Equilibrium centrifugation of proteins in acidic solutions. Beta-lactoglobulin A in aqueous acetic, propionic, and butyric acids.

Short chain aliphatic acids are almost neutrally buoyant in aqueous solutions, and preferential interaction of macromolecules with these solvent components should not greatly affect apparent molecular weights determined by equilibrium ultracentrifugation. The feasibility of molecular weight estimations using native, neutral pH values of partial specific volume has been tested: equilibrium ultracentrifugation of β-lactoglobulin A (β-LgA) has been carried out in aqueous acetic, propionic, and butyric acids in the absence of any other added electrolyte. These solutions are highly nonideal because of the extreme Donnan effect. Apparent molecular weights estimated at infinite dilution using the native neutral pH value of the partial specific volume, $\text{v}{}_{\mathrm{p}}$, differed by less than 5% from the monomer formula weight. The 10 m acids appear to be least effective as dissociating agents for β-LgA, with a weak reversible monomer-dimer association suggested in 10 m acetic acid, with significant heterogeneity apparent in 10 m propionic acid, and with a lack of direct solubility in 10 m butyric acid. All the 0.1 m acids and all the 1 m acids were essentially equally effective as dissociating agents, with the exception of 1 m butyric acid which dissolved β-LgA only slowly to give significantly heterogeneous solutions. From these results and from our previous experiments with aldolase (6), it appears feasible to use the native values of $\text{v}{}_{\mathrm{p}}$ to obtain estimates of molecular weights of proteins in aqueous organic acids as dissociating agents.     

Purification and properties of ATPase inhibitor from rat liver mitochondria

(1) The ATPase inhibitor protein has been isolated from rat liver mitochondria in purified form. The molecular weight determined by sodium dodecyl sulfate gel electrophoresis is approximately 9500, and the isoelectric point is 8.9.

(2) The protein inhibits both the soluble ATPase and the particle-bound ATPase from rat liver mitochondria. It also inhibits ATPase activities of soluble F₁, and inhibitor-depleted submitochondrial particles derived from bovine heart mitochondria.

(3) On particle-bound ATPase the inhibitor has its maximal effect if incubated in the presence of Mg²⁺. ATP at slightly acidic pH.

(4) The inhibitor has a minimal effect on P_i-ATP exchange activity in sonicated submitochondrial particles. However, unexpectedly the inhibitor greatly stimulates P_i-ATP exchange activity in whole mitochondria while the low ATPase activity of the mitochondria is not affected. The possible mechanism of action of the inhibitor on intact mitochondria is offered.     

Differentiation-associated changes in membrane proteins of mouse myeloid leukemia cells

The mouse myeloid leukemia cell line (M1) is known to differentiate in vitro into macrophages and granulocytes upon treatment with various inducer including mouse ascitic fluid. Changes of cell surface proteins during differentiation of M1 cells were analyzed by the lactoperoxidase-catalyzed radioiodination method and SDS-polycrylamide slab gel electrphoresis. Treatment of the cells with ascitic fluid changed the electrophoretic pattern of the iodinated proteins, the prominent change being the appearance of a new protein with a molecular weight of 180 000 (P180). Iodinated P180 was also detected in normal macrophages in granulocytes, which are similar to differentiated M1 cells. This protein was metabolically labeled with l-[¹⁴C]fucose, increasing with the period of the treatment. P180 was not expressed on ascitic fluid-treatment of a resistant clone of M1 cells that could not be induced to differentiate. These results indicate that P180 is a glycoprotein that is exposed on the outer surface of differentiated M1 cells, and that its expression is associated with differentiation of the cells.P180 was solubilized from ¹²⁵I-labeled macrophages with detergents bound to concanavalin A-Sepharose. This suggests that P180 is one of the receptors for concanavalin A. Therefore, P180 may contribute partly to the increases in agglutinability by concanavalin A and in the number of concanavalin A binding sites on the surface of M1 cells, which are known to be associated with differentiation of M1 cells.     

miR-125b-5p促进分枝杆菌在宿主细胞和小鼠体内存活的研究

通过观察miR-125b-5p对分枝杆菌在宿主细胞和小鼠体内存活情况的影响,探究其在抗结核免疫过程中的作用。采用不同培养基对分枝杆菌进行培养并计数;以1640培养基加10%胎牛血清培养所有实验用细胞。将终浓度50 nmol/L的miR-125b-5p 模拟物、miR-125b-5p 抑制剂及磷酸盐缓冲液(PBS)对照加入细胞后,在不同时间点收集细胞。用分枝杆菌分别感染宿主细胞(A549、THP-1和RAW264.7)以及C57BL/6小鼠。采用定量聚合酶链反应检测miR-125b-5p的表达量。结果miR-125b-5p在分枝杆菌感染的多种宿主细胞及小鼠中都显著上调表达,其中小鼠肺部的表达量提高了约15倍。分别转染模拟物和抑制剂后,再用分枝杆菌感染细胞,结果发现miR-125b-5p可促进分枝杆菌在宿主细胞内的生长。当miR-125b-5p抑制剂注射到卡介苗(BCG)感染的小鼠体内时,小鼠体内的细菌载量显著降低(P<0.05)。本研究证明miR-125b-5p可调控分枝杆菌在宿主细胞及小鼠体内的生长,在抗结核免疫过程中发挥了重要作用。进一步对其作用机制的深入研究将为临床结核病的治疗提供理论指导。     

Seed dormancy and germination in Dodonaea viscosa (Sapindaceae) from south-western Saudi Arabia

Dodonaea viscosa (Sapindaceae) is widespread in the mountainous highlands of the southwestern part of Kingdom of Saudi Arabia, where it is a medicinally important species for the people in Saudi Arabia. Seeds of this species were collected from Mount Atharb in Al-Baha region, at an altitude of 2100 m. The aims of this study were to determine if the seeds of D. viscosa have physical dormancy (i.e. a water-impermeable seed coat) and, if so, what treatments would break dormancy, and what conditions promote germination after dormancy has been broken. The dormancy-breaking treatments included: soaking of seeds in concentrated sulfuric acid (H₂SO₄) for 10 min, immersion in boiling water for 10 min and exposure to 50 °C for 1 min. After seeds had been pre-treated with H₂SO₄, to break dormancy, they were incubated at constant temperatures from 5 to 35 °C, under 12-h photoperiods or in continuous darkness, and germination recorded. Salinity tolerance was investigated by incubating acid-scarified seeds in different concentrations of mM NaCl in the light at 25 °C.Untreated seeds had low final germination 30%. Seeds that had been acid-scarified, immersed in boiling water or exposed to 50 °C all achieved 91% subsequently when incubated at 25 °C. Thus, seeds of this species in Saudi Arabia have physical dormancy, which can be broken by all three treatments designed to increase the permeability of the testa. After pre-treatment, there was a broad optimum constant temperature for germination that ranged between 5 and 25 °C but germination was inhibited by higher temperatures (30 and 35 °C). Light had little effect on this germination response. Scarified seeds were also sensitive to salinity, with the highest germination in distilled water and complete inhibition in 400 mM NaCl. Seeds that failed to germinate in saline treatments were mostly able to germinate on transfer to distilled water, suggesting osmotic inhibition.     

外源水杨酸对NaCl胁迫下白榆幼苗光合作用及离子分配的影响

以1年生白榆幼苗为研究对象,设置0、0.5、1.0和2.0 mmol·L^-1水杨酸(SA)与0、50、100和150 mmol·L^-1 NaCl处理组合,考察盐胁迫下白榆幼苗生物量、光合色素含量、光合作用参数及根叶离子含量、分配、运输的情况,探讨外源SA对NaCl胁迫下白榆幼苗耐盐生理特征的影响。结果表明:(1)NaCl胁迫显著抑制了白榆幼苗的生长、光合色素含量及光合能力,并破坏了白榆体内离子平衡。(2)喷施外源SA使盐胁迫下白榆幼苗的干重和根冠比均不同程度升高,0.5和2.0 mmol·L^-1 SA不同程度提高了50和100 mmol·L^-1 NaCl处理组幼苗叶片光合色素含量。(3)0.5 mmol·L^-1 SA显著提升了50 mmol·L^-1 NaCl处理组白榆幼苗的净光合速率(Pn)、气孔导度(Gs)和蒸腾速率(Tr),1.0和2.0 mmol·L^-1 SA对150 mmol·L^-1 NaCl处理组幼苗净光合速率改善效果较好,外源SA对100 mmol·L^-1 NaCl处理组幼苗的光合作用参数无显著影响。(4)NaCl胁迫下,外源SA处理的白榆幼苗叶和根Na^+含量及Na^+/K^+、Na^+/Ca^2+和Na^+/Mg^2+显著降低,离子选择运输系数SK,Na、SCa,Na和SMg,Na升高,从而促进了幼苗K^+、Ca^2+和Mg^2+由根向叶片的转运;隶属函数分析发现对白榆幼苗叶和根中离子含量改善效果最好的SA浓度分别为1.0和2.0 mmol·L^-1。因此,适宜浓度的外源水杨酸能够有效改善NaCl胁迫下白榆幼苗的光合能力,有效调节白榆幼苗体内离子状态,从而增强白榆对NaCl胁迫的抗性。     

Response of greenhouse tomato to salt stress and K+ supplement

Abstract

The effect of NaCl salinity and potassium supplement on growth, tissue ion concentration, photosynthesis, yield and fruit quality characteristics of tomato plants was studied. Tomato plants, hyb. Belladonna, were grown in 8.5 l pots, filled with 1:3 sand:perlite mixture and irrigated with a half-strength Hoagland solution through a closed hydroponic system. Six irrigation treatments were applied, including combinations of 3 salinity (0, 35 and 70 mM NaCl) and two potassium levels (K1: 200 ppm and K2: 400 ppm) in the nutrient solution. Salinity reduced photosynthesis resulting in reduced plant height and dry weight. Yield was reduced by 25% and 69% at 35 and 70 mM, respectively, as compared to control plants (0 mM NaCl). Both total soluble solids and titratable acidity of the fruit increased with increasing salinity and K levels. The application of high potassium level (K2) reduced the concentration of Na and increased that of K in the leaves and roots of the plants, as compared to K1 treatment. Toxicity symptoms were mostly observed in the leaves of 70K1 plants, while no visual symptoms of toxicity were observed in 70K2 treatment. Despite the positive effects of potassium supplement in reducing Na concentration and the absence of toxicity symptoms in the leaves, plant growth was not improved, while leaf photosynthesis was reduced. Furthermore, no positive effects in the percentage of marketable fruit, mean fruit weight and yield were observed in the plants receiving extra K.     

<i>Trichoderma</i>-Induced Improvement in Growth,Photosynthetic Pigments,Proline, and Glutathione Levels in <i>Cucurbita pepo</i> Seedlings under Salt Stress

Salt stress is one of the major abiotic stress in plants. However, traditional approaches are not always efficient in conferring salt tolerance. Experiments were conducted to understand the role of Trichoderma spp. (T. harzianum and T. viride) in growth, chlorophyll (Chl) synthesis, and proline accumulation of C. pepo exposed to salinity stress. There were three salt stress (50, 100, and 150 mM NaCl) lavels and three different Trichoderma inoculation viz. T. harzianum, T. viride, and T. harzianum + T. viride. Salt stress significantly declined the growth in terms of the shoot and root lengths; however, it was improved by the inoculation of Trichoderma spp. C. pepo inoculated with Trichoderma exhibited increased synthesis of pigments like chl a, chl b, carotenoids, and anthocyanins under normal conditions. It was interesting to observe that such positive effects were maintained under salt-stressed conditions, as reflected by the amelioration of the salinity-mediated decline in growth, physiology and antioxidant defense. The inoculation of Trichoderma spp. enhanced the synthesis of proline, glutathione, proteins and increased the relative water content. In addition, Trichoderma inoculation increased membrane stability and reduced the generation of hydrogen peroxide. Therefore, Trichoderma spp. can be exploited either individually or in combination to enhance the growth and physiology of C. pepo under saline conditions.     

Salinity stress response of non-transformed and AtCKX transgenic centaury (Centaurium erythraea Rafn.) shoots and roots grown in vitro

Common centaury (Centaurium erythraea Rafn.) is a plant species that can inhabit saline soils. It is known as a plant with high spontaneous regeneration potential in vitro. In the present work we evaluated shoots and roots salinity tolerance of non-transformed and three AtCKX transgenic centaury lines to graded NaCl concentrations (0, 50, 100, 150, 200 mM) in vitro. Overexpression of AtCKX genes in transgenic centaury plants resulted in an altered cytokinins (CKs) profile leading to a decline of bioactive CK levels and, at the same time, increased contents of storage CK forms, inactive CK forms and/or CK nucleotides. Significant increment of fresh shoot weight was obtained in shoots of non-transformed and AtCKX1 transgenic line only on medium supplemented with 50 mM NaCl. However two analysed AtCKX2 transgenic lines reduced shoot growth at all NaCl concentrations. In general, centaury roots showed higher tolerance to salinity than shoots. Non-transformed and AtCKX1 transgenic lines tolerated up to 100 mM NaCl without change in frequency of regeneration and number of regenerated plants. Roots of two analysed AtCKX2 transgenic lines showed different regeneration potential under salt stress. Regeneration of transgenic AtCKX2-26 shoots even at 200 mM NaCl was recorded. Salinity stress response of centaury shoots and roots was also evaluated at biochemical level. Free proline, malondialdehyde and hydrogen peroxide content as well as antioxidative enzymes activities were investigated in shoots and roots after 1, 2, 4 and 8 weeks. In general, adition of NaCl in culture medium elevated all biochemical parameters in centaury shoots and in roots. Considering that all analysed AtCKX transgenic centaury lines showed altered salt tolerance to graded NaCl concentrations in vitro it can be assumed that CKs might be involved in plant defence to salt stress conditions.