Effects of saline and osmotic stress on proline and sugar accumulation in Populus euphratica in vitro

The use of in vitro shoot cultures to evaluate osmotic and salt tolerance and the effects of salt and mannitol in the medium on proline and sugar accumulation were investigated in two poplar species, P. euphratica and P. alba cv. Pyramidalis × P. tomentosa. Shoot length, leaf number, whole plant dry weight, and the accumulation of proline and total soluble sugars in leaves were quantified after 2 weeks. All P. euphratica plantlets survived at all levels of mannitol and NaCl, while the mortality of P. alba cv. Pyramidalis × P. tomentosa increased both at the mannitol and the NaCl treatments. A significant increase in proline accumulation was observed in both young and mature P. euphratica leaves at 200 mM mannitol and above, and at 150 mM NaCl and above. The total soluble sugar content increased in young P. euphratica leaves at 250 mM NaCl; however, it decreased in the mature leaves. Similar increases of the total soluble sugar content were not seen in P. alba cv. Pyramidalis × P. tomentosa plants in response to either mannitol or NaCl treatment. Our results suggest that accumulated proline and sugars promote osmotic and salt tolerance. The effects of accumulated proline and total soluble sugars on leaves are discussed in relation to growth and osmotic adjustment. This revised version was published online in August 2006 with corrections to the Cover Date.     

Taxane Production in Suspension Culture of <Emphasis Type="Italic">Taxus</Emphasis> × <Emphasis Type="Italic">Media</Emphasis> var. <Emphasis Type="Italic">Hicksii</Emphasis> Carried Out in Flasks and Bioreactor

Paclitaxel and 10-deacetylbaccatin III (10-DAB III) were produced in suspension cultures of Taxus × media var. Hicksii grown in shake-flasks and in a 7-l bioreactor reaching, in the bioreactor, 4.4 mg l⁻¹ (on day 14) and 37.5 mg l⁻¹ (on day 11). In shake-flasks the highest total content of paclitaxel and 10-DAB III was 7.3 mg l⁻¹ (on day 4) and 8.8 mg l⁻¹ (on day 18). Phenylalanine, at 0.05 mM, increased paclitaxel accumulation in cells cultivated in bioreactor and flasks 30-fold and 9-fold (from 0.02 mg l⁻¹ to 0.6 mg l⁻¹ and to 0.2 mg l⁻¹, respectively). The 10-DAB III content in cells from flasks was increased from 0.4 mg l⁻¹ to 1.6 mg l⁻¹.     

不同温度条件下水稻种子活力QTL的定位分析

为了揭示基因型与环境温度之间的互作对种子活力的影响,利用1个粳籼交来源的重组自交系群体,采用纸卷法在15、20和25℃条件下进行发芽试验,考察了发芽率、芽长、根长及干重等4个种子活力相关性状。结合一张含有198个DNA标记的连锁图谱,用作图软件QTL Mapper1.0定位与种子活力相关的QTL。共检测到34个主效应QTL。这些QTL中的绝大多数(82％)成簇分布于第3、5和8号染色体的5个不同染色体区段上,分别被命名为QTL qSV-3-1、qSV-3-2、qSV-5、qSV-8-1和qSV-8-2。其中,QTL qSV-3-1、qSV-3-2和qSV-8-1对种子活力的效应大小和方向在3个温度条件下均较一致;而QTL qSV-5和qSV-8-2主要在20和25℃条件下起作用,在15℃低温条件下作用甚微或不起作用。表明种子活力QTL具有显著的基因型与环境温度之间的互作,而且这种互作具有明显的QTL特异性。芽长是唯一同时受5个与种子活力高度相关的染色体区段共同影响的指标,因此,相对而言,作为水稻种子活力的测定指标,芽长是最具有代表性的。     

Temporary flooding increases iron phytoavailability in calcareous Vertisols and Inceptisols

Temporary soil flooding before cultivation alleviates iron chlorosis in crops grown on some calcareous Mexican Vertisols. In order to investigate the effectiveness of such practice we carried out experiments with ten calcareous Vertisols from Mexico and eight calcareous Inceptisols from Spain. In an incubation experiment, we studied the release of Fe²⁺ into the solution of soil suspensions in sealed vials with 5 m M CaCl₂. In a pot experiment, we measured the leaf SPAD value (i.e. an estimate of leaf chlorophyll concentration) of lupin and strawberry sequentially grown on a soil-sand mixture previously flooded for 30 days (SPAD_f value) and on a non-flooded (control) mixture (SPAD_c value). The amount of Fe²⁺ released by the soil at day 58 and the increase in oxalate-extractable Fe (Fe_o) upon incubation in vials were larger on average for the Inceptisols than for the Vertisols. The SPAD_c values for lupin and strawberry were (i) larger for the Vertisols than for the Inceptisols (probably because the Vertisols contain little carbonate and induce less Fe chlorosis than the Inceptisols) and (ii) correlated with Fe_o, and with citrate/ascorbate- and DTPA-extractable Fe (Fe_ca, Fe_DTPA). The SPAD_f-SPAD_c differencewas (i) much larger for the Inceptisols than for the Vertisols and (ii) correlated with the increases in Fe_o and Fe_ca caused by flooding and with the amount of Fe²⁺ released in the incubation experiment. We hypothesize that the weak response of the Vertisols to flooding was partly a result of their history including flooding episodes in the field, so a steady state had been reached in which the pool of Fe compounds undergoing reductive dissolution and reprecipitating upon oxidation as poorly crystalline Fe oxides (the main source of phytoavailable Fe) remained relatively constant and thus changed little after pot flooding. The Inceptisols, which had never been flooded in the field, were capable of releasing Fe from sources other than poorly crystalline Fe oxides upon flooding, thus making this treatment effective against Fe chlorosis. Our results point to the need to further study those soil chemical and mineralogical properties that are related to increases in Fe phytoavailability upon temporary soil flooding.     

Recombinant expression,purification, and characterization of polyphenol oxidase 2 (VvPPO2) from “Shine Muscat” (Vitis labruscana Bailey × Vitis vinifera L.)

Polyphenol oxidases (PPOs) catalyze browning reactions in various plant organs, therefore controlling the reactions is important for the food industry. PPOs have been assumed to be involved in skin browning of white grape cultivars; however, the molecular mechanism underlying PPO-mediated browning process remains elusive. We have recently identified a new PPO gene named VvPPO2 from “Shine Muscat” (Vitis labruscana Bailey × V. vinifera L.), and have shown that the gene is transcribed at a higher level than the previously identified VvPPO1 in browning, physiologically disordered berry skins at the maturation stage. In this study, we expressed VvPPO2 in Escherichia coli and, using the purified preparation, revealed unique physicochemical characteristics of the enzyme. Our study opens up a way to not only understand the berry skin browning process but also to elucidate the enzymatic maturation process of grape PPOs.     

The effect of daily exposure to low hardening temperature on plant vital activity

Phenomenological responses of plants to daily short-term exposure to low hardening temperature was studied under chamber and field conditions. Experiments were carried out on cucumber (Cucumis sativus L.), barley (Hordeum vulgare L.), marigolds (Tagetes L.), and petunia (Petunia × hybrida) plants. The obtained data demonstrated a similar pattern of response in all studied plant species to different variants of exposure to low hardening temperature. The main features of plant response to daily short-term exposure to low hardening temperature include: a higher increment in cold tolerance (cf. two-or threefold increase relative to constant low hardening temperature) that peaked on day 5 (cf. day 2 at constant low hardening temperature) and was maintained for 2 weeks (cf. 3–4 days at constant low hardening temperature); a simultaneous increase in heat tolerance (cf. twofold relative to constant low hardening temperature) maintained over a long period (cf. only in the beginning of the exposure to constant low hardening temperature); a sharp drop in the subsequent cold tolerance after plant incubation in the dark (cf. a very low decrease in cold tolerance following the exposure to constant low hardening temperature); a combination of high cold tolerance and high photochemical activity of the photosynthetic apparatus (cf. a low non-photochemical quenching at constant low hardening temperature); and the capacity to increase cold tolerance in response to repeated short-term exposures to low hardening temperature in plants grown outdoors (cf. a gradual increase after repeated exposure to constant low hardening temperature). Possible mechanisms underlying the plant response to daily short-term exposure to low temperature are proposed.     

Purification and characterization of levoglucosan kinase from <Emphasis Type="Italic">Lipomyces starkeyi</Emphasis> YZ-215

A protein named as levoglucosan kinase (EC 2.7.-.-)was purified to homogeneity from a wild isolated strain of Lipomyces starkeyi YZ-215. The protein was purified approximately 30-fold by conventional ammonium sulphate fractionation followed by Resource Q chromatography and two steps of Superdex 200 chromatography, and its physical and kinetic properties were investigated. The purified enzyme showed a molecular weight of 48 kDa by SDS-PAGE and 47.7 kDa by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), respectively. The enzyme was stable at pH 7–10 and showed maximum activity at 30°C and pH 9.0. Kinetic constants (apparent K _m values) for levoglucosan and ATP were 68.6 ± 13.7 mM and 0.68 ± 0.06 mM, respectively. After in-gel digestion by trypsin, three peptides were sequenced and analyzed by electrospray ionization quadrupole time-of-flight tandem mass spectrometry (ESI-Q-TOF MS/MS). Data of the amino acid sequences indicated that this protein might be a novel kinase. The purification of levoglucosan kinase from L. starkeyi YZ-215 represented a fundamental step to provide insights into the efficient utilization of cellulosic pyrolysate by bioconversion.     

Growth-suppressive function of human connexin32 in a conditional immortalized mouse hepatocyte cell line

Mouse hepatocytes immortalized with a temperature-sensitive allele of the SV40 large T-antigen (CHST8 cells) were found to lack the high expression of the gap junction proteins Cx26 and Cx32 that characterizes normal mouse hepatocytes, expressing instead Cx43 and Cx45 at minimal levels. In order to examine the growth suppressive function of Cx32 on hepatocytes, we transfected these CHST8 cells with human Cx32 complementary deoxyribonucleic acid and measured the growth rates at 33, 37, and 39 degrees C. Expression of human Cx32 and its messenger ribonucleic acid in the stable cell lines was confirmed by immunocytochemistry and by Western and Northern blots analyses. Dye transfer following lucifer yellow injection into the transfectants was extensive; Cx32 channels displayed unitary conductances of about 70 pS and were moderately voltage sensitive. When cultured at 33 and 39 degrees C, growth rates of both parental cells and transfectants were of the same level. When examined at 37 degrees C, growth rate of the transfectant, which highly expressed Cx32 at the membranes, was significantly decreased compared to the parental cells. However, no changes in the expression of Cx32 protein in the transfectants were observed between 33 and 37 degrees C. These results suggest that Cx32 expression could inhibit hepatocyte growth in vitro using the conditional immortalized cells. Cx32 transfectants using a conditional immortalized mouse hepatocyte may be useful for examining the mechanisms of growth and differentiation in hepatocytes by gap junction expression.     

QTLs for agronomic traits from a Mediterranean barley progeny grown in several environments

In order to identify quantitative trait loci (QTLs) controlling agronomic trait variation and their consistency under Mediterranean conditions in barley, a progeny of 167 recombinant inbred lines (RILs) and the parents Tadmor and Er/Apm, originating from the Mediterranean basin, were grown under Mediterranean conditions in 1995, 1996, 1997 and 1999. For the 2 first years (M95 and G96), one replicate was grown, but for the latter (M97 and M99) two rainfed (rain) and two irrigated (ir) replicates were produced. M95, G96, M97rain, M97ir, M99rain and M99ir were considered as six different environments and were compared in terms of their meteorological conditions and water supply. Grain yield and yield components were assessed, as well as heading date and plant height. Highly significant differences were noted between environments. QTLs were obtained from each environment separately and from a multiple environment analysis (simple interval mapping and simplified composite interval mapping). Despite heterogeneity between environments, numerous QTLs were common to several environments. This was particularly true for traits like plant height and thousand-grain weight. The most reliable QTLs which explained the largest part of the phenotypic variation were obtained for plant height on chromosomes 3 (3H) and 6 (6H). The multiple-environment analysis provided an opportunity to identify consistent QTLs for agronomic traits over six Mediterranean environments. A total of 24 consistent QTLs were detected. Out of these, 11 presented main effects, seven presented QTL×E interaction, and six presented both effects. In addition, 18 of the consistent QTLs were common to other published work and six seemed specific to this study. These latter QTLs could be involved in Mediterranean adaptive specificities or could be specific to the studied genetic background. Finally, when the rainfed and the irrigated environments of M97 were considered separately, a total of 16 QTLs presenting main effects over the two water conditions were identified, whereas five QTLs seemed dependent on the water conditions. Received: 31 January 2001 / Accepted: 19 February 2001