Reduction in SBPase Activity by Antisense RNA in Transgenic Rice Plants: Effect on Photosynthesis,Growth, and Biomass Allocation at Different Nitrogen Levels

Rice cultivar zhonghua11 (Oryza sativa L. ssp. japonica) plants with decreased sedoheptulose-1, 7-bisphosphatase (SBPase) were obtained by transformation with the rice SBPase antisense gene under the control of the maize ubiquitin promoter. The transgenic and wild-type plants were grown at different nitrogen levels (0.1, 1, or 10 mM NH₄NO₃). Growth rates of the seedlings were measured by the changes in dry weight, and the photosynthetic carbon reduction activities and the potential efficiency of photosystem II were measured by CO₂ assimilation and F _v/F _m, respectively. At low N, there are strong effects on growth and photosynthesis when SBPase was reduced by genetic manipulation. Decreased SBPase activity led to a decrease in the amount of starch accumulated in the leaves at all N levels and the decrease was much more prominent in low N than that in high N, but the starch allocation between shoot and root was unaltered. The analysis of chlorophyll fluorescence and SBPase activity indicated that the decrease of growth and photosynthesis at different N levels were not related to the function of PSII but to the activity of SBPase. Western blot analysis showed the content of SBPase in thylakoid membranes was much more than in the stroma fractions in transgenic plants at low N. Results suggested that low N in addition to a 34% decrease in SBPase activity is sufficient to diminish photosynthesis and limit biomass production. Decreased SBPase activity may reduce the N use efficiency of photosynthesis and growth and alter biomass allocation.     

Palmitoylation modification of Gαo depresses its susceptibility to GAP-43 activation

Interaction between GAP-43 (growth associated protein-43) and Gα_o (alpha subunit of Go protein) influences the signal transduction pathways leading to differentiation of neural cells. GAP-43 is known to increase guanine nucleotide exchange by Gα_o, which is a major component of neuronal growth cone membranes. However, it is not clear whether GAP-43 stimulation is related to the Gα_o palmitoylation or the conversion of Gα_o from oligmers to monomers, which was shown to be a necessary regulatory factor in GDP/GTP exchange of Gα_o. Here we expressed and purified GAP-43, GST-GAP-43 and Gα_o proteins, detected their stimulatory effect on [³⁵S]-GTPγS binding of Gα_o. It was found that the EC₅₀ of both GAP-43 and GST-GAP-43 activation were tenfold lower in case of depalmitoylated Gα_o than palmitoylated Gα_o. Non-denaturing gel electrophoresis and p-PDM cross-linking analysis revealed that addition of GST-GAP-43 induced disassociation of depalmitoylated Gα_o from oligomers to monomers, but did not influence the oligomeric state of palmitoylated Gα_o, which suggests that palmitoylation is a key regulatory factor in GAP-43 stimulation on Gα_o. These results indicated the interaction of GAP-43 and Gα_o could accelerate conversion of depalmitoylated Gα_o but not palmitoylated Gα_o from oligomers to monomers, so as to increase the GTPγS binding activity of Gα_o. Results here provide new evidence about how signaling protein palmitoylation is involved in the G-protein-coupled signal transduction cascade, and give a useful clue on the participation of GAP-43 in G-protein cycle by its preferential activation of depalmitoylated Gα_o.     

Hitchhiking of Cu/Zn Superoxide Dismutase to Peroxisomes – Evidence for a Natural Piggyback Import Mechanism in Mammals

Most newly synthesized peroxisomal proteins are imported in a receptor-mediated fashion, depending on the interaction of a peroxisomal targeting signal (PTS) with its cognate targeting receptor Pex5 or Pex7 located in the cytoplasm. Apart from this classic mechanism, heterologous protein complexes that have been proposed more than a decade ago are also to be imported into peroxisomes. However, it remains still unclear if this so-called piggyback import is of physiological relevance in mammals. Here, we show that Cu/Zn superoxide dismutase 1 (SOD1), an enzyme without an endogenous PTS, is targeted to peroxisomes using its physiological interaction partner 'copper chaperone of SOD1' (CCS) as a shuttle. Both proteins have been identified as peroxisomal constituents by 2D-liquid chromatography mass spectrometry of isolated rat liver peroxisomes. Yet, while a major fraction of CCS was imported into peroxisomes in a PTS1-dependent fashion in CHO cells, overexpressed SOD1 remained in the cytoplasm. However, increasing the concentrations of both CCS and SOD1 led to an enrichment of SOD1 in peroxisomes. In contrast, CCS-mediated SOD1 import into peroxisomes was abolished by deletion of the SOD domain of CCS, which is required for heterodimer formation. SOD1/CCS co-import is the first demonstration of a physiologically relevant piggyback import into mammalian peroxisomes.     

Non-Additive Effects of Water and Nitrogen Addition on Ecosystem Carbon Exchange in a Temperate Steppe

Changes in precipitation and nitrogen (N) deposition can influence ecosystem carbon (C) cycling and budget in terrestrial biomes, with consequent feedbacks to climate change. However, little is known about the main and interactive effects of water and N additions on net ecosystem C exchange (NEE). In a temperate steppe of northern China, a field-manipulated experiment was conducted to evaluate the responses of NEE and its components to improve N and water availability from 2005 to 2008. The results showed that both water and N additions stimulated gross ecosystem productivity (GEP), ecosystem respiration (ER), and NEE. Water addition increased GEP by 17%, ER by 24%, and NEE by 11% during the experimental period, whereas N addition increased GEP by 17%, ER by 16%, and NEE by 19%. The main effects of both water and N additions changed with time, with the strongest water stimulation in the dry year and a diminishing N stimulation over time. When water and N were added in combination, there were non-additive effects of water and N on ecosystem C fluxes, which could be explained by the changes in species composition and the shifts of limiting resources from belowground (water or N) to aboveground (light). The positive water and N additions effects indicate that increasing precipitation and N deposition in the future will favor C sequestration in the temperate steppe. The non-additive effects of water and N on ecosystem C fluxes suggest that multifactor experiments are better able to capture complex interactive processes, thus improving model simulations and projections.     

HisE11 and HisF8 Provide Bis-histidyl Heme Hexa-coordination in the Globin Domain of Geobacter sulfurreducens Globin-coupled Sensor

Among heme-based sensors, recent phylogenomic and sequence analyses have identified 34 globin coupled sensors (GCS), to which an aerotactic or gene-regulating function has been tentatively ascribed. Here, the structural and biochemical characterization of the globin domain of the GCS from Geobacter sulfurreducens (GsGCS¹⁶²) is reported. A combination of X-ray crystallography (crystal structure at 1.5 Å resolution), UV-vis and resonance Raman spectroscopy reveals the ferric GsGCS¹⁶² as an example of bis-histidyl hexa-coordinated GCS. In contrast to the known hexa-coordinated globins, the distal heme-coordination in ferric GsGCS¹⁶² is provided by a His residue unexpectedly located at the E11 topological site. Furthermore, UV-vis and resonance Raman spectroscopy indicated that ferrous deoxygenated GsGCS¹⁶² is a penta-/hexa-coordinated mixture, and the heme hexa-to-penta-coordination transition does not represent a rate-limiting step for carbonylation kinetics. Lastly, electron paramagnetic resonance indicates that ferrous nitrosylated GsGCS¹⁶² is a penta-coordinated species, where the proximal HisF8-Fe bond is severed.     

上海地区番茄黄化曲叶病毒病的鉴定及嫁接接种法研究

番茄黄化曲叶病毒(tomato yellow leafcurl virus,TYLCV)是一种由烟粉虱(Bemisia tabaci)和嫁接传播的双生病毒,在热带、亚热带地区给番茄生产造成严重威胁.根据番茄黄化曲叶病毒的保守序列设计一对引物,运用PCR技术从上海地区的感病番茄中扩增出一条575bp的特异带,而健康植株无此带.测序表明该序列与番茄黄化曲叶病毒具有极高的同源性(97%～99%).将健康接穗嫁接到感染番茄黄化曲叶病毒的番茄砧木上,间隔15 d和30 d,分别提取接穗的DNA,并用PCR法检测病毒,发现嫁接15 d后在部分接穗中检测到TYLCV病毒,嫁接30 d后在所有的接穗中均检测到病毒,因此,嫁接法可以作为番茄黄化曲叶病毒病的接种鉴定方法.     

Twenty-one novel tri- and tetranucleotide microsatellite loci for the Amur tiger (Panthera tigris altaica)

Amur tiger is the largest subspecies of tiger in the world and his conservation has also received much attention. In this study, we isolated and characterized twenty-one tri- and tetranucleotide microsatellite markers from this species. The number of alleles for each locus ranged from two to nine in a group of 60 individuals and the observed and expected heterozygosities were 0.333–0.917 and 0.302–0.822, respectively. The overall discrimination power and exclusion probabilities in parentage and paternity testing for these markers were 1.00, 0.9947 and 0.9999, respectively, indicating high-resolution power of microsatellite markers.