Solution structure of the core SMN-Gemin2 complex

In humans, assembly of spliceosomal snRNPs (small nuclear ribonucleoproteins) begins in the cytoplasm where the multi-protein SMN (survival of motor neuron) complex mediates the formation of a seven-membered ring of Sm proteins on to a conserved site of the snRNA (small nuclear RNA). The SMN complex contains the SMN protein Gemin2 and several additional Gemins that participate in snRNP biosynthesis. SMN was first identified as the product of a gene found to be deleted or mutated in patients with the neurodegenerative disease SMA (spinal muscular atrophy), the leading genetic cause of infant mortality. In the present study, we report the solution structure of Gemin2 bound to the Gemin2-binding domain of SMN determined by NMR spectroscopy. This complex reveals the structure of Gemin2, how Gemin2 binds to SMN and the roles of conserved SMN residues near the binding interface. Surprisingly, several conserved SMN residues, including the sites of two SMA patient mutations, are not required for binding to Gemin2. Instead, they form a conserved SMN/Gemin2 surface that may be functionally important for snRNP assembly. The SMN-Gemin2 structure explains how Gemin2 is stabilized by SMN and establishes a framework for structure-function studies to investigate snRNP biogenesis as well as biological processes involving Gemin2 that do not involve snRNP assembly.     

Land Use and Land Cover Change Dynamics across the Brazilian Amazon: Insights from Extensive Time-Series Analysis of Remote Sensing Data

Throughout the Amazon region, the age of forests regenerating on previously deforested land is determined, in part, by the periods of active land use prior to abandonment and the frequency of reclearance of regrowth, both of which can be quantified by comparing time-series of Landsat sensor data. Using these time-series of near annual data from 1973–2011 for an area north of Manaus (in Amazonas state), from 1984–2010 for south of Santarém (Pará state) and 1984–2011 near Machadinho d’Oeste (Rondônia state), the changes in the area of primary forest, non-forest and secondary forest were documented from which the age of regenerating forests, periods of active land use and the frequency of forest reclearance were derived. At Manaus, and at the end of the time-series, over 50% of regenerating forests were older than 16 years, whilst at Santarém and Machadinho d’Oeste, 57% and 41% of forests respectively were aged 6–15 years, with the remainder being mostly younger forests. These differences were attributed to the time since deforestation commenced but also the greater frequencies of reclearance of forests at the latter two sites with short periods of use in the intervening periods. The majority of clearance for agriculture was also found outside of protected areas. The study suggested that a) the history of clearance and land use should be taken into account when protecting deforested land for the purpose of restoring both tree species diversity and biomass through natural regeneration and b) a greater proportion of the forested landscape should be placed under protection, including areas of regrowth.     

The stimulatory effect of chronic lithium treatment on basal thyrotropin secretion in rats: In vivo antagonism by methylparaben

Chronic treatment of rats with lithium chloride was examined in order to determine its effect on hypothalamic monoamine and metabolite content, basal thyrotropin (TSH) secretion and thyroid function. The hypothalamic concentrations of noradrenaline (NA), dopamine (DA) and its metabolites, dihydroxyphenylacetic acid. (DOPAC) and homovanillic acid (HVA) in the lithium treated rats remained unaltered when compared to control levels. NA turnover and the NA metabolite, 3-methoxy-4-hydroxyphenylglycol (total MHPG), were significantly lower (p<0.01), whereas both serotonin (5-HT) and its metabolite, 5-hydroxyindole-3-acetic acid (5-HIAA), were significantly higher (p<0.01 and p<0.02, respectively) in the lithium treated rat hypothalami than in controls. Chronic lithium treatment significantly elevated basal TSH levels (p<0.05). This effect was antagonized by methylp-hydroxybenzoate (methylparaben, p<0.01), which did not itself affect basal TSH levels. Free serum T₃ and T₄ levels were not significantly affected by chronic lithium treatment, although T₄ tended to be slightly lower than control levels. The monoamine changes observed in the hypothalamus of lithium treated rats did not appear to account for the elevated TSH levels observed in these rats since NA activity which is generally regarded as stimulatory was decreased and 5-HT which has an inhibitory effect on TSH secretion, was increased. The elevated TSH levels may have been due to a reduced negative feedback inhibition of TSH release by the mildly reduced circulating T₄ levels caused by chronic lithium treatment. A further possibility is that the pituitary cGMP (and hence TSH) response to TRH may have been enhanced by chronic lithium treatment and methylparaben may have antagonized this effect.     

Auto-align - multi-modality fluorescence microscopy image co-registration

Multi-modality microscopes incorporate multiple microscopy techniques into one module, imaging through a common objective lens. Simultaneous or consecutive image acquisition of a single specimen, using multiple techniques, increases the amount of measurable information available. In order to benefit from each modality, it is necessary to accurately co-register data sets. Intrinsic differences in the image formation process employed by each modality result in images which possess different characteristics. In addition, as a result of using different measurement devices, images often differ in size and can suffer relative geometrical deformations including rotation, scale and translation, making registration a complex problem. Current methods generally rely on manual input and are therefore subject to human error. Here, we present an automated image registration tool for fluorescence microscopy. We show that it successfully registers images obtained via total internal reflection fluorescence (TIRF), or epi-fluorescence, and confocal microscopy. Furthermore, we provide several other applications including channel merging following image acquisition through an emission beam splitter, and lateral stage drift correction. We also discuss areas of membrane trafficking which could benefit from application of Auto-Align. Auto-Align is an essential item in the advanced microscopist's toolbox which can create a synergy of single or multi-modality image data.     

A simple rapid immunoassay for S-adenosylhomocysteine in plasma

The measurement of plasma S-adenosylhomocysteine is a more sensitive indicator of the risk for vascular disease than is plasma homocysteine. Because the level of S-adenosylhomocysteine is normally in the nanomolar range, it has been difficult to measure and necessitated the development of complex fluorometric and mass-spectrophotometric methods. We have now adapted an existing immunoassay used for the measurement of homocysteine to the measurement of S-adenosylhomocysteine in plasma. This assay is sensitive down to the level of less than 0.1 pmol, and there is no interference by S-adenosylmethionine. The assay is carried out in microplates, allows the measurement of 12 samples per plate and can easily be carried out in a 4-h period. The method is applicable to plasma samples having S-adenosylhomocysteine concentrations ranging from 10 to 150 nM without dilution. The mean value for 16 normal subjects by this method was 18.9±1.4 nM (S.E.M.), compared with 17.8±1.4 nM obtained by a previously described method using two high-performance liquid chromatography columns with fluorescence derivatization. Mean values for seven cirrhotic patients were 46.5±3.3 nM by this new method compared with 44.6±5.3 by the former method. The ease and speed of this method should allow the widespread measurement of this important metabolite in laboratories without access to sophisticated equipment.     

Identifying decomposition products in extracts of cellular metabolites

Most methods of analyzing intracellular metabolites require extraction of metabolites from the cells. A concern in these methods is underestimation of metabolite levels due to incomplete extraction. In comparing extraction methods, then, it would seem that the best method for extracting a particular metabolite is the one that gives the largest yield. In extracting Escherichia coli with different methanol:water mixtures, we observed that >or=50% water gave an increased yield of nucleosides and bases compared with 相似文献