caspase-3、bax、bcl-2及c-kit在中华大蟾蜍脊髓损伤后的表达变化

目的：探究高等动物脊髓损伤修复困难的原因。方法：利用免疫组化方法检验中华大蟾蜍脊髓损伤后凋亡相关基因及c—kit的表达。结果：①损伤后促凋亡因子（easpase-3及bax）随时间变化的规律为先升高再降低;②抑制凋亡因子bcl-2表达规律为先降低后升高;③bcl-2／bax比值的变化趋势与easpase-3的相反,bel-2/bax比值的升高可能抑制caspase-3表达;④c—kit表达呈现先增高再降低的规律,c-kit表达出现峰值时bcl-2表达增加,bax减少。结论：这些差异可能能够促进蟾蜍神经损伤后修复,也是低等脊椎动物较高等脊椎动物再生能力强的原因之一,为哺乳类脊髓损伤后修复方法研究提供一定的理论依据。     

PIF3 Is Involved in the Primary Root Growth Inhibition of Arabidopsis Induced by Nitric Oxide in the Light

PHYTOCHROME INTERACTING FACTOR3 （PIF3） is an important component in the phytochrome signaling pathway and mediates plant responses to various environmental conditions. We found that PIF3 is involved in the inhibition of root growth of Arabidopsis thaliana seedlings induced by nitric oxide （NO） in light. Overexpression of PIF3 partially alleviated the inhibitory effect of NO on root growth, whereas the pif3-1 mutant displayed enhanced sensitivity to NO in terms of root growth. During phytochrome signaling, the photoreceptor PHYB mediates the degradation of PIF3. We found that the phyB-9 mutant had a similar phenotype to that of PIF3ox in terms of responsiveness to NO. Furthermore, NO treatment promoted the accumulation of PHYB, and thus reduced PIF3 content. Our results further show that the activity of PIF3 is regulated by the DELLA protein RGL3[RGA （repressor of gal-3） LIKE 3]. Therefore, we speculate that PIF3 lies downstream of PHYB and RGL3, and plays an important role in the inhibitory effect of NO on root growth of Arabidopsis seedlings in light.     

Salt Stress in Arabidopsis： Lipid Transfer Protein AZI1 and Its Control by Mitogen-Activated Protein Kinase MPK3

A plant＇s capability to cope with environmental challenges largely relies on signal transmission through mitogen-activated protein kinase （MAPK） cascades. In Arabidopsis thaliana, MPK3 is particularly strongly associated with numerous abiotic and biotic stress responses. Identification of MPK3 substrates is a milestone towards improving stress resistance in plants. Here, we characterize AZI1, a lipid transfer protein （LTP）-related hybrid proline-rich protein （HyPRP）, as a novel target of MPK3. AZI1 is phosphorylated by MPK3 in vitro. As documented by co-immunoprecipitation and bimolecular fluorescence complementation experiments, AZI1 interacts with MPK3 to form protein complexes in planta. Furthermore, null mutants of azil are hypersensitive to salt stress, while AZIl-overexpressing lines are markedly more tolerant. AZI1 overexpression in the mpk3 genetic background partially alleviates the salt-hypersensitive phenotype of this mutant, but functional MPK3 appears to be required for the full extent of AZIl-conferred robustness. Notably, this robustness does not come at the expense of normal development. Immunoblot and RT-PCR data point to a role of MPK3 as positive regulator of AZI1 abundance.     

Open and Closed： The Roles of Linker Histones in Plants and Animals

Histones package DNA in all eukaryotes and play key roles in regulating gene expression. Approximately 150 base pairs of DNA wraps around an octamer of core histones to form the nucleosome, the basic unit of chromatin. Linker histones compact chromatin further by binding to and neutralizing the charge of the DNA between nucleosomes. It is well established that chromatin packing is regulated by a complex pattern of posttranslational modifications （PTMs） to core histones, but linker histone function is less well understood. In this review, we describe the current understand- ing of the many roles that linker histones play in cellular processes, including gene regulation, cell division, and devel- opment, while putting the linker histone in the context of other nuclear proteins. Although intriguing roles for plant linker histones are beginning to emerge, much of our current understanding comes from work in animal systems. Many unanswered questions remain and additional work is required to fully elucidate the complex processes mediated by linker histones in plants.     

姜黄素对大鼠脑缺氧缺血损伤时脑组织MDA变化、caspase-3表达及细胞凋亡的影响

目的:探讨姜黄素对大鼠脑缺氧缺血损伤时脑组织MDA变化、caspase-3表达及细胞凋亡的影响。方法:健康SD雄性大鼠48只,随机分为假手术对照组(SH组)、脑缺氧缺血组(HI组)、姜黄素组(CU组)、溶剂对照组(SC组);生化方法检测脑组织丙二醛(MDA)含量;免疫组织化学测定大脑皮质caspase-3的表达;电镜观察大脑皮质形态学结构变化。结果:姜黄素可使脑组织MDA含量明显减低,并且抑制caspase-3蛋白的表达;神经元细胞凋亡减轻。结论:细胞凋亡参与了大脑缺氧缺血损伤的发生,姜黄素可能通过减低MDA含量、下调caspase-3的表达抑制细胞凋亡,从而减轻脑缺氧缺血性损伤。     

Plant E3 Ligases： Flexible Enzymes in a Sessile World

Since its discovery in the late 1970s, the ubiquitin proteasome pathway appears to be omnipresent in many research fields. Although originally discovered in animals, the pathway has a very central role in plants, which may be correlated to their sessile lifestyle. E3 ligases function as flexible and highly diverse key regulators within the path- way by targeting substrate proteins for ubiquitylation, and often proteolytic degradation via the 26S proteasome. This review provides a concise overview on the most common classes of E3 ligases so far described in plants, and emphasizes recent findings regarding these interesting and flexible enzymes and their diverse functions in plant biology.     

缺血后适应对局灶性脑缺血/再灌注大鼠caspase-3表达的影响

目的：探讨缺血后适应对大鼠局灶性脑缺血/再灌注损伤后caspase-3表达的影响。方法：大脑中动脉线拴法复制大鼠局灶性脑缺血/再灌注损伤动物模型。将30只雄性SD大鼠随机分为3组（n=10）：假手术组（sham组）、缺血/再灌注（I/R）组和缺血后适应（IP）组。利用原位缺口末端标记法观察神经细胞凋亡的变化。应用Western blot检测大鼠局灶性脑缺血/再灌注损伤后caspase-3蛋白表达水平的变化。结果：大鼠脑缺血/再灌注后凋亡细胞数量和caspase-3蛋白表达水平均显著升高,而缺血后适应组凋亡细胞数量和caspase-3蛋白表达水平均显著低于缺血/再灌注组（P〈0.01）。结论：缺血后适应可抑制大鼠脑缺血/再灌注后细胞凋亡的发生,此作用可能与下调caspase-3蛋白表达有关。     

Mechanism of immunomodulatory drugs＇ action in the treatment of multiple myeloma

Although immunomodulatory drugs （IMiDs）, such as thal- idomide, lenalidomide, and pomalidomide, are widely used in the treatment of multiple myeloma （MM）, the molecular mechanism of IMiDs＇ action is largely unknown. In this review, we will summarize recent advances in the application of IMiDs in MM cancer treatment as well as their effects on immunomodulatory activities, anti-angiogenic activities, intervention of cell surface adhesion molecules between myeloma cells and bone marrow stromal cells, anti-inflam- matory activities, anti-proliferation, pro-apoptotic effects, cell cycle arrest, and inhibition of cell migration and metastasis. In addition, the potential IMiDs＇ target protein, IMiDs＇ target protein＇s functional role, and the potential molecular mechanisms of IMiDs resistance will be discussed. We wish, by presentation of our naive discussion, that this review article will facilitate further investigation in these fields.     

~ffects of endothelial microvesicles induced by A23187 on H9c2 cardiomyocytes

Objective To investigate the effects of endothelial microvesicles （EMVs） induced by calcium ionophore A23187 on H9c2 cardiomyocytes. Methods Human umbilical vein endothelial cells （HUVECs） were treated with 10 μmol/L A23187 for 30 min. EMVs from HUVECs were isolated by ultracentrifugation from the conditioned culture medium. EMVs were characterized using 1 and 2 Ilm latex beads and anti- PE-CD144 antibody by flow cytometry. For functional research, EMVs at different concentrations were co- cultured with H9c2 cardiomyocytes for 6 h. Cell viability of H9c2 cells and the activity of LDH leaked from H9c2 cells were tested by colorimetry. Moreover, apoptosis of H9c2 cells was observed through Hoechst 33258 staining and tested by FITC-Annexin V/Pl double staining. Results EMVs were induced by A23187 on HUVECs, and isolated by ultracentrifugation. We identified the membrane vesicles （〈 1 μm） induced by A23187 were CD144 positive. In addition, the EMVs could significantly reduce the viability of H9c2 cells, and increase LDH leakage from H9c2 cells in a dose dependent manner （P〈0.05）. Condensed nuclei could be observed with the increasing concentrations of EMVs through Hoechst 33258 staining. Furthermore, increased apoptosis rates of H9c2 cells could be assessed through FITC-Annexin V/PI double staining by flow cytometry. Conclusion Microvesicles could be released from HUVECs after induced by A23187 through calcium influx, and these EMVs exerted a pro-apoptotic effect on H9c2 cells by induction of apoptosis.     

Cell Wall,Cytoskeleton, and Cell Expansion in Higher Plants

To accommodate two seemingly contradictory biological roles in plant physiology, providing both the rigid structural support of plant cells and the adjustable elasticity needed for cell expansion, the composition of the plant cell wall has evolved to become an intricate network of cellulosic, hemicellulosic, and pectic polysaccharides and protein. Due to its complexity, many aspects of the cell wall influence plant cell expansion, and many new and insightful observations and technologies are forthcoming. The biosynthesis of cell wall polymers and the roles of the variety of proteins involved in polysaccharide synthesis continue to be characterized. The interactions within the cell wall polymer network and the modification of these interactions provide insight into how the plant cell wall provides its dual function. The complex cell wall architecture is controlled and organized in part by the dynamic intracellular cytoskeleton and by diverse trafficking pathways of the cell wall polymers and cell wall-related machinery. Meanwhile, the cell wall is continually influenced by hormonal and integrity sensing stimuli that are perceived by the cell. These many processes cooperate to construct, maintain, and manipulate the intricate plant cell wall--an essential structure for the sustaining of the plant stature, growth, and life.     

Dissecting the Heat Stress Response in Chlamydomonas by Pharmaceutical and RNAi Approaches Reveals Conserved and Novel Aspects

To study how conserved fundamental concepts of the heat stress response （HSR） are in photosynthetic eukaryotes, we applied pharmaceutical and antisense/amiRNA approaches to the unicellular green alga Chlamydomonas reinhardtii. The Chlamydomonas HSR appears to be triggered by the accumulation of unfolded proteins, as it was induced at ambient temperatures by feeding cells with the arginine analog canavanine. The protein kinase inhibitor staurosporine strongly retarded the HSR, demonstrating the importance of phosphorylation during activation of the HSR also in Chlamydomonas. While the removal of extracellular calcium by the application of EGTA and BAPTA inhibited the HSR in moss and higher plants, only the addition of BAPTA, but not of EGTA, retarded the HSR and impaired thermotoler- ance in Chlamydomonas. The addition of cycloheximide, an inhibitor of cytosolic protein synthesis, abolished the attenu- ation of the HSR, indicating that protein synthesis is necessary to restore proteostasis. HSP90 inhibitors induced a stress response when added at ambient conditions and retarded attenuation of the HSR at elevated temperatures. In addition, we detected a direct physical interaction between cytosolic HSP90A/HSP70A and heat shock factor 1, but surprisingly this interaction persisted after the onset of stress. Finally, the expression of antisense constructs targeting chloroplast HSP70B resulted in a delay of the cell＇s entire HSR, thus suggesting the existence of a retrograde stress signaling cascade that is desensitized in HSP7OB-antisense strains.     

Intracellular and Extracellular Phosphatidylinositol 3-Phosphate Produced by Phytophthora Species Is Important for Infection

RxLR effectors produced by Phytophthora pathogens have been proposed to bind to phosphatidylinositol 3-phosphate （Ptdlns（3）P） to mediate their translocation into host cells and/or to increase their stability in planta. Since the levels of Ptdlns（3）P in plants are low, we examined whether Phytophthora species may produce Ptdlns（3）P to pro- mote infection. We observed that Ptdlns（3）P-specific GFP biosensors could bind to P. parasitica and P. sojae hyphae dur- ing infection of Nicotiana benthamiana leaves transiently secreting the biosensors, suggesting that the hyphae exposed Ptdlns（3）P on their plasma membrane and/or secreted Ptdlns（3）R Silencing of the phosphatidylinositol 3-kinases （PI3K） genes, treatment with LY294002, or expression of Ptdlns（3）pobinding proteins by P. sojae reduced the virulence of the pathogen on soybean, indicating that pathogen-synthesized Ptdlns（3）P was required for full virulence. Secretion of Ptdlns（3）P-binding proteins or of a PI3P-5-kinase by N. benthamiana leaves significantly increased the level of resist- ance to infection by P. parasitica and P. capsici. Together, our results support the hypothesis that Phytophthora species produce external Ptdlns（3）P to aid in infection, such as to promote entry of RxLR effectors into host cells. Our results derived from P. sojae RxLR effector Avrlb confirm that both the N-terminus and the C-terminus of this effector can bind Ptdlns（3）P.     

14-3-3 Binding to Cyclin Y contributes to cyclin Y/CDK14 association

Cyclin Y is a highly conserved cyclin among eumetazoans, yet its function and regulation are poorly understood. To search for Cyclin Y-interacting proteins, we screened a yeast two-hybrid library using human Cyclin Y （CCNY） as a bait and identified the following interactors： CDK14 and four members of the 14-3-3 family （ε,β,η,τ）. The interaction between CCNY and 14-3-3 proteins was confirmed both in vitro and in vivo. The results showed that Ser-100 and Ser-326 residues in CCNY were crucial for 14-3-3 binding. Interestingly, binding of CCNY to 14-3-3 significantly enhanced the association between CCNY and CDK14. Our findings may add a new layer of regulation of CCNY binding to its kinase partner.     

PRCl Marks the Difference in Plant PcG Repression

ABSTRACT From mammals to plants, the Polycomb Group （PcG） machinery plays a crucial role in maintaining the repres- sion of genes that are not required in a specific differentiation status. However, the mechanism by which PeG machinery mediates gene repression is still largely unknown in plants. Compared to animals, few PcG proteins have been identi- fied in plants, not only because just some of these proteins are clearly conserved to their animal counterparts, but also because some PcG functions are carried out by plant-specific proteins, most of them as yet uncharacterized. For a long time, the apparent lack of Polycomb Repressive Complex （PRC）I components in plants was interpreted according to the idea that plants, as sessile organisms, do not need a long-term repression, as they must be able to respond rapidly to environmental signals; however, some PRC1 components have been recently identified, indicating that this may not be the case. Furthermore, new data regarding the recruitment of PcG complexes and maintenance of PcG repression in plants have revealed important differences to what has been reported so far. This review highlights recent progress in plant PcG function, focusing on the role of the putative PRC1 components.     

Redox Regulation of Arabidopsis Mitochondrial Citrate Synthase

Citrate synthase has a key role in the tricarboxylic （TCA） cycle of mitochondria of all organisms, as it cata- lyzes the first committed step which is the fusion of a carbon-carbon bond between oxaloacetate and acetyl CoA. The regulation of TCA cycle function is especially important in plants, since mitochondrial activities have to be coordinated with photosynthesis. The posttranslational regulation of TCA cycle activity in plants is thus far almost entirely unexplored. Although several TCA cycle enzymes have been identified as thioredoxin targets in vitro, the existence of any thioredoxin-dependent regulation as known for the Calvin cycle, yet remains to be demonstrated. Here we have investigated the redox regulation of the Arabidopsis citrate synthase enzyme by site-directed mutagenesis of its six cysteine residues. Our results indicate that oxidation inhibits the enzyme activity by the formation of mixed disulfides, as the partially oxidized citrate synthase enzyme forms large redox-dependent aggregates. Furthermore, we were able to demonstrate that thioredoxin can cleave diverse intraas well as intermolecular disulfide bridges, which strongly enhances the activity of the enzyme. Activity measurements with the cysteine variants of the enzyme revealed important cysteine residues affecting total enzyme activity as well as the redox sensitivity of the enzyme.     

Organelle pH in the Arabidopsis Endomembrane System

The pH of intracellular compartments is essential for the viability of cells. Despite its relevance, little is known about the pH of these compartments. To measure pH in vivo, we have first generated two pH sensors by combining the improved-solubility feature of solubility-modified green fluorescent protein （GFP） （smGFP） with the pH-sensing capabil- ity of the pHluorins and codon optimized for expression in Arabidopsis. PEpHluorin （plant-solubility-modified ecliptic pHluorin） gradually loses fluorescence as pH is lowered with fluorescence vanishing at pH 6.2 and PRpHluorin （plant- solubility-modified ratiomatric pHluorin）, a dual-excitation sensor, allowing for precise measurements. Compartment- specific sensors were generated by further fusing specific sorting signals to PEpHluorin and PRpHluorin. Our results show that the pH of cytosol and nucleus is similar （pH 7.3 and 7.2）, while peroxisomes, mitochondrial matrix, and plastidial stroma have alkaline pH. Compartments of the secretory pathway reveal a gradual acidification, spanning from pH 7.1 in the endoplasmic reticulum （ER） to pH 5.2 in the vacuole. Surprisingly, pH in the trans-Golgi network （TGN） and mul- tivesicular body （MVB） is, with pH 6.3 and 6.2, quite similar. The inhibition of vacuolar-type H＋-ATPase （V-ATPase） with concanamycin A （ConcA） caused drastic increase in pH in TGN and vacuole. Overall, the PEpHluorin and PRpHluorin are excellent pH sensors for visualization and quantification of pH in vivo, respectively.     

The evolutionary ecology of symbiont-conferred resistance to parasitoids in aphids

Abstract Aphids may harbor a wide variety of facultative bacterial endosymbionts. These symbionts are transmitted maternally with high fidelity and they show horizontal trans-mission as well, albeit at rates too low to enable infectious spread. Such symbionts need to provide a net fitness benefit to their hosts to persist and spread. Several symbionts haveachieved this by evolving the ability to protect their hosts against parasitoids. Reviewing empirical work and some models, I explore the evolutionary ecology of symbiont-conferredresistance to parasitoids in order to understand how defensive symbiont frequencies are maintained at the intermediate levels observed in aphid populations. I further show thatdefensive symbionts alter the reciprocal selection between aphids and parasitoids by augmenting the heritable variation for resistance, by increasing the genetic specificity of thehost-parasitoid interaction, and by inducing environment-dependent trade-offs. These effects are conducive to very dynamic, symbiont-mediated coevolution that is driven by frequency-dependent selection. Finally I argue that defensive symbionts represent a problem for biological control of pest aphids, and I propose to mitigate this problem byexploiting the parasitoids＇ demonstrated ability to rapidly evolve counteradaptations to symbiont-conferred resistance.     

How does the host-specialized aphid deal with food deficiency？

Aphis gossypii Glover shows obvious host specialization, with cucurbit- and cotton-specialized biotypes or host races in many regions. Because its annual natal hostcrops senesce earlier the cucurbit-specialized biotype may suffer food deficiency. The method this biotype uses to overcome this challenge is still poorly understood. In orderto understand the potential of the cucurbit-specialized biotype aphids in host shift and usage, the performance of this biotype on cotton （Gossypium hirsutum）, a common butpoor quality host plant, was explored in this study. The cucurbit-specialized aphids could establish populations on cotton only when these plants had at least nine leaves, and subsequent populations developed rather slowly. The presence of whitefly populations on cotton improved the success rate of cucurbit-specialized aphids. The cucurbit-specialized aphidswere mainly distributed on the older leaves of cotton, with only a few settling on the upper leaves. The cucurbit-specialized aphids reared on cotton for 40, 54 and 61 days stillmaintained strong preference for their natal host plant, cucumber （Cucumis sativus）, rather than cotton, and their net reproductive rates and intrinsic rates of natural increase weredramatically lower when they were transferred onto new six-leaf cotton plants or detached leaves. Therefore, we concluded that the cucurbit-specialized aphids have the potentialto utilize mature or whitefly-stressed cotton plants, but that this feeding experience on cotton did not alter their specialization for cucurbits. Some cotton plants could act as atemporary host for the cucurbit-specialized aphids to overcome food deficiency arising from senescing cucurbits.