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Hepatocytes metabolize energy-rich cytoplasmic lipid droplets (LDs) in the lysosome-directed process of autophagy. An organelle-selective form of this process (macrolipophagy) results in the engulfment of LDs within double-membrane delimited structures (autophagosomes) before lysosomal fusion. Whether this is an exclusive autophagic mechanism used by hepatocytes to catabolize LDs is unclear. It is also unknown whether lysosomes alone might be sufficient to mediate LD turnover in the absence of an autophagosomal intermediate. We performed live-cell microscopy of hepatocytes to monitor the dynamic interactions between lysosomes and LDs in real-time. We additionally used a fluorescent variant of the LD-specific protein (PLIN2) that exhibits altered fluorescence in response to LD interactions with the lysosome. We find that mammalian lysosomes and LDs undergo interactions during which proteins and lipids can be transferred from LDs directly into lysosomes. Electron microscopy (EM) of primary hepatocytes or hepatocyte-derived cell lines supports the existence of these interactions. It reveals a dramatic process whereby the lipid contents of the LD can be “extruded” directly into the lysosomal lumen under nutrient-limited conditions. Significantly, these interactions are not affected by perturbations to crucial components of the canonical macroautophagy machinery and can occur in the absence of double-membrane lipoautophagosomes. These findings implicate the existence of an autophagic mechanism used by mammalian cells for the direct transfer of LD components into the lysosome for breakdown. This process further emphasizes the critical role of lysosomes in hepatic LD catabolism and provides insights into the mechanisms underlying lipid homeostasis in the liver.

SPOP mutation disables 53BP1 ubiquitination and exclusion from DNA double-strand break sites and causes genome instability., 53BP1 activates nonhomologous end joining (NHEJ) and inhibits homologous recombination (HR) repair of DNA double-strand breaks (DSBs). Dissociation of 53BP1 from DSBs and consequent activation of HR, a less error-prone pathway than NHEJ, helps maintain genome integrity during DNA replication; however, the underlying mechanisms are not fully understood. Here, we demonstrate that E3 ubiquitin ligase SPOP promotes HR during S phase of the cell cycle by excluding 53BP1 from DSBs. In response to DNA damage, ATM kinase–catalyzed phosphorylation of SPOP causes a conformational change in SPOP, revealed by x-ray crystal structures, that stabilizes its interaction with 53BP1. 53BP1-bound SPOP induces polyubiquitination of 53BP1, eliciting 53BP1 extraction from chromatin by a valosin-containing protein/p97 segregase complex. Our work shows that SPOP facilitates HR repair over NHEJ during DNA replication by contributing to 53BP1 removal from chromatin. Cancer-derived SPOP mutations block SPOP interaction with 53BP1, inducing HR defects and chromosomal instability.

The membrane deforming dynamin family members MxA and MxB are large GTPases that convey resistance to a variety of infectious viruses. During viral infection, Mx proteins are known to show markedly increased expression via an interferon-responsive promoter to associate with nuclear pores. In this study we report that MxB is an inner mitochondrial membrane GTPase that plays an important role in the morphology and function of this organelle. Expression of mutant MxB or siRNA knockdown of MxB leads to fragmented mitochondria with disrupted inner membranes that are unable to maintain a proton gradient, while expelling their nucleoid-based genome into the cytoplasm. These findings implicate a dynamin family member in mitochondrial-based changes frequently observed during an interferon-based, anti-viral response., Mx proteins belong to the dynamin family of large GTPases and are highly induced by interferon in virally infected cells. The authors show that uninfected immune cells and hepatocytes also express MxB protein that associates with mitochondria to alter the morphology and genome of this organelle.

Primary sclerosing cholangitis (PSC) is an incurable, fibroinflammatory biliary disease for which there is no effective pharmacotherapy. We recently reported cholangiocyte senescence as an important phenotype in PSC while others showed that portal macrophages accumulate in PSC. Unfortunately, our ability to explore cholangiocyte senescence and macrophage accumulation has been hampered by limited in vitro models. Thus, our aim was to develop and characterize a three dimensional model (3D) of normal and diseased bile ducts (cholangioids) starting with normal human cholangiocytes (NHC), senescent NHC (NHC-sen), and cholangiocytes from PSC patients. In 3D culture, NHCs formed spheroids of ~5000 cells with a central lumen of ~150 μm. By confocal microscopy and western blot, cholangioids retained expression of cholangiocyte proteins (cytokeratin 7/19) and markers of epithelial polarity (secretin receptor and GM130). Cholangioids are functionally active, and upon secretin stimulation, luminal size increased by ~ 80%. Cholangioids exposed to hydrogen peroxide exhibited cellular senescence (SA-βGal and γH2A.x expression) and the senescence associated secretory phenotype (SASP; increased IL-6, p21, β;-Gal and p16 expression). Furthermore, cholangioids derived from NHC-sen or PSC patients were smaller and had slower growth than the controls. When co-cultured with THP-1 macrophages, the number of macrophages associated with NHC-sen or PSC cholangioids was 5 to7-fold greater compared to co-culture with non-senescent NHC. We observed that NHC-sen and PSC cholangioids release greater numbers of extracellular vesicles (ECVs) compared to controls. Moreover, conditioned media from NHC-sen cholangioids resulted in an ~2-fold increase in macrophage migration. In summary, we developed a method to normal and diseased cholangioids, characterized them morphologically and functionally, showed that they can be induced to senescence and SASP, and demonstrated both ECV release and macrophage attraction. This novel model mimics several features of PSC and thus will be useful for studying the pathogenesis of PSC and potentially identifying new therapeutic targets.

Background and aims Hepatocytes play a central role in storage and utilization of fat by the liver. Selective breakdown of lipid droplets (LDs) by autophagy (also called lipophagy) is a key process utilized to catabolize these lipids as an energy source. How the autophagic machinery is selectively targeted to LDs, where it mediates membrane engulfment and subsequent degradation, is unclear. Recently, we have reported that two distinct GTPases, the mechanoenzyme, dynamin2 (Dyn2), and the small regulatory Rab GTPase, Rab10, work independently at distinct steps of lipophagy in hepatocytes. Approach and results In an attempt to understand how these proteins are regulated and recruited to autophagic organelles, we performed a nonbiased biochemical screen for Dyn2-binding partners and found that Dyn2 actually binds Rab10 directly through a defined effector domain of Rab10 and the middle domain of Dyn2. These two GTPases can be observed to interact transiently on membrane tubules in hepatoma cells and along LD-centric autophagic membranes. Most important, we found that a targeted disruption of this interaction leads to an inability of cells to trim tubulated cytoplasmic membranes, some of which extend from lipophagic organelles, resulting in LD accumulation. Conclusions This study identifies a functional, and direct, interaction between Dyn2 and a regulatory Rab GTPase that may play an important role in hepatocellular metabolism.

Single-phase monoclinic aluminum–gallium oxide powders, β−(AlxGa1−x)2O3, have been produced by solution combustion synthesis for Al fraction 0 ≤ x

Thin-film photovoltaic (PV) devices can be fabricated using a solution-based synthesis procedure in which metal-chalcogenide nanocrystals with aliphatic coordinating ligands are suspended in a solvent to produce a printable ink (NC-ink). However, the aliphatic ligands that are used to solubilize and stabilize the nanocrystals operate as a significant source of carbon impurities that are incorporated into the final device absorber layer. Despite the ubiquity of this technique and the fact that carbon defects have been reported to be found across a spectrum of devices, the structure, properties, and influence of the carbon on PV device performance remain relatively unexplored. Our findings indicate that these organic ligands undergo a pyrolysis reaction during annealing, producing an electrically conductive, graphitic carbon that also reacts with chalcogens (S or Se) to produce heterocyclic moieties. In this work, we used oleylamine (OLA) and dodecylamine (DDA) to fabricate Cu2ZnSn(SxSe1–x)4 (CZTSSe) photov...

The mechanisms by which epithelial cells regulate clathrin-mediated endocytosis (CME) of transferrin are poorly defined and generally viewed as a constitutive process that occurs continuously without regulatory constraints. In this study, we demonstrate for the first time that endocytosis of the transferrin receptor is a regulated process that requires activated Src kinase and, subsequently, phosphorylation of two important components of the endocytic machinery, namely, the large GTPase dynamin 2 (Dyn2) and its associated actin-binding protein, cortactin (Cort). To our knowledge these findings are among the first to implicate an Src-mediated endocytic cascade in what was previously presumed to be a nonregulated internalization process.

Cortactin is an actin-binding protein that is overexpressed in many cancers and is a substrate for both tyrosine and serine/threonine kinases. Tyrosine phosphorylation of cortactin has been observed to increase cell motility and invasion in vivo, although it has been reported to have both positive and negative effects on actin polymerization in vitro. In contrast, serine phosphorylation of cortactin has been shown to stimulate actin assembly in vitro. Currently, the effects of cortactin serine phosphorylation on cell migration are unclear, and furthermore, how the distinct phospho-forms of cortactin may differentially contribute to cell migration has not been directly compared. Therefore, we tested the effects of different tyrosine and serine phospho-mutants of cortactin on lamellipodial protrusion, actin assembly within cells, and focal adhesion dynamics. Interestingly, while expression of either tyrosine or serine phospho-mimetic cortactin mutants resulted in increased lamellipodial protrusion and cell migration, these effects appeared to be via distinct processes. Cortactin mutants mimicking serine phosphorylation appeared to predominantly affect actin polymerization, whereas mutation of cortactin tyrosine residues resulted in alterations in focal adhesion turnover. Thus these findings provide novel insights into how distinct phospho-forms of cortactin may differentially contribute to actin and focal adhesion dynamics to control cell migration.

Passive grain boundaries (GBs) are essential for polycrystalline solar cells to reach high efficiency. However, the GBs in Cu2ZnSn(S,Se)4 have less favorable defect chemistry compared to CuInGaSe2. Here, using scanning probe microscopy we show that lithium doping of Cu2ZnSn(S,Se)4 changes the polarity of the electric field at the GB such that minority carrier electrons are repelled from the GB. Solar cells with lithium-doping show improved performance and yield a new efficiency record of 11.8% for hydrazine-free solution-processed Cu2ZnSn(S,Se)4. We propose that lithium competes for copper vacancies (forming benign isoelectronic LiCu defects) decreasing the concentration of ZnCu donors and competes for zinc vacancies (forming a LiZn acceptor that is likely shallower than CuZn). Both phenomena may explain the order of magnitude increase in conductivity. Further, the effects of lithium doping reported here establish that extrinsic species are able to alter the nanoscale electric fields near the GBs in Cu2ZnSn(S,Se)4. This will be essential for this low-cost Earth abundant element semiconductor to achieve efficiencies that compete with CuInGaSe2 and CdTe.

Field experiments were conducted on two North Carolina research stations in 1999, 2000, and 2001; on-farm in Lenoir, Wayne, and Wilson counties, NC, in 2002; and on-farm in Port Royal, VA, in 2000, 2001, and 2002 to evaluate possible gains from site-specific herbicide applications at these locations. Fields were scouted for weed populations using custom software on a handheld computer linked to a Global Positioning System. Scouts generated field-specific sampling grids and recorded weed density information for each grid cell. The decision aid HADSS™ (Herbicide Application Decision Support System) was used to estimate expected net return and yield loss remaining after treatment in each sample grid of every field under differing assumptions of weed size and soil moisture conditions, assuming the field was planted with either conventional or glyphosate-resistant (GR) soybean. The optimal whole-field treatment (that treatment with the highest expected net return summed across all grid cells within a field) resulted in average theoretical net returns of $79/ha (U.S. dollars) and $139/ha for conventional and GR soybean, respectively. When the most economical treatment for each grid cell was used in site-specific weed management, theoretical net returns increased by $13/ha (conventional) and $4.50/ha (GR), and expected yield loss after treatment was reduced by 10.5 and 4%, respectively, compared with the whole-field optimal treatment. When the most effective treatment for each grid cell was used in site-specific weed management, theoretical net returns decreased by $18/ha (conventional) and $4/ha (GR), and expected yield loss after treatment was reduced by 27 and 19%, respectively, compared with the whole-field optimal treatment. Site-specific herbicide applications could have reduced the volume of herbicides sprayed by as much as 70% in some situations but increased herbicide amounts in others. On average, the whole-field treatment was optimal in terms of net return for only 35% (conventional) and 57% (GR) of grid cells.

Proportions of marine vs. terrestrial resources in prehistoric human diets in the southern Mariana Islands (Guam, Rota, Saipan), Micronesia, have been estimated by analysis of stable isotope ratios of carbon and nitrogen in bone collagen and of carbon in apatite. The isotopic composition of marine and terrestrial food resources from the Marianas have also been determined. Experimental evidence shows that collagen carbon isotopes mainly reflect those of dietary protein sources and thus overestimate the contribution of marine animal foods. Marine protein consumption apparently ranges from approximately 20% to approximately 50% on these islands. Experiments also demonstrate the carbon isotope ratio of bone apatite carbonate accurately reflects that of the whole diet. Carbonate carbon isotope data suggest some individuals consumed significant amounts of 13C-enriched (C4) plants or seaweeds. Sugar cane is an indigenous C4 crop and seaweeds are eaten throughout the Pacific, but they have not been considered by archaeologists to have been prehistoric dietary staples. Apatite carbon isotope analysis has apparently identified previously unrecognized prehistoric dietary adaptations in the Mariana Islands, but this must be confirmed by archaeobotanical evidence.

Ultrastructural, immunocytochemical and UV absorption spectroscopy techniques were used to elucidate the progressive changes that occur within woody cell walls during decay by Ceriporiopsis subvermispora. After only 2 weeks of incubation, uranyl acetate staining revealed a diffuse electron dense zone in the secondary wall near hyphae and around the outer circumference of the wall. The extent of cell wall staining increased with longer fungal incubation. No staining occurred in sound unaltered cell walls. Proteins of varying molecular weights (insulin, 5730 Da; myoglobin, 17 600 Da; ovalbumin, 44 287 Da) were infiltrated into sound and decayed wood followed by immunogold labelling and transmission electron microscopy. Insulin readily penetrated into the outer most regions of secondary walls of wood cells after 2 weeks of decay. Myoglobin was first observed to penetrate cell walls at 4 weeks of degradation and ovalbumin was found after 8 weeks in wood with advanced stages of decay where extensive cell wall disruption was evident. None of the proteins used were localized within cell walls of untreated, control wood samples. UV microspectrophotometry demonstrated a progressive loss of absorbance at 240 and 280 nm within the secondary walls and middle lamellae at various sampling times throughout the duration of the decay study.

A detailed computer model which predicts etch rates and spatial uniformity for chemical vapor etching of GaAs with CH3I in a vertical rotating-disk organometallic vapor phase epitaxy (OMVPE) reactor has been developed. Etch rate predictions compare favorably with experiments performed in the vertical reactor at several temperatures ranging from 525 to 590°C and CH3I mole fractions from 0.005 to 0.050 in an H2 carrier gas. Agreement between the data and the model verifies our earlier assertion that gas phase decomposition of CH3I is the rate-limiting step, and that the etchant species are I and to a lesser extent CH3 radicals. Predictions further show that radial etch rate uniformity improves from a 5% variation over 50% of the substrate at low rotation (20–100 rpm) to a 5% variation over 80% of the substrate at 500 rpm. Increasing rotation rate also reduces the etch rate, a result that is in contrast to earlier results for OMVPE growth from TMGa and AsH3. These contrasting results arise because growth is rate limited by gas phase diffusion of TMGa, whereas CH3I vapor etching is rate limited by gas phase decomposition of CH3I.

CH3I vapor etching of masked and patterned GaAs substrates has been experimentally investigated. For GaAs samples masked with silicon nitride stripes that are wider than 30 tzm, the etch depth increased compared to unmasked samples, the magnitude of which increased with increasing mask width. Etching of bulk substrates of (lll)Ga and (lll)As GaAs revealed a dependence of etch rate on crystal orientation, with (lll)Ga > (100)GaAs > (lll)As. Increasing etch temperature reduced the orientation dependence of etch rates. Orientation dependence of etch rates was also observed on non-planar GaAs substrates patterned to expose different orientations on wet-etched groove structures. In this case, etch rate differences between the different orientations were amplified when compared to the bulk substrate results. Finally, it was found that the extent of mask undercutting depended on the direction of mask stripes in a fashion consistent with the orientation reactivity results. Mask stripes on (100)GaAs oriented in the [011] direction were severely undercut whereas stripes oriented in the [011] direction were undercut less.

Animal glue in combination with other chemicals is often used as a leveling agent in the copper electroplating industry. The control of the glue concentration in the electrolyte is critical to the quality of copper produced. A quantitative galvanostatic technique for glue analysis in copper electrolyte containing lignin sulfonate and Cl− was developed. The kinetics of glue hydrolysis in industrial electrolytes was studied and found to follow first-order reaction kinetics, with sulfuric acid acting as a catalyst. The dependence of the glue hydrolysis rate constant on temperature follows the Arrhenius equation. By adding fresh glue to the electrolyte, the glue activity first rises and then falls. This effect can be explained by the presence of long-chain molecules in the glue which are less active but hydrolyze into the more active medium-sized molecules. A mathematical model of this process shows good agreement with experimental data. The bulk of the electrolyte flow in the INCO commercial electrolytic plating cell bypasses the electrodes, probably across the bottom of the cell. The electrolyte circulation between electrodes is not very intense. A simple equation for the glue concentration calculation in the cell inlet and outlet, depending on the glue addition rate, was derived.

Atrial fibrillation (AF) is the most common cardiac arrhythmia in the western world. Genetic variants in the cardiac I Kr channel have been identified to influence ventricular repolarization. The aim of this work is to investigate the effect of the mutation N588K on atrial repolarization and the predisposition to AF. Experimental data of N588K mutated hERG channel were incorporated in an atrial ionic model using parameter fitting. The effects of the mutation were analyzed in cell and tissue. N588K showed a gain of function effect, causing a rapid repolarization and a shortening of the action potential duration. Computer simulations of a schematic right atrial geometry were used to investigate the excitation conduction properties. The effective refractory period of mutant cells were reduced from 317 to 233 ms at 1 Hz . The conduction velocity is not significantly influenced by the mutation. Nevertheless, the wavelength of mutant cells is for all frequencies smaller, indicating that the mutation N588K predisposes the initiation and perpetuation of AF.

The validity of using carbon isotopes in biological apatite from bone for dietary studies has been questioned because of diagenetic alterations and supposed exchange of carbon from the burial environment. Laboratory and natural experiments have been devised to test the integrity of carbon isotope ratios in biological apatite with respect to isotopic exchange of carbon with natural waters. The laboratory experiment subjected a sample of bone to a synthetic groundwater whose carbon isotopic composition had been adjusted to +9750‰. Analyses indicate that simple carbonates exchanged rapidly with the groundwater, but that the actual biological apatite exchanged very little, if any, of its carbon, even with 2 years exposure. The natural experiment involved analysis of mammoth and mastodon teeth that had been submerged in sea water for 11,000 years. With proper cleaning, the original dietary signals were well-preserved despite the long exposure to oceanic dissolved carbonate of much different isotopic composition. These experiments suggest that properly cleaned biological apatite is an appropriate analytical phase for dietary studies on bone or teeth as old as 10,000 years.

The large GTPase dynamin (Dyn2) has been demonstrated by us and others to interact with several different actin-binding proteins. To define how Dyn2 might participate in actin dynamics in livings cells we have expressed green fluorescent protein (GFP)-tagged Dyn2 in cultured cells and observed labeling of comet-like vesicles and macropinosomes. The comet structures progressed with a constant velocity and were reminiscent of actin comets associated with motile vesicles in cells expressing type I phosphatidylinositol phosphate 5-kinases. Based on these observations we sought to determine whether Dyn2 is an integral component of actin comets. Cells expressing type I phosphatidylinositol phosphate 5-kinase and Dyn2-GFP revealed a prominent colocalization of Dyn2 and actin in comet structures. Interestingly, comet formation and motility were normal in cells expressing wild-type Dyn2-GFP but altered markedly in Dyn2 mutant-expressing cells. Dyn2K44A-GFP mutant cells displayed a significant reduction in comet number, length, velocity, and efficiency of movement. In contrast, comets in cells expressing Dyn2ΔPRD-GFP appeared dark and did not incorporate the mutant Dyn2 protein, indicating that the proline-rich domain (PRD) is required for Dyn2 recruitment. Further, these comets were significantly longer and slower than those in control cells. These findings demonstrate a role for Dyn2 in actin-based vesicle motility.

Site occupancy and cation binding states in the proposed thermoelectric n-type oxide SrxBa1−xNb2O6 (SBN100x) were investigated using X-ray photoelectron spectroscopy (XPS). Sr 3d XPS spectra from unreduced polycrystalline SBN100x with various compositions contained two distinct spin-orbit doublets corresponding to Sr occupying either A1 or A2 positions in the SBN lattice; the higher binding energy state was associated with Sr ions at A2 sites, presumably due to their increased coordination over Sr at A1 sites. To gain insight into optimizing the thermoelectric properties of reduced SBN, sintered SBN50 specimens were reduced in Ar/H2 or N2/H2 ambient. A decrease in the average Nb valence was observed in Nb 3d photoemission through the growth of low-binding energy components after reduction in either environment; evidence of surface NbN formation was apparent with longer reducing times in N2/H2. Both the single-component Ba 3d emission and the A2 component of the Sr 3d spectra show shifting to lower binding...

With the objective of developing an improved process for in situ etching of GaAs‐based materials in organometallic vapor phase epitaxy reactors, GaAs wafers and AlxGa1−xAs epilayers have been etched with CH3I vapor in a horizontal reactor operated at atmospheric pressure with H2 or He carrier gas. For a H2 flow rate of 2.1 s lpm, etching temperatures from 400 to 625 °C, and CH3I mol fractions (yCH3I)from 0.0012 to 0.015, the measured GaAs etch rate r (in A min−1) is given by r = k0 y0.83CH3I exp[ − 45(kcal mol−1)/RT] with k0=3.2×1016 A min−1. The value of k0 depends on the type of carrier gas, flow rate, total pressure, and reactor geometry. The etch rate appears to be controlled mainly by the decomposition of CH3I to CH3 and I, for which the activation energy has been reported to be 43.5 kcal mol−1. The etch rate of AlxGa1−xAs epilayers with x up to 0.7, which was measured at 480 °C with yCH3I= 0.015, does not depend on Al content. The surface morphology of etched GaAs wafers improves with decreasing tem...

Summary The maize mutation high chlorophyll fluorescence 60-muTable 1 (hcf60-m1), generated through Activator (Ac) tagging, has insufficient photosynthetic electron transport. Here we show that the Hcf60 gene encodes a protein with substantial amino acid similarity to plant plastid and bacterial ribosomal small subunit protein 17 (RPS17) proteins. The lack of detectable HCF60 transcripts in mutant leaves, and insertion of the transposed Ac element 17 bp upstream of the start of translation in the mutated locus, suggest that little if any RPS17 is produced. The mutant phenotype is consistent with reduced plastid translation. Seedling lethal hcf60-m1 plants display temperature and light-dependent chlorophyll deficiencies, a depletion of plastid rRNA pools, and few high-molecular-weight polysomal complexes. Growth under moderate light conditions (27°C, 100 μE m−2 sec−1) allows for substantial chlorophyll accumulation in mutant leaves, yet the number of functional photosystem II complexes appears low. Nevertheless, the presence of a limited but intact C4 system indicates that some plastid translation occurs. Table 1. . Leaf chlorophyll content under different growth conditions 17°C, 100 μE m−2 sec−1 27°C, 100 μE m−2 sec−1 Greenhouse 1st leaf 1st leaf 2nd leaf 1st leaf WT hcf60 WT hcf60 WT hcf60 WT hcf60 Total chlorophyll 0.385 ± 0.42 >0.002 ±0.00017 1.11 ±0.15 0.79 ± 0.1 1.3 ±0.2 0.71 ± 0.17 0.89 ±0.077 0.026 ± 0.012 μg mg−1 tissue n = 4 n = 4 n= 7 n= 7 n= 7 n= 7 n = 4 n = 3 Total chlorophyll 100 >0.5 100 71 100 55 100 2.9 % WT Chlorophyll a/b 2.12 ±0.2 – 2.54 ±0.22 1.89 ±0.16 2.65 ±0.26 1.97 ±0.12 3.02 ±0.11 2.87 ±0.23

Ethanol has been predicted to alter vesicle-based protein traffic in hepatocytes, in part, via a disruption of the microtubule (MT) cytoskeleton. However, information on the effects of chronic ethanol exposure on MT function in vivo is sparse. Therefore the goal of this study was to test for ethanol-induced changes in rat liver tubulin expression, assembly, and cellular organization, using molecular, biochemical and morphological methods. The results of this study showed that tubulin mRNA and protein levels were not altered by ethanol. Tubulin, isolated from control and ethanol-fed rats, showed similar polymerization characteristics as assessed by calculation of the critical concentration for assembly and morphological structure. In contrast, the total amount of assembly-competent tubulin was reduced in livers from ethanol-fed rats compared with control rats when assessed by quantitative immunoblot analysis using a tubulin antibody. In addition, we observed that MT regrowth and organization in cultured hepatocytes treated with cold and nocodazole was markedly impaired by chronic ethanol exposure. In summary, these results indicate that tubulin levels in liver are not reduced by ethanol exposure. While there is a substantial amount of tubulin protein capable of assembling into functional MTs in ethanol-damaged livers, a marked portion of this tubulin is polymerization incompetent. This may explain why these hepatocytes exhibit a reduced number of MTs with an altered organization.

The sulfidation reaction kinetics of fine particles of zinc titanate and zinc oxide with H{sub 2}S were studied in order to test the potential of the sorbent injection hot-gas desulfurization process. Fine sorbent particles with diameter between 0.3 and 60 {mu}m were sulfided with H{sub 2}S and/or reduced with H{sub 2} in a laminar flow reactor over the temperature range of 500-900{degrees}C. Sulfidation/reduction conversion was compared for different particle sizes and sorbents with various porosities and atomic ratios of Zn and Ti. In reduction of ZnO with H{sub 2} and without H{sub 2}S, significant amount of Zn was formed and vaporized, while the presence of H{sub 2}S suppressed elemental Zn formation. This suggests that H{sub 2}S may suppress the surface reduction of ZnO and/or gaseous Zn may react with H{sub 2}S homogeneously and form fine particles of ZnS. Formation and vaporization of elemental Zn from zinc titanate sorbents was slower than from zinc oxide with and without H{sub 2}S.

A reproducible system for the generation of fertile, transgenic maize plants has been developed. Cells from embryogenic maize suspension cultures were transformed with the bacterial gene bar using microprojectile bombardment. Transformed calli were selected from the suspension cultures using the herbicide bialaphos. Integration of bar and activity of the enzyme phosphinothricin acetyltransferase (PAT) encoded by bar were confirmed in all bialaphos-resistant callus lines. Fertile transformed maize plants (R0) were regenerated, and of 53 progeny (R1) tested, 29 had PAT activity. All PAT-positive progeny analyzed contained bar. Localized application of herbicide to leaves of bar-transformed R0 and R1 plants resulted in no necrosis, confirming functional activity of PAT in the transgenic plants. Cotransformation experiments were performed using a mixture of two plasmids, one encoding PAT and one containing the nonselected gene encoding [beta]-glucuronidase. R0 plants regenerated from co-transformed callus expressed both genes. These results describe and confirm the development of a system for introduction of DNA into maize.

Chloroplasts isolated from tall fescue, Festuca arundinacea Schreb., showed high rates of electron transport, comparable to rates observed for spinach chloroplasts.Chloroplasts were well coupled and rates of electron transport from water to methyl viologen (photosystem II and I) were increased two to five times when ADP and inorganic phosphate or methylamine (uncoupler) were added to the reaction mixture. Ratios of P:2e for photosystem II plus I were found to be near 1.2. Electron transport rates from water to p-phenylenediamine or 2,6-dichlorobenzoquinone (photosystem II) were over 300 micromoles O(2) per hour per milligram chlorophyll, while P:2e ratios were found to be over 0.5. The highest rates of electron transport were found in electron flow from diaminodurene to methyl viologen (photosystem I) and P:2e ratios remained near 0.5.Light intensity saturation curves for photosystem II and I, as well as the photosystems independently, resembled curves for spinach, with saturation of electron transport in photosystem I and photosystem II separately occurring at 35% of the available light intensity (6000 microeinsteins per square meter per second). Photosystem II and I in sequence were saturated at about half this light intensity.

We have observed the dynamics of sarcomere shortening and the diffracting action of single, functionally intact, unattached cardiac muscle cells enzymatically isolated from the ventricular tissue of adult rats. Sarcomere length was measured either (a) continuously by a light diffraction method or (b) by direct inspection of the cell's striated image as recorded on videotape or by cinemicroscopy (120--400 frames/s). At physiological levels of added CaCl2 (0.5--2.0 mM), many cells were quiescent (i.e., they did not beat spontaneously) and contracted in response to electrical stimulation (less than or equal to 1.0-ms pulse width). Sarcomere length in the quiescent, unstimulated cells (1.93 +/- 0.10 [SD] micrometers), at peak shortening (1.57 +/- 0.13 micrometers, n = 49), and the maximum velocity of sarcomere shortening and relengthening were comparable to previous observations in intact heart muscle preparations. The dispersion of light diffracted by the cell remained narrow, and individual striations remained distinct and laterally well registered throughout the shortening-relengthening cycle. In contrast, appreciable nonuniformity and internal buckling were seen at sarcomere lengths < 1.8 micrometers when the resting cell, embedded in gelatin, was longitudinally compressed These results indicate (a) that shortening and relengthening is characterized by uniform activation between myofibrils within the cardiac cell and (b) that physiologically significant relengthening forces in living heart muscle originate at the level of the cell rather than in extracellular connections. First-order diffracted light intensity, extremely variable during sarcomere shortening, was always greatest during midrelaxation preceding the onset of a very slow and uniform phase of sarcomere relengthening.

Ubiquitous soybean urease, as opposed to the seed-specific urease, designates the seemingly identical ureolytic activities of suspension cultures and leaves. It also appears to be the basal urease in developing seeds of a variety, Itachi, which lacks the seed-specific urease (Polacco, Winkler 1984 Plant Physiol 74: 800-804). On native polyacrylamide gels the ureolytic activities in crude extracts of these three tissues comigrate as determined by assays of gel slices. At this level of resolution the ubiquitous urease also migrates with or close to the fast (trimeric) form of the seed-specific urease. The ubiquitous urease was purified approximately 100-fold from suspension cultures of two cultivars (Itachi and Prize) as well as from developing seeds of Itachi. These partially purified preparations allowed visualization of native urease on polyacrylamide gels by activity staining and of urease subunits on denaturing lithium dodecyl sulfate gels by electrophoretic transfer to nitrocellulose and immunological detection (“Western Blot”). The ubiquitous urease holoenzyme migrates slightly less rapidly than the fast seed urease in native gels; its subunit migrates slightly less rapidly than the 93.5 kilodaltons subunit of either the fast or slow (hexameric) seed enzyme. The ubiquitous urease elutes from an agarose A-0.5 meter column with the fast form of the seed urease species suggesting that the ubiquitous urease, like the fast seed urease, exists as a trimeric holoenzyme. The soybean cultivar, Prize, produces the hexameric seed urease; yet its ubiquitous urease (from leaf and suspension culture) is trimeric. The pH dependence of the ureolytic activity of seed coats of both seed urease-negative (Itachi) and seed urease-positive (Williams) cultivars suggests that this activity is exclusively the ubiquitous urease. Its relatively higher levels in seed coats than in embryos of Itachi suggests that the ubiquitous urease is involved in degradation of urea derived from ureides. Consistent with a ureide origin for urea is the observation that addition of a urease inhibitor, phenylphosphordiamidate, to extracts of developing Itachi seeds (seed coat plus embryo) results in accumulation of urea from allantoic acid.

Lead-zinc and barite deposits in central Nova Scotia generally occur at the Tournaisian-Visean (Mississippian) clastic carbonate-evaporite disconformity overlying Cambrian to Devonian metasedimentary and granitic rocks. The deposits are located at the southern margin of the > 10-km-deep Carboniferous Maritimes (Fundy-Magdalen) basin.Fluid inclusion homogenization temperatures of ore-stage to postore minerals range from 230 degrees to 130 degrees C and fluid salinity is > or = 24 equiv wt percent NaCl. A mineralization age of ca. 300 to 330 Ma is suggested by: (1) fission-track dates on zircon from clastic host rock at Gays River, (2) K-Ar dates on clays from altered clastic rock in close proximity to mineralization, and (3) Rb-Sr data on illite from samples collected over a 10-m section of Tournaisian sandstone (far away from mineralization). Lead and strontium isotope data indicate that Tournaisian rock of the Horton Group could have been a source (at ca. 300 Ma) of lead and radiogenic strontium in the ore-stage minerals at the larger deposits. Visean carbonate carbon and sulfate were the dominant sources of carbon and sulfur, respectively. Hydrogen and oxygen isotope data suggest that the mineralizing fluids were basinal brines. Nonbiogenic sulfate reduction at the deposits is the likely sulfide precipitation mechanism.The ore-forming fluids may have originated at a >5-km depth under Visean evaporites in the Magdalen basin and migrated rapidly through Tournaisian clastic rocks and the immediately overlying Pembroke breccia to the basin margins. A pulse of regional tectonism may have set off at least one massive, hydrofracturing, compaction-driven, fluid expulsion event by about 300 Ma.

Measurements of δ13C and 14C-activity were performed on vinegars from various known sources. Natural vinegar can be distinguished from petrochemical acetic acid by 14C-analysis: Natural vinegar currently gives values of > 112% of modern activity; petrochemical acetic acid yields values of 0% of modern activity. Apple cider vinegar can be distinguished from corn-derived vinegar by δ13C-analysis: Cider vinegar gives δ13C-values near — 26%o; corn-derived vinegars yield δ13C-values near - 10% δ13C-Analysis also can be applied, with some restrictions, to wine vinegars. These techniques are applied to a series of retail vinegars.

Samples of 6 food and feed ingredients and a purified protein, plactoglobulin, were analyzed by 7 laboratories to determine the concentrations of cysteine as cysteic acid and methionine as methionine sulfone. Samples were oxidized by reaction with performic acid before hydrolysis with 6N HC1. The free amino acids were then separated and measured by ion-exchange chromatography on dedicated amino acid analyzers. Each laboratory was provided with a detailed method as well as sealed vials containing solutions of standards. For the determination of cysteine as cysteic acid, the coefficients of variation between laboratories for duplicate samples ranged from 7.13 to 10.8% for the 6 ingredients. For the determination of methionine as methionine sulfone, the coefficients of variation between laboratories for duplicate samples ranged from 1.18 to 12.8% for the 6 ingredients. Cysteine and methionine recoveries were determined by analysis of β-Iactoglobulin and were based on expected levels of each amino acid from amino acid sequence data. The mean recovery of cysteine was 95% with a range of 91-101%. The mean recovery of methionine was 101% with a range of 98-106%. This method has been adopted official first action.

The second thermodynamic dissociation constants of N,N-bis(2-hydroxyethyl)-2-aminoethanesulfonic acid (hereafter known as “ bes ”), a useful secondary standard biochemical buffer within the pH range 6.6 to 7.4, have been determined from 278.15 to 328.15 K at intervals of 5 K (including 310.15 K), both in water and in 50 mass per cent methanol + water solvent mixtures. E.m.f. measurements in cells without liquid junction (using hydrogen and silver + silver bromide electrodes) were made. The pK2 (in water) for the process bes ± = bes − + H+ is expressed by the equation p K 2 = 1302.6( K T ) + 2.6850 + 0.00044289( T K ) , whereas that in 50 mass per cent methanol-water is expressed by p K 2 = 2198.7( K T ) − 2.8868 + 0.0085509( T K ) . The thermodynamic quantities associated with the ionization of bes in the two solvents have been calculated from the above pK2 expressions. The transfer thermodynamic functions for the dissociation process from water to 50 mass per cent methanol + water have also been evaluated. These results (obtained in water) are compared with those of compounds similar to bes .

The two thermodynamic dissociation constants of glycine at 11 temperatures from 5 to 55°C in 50 mass % methanol-water mixed solvent have been determined from precise emf measurements with hydrogen-silver bromide electrodes in cells without liquid junction. The first acidic dissociation constant (K 1)for the process HG+⇌H++G± is expressed as a function ofT(oK) by the equation pK 1 = 2043.5/T − 9.6504 + 0.019308T. At 25°C, pK 1is 2.961 in the mixed solvent, as compared with 2.350 in water, with ΔH°=1497 cal-mole−1, ΔG°=4038 cal-mole−1, ΔS°=−8.52 cal-°K−1-mole−1, and ΔC p o =−53 cal-°K−1-mole−1. The second acidic dissociation constant (K 2)for the process G±⇌H++G− over the temperature range studied is given by the equation pK 2 = 3627.1/T − 7.2371 + 0.015587T. At 25°C, pK 2is 9.578 in MeOH−H2O as compared with 9.780 in water, whereas ΔH° is 10,257 cal-mole−1, ΔG° is 13,063 cal-mole−1, ΔS° is −9.41 cal-°K−1-mole−1, and ΔC p o is −43 cal-°K−1-mole−1. The protonated glycine becomes weaker in 50 mass % methanol-water, whereas the second dissociation process becomes stronger despite the lower dielectric constant of the mixed solvent (ω=56.3 at 25°C).

The Temiscamie Fe formation of the Lake Albanel iron range in Mistassini territory, Quebec, is approximately 700 ft. thick and is divided into 6 members on the basis of lithology. The principal Fe-bearing minerals are siderite, magnetite, hematite, and iron silicates. A bulk chemical analysis of the Fe formation, excluding the thin lower argillite member, shows 29% of Total Fe and is similar in many respects to the average compositions of Fe formations of the Mesabi, Cuyuna, and Gogebic districts. The Temiscamie Fe formation is overlain by the Kallio slate and is underlain by the Boulder Bay quartzite and the upper and lower Albanel dolomites, the sequence constituting the Mistassini group. These rocks are underlain by sediments of the Papaskwasati group, which rest with angular unconformity on a basement of folded sediments and volcanics of the Sam Gunner group. The latter was earlier intruded by igneous rocks of the Takwa complex. The Papaskwasati and Mistassini sediments are relatively unmetamorphosed. They dip to the SE. and abut against high-grade metamorphic and igneous rocks of the Grenville subprovince along the Mistassini fault. The names Boulder Bay quartzite, Sam Gunner group, and Takwa complex are introduced. The Temiscamie Fe formation, on the basis of lithology, has been correlated with the Fe formation of the Labrador trough. Results of radioactivity dating are inconclusive but indicate the possibility of the Temiscamie Fe formation being younger than the Huronian Fe formations of the Lake Superior district. At the same time the radioactivity dates from northern Quebec and Labrador suggest a complicated geologic history for this region. Effects of the Grenville orogeny have been superimposed on rocks that were involved in 2 earlier major orogenies.

The thermal oxidative stability of poly(phosphonylureas) of the below from was studied. The products show significantly better thermal stability than their carbon counterparts. The product where R is C6H5— and R′ is 1.3-C6H4- has only 10% weight loss at 400°C and 40% weight loss at 600°C. The thermal degradation of the poly(phosphonylureas) followed using infrared spectroscopy. Possible initial degradation sites are discussed. Die thermo-oxidative Stabilitat von Polyphosphonylharnstoffen folgender Struktur wurde untersucht: Die Produkte sind thermisch viel stabiler als ihre Kohlenstoffanalogen. Das Produkt mit R gleich C6H5— und R′ gleich 1.3-C6H4— hat nur 10% Gewichtsverlust bei 400°C und 40% Gewichtsverlust bei 600°C. Der thermische Abbau der Polyphosphonylharnstoffe wurde durch Infrarotspektroskopie verfolgt. Es werden Reaktionen diskutiert, die wahrend des thermischen Abbaues ablaufen konnten.

Studies evaluating the distribution of stable carbon isotopes in bone and other tissues of both human and non-human animals have recently been reported1–6. Those investigations which examined the isotopic composition of bone have concentrated on the analysis of bone collagen and demonstrated that the 13C/12C ratios in bone collagen are directly related to the 13C/12C ratios of primary photosynthesizing plants in the diets of the animals concerned6–8. With regard to archaeological applications, such analyses have been limited to relatively young samples because of the instability of collagen in bone. We have extended the isotopic method of dietary analysis by using both the organic and inorganic phases of bone with equally good results. In the case of material over a few thousand years old, unless special conditions have preserved collagen, analysis of the organic phase of bone is no longer practical due to its deterioration9. The technique reported here allows dietary analysis of bone over 10,000 years old by using the inorganic phase, which is more stable in fossil material.

13C/12C ratios of the carboxyl and methyl carbons of acetic acid were determined by two published methods, using the thermal decomposition of sodium acetate and calcium acetate respectively. Comparison of the results obtained on a variety of samples revealed a systematic disagreement of 4.0% between the two methods. The source of this discrepancy is shown to be a fractionation associated with the calcium acetate method.

A cyclic leaking craniopharyngioma was found to be the source of recurrent fevers, meningismus, and coma. A sterile persistent neutrophilic pleocytosis with normal glucose and protein was the only clue in an extensive workup. Three head computed tomographic scans, including double contrast with 5-mm cuts, failed to locate the large suprasellar cyst outlined finally by head computed tomographic scan with metrizamide. A handful of case reports exist about leaking central nervous system cysts, but their presentation tended to be different: increased protein, lymphocytosis, and positive cranial imaging. It is believed this is a unique case that adds to the spectrum of chronic neutrophilic meningitis.