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229 records · Page 13

Furrow microtubules and localized exocytosis in cleaving Xenopus laevis embryos

In dividing Xenopus eggs, furrowing is accompanied by expansion of a new domain of plasma membrane in the cleavage plane. The source of the new membrane is known to include a store of oogenetically produced exocytotic vesicles, but the site where their exocytosis occurs has not been described. Previous work revealed a V-shaped array of microtubule bundles at the base of advancing furrows. Cold shock or exposure to nocodazole halted expansion of the new membrane domain, which suggests that these microtubules are involved in the localized exocytosis. In the present report, scanning electron microscopy revealed collections of pits or craters, up to approximately 1.5 micro m in diameter. These pits are evidently fusion pores at sites of recent exocytosis, clustered in the immediate vicinity of the deepening furrow base and therefore near the furrow microtubules. Confocal microscopy near the furrow base of live embryos labeled with the membrane dye FM1-43 captured time-lapse sequences of individual exocytotic events in which irregular patches of approximately 20 micro m(2) of unlabeled membrane abruptly displaced pre-existing FM1-43-labeled surface. In some cases, stable fusion pores, approximately 2 micro m in diameter, were seen at the surface for up to several minutes before suddenly delivering patches of unlabeled membrane. To test whether the presence of furrow microtubule bundles near the surface plays a role in directing or concentrating this localized exocytosis, membrane expansion was examined in embryos exposed to D(2)O to induce formation of microtubule monasters randomly under the surface. D(2)O treatment resulted in a rapid, uniform expansion of the egg surface via random, ectopic exocytosis of vesicles. This D(2)O-induced membrane expansion was completely blocked with nocodazole, indicating that the ectopic exocytosis was microtubule-dependent. Results indicate that exocytotic vesicles are present throughout the egg subcortex, and that the presence of microtubules near the surface is sufficient to mobilize them for exocytosis at the end of the cell cycle.

Non-NASA Center↗

Artificial meteor ablation studies - Olivine

Artificial meteor ablation has been performed on a Mg-rich olivine sample by means of an arc-heated plasma of ionized air. Experimental conditions simulated a meteor traveling about 12 km/sec at an altitude of 70 km. The mineral content of the original olivine was 98% olivine (including traces of olivine alteration products) and 2% chromite. The forsterite content of the original olivine was Fo 89 and increased to Fo 94 in the recrystallized olivine after ablation. In addition, lamella-like intergrowths of magnetite were prevalent. Wherever magnetite occurred, Mg increased and Fe decreased in the recrystallized olivine. The Fe for the magnetite exsolved from the original olivine crystals. Individual particles (i.e., spherules) were also characterized by magnetite intergrowths. Fusion crusts on the Allende and Murchison meteorites were compared with the fusion crust on the olivine sample. The Allende fusion crust consisted of a recrystallized olivine, richer in Mg and deficient in Fe in comparison with the original meteorite's olivine, and abundant magnetite grains.

Blanchard, M. B.↗

Subcellular targeting of nine calcium-dependent protein kinase isoforms from Arabidopsis

Calcium-dependent protein kinases (CDPKs) are specific to plants and some protists. Their activation by calcium makes them important switches for the transduction of intracellular calcium signals. Here, we identify the subcellular targeting potentials for nine CDPK isoforms from Arabidopsis, as determined by expression of green fluorescent protein (GFP) fusions in transgenic plants. Subcellular locations were determined by fluorescence microscopy in cells near the root tip. Isoforms AtCPK3-GFP and AtCPK4-GFP showed a nuclear and cytosolic distribution similar to that of free GFP. Membrane fractionation experiments confirmed that these isoforms were primarily soluble. A membrane association was observed for AtCPKs 1, 7, 8, 9, 16, 21, and 28, based on imaging and membrane fractionation experiments. This correlates with the presence of potential N-terminal acylation sites, consistent with acylation as an important factor in membrane association. All but one of the membrane-associated isoforms targeted exclusively to the plasma membrane. The exception was AtCPK1-GFP, which targeted to peroxisomes, as determined by covisualization with a peroxisome marker. Peroxisome targeting of AtCPK1-GFP was disrupted by a deletion of two potential N-terminal acylation sites. The observation of a peroxisome-located CDPK suggests a mechanism for calcium regulation of peroxisomal functions involved in oxidative stress and lipid metabolism.

Non-NASA Center↗

Changing patterns of localization of the tobacco mosaic virus movement protein and replicase to the endoplasmic reticulum and microtubules during infection

Tobacco mosaic virus (TMV) derivatives that encode movement protein (MP) as a fusion to the green fluorescent protein (MP:GFP) were used in combination with antibody staining to identify host cell components to which MP and replicase accumulate in cells of infected Nicotiana benthamiana leaves and in infected BY-2 protoplasts. MP:GFP and replicase colocalized to the endoplasmic reticulum (ER; especially the cortical ER) and were present in large, irregularly shaped, ER-derived structures that may represent "viral factories." The ER-derived structures required an intact cytoskeleton, and microtubules appeared to redistribute MP:GFP from these sites during late stages of infection. In leaves, MP:GFP accumulated in plasmodesmata, whereas in protoplasts, the MP:GFP was targeted to distinct, punctate sites near the plasma membrane. Treating protoplasts with cytochalasin D and brefeldin A at the time of inoculation prevented the accumulation of MP:GFP at these sites. It is proposed that the punctate sites anchor the cortical ER to plasma membrane and are related to sites at which plasmodesmata form in walled cells. Hairlike structures containing MP:GFP appeared on the surface of some of the infected protoplasts and are reminiscent of similar structures induced by other plant viruses. We present a model that postulates the role of the ER and cytoskeleton in targeting the MP and viral ribonucleoprotein from sites of virus synthesis to the plasmodesmata through which infection is spread.

NASA Discipline Plant Biology↗

The Vasimr Engine: Project Status and Recent Accomplishments

The development of the Variable Specific Impulse Magnetoplasma Rocket (VASIMR) was initiated in the late 1970s to address a critical requirement for fast, high-power interplanetary space transportation. While not being a fusion rocket, it nevertheless borrows heavily from that technology and takes advantage of the natural topology of open-ended magnetic systems. In addition to its high power density and high exhaust velocity, VASIMR is capable of "constant power throttling" a feature, which allows in-flight mission-optimization of thrust and specific impulse to enhance performance and reduce trip time. A NASA-led, research team, involving industry, academia and government facilities is pursuing the development of this concept in the United States. The technology can be validated, in the near term, in venues such as the International Space Station, where it can also serve as both a drag compensation device and a plasma contactor for the orbital facility. Other near-Earth applications in the commercial and scientific satellite sectors are also envisioned. This presentation covers the evolution of the VASIMR concept to its present status, as well as recent accomplishments in our understanding of the physics. Approaches and collaborative programs addressing the major technical challenges will also be presented.

ChangDiaz, Franklin R.↗

Automation of preparation of nonmetallic samples for analysis by atomic absorption and inductively coupled plasma spectrometry

For a rapid preparation of solutions intended for analysis by inductively coupled plasma emission spectrometry or atomic absorption spectrometry, an automatic device called Plasmasol was developed. This apparatus used the property of nonwettability of glassy C to fuse the sample in an appropriate flux. The sample-flux mixture is placed in a composite crucible, then heated at high temperature, swirled until full dissolution is achieved, and then poured into a water-filled beaker. After acid addition, dissolution of the melt, and filling to the mark, the solution is ready for analysis. The analytical results obtained, either for oxide samples or for prereduced iron ores show that the solutions prepared with this device are undistinguished from those obtained by manual dissolutions done by acid digestion or by high temperature fusion. Preparation reproducibility and analytical tests illustrate the performance of Plasmasol.

Wittmann, A.↗

The Arabidopsis SKU5 gene encodes an extracellular glycosyl phosphatidylinositol-anchored glycoprotein involved in directional root growth

To investigate how roots respond to directional cues, we characterized a T-DNA-tagged Arabidopsis mutant named sku5 in which the roots skewed and looped away from the normal downward direction of growth on inclined agar surfaces. sku5 roots and etiolated hypocotyls were slightly shorter than normal and exhibited a counterclockwise (left-handed) axial rotation bias. The surface-dependent skewing phenotype disappeared when the roots penetrated the agar surface, but the axial rotation defect persisted, revealing that these two directional growth processes are separable. The SKU5 gene belongs to a 19-member gene family designated SKS (SKU5 Similar) that is related structurally to the multiple-copper oxidases ascorbate oxidase and laccase. However, the SKS proteins lack several of the conserved copper binding motifs characteristic of copper oxidases, and no enzymatic function could be assigned to the SKU5 protein. Analysis of plants expressing SKU5 reporter constructs and protein gel blot analysis showed that SKU5 was expressed most strongly in expanding tissues. SKU5 was glycosylated and modified by glycosyl phosphatidylinositol and localized to both the plasma membrane and the cell wall. Our observations suggest that SKU5 affects two directional growth processes, possibly by participating in cell wall expansion.

NASA Discipline Plant Biology↗

A Spherical Torus Nuclear Fusion Reactor Space Propulsion Vehicle Concept for Fast Interplanetary Travel

A conceptual vehicle design enabling fast outer solar system travel was produced predicated on a small aspect ratio spherical torus nuclear fusion reactor. Initial requirements were for a human mission to Saturn with a greater than 5% payload mass fraction and a one way trip time of less than one year. Analysis revealed that the vehicle could deliver a 108 mt crew habitat payload to Saturn rendezvous in 235 days, with an initial mass in low Earth orbit of 2,941 mt. Engineering conceptual design, analysis, and assessment was performed on all ma or systems including payload, central truss, nuclear reactor (including divertor and fuel injector), power conversion (including turbine, compressor, alternator, radiator, recuperator, and conditioning), magnetic nozzle, neutral beam injector, tankage, start/re-start reactor and battery, refrigeration, communications, reaction control, and in-space operations. Detailed assessment was done on reactor operations, including plasma characteristics, power balance, power utilization, and component design.

Williams, Craig H.↗

Composite wall concept for high temperature turbine shrouds: Survey of low modulus strain isolator materials

Plasma sprayed yttria stabilized zirconium oxide turbine seal specimens, incorporating various low modulus porous metal strain isolator pads between the zirconium oxide and a dense metal substrate, were subjected to cyclic thermal shock testing. Specimens that had a low modulus pad composed of sintered FeNiCrAlY fibermetal survived 1000 thermal shock cycles without spalling of the ceramic. A figure of merit for the low modulus pad materials taking into consideration the elastic modulus, thermal conductivity, strength, and oxidation resistance of the pad was proposed, and showed reasonable agreement with the thermal shock results. A potential surface distress problem on the zirconium oxide, associated with nonuniform temperature distribution and rapid stress relaxation was identified. One approach to solving the surface distress problem through application of laser surface fusion of the zirconium oxide layer showed some promise, but improvements in the laser surface fusion process are necessary to prevent process associated damage to the ceramic.

Bill, R. C.↗

Laser-assisted patch clamping: a methodology

Laser microsurgery can be used to perform both cell biological manipulations, such as targeted cell ablation, and molecular genetic manipulations, such as genetic transformation and chromosome dissection. In this report, we describe a laser microsurgical method that can be used either to ablate single cells or to ablate a small area (1-3 microns diameter) of the extracellular matrix. In plants and microorganisms, the extracellular matrix consists of the cell wall. While conventional patch clamping of these cells, as well as of many animal cells, requires enzymatic digestion of the extracellular matrix, we illustrate that laser microsurgery of a portion of the wall enables patch clamp access to the plasma membrane of higher plant cells remaining situated in their tissue environment. What follows is a detailed description of the construction and use of an economical laser microsurgery system, including procedures for single cell and targeted cell wall ablation. This methodology will be of interest to scientists wishing to perform cellular or subcellular ablation with a high degree of accuracy, or wishing to study how the extracellular matrix affects ion channel function.

Non-NASA Center↗

Development of Ceramic Fibers for Reinforcement in Composite Materials

Refinements of the vertical arc fiberizing apparatus resulted in its ability to fiberize very different refractory glasses having wide ranges of properties. Although the apparatus, was originally designed as a laboratory research tool for the evaluation of many compositions daily, up to one quarter pound of fibers of a single composition could be produced in an 8-hour day. Fibers up to six and a half feet long were produced with the apparatus. Studies were conducted of two methods of fiberizing refractory glasses requiring rapid freezing from the melt. The first method consisted of fiberizing droplets of molten glass passing through an annular nozzle. The second method consisted of reconstructing the annular nozzle in. the shape of a horseshoe to achieve a shorter delay in blasting a molten droplet from the tip of a rod. Both methods were judged feasible for producing fibers of glasses requiring rapid freezing. The first method would be more amenable to volume fiber production. Studies of induction heating for fiber formation did not lead to its designation as a very efficient heating method. Problems. remain to be solved, in the design of a suitable susceptor for a higher heating rate, in protecting the susceptor from oxidation with an inert gas, in contamination of the melt from a refractory crucible, and in the protective radiation shielding of the induction concentrator coil. It is not considered practical to continue studies of this heating method. In the course of this program 151 refractory glass compositions were evaluated for fiber, forming characteristics. Of the various types of materials studied, the following showed promise in producing acceptable refractory fibers: sIlica- spinel (magnesium aluminate), silica- spinel-zirconia, silica-zirconia, silica-zinc spinel, aluminum phosphate glasses, and fluoride glasses. Compositions which did not produce acceptable fibers were high zirconia materials, barium spinels, and calcium aluminates. Improvements in the. testing apparatus for single fiber tensile strength increased the precision. of tests conducted on nine fibers. The highest mean tensile strength, a value of 295,000 pounds per square inch, was obtained with R-141 fibers. Treatment of R-74 fibers with anhydrous Linde A-1100 silane finish improved its mean fiber tensile strength by 25 percent. The lapse of time after fiber formation had no measurable effect on tensile strength. A static heating test conducted with various high melting fibers indicated that Fiberfrax and R-108 underwent no significant changes in bulk volume or resiliency on exposure to 2750 degrees Fahrenheit (1510 degrees Centigrade) in an oxidizing atmosphere. For fiber-resin composition fabrication, ten fiber materials were selected on the bases of high fiber yield, fusion temperature, and type of composition. Fiberfrax, a commercial ceramic fiber, was included for comparison. A new, more effective method of removing pellets from blown fibers was developed. The de-pelletized fibers were treated with a silane finish and felted into ten-inch diameter felts prior to resin impregnation. Composites containing 30 percent by weight of CTL 91-LD phenolic resin were molded under high pressure from the impregnated felts and post-cured to achieve optimum properties. Flexural strength, flexural modules of elasticity, and punch shear strength tests were conducted on the composite specimens. The highest average flexural strength obtained was 19,958 pounds per square inch with the R-74-fiber-resin composite. This compares very favorably with the military specification of 13,000 pounds per square inch flexural strength for randomly oriented fiber reinforced composites. The highest punch shear strength (11,509 pounds per square inch) was obtained with the R-89 fiber-resin composite. The effects of anhydrous fiber finishes on composite strength were not clearly indicated. Plasma arc tests at a heat flux of 550 British Thermal Units per square foot per second on eight composite materials indicated ablation rates generally equivalent to Fiberglas-Micarta No. 259-2. The composite reinforced with R-99 fibers had an average ablation rate of 0.008 inch per second and appears quite promising on the basis of these tests. Preliminary studies for processing fibers into yarn and fabric were conducted with R-99 fibers. The use of certain organic gums aided in 2 fabricating by hand several relatively strong yarns and a crude fabric swatch. This indicated the practicality of developing techniques for processing these fibers into yarn and fabric without significant damage to the fibers.

Gates, L. E.↗

Advanced Space Fission Propulsion Systems

Fission has been considered for in-space propulsion since the 1940s. Nuclear Thermal Propulsion (NTP) systems underwent extensive development from 1955-1973, completing 20 full power ground tests and achieving specific impulses nearly twice that of the best chemical propulsion systems. Space fission power systems (which may eventually enable Nuclear Electric Propulsion) have been flown in space by both the United States and the Former Soviet Union. Fission is the most developed and understood of the nuclear propulsion options (e.g. fission, fusion, antimatter, etc.), and fission has enjoyed tremendous terrestrial success for nearly 7 decades. Current space nuclear research and technology efforts are focused on devising and developing first generation systems that are safe, reliable and affordable. For propulsion, the focus is on nuclear thermal rockets that build on technologies and systems developed and tested under the Rover/NERVA and related programs from the Apollo era. NTP Affordability is achieved through use of previously developed fuels and materials, modern analytical techniques and test strategies, and development of a small engine for ground and flight technology demonstration. Initial NTP systems will be capable of achieving an Isp of ~900 s at a relatively high thrust-to-weight ratio. The development and use of first generation space fission power and propulsion systems will provide new, game changing capabilities for NASA. In addition, development and use of these systems will provide the foundation for developing extremely advanced power and propulsion systems capable of routinely and affordably accessing any point in the solar system. The energy density of fissile fuel (8 x 10(exp 13) Joules/kg) is more than adequate for enabling extensive exploration and utilization of the solar system. For space fission propulsion systems, the key is converting the virtually unlimited energy of fission into thrust at the desired specific impulse and thrust-to-weight ratio. This presentation will discuss potential space fission propulsion options ranging from first generation systems to highly advanced systems. Ongoing research that shows promise for enabling second generation NTP systems with Isp greater than 1000 s will be discussed, as will the potential for liquid, gas, or plasma core systems. Space fission propulsion systems could also be used in conjunction with simple (water-based) propellant depots to enable routine, affordable missions to various destinations (e.g. moon, Mars, asteroids) once in-space infrastructure is sufficiently developed. As fuel and material technologies advance, very high performance Nuclear Electric Propulsion (NEP) systems may also become viable. These systems could enable sophisticated science missions, highly efficient cargo delivery, and human missions to numerous destinations. Commonalities between NTP, fission power systems, and NEP will be discussed.

Houts, Michael G.↗

Caveolin-1 regulates shear stress-dependent activation of extracellular signal-regulated kinase

Fluid shear stress activates a member of the mitogen-activated protein (MAP) kinase family, extracellular signal-regulated kinase (ERK), by mechanisms dependent on cholesterol in the plasma membrane in bovine aortic endothelial cells (BAEC). Caveolae are microdomains of the plasma membrane that are enriched with cholesterol, caveolin, and signaling molecules. We hypothesized that caveolin-1 regulates shear activation of ERK. Because caveolin-1 is not exposed to the outside, cells were minimally permeabilized by Triton X-100 (0.01%) to deliver a neutralizing, polyclonal caveolin-1 antibody (pCav-1) inside the cells. pCav-1 then bound to caveolin-1 and inhibited shear activation of ERK but not c-Jun NH(2)-terminal kinase. Epitope mapping studies showed that pCav-1 binds to caveolin-1 at two regions (residues 1-21 and 61-101). When the recombinant proteins containing the epitopes fused to glutathione-S-transferase (GST-Cav(1-21) or GST-Cav(61-101)) were preincubated with pCav-1, only GST-Cav(61-101) reversed the inhibitory effect of the antibody on shear activation of ERK. Other antibodies, including m2234, which binds to caveolin-1 residues 1-21, had no effect on shear activation of ERK. Caveolin-1 residues 61-101 contain the scaffolding and oligomerization domains, suggesting that binding of pCav-1 to these regions likely disrupts the clustering of caveolin-1 or its interaction with signaling molecules involved in the shear-sensitive ERK pathway. We suggest that caveolae-like domains play a critical role in the mechanosensing and/or mechanosignal transduction of the ERK pathway.

Non-NASA Center↗