Effect of Cerium on Photosynthetic Pigments and Photochemical Reaction Activity in Soybean Seedling Under Ultraviolet-B Radiation Stress

Effects of cerium (Ce) on photosynthetic pigments and photochemical reaction activity in soybean (Glycine max L.) under ultraviolet-B (UV-B) radiation stress were studied under laboratory conditions. UV-B radiation caused the decrease in chlorophyll content, net photosynthetic rate, Hill reaction activity, photophosphorylation rate and Mg²⁺-ATPase activity. Ce (III) (20 mg L⁻¹) could alleviate UV-B-induced inhibition to these photosynthetic parameters because values of these photosynthetic parameters in Ce (III) + UV-B treatment were obviously higher than those with UV-B treatment alone. Dynamic changes of the above photosynthetic parameters show that Ce (III) could slow down the decrease rate of these photosynthetic parameters during a 5-day UV-B radiation and quicken the restoration during recovery period. The final restoration degree of five parameters mentioned above in leaves exposed to low level of UV-B radiation (0.15 W m²) was higher than that exposed to high level (0.45 W m²). Correlating net photosynthetic rate with other four parameters, we found that the regulating mechanisms Ce (ΠΙ) on photosynthesis under various level of UV-B radiation were not the same. The protective effects of Ce (III) on photosynthesis in plants were influenced by the intensity of UV-B radiation.     

In Vivo Esr Studies of Antioxidant Activity on Free Radical Reaction in Living Mice Under Oxidative Stress

In vivo antioxidant activity seems to be quite complicate due to multiple interaction with biomaterials and differs from results by in vitro experiments. In vivo estimation of antioxidant activity is performed by measuring TBA reactive substances in blood or hydrocarbon gases in breath, but these systems do not measure free radical reaction but the final products of oxidative reaction. In the present study, we applied in vivo ESR to evaluate antioxidant activity by monitoring the redox reaction of nitroxide radical and clearly found that the nitroxide is very susceptible to oxidative stress in vivo and quite useful to evaluate antioxidant activity non-invasively.     

Conserved,Disordered C Terminus of DnaK Enhances Cellular Survival upon Stress and DnaK in Vitro Chaperone Activity

The 70-kDa heat shock proteins (Hsp70s) function as molecular chaperones through the allosteric coupling of their nucleotide- and substrate-binding domains, the structures of which are highly conserved. In contrast, the roles of the poorly structured, variable length C-terminal regions present on Hsp70s remain unclear. In many eukaryotic Hsp70s, the extreme C-terminal EEVD tetrapeptide sequence associates with co-chaperones via binding to tetratricopeptide repeat domains. It is not known whether this is the only function for this region in eukaryotic Hsp70s and what roles this region performs in Hsp70s that do not form complexes with tetratricopeptide repeat domains. We compared C-terminal sequences of 730 Hsp70 family members and identified a novel conservation pattern in a diverse subset of 165 bacterial and organellar Hsp70s. Mutation of conserved C-terminal sequence in DnaK, the predominant Hsp70 in Escherichia coli, results in significant impairment of its protein refolding activity in vitro without affecting interdomain allostery, interaction with co-chaperones DnaJ and GrpE, or the binding of a peptide substrate, defying classical explanations for the chaperoning mechanism of Hsp70. Moreover, mutation of specific conserved sites within the DnaK C terminus reduces the capacity of the cell to withstand stresses on protein folding caused by elevated temperature or the absence of other chaperones. These features of the C-terminal region support a model in which it acts as a disordered tether linked to a conserved, weak substrate-binding motif and that this enhances chaperone function by transiently interacting with folding clients.     

Effects of Osmoprotectants upon NaCl Stress in Rice

Plants accumulate a number of osmoprotective substances in response to NaCl stress, one of them being proline (Pro). While characterizing some of the changes in solute accumulation in NaCl-stressed rice (Oryza sativa L.), we identified several other potential osmoprotectants. One such substance, trehalose, begins to accumulate in small amounts in roots after 3 d. We performed a series of experiments to compare the effects of Pro and trehalose on ion accumulation to determine whether the two chemicals protect the same physiological processes. We found that Pro either has no effect or, in some cases, exasperates the effect of NaCl on growth inhibition, chlorophyll loss, and induction of a highly sensitive marker for plant stress, the osmotically regulated salT gene. By contrast, low to moderate concentrations of trehalose reduce Na+ accumulation, salT expression, and growth inhibition. Somewhat higher concentrations (10 mM) prevent NaCl-induced loss of chlorophyll in blades, preserve root integrity, and enhance growth. The results of this study indicate that during osmotic stress trehalose or carbohydrates might be more important for rice than Pro.     

基于SOS反应及氧化应激反应相关启动子的辐射生物传感器研究

近年来的动物实验结果表明电磁辐射的危害主要是具有神经系统毒性、诱发肿瘤和生殖系统损伤等,广域、隐蔽和累积效应是辐射的特点,除对机体进行直接损伤外,还可导致间接损伤,即通过产生活性氧(ROS)和自由基攻击生物大分子。为了迅速和简便地检测辐射毒性的大小建立了新型的辐射生物传感器,构建了携带SOS反应和氧化应激反应相关的SulA、RecA、Cda和SoxR四种启动子融合经过密码子简并性优化的增强型绿色荧光蛋白(enhanced green fluorescent protein,EGFP)报告因子的工程菌传感器,并对这些生物传感器进行了γ射线辐照处理,筛选出了针对γ射线响应较好的,优选RecA工程菌传感器。利用PCR和Overlap PCR克隆获得了启动子-报告因子融合基因,并插入表达载体PUC19中,转化入宿主大肠杆菌DH5α,通过提取质粒进行双酶切和测序验证后,将构建成功的工程菌传感器首先进行化学毒性试剂刺激,一旦化学试剂刺激结果阳性便进行物理辐射刺激。结果显示,构建成功的4种工程菌传感器均对物理辐射产生应答,且随物理辐射剂量的增加(0～30Gy),绿色荧光强度逐渐增强。运用合成生物学手段,成功建立基于生物损伤修复效应和氧化应激反应的辐射生物传感器,具有制备简便、结果可视性等优点,能满足快速、广范围、在线监测的需求,在细胞毒性物、辐射环境乃至空间射线的损伤能力测定方面具有良好的应用前景。     

A Critical Examination of Escherichia coli Esterase Activity

The ability of Escherichia coli to grow on a series of acetylated and glycosylated compounds has been investigated. It is surmised that E. coli maintains low levels of nonspecific esterase activity. This observation may have ramifications for previous reports that relied on nonspecific esterases from E. coli to genetically encode nonnatural amino acids. It had been reported that nonspecific esterases from E. coli deacetylate tri-acetyl O-linked glycosylated serine and threonine in vivo. The glycosylated amino acids were reported to have been genetically encoded into proteins in response to the amber stop codon. However, it is our contention that such amino acids are not utilized in this manner within E. coli. The current results report in vitro analysis of the original enzyme and an in vivo analysis of a glycosylated amino acid. It is concluded that the amber suppression method with nonnatural amino acids may require a caveat for use in certain instances.The central question addressed in this paper is whether the glycosylated amino acids GlcNAc-Ser and GalNAc-Thr have been genetically encoded into proteins in vivo (1, 2). The reports for the incorporation of these two amino acids are unique from all other reports (3) that have incorporated unnatural amino acids using the recoded UAG codon and Methanococcus jannaschii orthogonal pairs in that these two amino acids required further processing by the host organism before incorporation (see Fig. 1). Here we posit that the primary barrier to their incorporation would appear to be the fact that the host organism used in the original reports, Escherichia coli, maintains very low levels of nonspecific esterase activity. In fact, the original reports used citations from mammalian biology to substantiate the nonspecific esterase mechanism (see below).Open in a separate windowFIGURE 1.Proposed product of an esterase with GlcNAc-Ser and other esterase substrates discussed in this study.E. coli is likely the most thoroughly studied microorganism. This is especially true in regard to carbohydrate and amino acid uptake and utilization (4). Therefore, it should not be surprising that it has long been known that esterified carbon sources are not metabolized by E. coli in standard assays used to probe for microorganism lipase and esterase activity (5). Such results and our current analysis underscore the limitations of the reports that triacetyl O-linked glycosylated amino acids (GlcNAc-Ser and GalNAc-Thr) were deacetylated in E. coli by endogenous “nonspecific” esterases. The deacetylated amino acids were then believed to have been genetically encoded into full-length proteins in vivo (1, 2).In these previous studies the glycosylated amino acids were provided to the growth media as their tetraacetate analogs, and it was construed from the mass spectra and lectin binding assays that the ester groups of the saccharide had all been hydrolyzed. The notion that E. coli rapidly hydrolyzes a simple ester is not easily reconciled with what is commonly observed when the ester functional group is introduced into cultures of E. coli. For example, we were prompted by reports that claimed to have harvested β-hydroxy esters from E. coli (6). There was nothing in such a report to indicate that the E. coli strain used had undergone a drastic genetic modification beyond the introduction of one enzyme derived from yeast. The enzyme from yeast was expressed in E. coli to asymmetrically reduce β-keto esters to the corresponding β-hydroxy esters. The reduction was accomplished in 87% yield and was performed in whole cells. It stands to reason that such a report having claimed to extract significant amounts of an esterified product would not be possible if E. coli maintained even moderate levels of nonspecific esterase activity. The fact that E. coli maintains low levels of endogenous esterases and lipases has been quite pivotal for a number of studies that have used this organism as the host to express esterase genes in vivo (see below).Nonspecific esterase activity is common in eukaryotic organisms, for example, our ability to hydrolyze triacylglycerides to access an important energy source, but this stands in stark contrast to E. coli where it is possible to directly extract O-acetylated oligosaccharides (7) and other simple esters (6) in high yields. These reports are consistent with the observation that UDP-2,3-diacylglucosamine accumulates in E. coli when genes from lipid biosynthesis are deleted (8). E. coli is also the preferred host for evaluating esterase and lipase activity when screening genes from cultured and uncultured organisms (9, 10). Screening for lipase activity from various microorganisms is often performed on tributyrin agar plates (11). The results are typically the same as for triacetin, and it is repeatedly observed that E. coli does not naturally grow on triesters of glycerol (12, 13). These and many other similar esterase screens (14) would not have been feasible if E. coli produced even moderate levels of a lipase or nonspecific esterase.In the present article we use a combination of our current findings and a thorough review of the relevant literature to conclude that E. coli may not maintain sufficient levels of nonspecific esterase activity to permit the in vivo incorporation of the glycosylated amino acids by the mechanism reported (Fig. 1). Our conclusion is further supported by isothermal calorimetry measurements of Zhang et al. (1) original enzyme showing it retains considerable wild-type activity. We also show that the amino acid GlcNAc-Ser appears to be metabolized in E. coli.     

发育心肌和心肌干细胞分化后心肌特异蛋白表达的比较研究

为探讨MCSC移植修复缺血性心肌的可能性,观察了大鼠发育心肌和MCSC分化为心肌细胞的cTnT和Cx-43表达特点。结果表明从胚胎发育至成年,心肌cTnT mRNA表达逐渐上调;用BMP-2诱导1周的MCSC可检测到cTnT mRNA表达,3～4周表达显著升高,诱导2周能观察到cTnT蛋白,3～4周可见cTnT呈现横纹样结构。胚胎第11 d心肌可检测到Cx-43 mRNA表达,生后7 d达高峰,以后逐渐降低,17 d趋于稳定。胚胎期Cx-43蛋白多分布于肌膜下,生后10 d位于细胞连接处。诱导的MCSC中Cx-43 mRNA表达特征与cTnT mRNA相似,2周可观察到Cx-43蛋白,3～4周可见Cx-43位于相邻细胞连接处。本研究结果提示,在BMP-2诱导下MCSC可分化为心肌细胞,表现为成熟心肌细胞的结构特征。MCSC有望成为治疗缺血性心脏病的理想种子细胞。     

Secondary Rhythms of Cardiac Activity in Rat Ontogeny

Endogenous periodic oscillations of the heart beat rate are described in rat pups aged 3–4, 7–8, 10–11, 13–14, 21–22 days and 1.5 month after birth. These oscillations have all characteristic features established earlier for the secondary rhythms of endogenous contractile activity in the wall of various regions of the gastrointestinal tract and for bursts of spontaneous somatomotor excitation in the early postnatal ontogeny of rats: a multi-stage organization, inconstancy, irregularity of components. In frequency spectra of secondary oscillations of the heart rate obtained by means of fast Fourier transform of R–R intervals of the periodogram, age-related changes of the spectral frequency power are demonstrated in 4 ranges, 0.01–0.03, 0.03–0.1, 0.1–1.0, and 1.0–2.5 Hz, which correspond to the about-one-minute, decasecond, and about-one-second waves of the heart rhythm oscillations and to sinus arrhythmia. It is shown that the dominating frequencies of the secondary rhythms in each range do not have regular age-related changes, which is characteristic of all endogenous secondary rhythms.     

Orexin神经元调节应激反应的研究进展

orexin是下丘脑的一种重要神经肽,在摄食、睡眠和药物成瘾等生理心理过程中起着重要作用.近年来发现下丘脑Orexin能神经纤维密集地投向参与应激调控的脑区;orexin基因敲除的老鼠表现为防御反应迟钝;另外,侧脑室微量注射orexin可导致血浆中ACTH水平的上升并诱导出应激样呼吸-心血管反应和行为反应;因此,下丘脑及中枢orexin系统对应激的调控可能起关键作用.其可能机制为强烈刺激能活化下丘脑orexin神经元,诱导中枢神经系统orexin的释放,从而激活CRF通路及蓝斑-交感-肾上腺髓质系统,提高血浆皮质酮与去甲肾上腺素的水平,诱导应激反应,维持机体的稳定.     

Rubisco Activity: Effects of Drought Stress

Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) activityis modulated in vivo either by reaction with CO₂ and Mg²⁺ tocarbamylate a lysine residue in the catalytic site, or by thebinding of inhibitors within the catalytic site. Binding ofinhibitors blocks either activity or the carbamylation of thelysine residue that is essential for activity. At night, inmany species, 2-carboxyarabinitol-1-phosphate (CA1P) is formedwhich binds tightly to Rubisco, inhibiting catalytic activity.Recent work has shown that tight-binding inhibitors can alsodecrease Rubisco activity in the light and contribute to theregulation of Rubisco activity. Here we determine the influencethat such inhibitors of Rubisco exert on catalytic activityduring drought stress. In tobacco plants, ‘total Rubiscoactivity’, i.e. the activity following pre-incubationwith CO₂ and Mg²⁺, was positively correlated with leaf relativewater content. However, ‘total Rubisco activity’in extracts from leaves with low water potential increased markedlywhen tightly bound inhibitors were removed, thus increasingthe number of catalytic sites available. This suggests thatin tobacco the decrease of Rubisco activity under drought stressis not primarily the result of changes in activation by CO₂and Mg²⁺ but due rather to the presence of tight-binding inhibitors.The amounts of inhibitor present in leaves of droughted tobaccobased on the decrease in Rubisco activity per mg soluble proteinwere usually much greater than the amounts of the known inhibitors(CA1P and ‘daytime inhibitor’) that can be recoveredin acid extracts. Alternative explanations for the differencebetween maximal and total activities are discussed.