基于生境质量和生态响应的莱州湾生态环境质量评价

莱州湾沿岸海域未来10a是山东省集约用海规划的重要区域。为了降低和减少用海工程对海洋生态系统可能产生的不利影响,确定集约用海适宜规模和科学选址,应用已构建的集约用海对海洋生态影响评价指标体系和方法,从生境质量和生态响应两方面对莱州湾2004年至2012年的生态环境质量状况进行了评价。结果表明:位于莱州湾西部和南部近岸海域的Ⅰ区和Ⅳ区生态环境质量达到"差"的等级,生境质量总体上处于轻度到中度污染水平,水质和生物质量污染较严重,生态受到中度到严重扰动,其中浮游植物和浮游动物密度偏离标准值较大,大型底栖生物密度和生物量总体偏离标准值不大,但其物种组成已发生了明显变化,鱼卵和仔鱼数量急剧下降。在这两个区域要严格控制集约用海工程规模和新的污染排放,密切关注其对海洋生态的影响。位于莱州湾中部和东部的Ⅱ区和Ⅲ区生态环境质量接近"一般"等级,生境质量总体上处于轻度污染水平,水质受到一定程度污染,生态处于中度扰动状况。总体上,2011年和2012年莱州湾生态环境质量综合指数E值明显高于其他年份,生态环境有明显好转趋势。     

巴西橡胶树胶乳中DUF1262结构域蛋白结构与基因表达分析

为了揭示天然橡胶生物合成酶互作蛋白结构及其在天然橡胶生物合成过程中的功能。本研究以橡胶树胶乳橡胶粒子总蛋白为研究对象,采用免疫共沉淀实验技术以天然橡胶合成关键酶顺式-异戊二烯基转移酶（CPT）抗体从胶乳中捕获了1个含DUF1262结构域的未知功能蛋白。生物信息学分析表明橡胶树基因组中包含50个编码含DUF1262结构域蛋白的基因序列;蛋白质相互作用网络分析表明DUF1262结构域蛋白可能参与调节信号转导或转录调控等过程;荧光定量PCR结果表明编码该蛋白基因的转录本在根、叶、花、枝和胶乳等组织中广泛分布,但在胶乳中表达较低,在树皮表达较高;水杨酸、脱落酸、过氧化氢及干旱处理可增强该基因在叶片中的转录水平。本研究证明DUF1262参与橡胶树逆境反应等生理过程,为揭示胶乳生物合成调控机制提供新线索。     

CURVED CHIMERIC PALEA 1 encoding an EMF1‐like protein maintains epigenetic repression of OsMADS58 in rice palea development

Floral organ specification is controlled by various MADS‐box genes in both dicots and monocots, whose expression is often subjected to both genetic and epigenetic regulation in Arabidopsis thaliana. However, little information is known about the role of epigenetic modification of MADS‐box genes during rice flower development. Here, we report the characterization of a rice gene, CURVED CHIMERIC PALEA 1 (CCP1) that functions in palea development. Mutation in CCP1 resulted in abnormal palea with ectopic stigmatic tissues and other pleiotropic phenotypes. We found that OsMADS58, a C‐class gene responsible for carpel morphogenesis, was ectopically expressed in the ccp1 palea, indicating that the ccp1 palea was misspecified and partially acquired carpel‐like identity. Constitutive expression of OsMADS58 in the wild‐type rice plants caused morphological abnormality of palea similar to that of ccp1, whereas OsMADS58 knockdown by RNAi in ccp1 could rescue the abnormal phenotype of mutant palea, suggesting that the repression of OsMADS58 expression by CCP1 is critical for palea development. Map‐based cloning revealed that CCP1 encodes a putative plant‐specific EMBRYONIC FLOWER1 (EMF1)‐like protein. Chromatin immunoprecipitation assay showed that the level of the H3K27me3 at the OsMADS58 locus was greatly reduced in ccp1 compared with that in the wild‐type. Taken together, our results show that CCP1 plays an important role in palea development through maintaining H3K27me3‐mediated epigenetic silence of the carpel identity‐specifying gene OsMADS58, shedding light on the epigenetic mechanism in floral organ development.     

Toll样受体4(TLR4)基因剔除小鼠构建及初步表型分析

目的 利用CRISPR/Cas9技术构建Toll样受体4(TLR4)基因敲除小鼠模型,并观察突变小鼠对革兰氏阴性细菌脂多糖(LPS)刺激响应的变化。方法 针对TLR4基因外显子2设计并合成1对sgRNA片段,与编码Cas9的mRNA混合后通过受精卵显微注射方法,建立TLR4基因敲除小鼠,通过繁育获得基因敲除纯合子小鼠(TLR4^-/-小鼠);通过LPS刺激,分析TLR4^-/-小鼠对炎症应激的反应情况,并在分子和病理水平上和野生型对照(WT)进行比较。结果 PCR及测序检测表明TLR4基因外显子2在小鼠基因中被成功敲除;给予LPS刺激后,IL1β、IL6、MyD88、iNOS及TNFa等炎症因子的表达在野生型小鼠的心、肝和肺组织中显著上调,而在TLR4^-/-小鼠中则几乎没有变化;血生化指标显示LPS刺激后WT小鼠血清中的尿素(Urea)和肌酐(Cre)水平显著升高,而TLR4^-/-小鼠刺激前后无显著变化,病理分析同样发现TLR4^-/-小鼠能够抵抗LPS对肾组织的损伤。结论 利用CRISPR/Cas9技术成功构建了TLR4基因剔除小鼠模型,TLR4的缺失能够降低IL1β、IL6、MyD88、iNOS及TNFa炎症因子对LPS刺激的响应,抑制LPS引起的炎症反应及对组织的损伤。     

Two Arabidopsis cytochrome P450 monooxygenases, CYP714A1 and CYP714A2, function redundantly in plant development through gibberellin deactivation

The rice gene ELONGATED UPPERMOST INTERNODE1 (EUI1) encodes a P450 monooxygenase that epoxidizes gibberellins (GAs) in a deactivation reaction. The Arabidopsis genome contains a tandemly duplicated gene pair ELA1 (CYP714A1) and ELA2 (CYP714A2) that encode EUI homologs. In this work, we dissected the functions of the two proteins. ELA1 and ELA2 exhibited overlapping yet distinct gene expression patterns. We showed that while single mutants of ELA1 or ELA2 exhibited no obvious morphological phenotype, simultaneous elimination of ELA1 and ELA2 expression in ELA1-RNAi/ela2 resulted in increased biomass and enlarged organs. By contrast, transgenic plants constitutively expressing either ELA1 or ELA2 were dwarfed, similar to those overexpressing the rice EUI gene. We also discovered that overexpression of ELA1 resulted in a severe dwarf phenotype, while overexpression of ELA2 gave rise to a breeding-favored semi-dwarf phenotype in rice. Consistent with the phenotypes, we found that the ELA1-RNAi/ela2 plants increased amounts of biologically active GAs that were decreased in the internodes of transgenic rice with ELA1 and ELA2 overexpression. In contrast, the precursor GA(12) slightly accumulated in the transgenic rice, and GA(19) highly accumulated in the ELA2 overexpression rice. Taken together, our study strongly suggests that the two Arabidopsis EUI homologs subtly regulate plant growth most likely through catalyzing deactivation of bioactive GAs similar to rice EUI. The two P450s may also function in early stages of the GA biosynthetic pathway. Our results also suggest that ELA2 could be an excellent tool for molecular breeding for high yield potential in cereal crops.     

BENT UPPERMOST INTERNODE1 encodes the class II formin FH5 crucial for actin organization and rice development

The actin cytoskeleton is an important regulator of cell expansion and morphogenesis in plants. However, the molecular mechanisms linking the actin cytoskeleton to these processes remain largely unknown. Here, we report the functional analysis of rice (Oryza sativa) FH5/BENT UPPERMOST INTERNODE1 (BUI1), which encodes a formin-type actin nucleation factor and affects cell expansion and plant morphogenesis in rice. The bui1 mutant displayed pleiotropic phenotypes, including bent uppermost internode, dwarfism, wavy panicle rachis, and enhanced gravitropic response. Cytological observation indicated that the growth defects of bui1 were caused mainly by inhibition of cell expansion. Map-based cloning revealed that BUI1 encodes the class II formin FH5. FH5 contains a phosphatase tensin-like domain at its amino terminus and two highly conserved formin-homology domains, FH1 and FH2. In vitro biochemical analyses indicated that FH5 is capable of nucleating actin assembly from free or profilin-bound monomeric actin. FH5 also interacts with the barbed end of actin filaments and prevents the addition and loss of actin subunits from the same end. Interestingly, the FH2 domain of FH5 could bundle actin filaments directly and stabilize actin filaments in vitro. Consistent with these in vitro biochemical activities of FH5/BUI1, the amount of filamentous actin decreased, and the longitudinal actin cables almost disappeared in bui1 cells. The FH2 or FH1FH2 domains of FH5 could also bind to and bundle microtubules in vitro. Thus, our study identified a rice formin protein that regulates de novo actin nucleation and spatial organization of the actin filaments, which are important for proper cell expansion and rice morphogenesis.     

基于ISSR标记和线粒体Cyt b基因分析高原鼠兔的遗传多样性及其遗传分化

高原鼠兔(Ochotona curzoniae)是青藏高原的特有动物和关键种。本实验应用ISSR分子标记和细胞色素b(Cytochrome b,Cytb)基因序列分析了雅鲁藏布江两岸高原鼠兔4个种群的遗传多样性和遗传分化。结果显示,两种标记所得到的4个种群的遗传多样性都较高,并以Cytb基因为指标的遗传多样性水平在种群间体现出较大差异。在ISSR标记中,分子方差分析(AMOVA)表明种群间的遗传变异为13.50%,种群分化较低且UPGMA聚类时江北岸与南岸的种群有交叉;而在Cytb基因中遗传变异主要发生在种群间,占79.24%,种群间存在着显著的遗传分化,且江北岸和南岸的两个种群分别聚为一类。研究结果表明,mtDNA基因在反映种群遗传多样性和遗传分化上较ISSR分子标记相对具有优势。     

The rice hydroperoxide lyase OsHPL3 functions in defense responses by modulating the oxylipin pathway

As important signal molecules, jasmonates (JAs) and green leaf volatiles (GLVs) play diverse roles in plant defense responses against insect pests and pathogens. However, how plants employ their specific defense responses by modulating the levels of JA and GLVs remains unclear. Here, we describe identification of a role for the rice HPL3 gene, which encodes a hydroperoxide lyase (HPL), OsHPL3/CYP74B2, in mediating plant‐specific defense responses. The loss‐of‐function mutant hpl3‐1 produced disease‐resembling lesions spreading through the whole leaves. A biochemical assay revealed that OsHPL3 possesses intrinsic HPL activity, hydrolyzing hydroperoxylinolenic acid to produce GLVs. The hpl3‐1 plants exhibited enhanced induction of JA, trypsin proteinase inhibitors and other volatiles, but decreased levels of GLVs including (Z)‐3‐hexen‐1‐ol. OsHPL3 positively modulates resistance to the rice brown planthopper [BPH, Nilaparvata lugens (Stål)] but negatively modulates resistance to the rice striped stem borer [SSB, Chilo suppressalis (Walker)]. Moreover, hpl3‐1 plants were more attractive to a BPH egg parasitoid, Anagrus nilaparvatae, than the wild‐type, most likely as a result of increased release of BPH‐induced volatiles. Interestingly, hpl3‐1 plants also showed increased resistance to bacterial blight (Xanthomonas oryzae pv. oryzae). Collectively, these results indicate that OsHPL3, by affecting the levels of JA, GLVs and other volatiles, modulates rice‐specific defense responses against different invaders.     

Construction of Stress Responsive Synthetic Promoters and Analysis of Their Activity in Transgenic Arabidopsis thaliana

To obtain strong inducible promoters to drive abiotic stress-inducible transgene expression with minimal negative effects, we constructed three artificial synthetic promoters (EKCM, EKCRM, and ECCRM) comprising multiple cis-acting stress-response elements. Each promoter was fused independently to the β-glucuronidase (GUS) reporter gene, and GUS expression was analyzed in stable expression systems in Arabidopsis thaliana. T2 transgenic progenies showed integration of the promoter-GUS construct in their genome. RT-PCR assays and histochemical staining analysis showed that GUS expression driven by each promoter increased under desiccation, cold, and high salt conditions. The activity of synthetic promoters, assessed by fluorometric quantitative analysis of GUS enzyme activity, was significantly higher than that of the rd29A promoter under various stress treatments. The most powerful promoter, EKCM, allowed about 1.29-fold in GUS activity relative to the rd29A promoter, on average, under dehydration conditions. All three synthetic promoters could drive stress-inducible GUS expression in different organs of transgenic Arabidopsis. These synthetic promoters represent valuable tools for improving the stress tolerance of crops.     

系统性红斑狼疮患者血清中IL-17 的检测及临床意义

目的:通过检测系统性红斑狼疮(SLE)患者外周血中IL-17的水平,探讨其临床意义。方法:用酶联免疫吸附法(enzymelinked immunosorbent assay,ELISA)检测61例SLE患者及30例健康人血清IL-17水平,并收集整理SLE患者的临床资料及实验室数据,分析其与临床的关系。结果:活动期SLE患者血清IL-17水平明显高于正常对照组(P<0.01),与SLE非活动组相比,差异亦有统计学意义(P<0.01)。但SLE非活动组与正常对照组间无统计学意义。结论:IL-17水平在活动期SLE患者血清中表达明显增高,且与SLEDAI评分呈正相关,提示IL-17可能参与了SLE疾病的病理过程,可能与疾病活动的关系密切。