Influence of NaCl, Cd(NO3)2 and air humidity on transpiration of Tamarix aphylla

Transpiration rates of young Tamarix aphylla (L.) Karst, plants grown in hydroponics were measured under NaCl- and Cd(NO₃)₂-stress. Transpiration rates were negatively correlated with the relative humidity of the ambient air at all NaCl concentrations investigated. Low and intermediate concentrations of Cd²⁺ (45 and 90 μ M , respectively) in the medium caused an increase in transpiration rates. This was particularly pronounced at low levels of relative humidity. At 180 μ M Cd²⁺, transpiration rates dropped, probably as a result of root damage due to Cd²⁺ toxicity. Since the transpiration rates differed by a factor of ca 3 between day and night, it is concluded that the stomata did not lose their ability to regulate transpiration under the influence of NaCl or of Cd(NO₃)₂. The transpiration behaviour of T. aphylla indicates that the effect of water vapour pressure (presented as relative humidity) on the degree of stomatal opening is small. Under conditions of ample water supply transpiration follows the evaporative demand of the ambient air and is influenced by the water uptake capacity of the root system as well as by other environmental factors, e.g. light.     

Enhanced accumulation of starch and total carbohydrates in alginate-immobilized Chlorella spp. induced by Azospirillum brasilense: II. Heterotrophic conditions

The effect of the bacterium Azospirillum brasilense jointly immobilized with Chlorella vulgaris or C. sorokiniana in alginate beads on total carbohydrates and starch was studied under dark and heterotrophic conditions for 144h in synthetic growth medium supplemented with either d-glucose or Na-acetate as carbon sources. In all treatments, enhanced total carbohydrates and starch content per culture and per cell was obtained after 24h; only jointly immobilized C. vulgaris growing on d-glucose significantly increased total carbohydrates and starch content after 96h. Enhanced accumulation of carbohydrate and starch under jointly immobilized conditions was variable with time of sampling and substrate used. Similar results occurred when the microalgae was immobilized alone. In both microalgae growing on either carbon sources, the bacterium promoted accumulation of carbohydrates and starch; when the microalgae were immobilized alone, they used the carbon sources for cell multiplication. In jointly immobilized conditions with Chlorella spp., affinity to carbon source and volumetric productivity and yield were higher than when Chlorella spp. were immobilized alone; however, the growth rate was higher in microalgae immobilized alone. This study demonstrates that under heterotrophic conditions, A. brasilense promotes the accumulation of carbohydrates in two strains Chlorella spp. under certain time-substrate combinations, producing mainly starch. As such, this bacterium is a biological factor that can change the composition of compounds in microalgae in dark, heterotrophic conditions.     

Differential association of microRNAs with polysomes reflects distinct strengths of interactions with their mRNA targets

While microRNAs have been shown to copurify with polysomes, their relative fraction in the translation pool (polysome occupancy) has not yet been measured. Here, we introduce a high-throughput method for quantifying polysome occupancies of hundreds of microRNAs and use it to investigate factors affecting these occupancies. Analysis in human embryonic stem cells (hESCs) and foreskin fibroblasts (hFFs) revealed microRNA-specific preferences for low, medium, or high polysome occupancy. Bioinformatics and functional analysis based on overexpression of endogenous and chimeric microRNAs showed that the polysome occupancy of microRNAs is specified by its mature sequence and depends on the choice of seed. Nuclease treatment further suggested that the differential occupancy of the microRNAs reflects interactions with their mRNA targets. Indeed, analysis of microNRA•mRNA duplexes showed that pairs involving high occupancy microRNAs exhibit significantly higher binding energy compared to pairs with low occupancy microRNAs. Since mRNAs reside primarily in polysomes, strong interactions lead to high association of microRNAs with polysomes and vice versa for weak interactions. Comparison between hESCs and hFFs data revealed that hESCs tend to express lower occupancy microRNAs, suggesting that cell type–dependent translational features may be affected by expression of a particular set of microRNAs.     

Ethanol causes the redistribution of L1 cell adhesion molecule in lipid rafts

Fetal alcohol spectrum disorder is estimated to affect 1% of live births. The similarities between children with fetal alcohol syndrome and those with mutations in the gene encoding L1 cell adhesion molecule (L1) implicates L1 as a target of ethanol developmental neurotoxicity. Ethanol specifically inhibits the neurite outgrowth promoting function of L1 at pharmacologic concentrations. Emerging evidence shows that localized disruption of the lipid rafts reduces L1-mediated neurite outgrowth. We hypothesize that ethanol impairment of the association of L1 with lipid rafts is a mechanism underlying ethanol's inhibition of L1-mediated neurite outgrowth. In this study, we examine the effects of ethanol on the association of L1 and lipid rafts. We show that, in vitro, L1 but not N-cadherin shifts into lipid rafts following treatment with 25 mM ethanol. The ethanol concentrations causing this effect are similar to those inhibiting L1-mediated neurite outgrowth. Increasing chain length of the alcohol demonstrates the same cutoff as that previously shown for inhibition of L1-L1 binding. In addition, in cerebellar granule neurons in which lipid rafts are disrupted with methyl-beta-cyclodextrin, the rate of L1-mediated neurite outgrowth on L1-Fc is reduced to background rate and that this background rate is not ethanol sensitive. These data indicate that ethanol may inhibit L1-mediated neurite outgrowth by retarding L1 trafficking through a lipid raft compartment.     

The dynamic of the t-haplotype in wild populations of the house mouse Mus musculus domesticus in Israel

The t-haplotype, a variant of the proximal part of the mouse chromosome 17, is composed of at least four inversions and is inherited as a single genetic unit. The haplotype causes embryonic mortality or male sterility when homozygous. Genes within the complex are responsible for distortion of Mendelian transmission ratio in males. Thus, the t-haplotype in heterozygous males is transferred to over 95% of the progeny. We examined the dynamic and behavior of the t-haplotype in wild populations of the house mouse in Israel. The Israeli populations show high frequency (15%–20%) of both partial and complete t-carrying mice, supporting the suggestion that the t-complex evolved in the M. domesticus line in the Israeli region. In one population that had the highest frequency of t-carrying individuals, we compared the level of gene diversity between t-carrying and normal mice in the marker’s loci: H-2 locus of the major histocompatibility complex (MHC) on the t-haplotype of chromosome 17, three microsatellites on other chromosomes, and the mitochondrial D-loop. Genetic variability was high in all tested loci in both t and (+) mice. All t mice carried the same chromosome and showed the same H-2 haplotype. While t-carrying mice showed significant H-2 heterozygotes access, (+) mice expressed significant H-2 heterozygote deficiency. There were no differences in the level of gene diversity between t and (+) mice in the other loci. Heterozygosity level at the MHC may be an additional factor in the selective forces balancing the t-haplotype polymorphism.     

Genetic structure of a galling aphid Slavum wertheimae and its host tree Pistacia atlantica across an Irano-Turanian distribution: from fragmentation to speciation?

The Irano-Turanian distribution zone in the Levant crossroad is fragmented along different phyto- and zoogeographic and climatic regions, a relict of wider distribution in moister conditions during the Pleistocene and the Holocene. We examined the effect of the disjunct Irano-Turanian distribution among distinct mesic and xeric habitats on the genetic structure of the gall-forming aphid Slavum wertheimae and its obligate host tree Pistacia atlantica in Israel and Jordan. The genetic study included amplified fragment length polymorphism analysis of the trees and aphids and sequence analysis of fragments of the mitochondrial genes cytochrome oxidase I and II (COI and COII) of the aphids. P. atlantica trees did not show any differentiation or genetic structure among climatic regions. S. wertheimae aphids in Israel exhibited two distinct phylogenetic groups, one occupying the mesic region in the north and the other inhabiting the xeric south. The Jordanian aphids clustered within the Israeli northern populations. The results suggest that while the fragmented Irano-Turanian distribution in the Levant does not affect the genetic structure of P. atlantica trees, it promotes genetic differentiation among the aphids?? populations and may initiate an allopatric speciation.     

Homeobox-clock protein interaction in zebrafish. A shared mechanism for pineal-specific and circadian gene expression

In non-mammalian vertebrates, the pineal gland is photoreceptive and contains an intrinsic circadian oscillator that drives rhythmic production and secretion of melatonin. These features require an accurate spatiotemporal expression of an array of specific genes in the pineal gland. Among these is the arylalkylamine N-acetyltransferase, a key enzyme in the melatonin production pathway. In zebrafish, pineal specificity of zfaanat2 is determined by a region designated the pineal-restrictive downstream module (PRDM), which contains three photoreceptor conserved elements (PCEs) and an E-box, elements that are generally associated with photoreceptor-specific and rhythmic expression, respectively. Here, by using in vivo and in vitro approaches, it was found that the PCEs and E-box of the PRDM mediate a synergistic effect of the photoreceptor-specific homeobox OTX5 and rhythmically expressed clock protein heterodimer, BMAL/CLOCK, on zfaanat2 expression. Furthermore, the distance between the PCEs and the E-box was found to be critical for PRDM function, suggesting a possible physical feature of this synergistic interaction. OTX5-BMAL/CLOCK may act through this mechanism to simultaneously control pineal-specific and rhythmic expression of zfaanat2 and possibly also other pineal and retinal genes.     

FXYD proteins stabilize Na,K-ATPase: amplification of specific phosphatidylserine-protein interactions

FXYD proteins are a family of seven small regulatory proteins, expressed in a tissue-specific manner, that associate with Na,K-ATPase as subsidiary subunits and modulate kinetic properties. This study describes an additional property of FXYD proteins as stabilizers of Na,K-ATPase. FXYD1 (phospholemman), FXYD2 (γ subunit), and FXYD4 (CHIF) have been expressed in Escherichia coli and purified. These FXYD proteins associate spontaneously in vitro with detergent-soluble purified recombinant human Na,K-ATPase (α1β1) to form α1β1FXYD complexes. Compared with the control (α1β1), all three FXYD proteins strongly protect Na,K-ATPase activity against inactivation by heating or excess detergent (C₁₂E₈), with effectiveness FXYD1 > FXYD2 ≥ FXYD4. Heating also inactivates E₁ ↔ E₂ conformational changes and cation occlusion, and FXYD1 protects strongly. Incubation of α1β1 or α1β1FXYD complexes with guanidinium chloride (up to 6 m) causes protein unfolding, detected by changes in protein fluorescence, but FXYD proteins do not protect. Thus, general protein denaturation is not the cause of thermally mediated or detergent-mediated inactivation. By contrast, the experiments show that displacement of specifically bound phosphatidylserine is the primary cause of thermally mediated or detergent-mediated inactivation, and FXYD proteins stabilize phosphatidylserine-Na,K-ATPase interactions. Phosphatidylserine probably binds near trans-membrane segments M9 of the α subunit and the FXYD protein, which are in proximity. FXYD1, FXYD2, and FXYD4 co-expressed in HeLa cells with rat α1 protect strongly against thermal inactivation. Stabilization of Na,K-ATPase by three FXYD proteins in a mammalian cell membrane, as well the purified recombinant Na,K-ATPase, suggests that stabilization is a general property of FXYD proteins, consistent with a significant biological function.     

Dissecting eukaryotic translation and its control by ribosome density mapping

Translation of an mRNA is generally divided into three stages: initiation, elongation and termination. The relative rates of these steps determine both the number and position of ribosomes along the mRNA, but traditional velocity sedimentation assays for the translational status of mRNA determine only the number of bound ribosomes. We developed a procedure, termed Ribosome Density Mapping (RDM), that uses site-specific cleavage of polysomal mRNA followed by separation on a sucrose gradient and northern analysis, to determine the number of ribosomes associated with specified portions of a particular mRNA. This procedure allows us to test models for translation and its control, and to examine properties of individual steps of translation in vivo. We tested specific predictions from the current model for translational control of GCN4 expression in yeast and found that ribosomes were differentially associated with the uORFs elements and coding region under different growth conditions, consistent with this model. We also mapped ribosome density along the ORF of several mRNAs, to probe basic kinetic properties of translational steps in yeast. We found no detectable decline in ribosome density between the 5′ and 3′ ends of the ORFs, suggesting that the average processivity of elongation is very high. Conversely, there was no queue of ribosomes at the termination site, suggesting that termination is not very slow relative to elongation and initiation. Finally, the RDM results suggest that less frequent initiation of translation on mRNAs with longer ORFs is responsible for the inverse correlation between ORF length and ribosomal density that we observed in a global analysis of translation. These results provide new insights into eukaryotic translation in vivo.