Water relations and leaf chemistry of Chrysothamnus nauseosus ssp. consimilis (Asteraceae) and Sarcobatus vermiculatus (Chenopodiaceae)

At Mono Lake, California, we investigated field water relations, leaf and xylem chemistry, and gas exchange for two shrub species that commonly co-occur on marginally saline soils, and have similar life histories and rooting patterns. Both species had highest root length densities close to the surface and have large tap roots that probably reach ground water at 3.4-5.0 m on the study site. The species differed greatly in leaf water relations and leaf chemistry. Sarcobatus vermiculatus had a seasonal minimum predawn xylem pressure potential (ψ_pd) of -2.7 MPa and a midday potential (ψ_md) of -4.1 MPa. These were significantly lower than for Chrysothamnus nauseosus, which had a minimum ψ_pd of -1.0 MPa and ψ_md of -2.2 MPa. Sarcobatus had leaf Na of up to 9.1 % and K up to 2.7 % of dry mass, and these were significantly higher than for Chrysothamnus which had seasonal maxima of 0.4% leaf Na and 2.4 % leaf K. The molar ratios of leaf K/Na, Ca/Na, and Mg/Na were substantially lower for Sarcobatus than for Chrysothamnus. Xylem ionic contents indicated that both species excluded some Na at the root, but that Chrysothamnus was excluding much more than Sarcobatus. The higher Na content of Sarcobatus leaves was associated with greater leaf succulence, lower calculated osmotic potential, and lower xylem pressure potentials. Despite large differences in water relations and leaf chemistry, these species maintained similar diurnal patterns and rates of photosynthesis and stomatal conductance to water vapor diffusion. Sarcobatus ψ_pd may not reflect soil moisture availability due to root osmotic and hydraulic properties.     

Minimizing the overlap problem in protein NMR: a computational framework for precision amino acid labeling

MOTIVATION: Recent advances in cell-free protein expression systems allow specific labeling of proteins with amino acids containing stable isotopes ((15)N, (13) C and (2)H), an important feature for protein structure determination by nuclear magnetic resonance (NMR) spectroscopy. Given this labeling ability, we present a mathematical optimization framework for designing a set of protein isotopomers, or labeling schedules, to reduce the congestion in the NMR spectra. The labeling schedules, which are derived by the optimization of a cost function, are tailored to a specific protein and NMR experiment. RESULTS: For 2D (15)N-(1)H HSQC experiments, we can produce an exact solution using a dynamic programming algorithm in under 2 h on a standard desktop machine. Applying the method to a standard benchmark protein, calmodulin, we are able to reduce the number of overlaps in the 500 MHz HSQC spectrum from 10 to 1 using four samples with a true cost function, and 10 to 4 if the cost function is derived from statistical estimates. On a set of 448 curated proteins from the BMRB database, we are able to reduce the relative percent congestion by 84.9% in their HSQC spectra using only four samples. Our method can be applied in a high-throughput manner on a proteomic scale using the server we developed. On a 100-node cluster, optimal schedules can be computed for every protein coded for in the human genome in less than a month. AVAILABILITY: A server for creating labeling schedules for (15)N-(1)H HSQC experiments as well as results for each of the individual 448 proteins used in the test set is available at http://nmr.proteomics.ics.uci.edu.     

Correlation between Nasal Membrane Permeability and Nasal Absorption Rate

The objective of this study was to investigate the relationship between in vitro permeability (P_app) values obtained from isolated nasal tissues and the absorption rates (k_a) of the same compounds following nasal administration in animals and humans. The P_app of a set of 11 drug compounds was measured using animal nasal explants and plasma time–concentration profiles for each of the same compounds following intravenous (IV) and intranasal (IN) administration were experimentally determined or obtained from literature reports. The plasma clearance was estimated from the IV plasma time–concentration profiles, and k_a was determined from the IN plasma time–concentration profiles using a deconvolution approach. The level of correlation between P_app and k_a was established using Pearson correlation analysis. A good correlation (r = 0.77) representing a point-to-point relationship for each of the compounds was observed. This result indicates that the nasal absorption for many drug candidates can be estimated from a readily measured in vitro P_app value.Key words: bioavailability, drug transport, nasal administration, nasal mucosa, permeability     

Dimethylsulfone as a growth substrate for novel methylotrophic species of Hyphomicrobium and Arthrobacter

Dimethylsulfone is a major product of the chemical oxidation in the atmosphere of the principal biogenic sulfur gas, dimethylsulfide, but no studies have been reported on the mechanisms for its microbiological degradation. Three novel strains of bacteria have been isolated from enrichment cultures provided with dimethylsulfone as the only carbon and energy substrate. These are novel facultatively methylotrophic species of Hyphonmicrobium and Arthobacter, capable of growth on a range of one-carbon substrates. Cell-free extracts contained activities of enzymes necessary for a reductive/oxidative pathway for dimethylsulfone degradation: membrane-bound-dimethylsulfone and dimethylsulfoxide reductases, dimethylsulfide monooxygenase, and methanethiol oxidase. Enzymatic evidence is also presented for the subsequent oxidation of formaldehyde by formaldehyde and formate dehydrogenases in the Hyphomicrobium strain and by a dissimilatory ribulose monophosphate cycle in the Arthrobacter strains. The strains also grew on dimethylsulfoxide and dimethylsulfide, and dimethylsulfide-grown bacteria oxidized dimethylsulfide and dimethylsulfoxide but not dimethylsulfone. Formaldehyde assimilation was effected in the Hyphomicrobium strain by the serine pathway, but enzymes of the ribulose monophosphate cycle for formaldehyde assimilation were present in the Arthrobacter strains grown on dimethylsulfone. In contrast, one of the Arthrobacter strains was shown to switch to the serine pathway during growth on methanol. Growth yields on dimethylsulfone and formaldehyde were consistent with the occurrence of the serine pathway in Hyphomicrobium strain S1 and the ribulose monophosphate cycle in Arthrobacter strain TGA, and with the proposed reductive pathway for dimethylsulfone degradation in both.     

Predawn disequilibrium between plant and soil water potentials in two cold-desert shrubs

Classical water relations theory predicts that predawn plant water potential should be in equilibrium with soil water potential (soil Ψ_w) around roots, and many interpretations of plant water status in natural populations are based on this expectation. We examined this expectation for two salt-tolerant, cold-desert shrub species in glasshouse experiments where frequent watering assured homogeneity in soil Ψ_w and soil-root hydraulic continuity and where NaCl controlled soil Ψ_w. Plant water potentials were measured with a pressure chamber (xylem Ψ_p) and thermocouple psychrometers (leaf Ψ_w). Soil Ψ_w was measured with in situ thermocouple psychrometers. Predawn leaf Ψ_w and xylem Ψ_p were significantly more negative than soil Ψ_w, for many treatments, indicating large predawn soil-plant Ψ_w disequilibria: up to 1.2 MPa for Chrysothamnus nauseosus (0 and 100 mm NaCl) and 1.8 MPa for Sarcobatus vermiculatus (0, 100, 300, and 600 mm NaCl). Significant nighttime canopy water loss was one mechanism contributing to predawn disequilibrium, assessed by comparison of xylem Ψ_p for bagged (to minimize transpiration) and unbagged canopies, and by gas exchange measurements. However, nighttime transpiration accounted for only part of the predawn disequilibrium. Other mechanisms that could act with nighttime transpiration to generate large predawn disequilibria are described and include a model of how leaf apoplastic solutes could contribute to the phenomenon. This study is among the first to conclusively document such large departures from the expectation of predawn soil-plant equilibrium for C₃ shrubs, and provides a general framework for considering relative contributions of nighttime transpiration and other plant-related mechanisms to predawn disequilibrium. Received: 12 November 1998 / Accepted: 5 May 1999     

Differences between Belclare and Suffolk ewes in fertilization rate, embryo quality and accessory sperm number after cervical or laparoscopic artificial insemination

Ewe breed has been shown to have a major effect on pregnancy rates following cervical AI using frozen-thawed semen. The main objective of this study was to examine the differences between purebred Belclare and Suffolk ewes (multiparous) in fertilization rate, number of accessory sperm and stage of embryo development on day 6 after cervical or laparoscopic AI with frozen-thawed semen. In experiment 1, Belclare and Suffolk ewes were synchronized for 12 days and were either cervically inseminated (year 1: n=28 and 31; year 2: n=16 and 15, respectively) or laparoscopically inseminated (year 2: n=13 and 14). In experiment 2, superovulated Belclare (n=4) and Suffolk (n=13) ewes were laparoscopically inseminated. All ewes were slaughtered 6 days after AI; oocytes/embryos were recovered, morphologically graded and stained to assess the number of cells and accessory spermatozoa. Data from both experiments were combined for statistical analysis. The proportion of ewes with fertilized oocytes was significantly higher following laparoscopic AI compared with cervical AI (54% versus 19%). More Belclare than Suffolk ewes yielded fertilized oocyte(s) after cervical AI (34% versus 10%, P<0.02) but there was no difference after laparoscopic AI (62% versus 60%). From the ewes that yielded at least one fertilized oocyte the proportion of Belclare ewes with embryos at the morula/blastocyst stage was significantly greater than for Suffolk ewes (94% versus 59%, P<0.02). A higher proportion of Belclare than Suffolk ewes had evidence of sperm reaching the site of fertilization following cervical AI (39% versus 15%, P<0.02) but there was no difference after laparoscopic AI (62% versus 64%, P>0.8). Amongst the ewes with evidence of sperm at the site of fertilization, laparoscopic AI resulted in a higher number of sperm per oocyte/embryo or per ewe than cervical AI (P<0.01). These results suggested that the difference in pregnancy rate between Suffolk and Belclare ewes following cervical AI was due to: (i) sperm traversing the cervix and uterus in a higher proportion of Belclare than Suffolk ewes, leading to a higher incidence of fertilization and (ii) the lower developmental competence of fertilized oocytes from Suffolk ewes.     

Genetics of psychosis; insights from views across the genome

The major psychotic illnesses, schizophrenia and bipolar disorder (BD), are among the most heritable common disorders, but finding specific susceptibility genes for them has not been straightforward. The reasons are widely assumed to include lack of valid phenotypic definition, absence of good theories of pathophysiology for candidate gene studies, and the involvement of many genes, each making small contributions to population risk. Within the last year or so, a number of genome wide association (GWAS) of schizophrenia and BD have been published. These have produced stronger evidence for association to specific risk loci than have earlier studies, specifically for the zinc finger binding protein 804A (ZNF804A) locus in schizophrenia and for the calcium channel, voltage-dependent, L type, alpha 1C subunit (CACNA1C) and ankyrin 3, node of Ranvier (ANK3) loci in bipolar disorder. The ZNF804A and CACNA1C loci appear to influence risk for both disorders, a finding that supports the hypothesis that schizophrenia and BD are not aetiologically distinct. In the case of schizophrenia, a number of rare copy number variants have also been detected that have fairly large effect sizes on disease risk, and that additionally influence risk of autism, mental retardation, and other neurodevelopmental disorders. The existing findings point to some likely pathophysiological mechanisms but also challenge current concepts of disease classification. They also provide grounds for optimism that larger studies will reveal more about the origins of these disorders, although currently, very little of the genetic risk of either disorder is explained.     

Differential scanning calorimetry of asparate transcarbamoylase and its isolate subunits.

The thermal denaturation of aspartate transcarbamoylas of Escherichia coli was investigated by differential scanning calorimetry. Isolated regulatory and catalytic subunits were heat denatured at 55 and 80 degrees C, respectively. In contrast, the intact enzyme was denatured in two steps. A small endotherm near 73 degrees C was assoicated with denaturation of the regulatory subunits and the major endotherm at 82 degrees C with denaturation of the catalytic subunits. Thus regulatory subunits are stabilized against heat denaturation by more than 17 degrees C when incorporated in the enzyme. Similar conclusions were obtained from measurements of the enthalpy of heat denaturation. Regulatory subunits yielded a much lower value of the enthalpy of denaturation, 1.91 cal/g, than that found for the catalytic subunit, 3.94 cal/g, or typical globular proteins (4 to 6 cal/g). When the regulatory subunits were incorporated into aspartate transcarbamoylase their enthalpy of denaturation was increased 125% (to 4.3 cal/g). The enthalpy of the catalytic subunits in the intact enzyme was increased 38% (enthalpy of denaturation of 5.43 cal/g). Stabilization of the isolated catalytic subunit as well as the intact enzyme was achieved by the addition of the bisubstrate analog N-(phosphonacetyl)-L-aspartate. Similarly the allosteric effectors, CTP and ATP, stabilized the isolated regulatory subunits or those subunits within the intact enzyme. However, the addition of the bisubstrate analog caused a decrease in the enthalpy of denaturation of the regulatory subunits within the enzyme. These results are consistent with other studies of the ligand-promoted conformational changes in the native enzyme.     

Synthesis and evaluation of 7-chloro-4-(piperazin-1-yl)quinoline-sulfonamide as hybrid antiprotozoal agents

A new series of 4-aminochloroquinoline based sulfonamides were synthesized and evaluated for antiamoebic and antimalarial activities. Out of the eleven compounds evaluated (F1–F11), two of them (F3 and F10) showed good activity against Entamoeba histolytica (IC₅₀ <5 μM). Three of the compounds (F5, F7 and F8) also displayed antimalarial activity against the chloroquine-resistant (FCR-3) strain of Plasmodium falciparum with IC₅₀ values of 2 μM. Compound F7, whose crystal structure was also determined, inhibited β-haematin formation more potently than quinine. To further understand the action of hybrid molecules F7 and F8, molecular docking was carried out against the homology model of P. falciparum enzyme dihydropteroate synthase (PfDHPS). The complexes showed that the inhibitors place themselves nicely into the active site of the enzyme and exhibit interaction energy which is in accordance with our activity profile data. Application of Lipinski ‘rule of five’ on all the compounds (F1–F11) suggested high drug likeness of F7 and F8, similar to quinine.     

No Evidence for Enrichment in Schizophrenia for Common Allelic Associations at Imprinted Loci

Most genetic studies assume that the function of a genetic variant is independent of the parent from which it is inherited, but this is not always true. The best known example of parent-of-origin effects arises with respect to alleles at imprinted loci. In classical imprinting, characteristically, either the maternal or paternal copy is expressed, but not both. Only alleles present in one of the parental copies of the gene, the expressed copy, is likely to contribute to disease. It has been postulated that imprinting is important in central nervous system development, and that consequently, imprinted loci may be involved in schizophrenia. If this is true, allowing for parent-of-origin effects might be important in genetic studies of schizophrenia. Here, we use genome-wide association data from one of the world’s largest samples (N = 695) of parent schizophrenia-offspring trios to test for parent-of-origin effects. To maximise power, we restricted our analyses to test two main hypotheses. If imprinting plays a disproportionate role in schizophrenia susceptibility, we postulated a) that alleles showing robust evidence for association to schizophrenia from previous genome-wide association studies should be enriched for parent-of-origin effects and b) that genes at loci imprinted in humans or mice should be enriched both for genome-wide significant associations, and in our sample, for parent-of-origin effects. Neither prediction was supported in the present study. We have shown, that it is unlikely that parent-of-origin effects or imprinting play particularly important roles in schizophrenia, although our findings do not exclude such effects at specific loci nor do they exclude such effects among rare alleles.