Transposition of Ac from the P Locus of Maize into Unreplicated Chromosomal Sites

We have analyzed donor and target sites of the mobile element Activator (Ac) that are altered as a result of somatic transposition from the P locus in maize. Previous genetic analysis has indicated that the two mitotic daughter lineages which result from Ac transposition from P differ in their Ac constitution at the P locus. Both lineages, however, usually contain transposed Ac elements which map to the same genetic position. Using methylation-sensitive restriction enzymes and genomic blot analysis, we identified Ac elements at both the donor P locus and Ac target sites and used this assay to clone the P locus and to identify transposed Ac elements. Daughter lineages were shown to be mitotic descendants from a single transposition event. When both lineages contained Ac genetic activity, they both contained a transposed Ac element on identical genomic fragments independent of the genetic position of the target site. This indicates that in the majority of cases, Ac transposition takes place after replication of the donor locus but before completion of replication at the target site.     

椎体静脉稀疏区注入骨水泥对PVP术中骨水泥渗漏的影响

目的:探讨椎体静脉稀疏区注入骨水泥对骨质疏松椎体压缩性骨折患者行经皮穿刺椎体成形术(percutaneous vertebroplasty,PVP)术中骨水泥渗漏的影响。方法:选择西安交通大学第二附属医院2014年1月至2018年6月收治的61例骨质疏松椎体压缩性骨折患者,根据骨水泥注入区域的不同,将所有患者分为A组(30例)及B组(31例),A组骨水泥注入区域为椎体静脉密集区(椎体中1/3平面处),B组骨水泥注入区域为椎体静脉稀疏区(椎体上1/3及下1/3平面处),对比两组的骨水泥渗漏率,术前、术后6个月时的视觉模拟评分(Visual analogue scale,VAS),治疗中的骨水泥用量、椎体高度恢复率及cobb角恢复度数。结果:B组的骨水泥渗漏率及骨水泥用量均明显低于A组(P0.05)。两组的VAS评分、椎体高度恢复率、cobb角恢复情况对比差异无统计学意义(P0.05)。结论:与椎体静脉密集区相比,在椎体静脉稀疏区注入骨水泥可显著降低骨质疏松椎体压缩性骨折患者PVP术中骨水泥渗漏率,椎体静脉稀疏区可作为PVP术中骨水泥注射的一个相对安全区域。     

Measurement of the Leakage of CO2 from Bundle-Sheath Cells of Leaves during C4 Photosynthesis

During C4 photosynthesis, CO2 is released in bundle-sheath cells by decarboxylation of C4 acids and then refixed via ribulose-1,5-bisphosphate carboxylase. In this study we examined the efficiency of this process by determining the proportion of the released CO2 that diffuses back to mesophyll cells instead of being refixed. This leak of CO2 was assessed by determining the amount of 14CO2 released from leaves during a chase in high [12CO2] following a 70-s pulse in 14CO2. A computer-based analysis of the time-course curve for 14CO2 release indicated a first-order process and provided an estimate of the initial velocity of 14CO2 release from leaves. From this value and the net rate of photosynthesis determined from the 14CO2 fixed in the pulse, the CO2 leak rate from bundle-sheath cells (expressed as a percentage of the rate of CO2 production from C4 acids) could be deduced. For nine species of Gramineae representing the different subgroups of C4 plants and two NAD-malic enzyme-type dicotyledonous species, the CO2 leak ranged between 8 and 14%. However, very high CO2 leak rates (averaging about 27%) were recorded for two NADP-malic enzyme-type dicotyledonous species of Flaveria. The results are discussed in terms of the efficiency of C4 photosynthesis and observed quantum yields.     

Dormancy Release,Germination, and Electrolyte Leakage from Apple Embryos during Stratification in the Presence of Sucrose

The dynamics of dormancy release during the stratification of apple (Malus domestica Borkh.) seeds was quantitatively described by three characteristics of seeds germination: the percentage of seeds that germinated by the tenth day, mean germination time, and the sum of seeds germinated in each of ten days (Timson's parameter), which allowed the assessment of the viability, the rate of dormancy release, and seed heterogeneity. We showed that apple seeds were characterized by a combined (physical and physiological) type of dormancy, with the seed coat and the embryo envelope being involved in the maintenance of physical dormancy. The addition of sucrose to the stratification medium accelerated the release of seed dormancy and improved all characteristics that determine seed germinability. Electrolyte leakage from embryos hardly changed during stratification, which agrees with the fact that all seeds remained viable throughout the entire period of dormancy. We assume that the release of seed dormancy is not a single-stage process.     

Energy-Dependent Solute Transport from the Apoplast into the Symplast of Leaves during Transpiration

Inorganic and organic salts, amino acids, sugars, and phosphate esters (concentrations usually 25 mM) were fed via the transpiration stream through the petiole into detached leaves of Lepidium sativum and Solanum tuberosum. While water was lost by transpiration, solutes did not accumulate in the apoplast. Uptake into leaf cells was indicated by stimulation of respiration and by changes of membrane potential and apoplastic pH. Apoplastic alkalinization (followed by transient acidification) and membrane depolarization (followed by repolarization) indicated energization of transport at the expense of the proton motive force (PMF) across the plasma membrane in all examined cases. Loss of ATP in the symplast during proton extrusion into the apoplast by the plasmalemma ATPase is thought to be responsible for stimulation of respiration. Even unphysiological solutes such as -morpholinoethane sulfonate (Mes), or potentially toxic salts such as CdCl₂ or AlCl₃, and metabolites involved in energy conservation such as AMP and NAD, were readily transported into leaf cells at the expense of metabolic energy. At the maximum stimulation of CO₂ release by D-serine (which is unlikely to be metabolized) respiration exceeded basal respiration by an average of 33%. Occasionally, and with other solutes, basal respiration was almost doubled. The ratio of transported solute to released extra CO₂ was 6.9 ± 1.1 (n = 11) in the case of D-serine. From this, maximum energized transport of D-serine was calculated to be close to 500 nmol/(m² leaf area s). Solute/CO₂ ratios similar to those observed with D-serine were also obtained for sucrose. Lower ones were observed with organic solutes such as L-glycine, pyruvate, malate or citrate where secondary metabolic conversions may contribute to CO₂ release.     

Proteins shifting from the cytoplasm into the nuclei during early embryogenesis of Drosophila melanogaster

Monoclonal antibodies have been used to study the distribution of several proteins in cleavage and blastoderm stages of Drosophila melanogaster. These antigens are known to be associated with hnRNA-containing particles in tissue culture cells. Protein blotting shows that they are present in the embryo 1 hr after egg deposition. A redistribution from the cytoplasm into the somatic nuclei can be observed during developmental stage $\text{12}\text{13}$, one stage prior to the formation of the cellular blastoderm. Yolk nuclei become stained by these antibodies at about the same time. The shift into pole cell nuclei, however, occurs $\text{1}\text{1}\text{2}$ hr later, during the migration of these cells into the posterior midgut rudiment.     

Transient entry of enterotoxin subunits into the periplasm occurs during their secretion from Vibrio cholerae.

Cholera toxin and heat-labile enterotoxin (LT) are structurally similar oligomeric proteins which are capable of being efficiently secreted from Vibrio cholerae. Here we report that these proteins transiently enter the periplasm of V. cholerae as they traverse the cell envelope to reach the extracellular milieu. Pulse-chase experiments on V. cholerae TRH7000 harboring an LT-encoding plasmid revealed that radiolabeled LT A and B subunits entered the periplasm rapidly, followed by their slow efflux (half-time, 13 min) into the medium. LT B-subunit efflux from the periplasm was calculated to be at a rate of ca. 170 monomers per min per cell (which is equivalent to 34 assembled LT holotoxin molecules per min per cell). These values were estimated to be sufficient to account for the increase in extracellular enterotoxin concentration during exponential cell growth. Thus, all enterotoxin subunits which are secreted into the medium can be assumed to be channelled via the periplasm. These findings led to an improved model of the pathway of toxin secretion by V. cholerae.     

Changes in the incorporation of carbon derived from glucose into cellular pools during the cell cycle of Saccharomyces cerevisiae

The rate of incorporation of 14C derived from [U-14C]glucose into cells of Saccharomyces cerevisiae X2180(1B) was investigated as a function of the cell cycle. After pulse-labelling of exponentially growing populations, centrifugal elutriation was used to isolate various cell fractions of increasing cell size, representing successive stages of the cell cycle. The total amount of 14C incorporated per cell was found to increase continuously during the cell cycle along with cellular protein content and Coulter counter cell volume. This pattern supports the model of exponential cell growth. In order to evaluate changes in intracellular carbon flow during the cell cycle, chemical extraction procedures were used to obtain four cellular fractions enriched in either low-molecular-mass components, lipid material, polysaccharides or proteins. The distribution of 14C among these cellular fractions varied during successive stages of the cell cycle, indicating cell-cycle-dependent fluctuations in intracellular carbon flow. During the G1 phase the flow of 14C into the low-molecular-mass pool increased markedly; concurrently, the rate of incorporation into the polysaccharide-enriched pool decreased.     

Growth inhibitors in oat grains. I. Leakage of inhibiting factors from oat grains during imbibition and the effect of these on germination and growth

Spikes of barley ( Hordeum vulgare L.) cultivar Bomi and high-lysine mutants Riso 1508 and Riso 56 were cultured on liquid media at varying N and sucrose levels. Bomi accumulated N in response to increasing N levels in the medium and a higher level was reached than in spikes of intact plants. The distribution of N in salt-soluble, hordein, and non-protein N fractions appeared to be normal. Endosperm dry weight and starch were lower than in intact plants and declined at higher N levels. A linear relationship was observed between starch content and the concentration of sucrose in the endosperm water. Uptake of culture medium by the spikes was affected by both N and sucrose concentration. The mutants had lower dry weights and starch contents, and higher sucrose contents than Bomi. At high N levels, the mutants accumulated less hordein, and more non-protein N than Bomi.     

Crystal structures of nitroalkane oxidase: insights into the reaction mechanism from a covalent complex of the flavoenzyme trapped during turnover

Nitroalkane oxidase (NAO) from Fusarium oxysporum catalyzes the oxidation of neutral nitroalkanes to the corresponding aldehydes or ketones with the production of H(2)O(2) and nitrite. The flavoenzyme is a new member of the acyl-CoA dehydrogenase (ACAD) family, but it does not react with acyl-CoA substrates. We present the 2.2 A resolution crystal structure of NAO trapped during the turnover of nitroethane as a covalent N5-FAD adduct (ES*). The homotetrameric structure of ES* was solved by MAD phasing with 52 Se-Met sites in an orthorhombic space group. The electron density for the N5-(2-nitrobutyl)-1,5-dihydro-FAD covalent intermediate is clearly resolved. The structure of ES was used to solve the crystal structure of oxidized NAO at 2.07 A resolution. The c axis for the trigonal space group of oxidized NAO is 485 A, and there are six subunits (1(1)/(2) holoenzymes) in the asymmetric unit. Four of the active sites contain spermine (EI), a weak competitive inhibitor, and two do not contain spermine (E(ox)). The active-site structures of E(ox), EI, and ES* reveal a hydrophobic channel that extends from the exterior of the protein and terminates at Asp402 and the N5 position on the re face of the FAD. Thus, Asp402 is in the correct position to serve as the active-site base, where it is proposed to abstract the alpha proton from neutral nitroalkane substrates. The structures for NAO and various members of the ACAD family overlay with root-mean-square deviations between 1.7 and 3.1 A. The homologous region typically spans more than 325 residues and includes Glu376, which is the active-site base in the prototypical member of the ACAD family. However, NAO and the ACADs exhibit differences in hydrogen-bonding patterns between the respective active-site base, substrate molecules, and FAD. These likely differentiate NAO from the homologues and, consequently, are proposed to result in the unique reaction mechanism of NAO.