Pharmacological analysis of nod factor-induced calcium spiking in Medicago truncatula. Evidence for the requirement of type IIA calcium pumps and phosphoinositide signaling

Bacterial Nod factors trigger a number of cellular responses in root hairs of compatible legume hosts, which include periodic, transient increases in cytosolic calcium levels, termed calcium spiking. We screened 13 pharmaceutical modulators of eukaryotic signal transduction for effects on Nod factor-induced calcium spiking. The purpose of this screening was 2-fold: to implicate enzymes required for Nod factor-induced calcium spiking in Medicago sp., and to identify inhibitors of calcium spiking suitable for correlating calcium spiking to other Nod factor responses to begin to understand the function of calcium spiking in Nod factor signal transduction. 2-Aminoethoxydiphenylborate, caffeine, cyclopiazonic acid (CPA), 2,5-di-(t-butyl)-1,4-hydroquinone, and U-73122 inhibit Nod factor-induced calcium spiking. CPA and U-73122 are inhibitors of plant type IIA calcium pumps and phospholipase C, respectively, and implicate the requirement for these enzymes in Nod factor-induced calcium spiking. CPA and U-73122 inhibit Nod factor-induced calcium spiking robustly at concentrations with no apparent toxicity to root hairs, making CPA and U-73122 suitable for testing whether calcium spiking is causal to subsequent Nod factor responses.     

Future role of neurologists.

Clinical neurologists in the health care system of the future should have a multifaceted role. Advances in the basic understanding of the nervous system and therapeutics of neurologic disease have created, for the first time in human history, an ethical imperative to correctly diagnose neurologic disease. In many situations, the neurologists may function as a consultant and principal physician for patients with primary nervous system disorders including Parkinson''s disease, multiple sclerosis, Alzheimer''s disease, epilepsy, migraine, cerebrovascular disease, movement disorders, and neuromuscular disease. Other important roles for neurologists include the training of future physicians, both neurologists and primary care physicians, the application of cost-effective approaches to care, and the support of health care delivery research and academic programs that link basic research efforts to the development of new therapy. To be successful, future residency training programs should include joint certification opportunities in both neurology and general medicine, and training programs for clinical investigators should be expanded. Despite its threats to specialists, managed care should also provide opportunities for new alliances among neurologists, other specialists, and primary care physicians that will both improve patient care and increase efficiency and cost-effectiveness.     

Silica fluxes and trapping in two contrasting natural impoundments of the upper Mississippi River

Human activities have altered riverine silica cycling and diminished the supply of silica to the oceans, but few rivers have been intensively monitored to evaluate the magnitude of these changes. In this study we measured dissolved silica (DSi) and amorphous silica (ASi) fluxes into and out of two large, culturally-impacted natural impoundments of the upper Mississippi River, Lakes St. Croix and Pepin, USA. ASi sedimentation rates and sediment–water fluxes of DSi were calculated for each lake, and a mass-balance approach was used to determine in-lake ASi production. ASi from terrestrial phytoliths in the lake sediments was determined to be only partially available to biotic recycling, and in-lake ASi dissolution was small relative to the total silica budgets. The river reaches upstream of the two lakes were found to have abundant DSi, and riverine diatom production was found to contribute significant amounts of ASi to each lake. The average total phosphorus concentration in Lake Pepin is four times that in Lake St. Croix but ASi production in Lake Pepin is only 2.3 times higher than in Lake St. Croix, indicating that diatom growth in Pepin is limited by factors such as turbidity. Lake St. Croix currently traps about 10% of the inflowing total bioavailable silica (TSi_b = DSi + ASi) while Lake Pepin traps closer to 20% of its inflowing TSi_b, clearly demonstrating the importance of silica retention in lakes and reservoirs along the land–ocean continuum.     

Solubilization, purification, and characterization of a truncated form of rat hepatic squalene synthetase.

Rat hepatic microsomal squalene synthetase (EC 2.5.1.21) was induced 25-fold by feeding rats with diet containing the hydroxymethylglutaryl-coenzyme A reductase inhibitor, fluvastatin, and cholestyramine, a bile acid sequestrant. A soluble squalene synthetase protein with an estimated mass of 32-35 kDa, as determined by gel filtration chromatography on Sephacryl S-200 column, was solubilized out of the microsomes by controlled proteolysis with trypsin. Approximately 25% of the activity was recovered in a soluble form. The enzyme was purified to homogeneity utilizing a series of column chromatography purification steps on DEAE-cellulose, hydroxylapatite, and phenyl-Sepharose sequentially. The purified enzyme showed a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Initial kinetic analysis indicated an S0.5 values for trans-farnesyl diphosphate of 1.0 microM and for NADPH of 40 microM. The Vmax with respect to trans-farnesyl diphosphate was calculated at 1.2 mumol/min/mg. NADH also serves as substrate for the reaction with S0.5 value of 800 microM. Western blot analysis utilizing rabbit antisera raised against the purified, trypsin-truncated enzyme showed a single band for the isolated solubilized enzyme at 32-33 kDa and a band for the intact microsomal enzyme at about 45-47 kDa.     

Circumpolar arctic tundra biomass and productivity dynamics in response to projected climate change and herbivory

Satellite remote sensing data have indicated a general ‘greening’ trend in the arctic tundra biome. However, the observed changes based on remote sensing are the result of multiple environmental drivers, and the effects of individual controls such as warming, herbivory, and other disturbances on changes in vegetation biomass, community structure, and ecosystem function remain unclear. We apply ArcVeg, an arctic tundra vegetation dynamics model, to estimate potential changes in vegetation biomass and net primary production (NPP) at the plant community and functional type levels. ArcVeg is driven by soil nitrogen output from the Terrestrial Ecosystem Model, existing densities of Rangifer populations, and projected summer temperature changes by the NCAR CCSM4.0 general circulation model across the Arctic. We quantified the changes in aboveground biomass and NPP resulting from (i) observed herbivory only; (ii) projected climate change only; and (iii) coupled effects of projected climate change and herbivory. We evaluated model outputs of the absolute and relative differences in biomass and NPP by country, bioclimate subzone, and floristic province. Estimated potential biomass increases resulting from temperature increase only are approximately 5% greater than the biomass modeled due to coupled warming and herbivory. Such potential increases are greater in areas currently occupied by large or dense Rangifer herds such as the Nenets‐occupied regions in Russia (27% greater vegetation increase without herbivores). In addition, herbivory modulates shifts in plant community structure caused by warming. Plant functional types such as shrubs and mosses were affected to a greater degree than other functional types by either warming or herbivory or coupled effects of the two.     

Land-use change,not climate,controls organic carbon burial in lakes

Lakes are a central component of the carbon cycle, both mineralizing terrestrially derived organic matter and storing substantial amounts of organic carbon (OC) in their sediments. However, the rates and controls on OC burial by lakes remain uncertain, as do the possible effects of future global change processes. To address these issues, we derived OC burial rates in ²¹⁰Pb-dated sediment cores from 116 small Minnesota lakes that cover major climate and land-use gradients. Rates for individual lakes presently range from 7 to 127 g C m^–2 yr^–1 and have increased by up to a factor of 8 since Euro-American settlement (mean increase: 2.8×). Mean pre-disturbance OC burial rates were similar (14–22 g C m^–2 yr^–1) across all land-cover categories (prairie, mixed deciduous and boreal forest), indicating minimal effect of the regional temperature gradient (approx. 4°C) on background carbon burial. The relationship between modern OC burial rates and temperature was also not significant after removal of the effect of total phosphorus. Contemporary burial rates were strongly correlated with lake-water nutrients and the extent of agricultural land cover in the catchment. Increased OC burial, documented even in relatively undisturbed boreal lake ecosystems, indicates a possible role for atmospheric nitrogen deposition. Our results suggest that globally, future land-cover change, intensification of agriculture and associated nutrient loading together with atmospheric N-deposition will enhance OC sequestration by lakes.     

Identification of a novel proline-arginine motif involved in CIN85-dependent clustering of Cbl and down-regulation of epidermal growth factor receptors

CIN85 is a multidomain adaptor protein implicated in Cbl-mediated down-regulation of receptor tyrosine kinases. CIN85 binding to Cbl is increased after growth factor stimulation and is critical for targeting receptor tyrosine kinases to clathrin-mediated endocytosis. Here we report the identification of a novel polyproline-arginine motif (PXXXPR), specifically recognized by the SH3 domains of CIN85 and its homologue CMS/CD2AP. This motif was indispensable for CIN85 binding to Cbl/Cbl-b, to other CIN85 SH3 domains' effectors, and for mediating an intramolecular interaction between the SH3-A domain and the proline-rich region of CIN85. Individual SH3 domains of CIN85 bound to PXXXPR peptides of Cbl/Cbl-b with micromolar affinities, whereas an extended structure of two or three SH3 domains bound with higher stoichiometry and increased affinity to the same peptides. This enabled full size CIN85 to simultaneously interact with multiple Cbl molecules, promoting their clustering in mammalian cells. The ability of CIN85 to cluster Cbl was important for ligand-induced stabilization of CIN85.Cbl.epidermal growth factor receptor complexes, as well as for epidermal growth factor receptor degradation in the lysosome. Thus, specific interactions of CIN85 SH3 domains with the PXXXPR motif in Cbl play multiple roles in down-regulation of receptor tyrosine kinases.