PEX5参与调控拟南芥根系抗逆性的初步研究

以拟南芥过氧化物酶体生成蛋白突变体pex5和野生型为试验材料,初步研究PEX5对拟南芥根系抗逆性的影响。结果表明,在MS基础培养基中,突变体pex5的根长相较于野生型根长显著变短;同时,拟南芥pex5突变体株高也低于野生型,说明PEX5基因功能缺失影响了拟南芥根系和茎的生长。pex5突变体在添加氯化钠或甘露醇的MS培养基上培养时,萌发和根伸长受到的抑制显著高于野生型,突变体幼苗的SOD和POD活性也显著低于野生型,说明PEX5参与调控拟南芥根系的抗逆性。本研究为解析PEX5在植物抗逆中的调控机制提供了重要参考。     

稻瘟病菌过氧化物酶体产生与降解的关键基因

过氧化物酶体(peroxisomes)是真核细胞中一类单层膜包被的细胞器,参与多种生化代谢.过氧化物酶体起源于内质网,过氧化物酶体形成相关的蛋白称为Peroxin,其编码基因通常写作PEX.细胞中过氧化物酶体的选择性消解称为过氧化物酶体自噬(pexophagy).参与细胞自噬(autophagy)的基因(ATG)大多参与过氧化物酶体自噬.近年来,丝状真菌中过氧化物酶体形成与降解机制的研究进展迅速,相关基因不断被鉴定.本文对相关研究进行了简要评述,并以稻瘟病菌为例,对丝状真菌基因组中可能的PEX和ATG基因进行了检索.发现稻瘟病菌中存在除PEX15,PEX17,PEX18,PEX21,PEX22,ATG19,ATG25,ATG30和ATG31之外的大多数PEX和ATG基因;同时,还存在多个丝状真菌特有的基因.说明过氧化物酶体的产生与消解在酵母、丝状真菌与哺乳动物之间相对保守,同时又各具特性.     

苜蓿假盘菌子囊孢子萌发特性及子实体超微结构观察

【背景】苜蓿假盘菌(Pseudopeziza medicaginis)侵染苜蓿叶片引起的褐斑病是苜蓿的重要病害之一。研究表明,该菌以子囊盘越冬,翌年释放子囊孢子完成初侵染,因此观察该菌子实体的超微结构特征,有利于进一步揭示子囊菌的侵染机制。【目的】明确P. medicaginis子囊孢子萌发、Ca2+信号途径是否参与调控附着胞形成;观察子囊孢子和子实体的超微结构特征。【方法】采用光学显微镜技术,研究不同碳、氮源和Ca2+/CaM信号途径对P. medicaginis子囊孢子萌发、附着胞形成的影响,并采用电镜技术观察子实体和子囊孢子的超微结构。【结果】碳、氮源有利于P. medicaginis子囊孢子的萌发,葡萄糖、麦芽糖、蔗糖、氨基乙酸、酵母粉、尿素、蛋白胨、硝酸铵、硝酸钠等碳、氮源诱导形成附着胞,以尿素诱导效果最明显;Ca2+不仅促进子囊孢子的萌发,显著诱导附着胞的形成,而且高于碳、氮源;10 mmol/L的Ca2+诱导效果最明显,随其浓度的增加,子囊孢子的萌发率和附着胞的形成率下降;低浓度Ca2+螯合剂EGTA诱导形成附着胞,≥1 mmol/L抑制子囊孢子的萌发,外源Ca2+或Ca2+载体A23187能够部分解除EGTA的抑制作用;磷酸酯酶抑制剂Neomycin、Ca2+通道阻断剂Nicardipine、CaM拮抗剂Chloropromazine完全抑制子囊孢子的萌发。成熟的P. medicaginis子实体突破寄主表皮,中间开裂露出子囊,子囊孔口无囊盖,呈横向裂口状。子囊孢子表面具有环形纹饰,细胞核色深,线粒体等细胞器数量多,脂类物质数量多且发达,胞内充满电子致密度高的物质。【结论】碳、氮源和Ca2+可诱导P. medicaginis子囊孢子形成附着胞,钙信号途径参与P. medicaginis子囊孢子的萌发和附着胞形成过程的调控。子实体突破寄主表皮外露呈开裂式,子囊顶端无囊盖,线粒体发达。该研究结果为P. medicaginis的深入研究提供重要细胞学事件。     

酵母过氧化物酶体的形成机制

过氧化物酶体是细胞中一种重要的细胞器。过氧化物酶体在细胞功能的发挥和人体健康方面有着重要作用。目前,以酵母过氧化物酶体为模型,研究过氧化物酶体的形成机制是研究热点。从过氧化物酶体起源、生成方式介绍最新研究进展,总结在酵母细胞中参与过氧化物酶体形成的必需基因(pex),及其编码Peroxin蛋白在过氧化物酶体形成过程中的作用,阐述酵母过氧化物酶体的形成途径和形成机制,展望了研究前景。     

稻瘟病菌微波诱发突变体的分析*

通过诱变获得突变体是研究稻瘟病菌变异机制的基础。本文用微波炉对稻瘟病菌分生孢子进行低强度短时间处理获得了一批形态发育和致病性突变体,并对它们进行了分析。突变体1-40-271菌落呈白色,产孢与萌发均正常,但萌发后即便在人工疏水表面上也不能形成附着胞,且丧失了致病性;突变体2-20-6菌落呈黄色,孢子萌发率为1%,萌发的孢子其附着胞形成率仅为0.01%,致病性减弱;突变体2-30-3菌落呈黄色,形成的附着胞大部分不正常,但致病性正常。Rep-PCR指纹分析发现,突变体2-20-6和2-30-3比其相应野生型少1条带,而突变体1-40-271与其野生型比较没有变化,说明微波可能造成稻瘟病菌基因组DNA缺失或点突变而发生变异。继代分析表明微波处理获得的稻瘟病菌形态和致病性突变体是稳定的。     

灰葡萄孢菌过氧化物酶体及细胞核的荧光蛋白标记

【目的】对灰葡萄孢菌(Botrytis cinerea)的细胞核和过氧化物酶体进行荧光蛋白标记,为研究其生长发育和侵染过程中细胞结构和细胞器动态提供基础。【方法】以绿色荧光蛋白(GFP)和红色荧光蛋白(DsRED、mCherry)为报告基因,利用根癌农杆菌介导转化(Agrobacterium tumefaciens mediated transformation,AtMT)将3种荧光蛋白标记载体分别导入灰葡萄孢菌标准菌株B05.10;通过PCR检测及荧光观察筛选和验证转化子,并进行单孢纯化;利用共聚焦显微镜记录细胞器荧光定位情况。【结果】获得了过氧化物酶体或细胞核稳定表达红、绿色荧光的重组单孢菌株,PCR验证表明标记基因成功整合入转化子基因组。在标记细胞核的菌株中,菌丝和孢子中可见多个明亮、圆形的荧光点,与DAPI染色共定位。标记过氧化物酶体的菌株中,菌丝和孢子中可见小点状绿色或红色荧光,在脂类物质诱导下荧光点数量明显增加,符合过氧化物酶体分布及动态特征。细胞壁染色结果显示,细胞壁染色产生的蓝色荧光与红、绿荧光蛋白的荧光互不干扰,标记效果良好。【结论】获得了理想的过氧化物酶体或细胞核荧光标记的灰葡萄孢菌菌株,为研究其细胞器动态以及生长发育与致病分子机制提供了参考和材料。     

稻瘟病菌微波诱发突变体的分析

通过诱变获得突变体是研究稻瘟病菌变异机制的基础。本文用微波炉对稻瘟病菌分生孢子进行低强度短时间处理获得了一批形态发育和致病性突变体,并对它们进行了分析。突变体1-40-271菌落呈白色,产孢与萌发均正常,但萌发后即便在人工疏水表面上也不能形成附着胞,且丧失了致病性;突变体2-20-6菌落呈黄色,孢子萌发率为1％,萌发的孢子其附着胞形成率仅为0．01％,致病性减弱;突变体2-30-3菌落呈黄色,形成的附着胞大部分不正常,但致病性正常。Rep-PCR指纹分析发现,突变体2-20-6和2-30-3比其相应野生型少1条带,而突变体1-40-271与其野生型比较没有变化,说明微波可能造成稻瘟病菌基因组DNA缺失或点突变而发生变异。继代分析表明微波处理获得的稻瘟病菌形态和致病性突变体是稳定的。     

2,4-D浓度对胡萝卜体细胞胚胎发生及其同工酶的影响

在含有不同浓度2,4-D的MS培养基上生长的胡萝卜组织,培养26天后,不含2,4-D培养基上的培养物,其酯酶同工酶谱比在含有2,4-D培养基上的培养物少一条酶带8(图3)。过氧化物酶同工酶谱中,发现前者出现的酶带1(图4)在后者的酶谱中看不到。 胡萝卜的培养组织和完整的小植物皆具有分泌过氧化物酶和酯酶的能力。总过氧化物酶活力的变化与愈伤组织、胚状体和小植物的形成有一定关系。     

菌株YN145对稻瘟病菌黑色素合成的影响及全基因组分析

【背景】枯草芽孢杆菌YN145是一株从湖南省桃江县的健康稻株中分离的细菌,前期研究中该菌对稻瘟病菌拮抗效果显著,在生物防治方面有很大的应用潜力。【目的】深入研究该菌株的生防机制并挖掘次级代谢产物基因资源。【方法】在4株稻瘟病菌生防菌中,选择其胞外抗菌物质抑制稻瘟病菌黑色素合成效果最佳的菌株YN145,采用紫外-可见分光光度计在波长400 nm处测定胞外和菌丝体内黑色素液的吸光度值,采用菌丝生长抑制平板法和分生孢子萌发抑制法测定抑菌活性。采用PacBio第三代测序和IlluminaHiSeq第二代测序相结合的技术对菌株YN145进行全基因组测序,并对测序数据进行组装,注释预测基因的功能,分析次级代谢产物合成基因簇。【结果】菌株YN145的胞外抗菌物质能较好地抑制稻瘟病菌黑色素合成、分生孢子萌发和菌丝生长。菌株YN145全基因组大小为4 167 871 bp,GC含量为43.86%,编码序列(coding sequence, CDS)数量为4 294个;共找到85个tRNA、30个rRNA和92个sRNA。同时预测到5个已知的次级代谢产物合成基因簇,分别编码合成bacillaene、bac...     

群体感应淬灭酶克隆表达及其对铜绿假单胞菌的影响

【背景】由于抗生素的大量使用,导致细菌耐药性越来越强,寻找新的抗细菌感染药物成为研究热点。【目的】克隆表达群体感应淬灭酶,探究其对铜绿假单胞菌毒力及致病性的影响。【方法】利用PCR技术从产群体感应淬灭酶的芽孢杆菌QSI-1基因组DNA中克隆出aiiA基因,将其克隆到表达载体pET30a并导入大肠杆菌E.coliBL21(DE3)中进行诱导表达,通过镍柱亲和层析获得纯化的N-酰基高丝氨酸内酯酶。用不同浓度的淬灭酶作用于铜绿假单胞菌,检测其对铜绿假单胞菌毒力因子产生以及生物膜形成能力的影响;以秀丽隐杆线虫为模型,考察其对线虫感染铜绿假单胞菌存活率的影响。【结果】克隆表达出群体感应淬灭酶,该酶能显著抑制铜绿假单胞菌毒力因子产生和生物膜的形成,并能降低铜绿假单胞菌对感染线虫的致死率。【结论】群体感应淬灭酶可作为一种能高效抑制细菌致病性的物质,为临床治疗细菌性感染提供新的策略。     

Characterization of SSEA-1 glycolipids from the brain of a patient with fucosidosis

Neutral glycolipids from the brain of a patient with Fucosidosis were analyzed and two complex glycolipids containing five and eight sugars were isolated from the cortical grey matter. These two glycolipids reacted with antibodies recognizing the SSEA-1 [Le^x(X)] carbohydrate determinant. SSEA-1 glycolipids are normally expressed in human embryonic brain but are found in only small amounts in postnatal human brain. The accumulation of the two SSEA-1 glycolipids in Fucosidosis brain thus represents a defect which affects the normal developmentally regulated decrease in postnatal, expression of these glycolipids, and may be a contributing factor in the abnormal brain development associated with the disease. Chemical characterization of the two isolated glycolipids by gas chromatographic and mass spectrometric analyses has identified the two glycolipids as lacto-N-fucopentaosylceramide (III) and difucosyl-neolactonorhexaosylceramide.Abbreviations DCl direct chemical ionization - FAB tastatiom bombardment - GC gas chromatography - GSLs glycosphingolipids - MS mass spectrometry - SSEA-1 stage specific embryonic antigen-1 - TLC thin layer chromatographys     

Structural Peculiarities of Na+,K+-ATPase Isozymes from the Calf Brain

Functionally active preparations of Na⁺,K⁺-ATPase isozymes from calf brain that contain catalytic subunits of three types (1, 2, and 3) were obtained using two approaches: a selective removal of contaminating proteins by the Jorgensen method and a selective solubilization of the enzyme with subsequent reconstitution of their membrane structure by the Esmann method. The ouabain inhibition constants were determined for the isozymes. The real isozyme composition of the Na⁺ pump from the grey matter containing glial cells and the brain stem containing neurons was determined. The plasma membranes of glial cells were shown to contain mainly Na⁺,K⁺-ATPase of the 11 type and minor amounts of isozymes of the 22(1) and the 31(2) type. The axolemma contains 21 and 31 isozymes. A carbohydrate analysis indicated that 11 enzyme preparations from the brain grey matter substantially differ from the renal enzymes of the same composition in the glycosylation of the 1 isoform. An enhanced sensitivity of the 3 catalytic subunit of Na⁺,K⁺-ATPase from neurons to endogenous proteolysis was found. A point of specific proteolysis in the amino acid sequence PNDNR⁴⁹² Y⁴⁹³ was localized (residue numbering is that of the human 3 subunit). This sequence corresponds to one of the regions of the greatest variability in 1-, 2-, 3-, and 4-subunits, but at the same time, it is characteristic of the 3 isoforms of various species. The presence of the 3 isoform of tubulin (cytoskeletal protein) was found for the first time in the high-molecular-mass Na⁺,K⁺-ATPase 31 isozyme complex isolated from the axolemma of brain stem neurons, and its binding to the 3 catalytic subunit was shown.     

Taxonomic status of the species of the green algal genusNeochloris

Komárek has recently reviewed the various species assigned to the green algal genusNeochloris Starr (Chlorococcales, Chlorococcaceae) and removed those with uninucleate vegetative cells to a new genus,Ettlia. Watanabe & Floyd, unaware ofKomárek's work, also reviewed the species ofNeochloris and distributed them among three genera—Neochloris, Chlorococcopsis gen. nov., andParietochloris gen. nov.—on the basis of details of the covering of the zoospore and the arrangement of the basal bodies of the flagellar apparatus. This paper reconciles these two treatments and makes additional recommendations at the ranks of genus, family, order, and class.     

Elucidation of Structure of Lipid A from the Marine Gram-Negative Bacterium Pseudoalteromonas haloplanktis ATCC 14393T

The chemical structure of lipid A, from the marine -proteobacterium Pseudoalteromonas haloplanktis 14393, a main product of lipopolysaccharide hydrolysis (1% AcOH), was determined using chemical methods and NMR spectroscopy. The lipid A was shown to be -1,6-glucosaminobiose 1,4-diphosphate acylated with two (R)-3-hydroxyalkanoic acid residues at C3 and C3 and amidated with one (R)-3-hydroxydodecanoyl and one (R)-3-dodecanoyloxydodecanoyl residue at N2 and N2, respectively.     

Phylogeny of cardinalfishes (Teleostei: Gobiiformes: Apogonidae) and the evolution of visceral bioluminescence

The cardinalfishes (Apogonidae) are a diverse clade of small, mostly reef-dwelling fishes, for which a variety of morphological data have not yielded a consistent phylogeny. We use DNA sequence to hypothesize phylogenetic relationships within Apogonidae and among apogonids and other acanthomorph families, to examine patterns of evolution including the distribution of a visceral bioluminescence system. In conformance with previous studies, Apogonidae is placed in a clade with Pempheridae, Kurtidae, Leiognathidae, and Gobioidei. The apogonid genus Pseudamia is recovered outside the remainder of the family, not as sister to the superficially similar genus Gymnapogon. Species sampled from the Caribbean and Western Atlantic (Phaeoptyx, Astrapogon, and some Apogon species) form a clade, as do the larger-bodied Glossamia and Cheilodipterus. Incidence of visceral bioluminescence is found scattered throughout the phylogeny, independently for each group in which it is present. Examination of the fine structure of the visceral bioluminescence system through histology shows that light organs exhibit a range of morphologies, with some composed of complex masses of tubules (Siphamia, Pempheris, Parapriacanthus) and others lacking tubules but containing chambers formed by folds of the visceral epithelium (Acropoma, Archamia, Jaydia, and Rhabdamia). Light organs in Siphamia, Acropoma, Pempheris and Parapriacanthus are distinct from but connected to the gut; those in Archamia, Jaydia, and Rhabdamia are simply portions of the intestinal tract, and are little differentiated from the surrounding tissues. The presence or absence of symbiotic luminescent bacteria does not correlate with light organ structure; the tubular light organs of Siphamia and chambered tubes of Acropoma house bacteria, those in Pempheridae and the other Apogonidae do not.     

The evolution of the Hox cluster: insights from outgroups

Two burgeoning research trends are helping to reconstruct the evolution of the Hox cluster with greater detail and clarity. First, Hox genes are being studied in a broader phylogenetic sampling of taxa: the past year has witnessed important new data from teleost fishes, onychophorans, myriapods, polychaetes, glossiphoniid leeches, ribbon worms, and sea anemones. Second, commonly accepted notions of animal relationships are being challenged by alternative phylogenetic hypotheses that are causing us to rethink the evolutionary relationships of important metazoan lineages, especially arthropods, annelids, nematodes, and platyhelminthes.     

Systematic significance of mature embryo of bamboos

The mature embryo of seven species belonging to five genera of Indian bamboos is described. In all these the basic pattern of embryo organisation is same: the scutellar and coleoptilar bundles are not separated by an internode, the epiblast is absent, the lower portion of the scutellum and the coleorhiza are separated by a cleft and the margins of embryonic leaves overlap. The features unique to fleshy fruited bamboos are: presence of a massive scutellum, the juxtaposition of plumule and radicle and the occurrence of a bud in the axil of the coleoptile. The fleshy fruit bearing bamboos should be classified into one group, the tribeMelocanneae. Evidence is provided to recognise additional groups in the subfamilyBambusoideae.     

N-andO-glycosylation of glycoprotein C synthesized by Herpes simplex virus type 1-infected ricin resistant cells

The progeny of Herpes simplex virus type 1 (HSV-1) grown in ricin-resistant 14 cells (Ric^R14) lackingN-acetylglucosaminyltransferase I was released in the extracellular medium at a very low rate. By using a monoclonal antibody immobilized on Sepharose we purified from HSV-1-infected Ric^R14 cells a viral glycoprotein (gC), which carries bothN-andO-linked oligosaccharides. Glycopeptides obtained from [³H]mannoselabeled gC by Pronase digestion were entirely susceptible to endo--N-acetylglucosaminidase H, and the major oligosaccharide released was Man₄GlcNAc. The accumulation of this high-mannose species was related to the enzymic defect of the host cells and to the long retention of the viral glycoprotein within the cells. The extent ofO-glycosylation evaluated in [¹⁴C]glucosamine-labeled gC from Ric^R14 cells as compared to that of gC from wild type cells did not appear to be significantly modified.Abbreviations Con A concanavalin A - BHK cells baby hamster kidney cells - HSV Herpes simplex virus