent the most comprehensive soil sample data sets of the African continent to date. Utilizing these point data sets in combination with a large number of covariates, we have generated a series of spatial predictions of soil properties relevant to the agricultural management—organic carbon, pH, sand, silt and clay fractions, bulk density, cation-exchange capacity, total nitrogen, exchangeable acidity, Al content and exchangeable bases (Ca, K, Mg, Na). We specifically investigate differences between two predictive approaches: random forests and linear regression. Results of 5-fold cross-validation demonstrate that the random forests algorithm consistently outperforms the linear regression algorithm, with average decreases of 15–75% in Root Mean Squared Error (RMSE) across soil properties and depths. Fitting and running random forests models takes an order of magnitude more time and the modelling success is sensitive to artifacts in the input data, but as long as quality-controlled point data are provided, an increase in soil mapping accuracy can be expected. Results also indicate that globally predicted soil classes (USDA Soil Taxonomy, especially Alfisols and Mollisols) help improve continental scale soil property mapping, and are among the most important predictors. This indicates a promising potential for transferring pedological knowledge from data rich countries to countries with limited soil data.     

Preparation of soybean seed samples for FT-IR microspectroscopy

Typical preparation of seed samples for infrared (IR) microspectroscopy involves imbibition of the seed for varying time periods followed by cryosectioning. Imbibition, however, may initiate germination even at 4° C with associated changes in the chemistry of the sample. We have found that it is possible to section seeds that are sufficiently hard, such as soybeans, on a standard laboratory microtome without imbibition. The use of dry sectioning of unimbibed seeds is reported here, as well as a comparison of different mounting media and modes of analysis. Glycerol, Tissue-Tek, and ethanol were used as mounting media, and the quality of the resulting spectra was assessed. Ethanol was the preferred mountant, because it dried quickly with no residue and thus did not interfere with the spectrum of interest. Analysis in transmission mode using barium fluoride windows to hold the samples was compared with transmission-reflection analysis with sections mounted on special infrared-reflecting slides. The two modes of analysis performed well in different regions of the spectrum. The mode of analysis (transmission vs. transmission-reflection) should be based on the components of greatest interest in the sample.     

Metabolism of kaurenoids by Gibberella fujikuroi in the presence of the plant growth retardant,N,N,N-trimethyl-1-methyl-(2′,6′,6′-trimethylcyclohex-2′-en-1′-yl)prop-2-enylammonium iodide

The plant growth retardant, N,N,N-trimethyl-1-methyl-(2′,6′,6′-trimethylcyclohex-2′-en-1′-yl)prop-2-enylammonium iodide, is shown to block gibberellin biosynthesis in Gibberella fujikuroi between mevalonate and ent-kaur-16-ene, probably by inhibiting ent-kaur-16-ene synthetase A-activity. In the presence of the plant growth retardant, cultures of the fungus incorporate (26.5%) added ent-[¹⁴C]-kaur-16-ene into gibberellin A₃. Under the same conditions kaur-16-ene, 13β-kaur-16-ene, and ent-kaur-15-ene are not metabolised to gibberellin analogues.     

Conserved structure of Snu13 from the highly reduced spliceosome of Cyanidioschyzon merolae

Structural and functional analysis of proteins involved in pre‐mRNA splicing is challenging because of the complexity of the splicing machinery, known as the spliceosome. Bioinformatic, proteomic, and biochemical analyses have identified a minimal spliceosome in the red alga Cyanidioschyzon merolae. This spliceosome consists of only 40 core proteins, compared to ～70 in S. cerevisiae (yeast) and ～150 in humans. We report the X‐ray crystallographic analysis of C. merolae Snu13 (CmSnu13), a key component of the assembling spliceosome, and present evidence for conservation of Snu13 function in this algal splicing pathway. The near identity of CmSnu13's three‐dimensional structure to yeast and human Snu13 suggests that C. merolae should be an excellent model system for investigating the structure and function of the conserved core of the spliceosome.     

Enzymes from seeds of Phaseolus vulgaris L.: Hydroxylation of gibberellins A20 and A1 and 2,3-dehydrogenation of gibberellin A20

A time-course study is described relating the enzyme activities for GA₂₀ metabolism with seed development in Phaseolus vulgaris L. Enzyme activity for the 3-hydroxylation of GA₂₀ to GA₁, and for the 2,3-desaturation of GA₂₀ to GA₅, was confined to the cotyledons and showed maximal specific activity at 21 d after anthesis. These enzyme activities co-occurred, together with a much lower level of activity for the 2-hydroxylation of GA₂₀ to GA₂₉. The observed rates of GA₁, GA₅ and GA₂₉ formation from GA₂₀ were constant under a range of incubation conditions. Enzyme activity for the conversion of GA₁ to GA₈ was detected only in embryos of seed from 40 d after anthesis. By deuterium-labelling and analysis of the products by gas chromatography-selected ion monitoring it was shown that 2-hydroxylation of GA₁ to GA₈ and 3-hydroxylation of GA₂₀ to GA₁ occur with retention of configuration and that the conversion of GA₂₀ to GA₅ occurs with loss of the 2- and 3-hydrogens. These results establish that GA₁ is not formed from GA₂₀ via GA₅.Abbreviations GA_n Gibberellin A_n - GC gas chromatography - HPLC high-performance liquid chromatography - MS mass spectrometry - SIM selected-ion monitoring - THO tritiated water     

THE EFFECT OF PHENOBARBITONE ANAESTHESIA UPON SOME ORGANIC PHOSPHATES, GLYCOLYTIC METABOLITES AND CITRIC ACID CYCLE-ASSOCIATED INTERMEDIATES OF THE RAT BRAIN

In order to study the effect of phenobarbitone anaesthesia upon the energy metabolism of the brain, organic phosphates, glycolytic metabolites and citric acid cycle intermediates were measured in rats anaesthetized with 175-200 mg/kg of phenobarbitone, and the results were compared to those obtained in rats anaesthetized with halo-thane or with nitrous oxide. An attempt was made to separate the effects of the phenobarbitone anaesthesia from those caused by the accompanying intracellular alkalosis by exposing one group of animals to hypercapnia of such a degree that normalization of the intracellular pH was achieved. Phenobarbitone anaesthesia did not alter the tissue concentrations of ATP, ADP or AMP, but led to a moderate increase in the phosphocreatine concentration. However, since this increase was reversed in the hypercapnic group it is concluded that it may be due partly to a pH-dependent shift in the creatine phosphokinase equilibrium. There was a decrease in the tissue concentrations of all measured substrates from pyruvate and onwards. The results indicate that phenobarbitone leads to a primary inhibition of glycolysis, which cannot be related to detectable changes in ATP, ADP or AMP. The resulting lowering of the tissue concentrations of a number of metabolic acids may be part of the explanation why barbiturate anaesthesia is associated with an intracellular alkalosis.     

Anticoagulant Therapy in Coronary Artery Disease: A Therapeutic Enigma

The authors'' experience with anticoagulant therapy in both the acute phase and the long-term management of myocardial infarction has proved disappointing. A review of the literature has failed to establish benefit when all patients with coronary artery disease are treated with anticoagulant drugs. A need for well-controlled studies still exists.