DnaK-dependent ribosome biogenesis in Escherichia coli? : competition for dominance between the alleles dnaK756 and dnaK ?+

The Escherichia coli chaperone DnaK is vital for many cellular functions, including ribosome biogenesis at high temperature. Thus, the dnaK756-ts (λ ^R ) mutant, at the non-permissive temperature, is inhibited at a late stage of ribosome assembly, yielding 21S, 32S and 45S precursor particles. This defect, unlike the λ resistance and thermosensitivity phenotypes, is not complemented by lysogenisation with a transducing phage λ dnaK ⁺ bearing the wild-type dnaK gene. However this dominant phenotype becomes recessive when dnaK ⁺ is expressed from a medium-copy-number plasmid. On the other hand, an excess of DnaK causes an unexpected dominant-lethal effect of the dnaK756 allele near non-permissive temperatures. This interplay between the dnaK ⁺ and dnaK756 alleles supports the idea of that DnaK oligomers form in the cell. Received: 28 April 1998 / Accepted: 24 July 1998     

乳酸乳球菌fabZ1和fabZ2基因功能的鉴定

大肠杆菌(Escherichia coli)是Ⅱ型脂肪酸合成系统的模式生物,3-羟基脂酰ACP脱水异构酶(FabA)是不饱和脂肪酸合成中的关键酶.生物信息学分析表明,乳酸乳球菌(Lactococcus lactis)的基因组中没有标注为3-羟基脂酰ACP脱水异构酶的基因,但有两个标注为3-羟基脂酰ACP脱水酶基因LlfabZ1和LlfabZ2,其编码的蛋白质与EcFabZ的相似性分别为41%和45.1%,且都具有3-羟基脂酰ACP脱水酶两个保守的α螺旋结构.用携带LlfabZ1和LlfabZ2的质粒载体遗传互补大肠杆菌fabA温度敏感突变株CY57,在42℃下不能恢复生长,但无细胞抽提物的结果显示LlFabZ1能够使反-2-癸烯酰ACP异构成顺-3-癸烯酰ACP,而LlFabZ2则不能.互补大肠杆菌fabZ突变株HW7显示,在诱导的条件下,含有LlfabZ2的转化子能够恢复生长,而LlfabZ1则不能.体外重建脂肪酸合成反应及蛋白质活性测定表明,LlFabZ1具有3-羟基脂酰ACP脱水异构酶功能,而LlFabZ2只具有3-羟基脂酰ACP脱水酶功能.另外,未得到LlfabZ1和LlfabZ2的突变株,表明LlFabZ1和LlFabZ2可能是乳酸乳球菌脂肪酸合成酶系中的必不可少的关键蛋白.上述结果证实了乳酸乳球菌fabZ1和fabZ2两个基因在脂肪酸合成中的功能.     

香菜CsEF 1α基因克隆与表达分析

翻译延伸因子EF 1α(elongation factor 1 alpha)是细胞中最丰富的蛋白质之一,其在确保mRNA正确解码以产生细胞蛋白质方面发挥重要作用。该研究采用RT PCR扩增方法克隆香菜CsEF 1α基因序列,利用生物信息学对CsEF 1α基因结构、序列特征及系统进化等进行分析,并采用qPCR探究CsEF 1α基因在香菜不同生长时期和非生物胁迫下的表达模式,为进一步揭示EF 1α基因调控机制的研究奠定基础。结果显示：(1)成功克隆获得香菜CsEF 1α基因序列;CsEF 1α基因包含1个1 344 bp的开放阅读框,编码447个氨基酸,分子式为C₂₂₀₂H₃₅₄₄N₅₉₄O₆₄₄S₂₀,蛋白质分子量为49.29 kD,等电点为9.12;氨基酸序列组成中赖氨酸数量最多(49个,占11.0%),色氨酸数量最少(3个,占0.7%);属碱性蛋白。(2)CsEF 1α蛋白主要由无规则卷曲(36.91%)和α 螺旋(30.43%)构成,定位于细胞质;系统进化树分析显示,CsEF 1α与胡萝卜、野生番茄、青蒿素和非洲菊的亲缘关系较接近;启动子分析包括4种植物生长发育元件、3种激素响应元件和3种胁迫响应元件。(3)qRT PCR结果显示,CsEF 1α基因的表达量随着香菜生长发育时间的延长而升高,并且与转录丰度的变化一致;CsEF 1α基因对4种不同非生物胁迫的响应表达模式有所差异;随着胁迫时间的延长,在盐胁迫下CsEF 1α基因表现出先升高后降低趋势,而在低温、高温和干旱胁迫下表现出先降低再升高的趋势。研究表明,CsEF 1α基因参与了香菜对非生物胁迫的应答,在香菜生长发育和非生物胁迫中具有重要调控作用。     

甘蓝型油菜RPD3/HDA1基因家族鉴定及早熟相关基因挖掘

该研究运用生物信息学方法鉴定甘蓝型油菜RPD3/HDA1基因家族,检测了其在‘黔油早2号’和‘中双11’中的表达水平及2个品种在低温(4℃)和ABA胁迫下该家族基因的表达特征,以探讨RPD3/HDA1基因在甘蓝型油菜中的潜在功能,为早熟油菜抗逆性遗传改良提供理论基础和候选基因。结果表明:(1)在甘蓝型油菜全基因组中共鉴定到28个RPD3/HDA1基因,将其命名为BnHDA1~BnHDA28,聚类为4个亚家族,同一亚家族成员的基因结构较为相似;在该基因家族中共检测到16对复制基因,均为片段重复。(2)顺式作用元件预测统计中共发现675个元件与植物激素、环境胁迫和光响应有关。(3)qRT-PCR分析显示,RPD3/HDA1基因在‘黔油早2号’中的表达量均高于‘中双11’;低温胁迫下,‘黔油早2号’和‘中双11’中RPD3/HDA1基因呈差异表达,与‘中双11’相比,RPD3/HDA1基因在‘黔油早2号’中的下调幅度较大;ABA处理后,RPD3/HDA1基因在2个品种中表达模式不一致,‘黔油早2号’中大部分RPD3/HDA1基因表达量较‘中双11’下调幅度小。研究认为,RPD3/HDA1基因可能在油菜开花中发挥调节作用,而且可能通过激素信号通路和防御信号通路参与油菜的生长发育和防御反应的调节。     

转录因子cpcR同源基因cpcR-c1和cpcR-t的功能鉴定

苏云金芽胞杆菌（Bacillus thuringiensis,Bt） LM1212菌株与典型的Bt菌株表型不同,可分化形成芽胞、形成细胞和晶体产生细胞。在LM1212菌株中,转录因子CpcR不仅参与了细胞分化过程,而且能够激活晶体蛋白基因cry35-like的启动子(P₃₅)。【目的】筛选cpcR同源基因,验证其生物学功能。【方法】本研究克隆了2个cpcR同源基因,来源于蜡样芽胞杆菌的cpcR-c1和来源于东洋芽胞杆菌的cpcR-t,将cpcR及其同源基因分别构建在pHT304-P₃₅-gfp、pHT304-P₃₅-lacZ报告载体上,获得的重组质粒转入无cpcR基因且无晶体蛋白基因的Bt HD73^–菌株中。利用激光共聚焦显微镜观察重组菌HD^–(cpcR-c1-P₃₅-gfp)和HD^–(cpcR-t-P₃₅-gfp)的细胞表型并进行芽胞计数实验。测定HD^–(cpcR-c1-P_35<...     

酵母PMT1和TED1基因在细胞壁应激反应中的作用

实验室前期研究表明,蛋白质O-甘露糖转移酶1 (Protein O-mannosyltransferase 1,Pmt1p) 和Ted1p（Traffcking of Emp24p/Erv25p-dependent cargo disrupted）在细胞寿命和内质网应激反应方面存在相互调控关系。进一步研究酵母Pmt1p和Ted1p在细胞壁应激反应中（诱导剂为荧光增白剂或刚果红）的作用。观察PMT1基因缺失（pmt1Δ）酵母菌株和TED1基因缺失（ted1Δ）酵母菌株,以及PMT1和TED1双基因缺失（pmt1Δ ted1Δ）酵母菌株在细胞壁应激反应条件下的克隆形成能力和细胞分裂增殖活性;qRT-PCR检测细胞壁应激反应通路中Slt2p、Ssd1p和Mpt5p等效应蛋白的转录水平。结果表明,在细胞壁应激反应条件下,与对照菌株的生长状态比较,pmt1Δ菌株生长缓慢,ted1Δ菌株生长较快;进一步缺失PMT1基因使得ted1Δ菌株生长缓慢。与对照菌株中效应蛋白的转录水平比较,pmt1Δ菌株和pmt1Δted1Δ菌株中SLT2、SSD1和MPT5基因的转录水平明显上调,ted1Δ菌株中的无明显变化;与pmt1Δ菌株比较,pmt1Δted1Δ菌株中SLT2和MPT5表达明显下调,SSD1表达无明显变化。缺失TED1基因增强酵母细胞对细胞壁应激反应的抵抗性;进一步缺失PMT1基因增强ted1Δ菌株对应激反应的敏感性,上调细胞壁应激反应通路中效应蛋白的转录表达水平。     

Synthesis and transporter binding properties of (R)-2β,3β- and (R)-2α,3α-diaryltropanes

(R)-2-Aryl-2-tropinone (9) was synthesized from (R)-2-carbomethoxy-3-tropinone (5) and was used as the key intermediate for the synthesis of (R)-2β,3β- and (R)-2α,3α-diaryltropanes. Inhibition of radioligand binding studies at the dopamine, serotonin, and norepinephrine transporters showed that the (R)-3β-(4-methylphenyl)-2β-phenyltropane (3b, RTI-422) possessed an IC₅₀ value of 1.96 nM at the dopamine transporter and was highly selective for this transporter relative to the serotonin and norepinephrine transporters.     

Overexpression of Arabidopsis thaliana Na+/H+ antiporter gene enhanced salt resistance in transgenic poplar (Populus?×?euramericana ‘Neva’)

Salinity is a major abiotic stress factor limiting plant growth and productivity. One possible method to enhance plant salt-resistance is to compartmentalize sodium ions away from the cytosol. In the present work, a vacuolar Na⁺/H⁺ antiporter gene AtNHX1 from Arabidopsis thaliana, was transferred into Populus × euramericana ‘Neva’ by Agrobacterium tumefaciens in order to enhance poplar salt-resistance. The results showed that the transgenic poplar were more resistant to NaCl than the wild-type (WT) in greenhouse condition. Compared with the WT, plant growth and photosynthetic capacity of the transgenic plants were enhanced, and the transgenic plants accumulated more Na⁺ and K⁺ in roots and leaves under the same NaCl condition, whereas malondialdehyde and relative electrical conductivity were lower. All of these properties of the transgenic poplar were likely to be a consequence of the overexpression of AtNHX1 caused Na⁺ sequestration in the vacuoles and improved K⁺ absorption, thus reducing their toxic effects. These results indicated overexpression of the AtNHX1 enhanced salt-resistance of poplar, and AtNHX1 played an important role in the compartmentation of Na⁺ into the vacuoles. Therefore, this study provides an effective way for improving salt resistance in trees.     

Characterization of the lys2 gene of Penicillium chrysogenum encoding α -aminoadipic acid reductase

A DNA fragment containing a gene homologous to LYS2 gene of Saccharomyces cerevisiae was cloned from a genomic DNA library of Penicillium chrysogenum AS-P-78. It encodes a protein of 1409 amino acids (Mr^ 154 859) with strong similarity to the S. cerevisiae (49.9% identity) Schizosaccharomycespombe (51.3% identity) and Candida albicans (48.12% identity) α-aminoadipate reductases and a lesser degree of identity to the amino acid-activating domains of the non-ribosomal peptide synthetases, including the α-aminoadipate-activating domain of the α-aminoadipyl-cysteinyl-valine synthetase of P. chrysogenum (12.4% identical amino acids). The lys2 gene contained one intron in the 5′-region and other in the 3′-region, as shown by comparing the nucleotide sequences of the cDNA and genomic DNA, and was transcribed as a 4.7-kb monocistronic mRNA. The lys2 gene was localized on chromosome III (7.5 Mb) in P. chrysogenum AS-P-78 and on chromosome IV (5.6 Mb) in strain P2, whereas the penicillin gene cluster is known to be located in chromosome I in both strains. The lys2-encoded protein is a member of the aminoacyladenylate-forming enzyme family with a reductase domain in its C-terminal region. Received: 26 January 1998 / Accepted: 4 May 1998     

Functional characterization of kanB by complementing in engineered Streptomycesfradiae ?neo6::tsr

A putative aminotransferase gene, kanB, lies in the biosynthetic gene cluster of Streptomyces kanamyceticus ATCC 12853 and has 66% identity with neo6 in neomycin biosynthesis. Streptomyces fradiae ∆neo6::tsr was generated by disrupting neo6 in the neomycin producer Streptomyces fradiae. Neomycin production was completely abolished in the disruptant mutant but was restored through self-complementation of neo6. S. fradiae HN4 was generated through complementation with kanB in Streptomyces fradiae ∆neo6::tsr. Based on metabolite analysis by ESI/MS and LC/MS, neomycin production was restored in Streptomyces fradiae HN4. Thus, like neo6, kanB also functions as a 2-deoxy-scyllo-inosose aminotransferase that has dual functions in the formation of 2-deoxy-scyllo-inosose (DOS). Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Singlet oxygen formation and chlorophyll a triplet excited state deactivation in the cytochrome b6f complex from Bryopsis?corticulans

We have attempted to investigate the correlation between the detergent-perturbed structural integrity of the Cyt b ₆ f complex from the marine green alga Bryopsis corticulans and its photo-protective properties, for which the nonionic detergents n-octyl-β-d-glucopyranoside (β-OG) and n-dodecyl-β-d-maltoside (β-DM), respectively, were used for the preparation of Cyt b ₆ f, and the singlet oxygen (¹O₂*) production as well as the triplet excited-state chlorophyll a (³Chl a*) formation and deactivation were examined by spectroscopic means. Near-infrared luminescence of ¹O₂ * (~1,270 nm) on photo-irradiation was detected for the β-OG preparation where the complex is mainly in oligomeric state, but not for the β-DM one in which the complex exists in dimeric form. Under anaerobic condition, photo-excitation of Chl a in the β-DM preparation generated ³Chl a* with a lower quantum yield of Φ_T ~ 0.02 and a longer lifetime of ~600 μs with respect to those as in the case of β-OG preparation, Φ_T ~ 0.12 and 200–300 μs. These results prove that the enzymatically active and intact Cyt b ₆ f complex on photo-excitation tends to produce little ³Chl a* or ¹O₂ *, which implies that the pigment–protein assembly of Cyt b ₆ f complex per se is crucial for photo-protection. F. Ma and X.-B. Chen contributed equally to this work.     

热带假丝酵母脂肪醛脱氢酶基因CtAld1和CtAld2的功能评价

【目的】热带假丝酵母是发酵法生产二元酸的重要工业菌株,具有较高的ω-氧化活性。脂肪醛脱氢酶在ω-氧化途径中起重要作用,催化脂肪醛生成脂肪酸,但其具体催化功能及对细胞生理影响还未被系统研究。本文通过删除脂肪醛脱氢酶基因CtAld1和CtAld2鉴定了其在ω-氧化途径中的功能。【方法】通过基因组信息挖掘获得热带假丝酵母脂肪醛脱氢酶基因CtAld1和CtAld2序列,在此基础上,通过同源重组敲除CtAld1和CtAld2基因。考察突变株的生长和胞内脂肪醛脱氢酶活性变化,并评价CtAld1和CtAld2基因敲除对细胞二元酸合成能力的影响。【结果】分别获得了热带假丝酵母突变株XZX-1(ΔCtAld1/ΔCtAld1)、XZX-2(ΔCtAld2/ΔCtAld2)和XZX-12(ΔCtAld1/ΔCtAld1,ΔCtAld2/ΔCtAld2)。在以十二烷为唯一碳源的培养基中,敲除CtAld2基因显著抑制细胞的生长,胞内脂肪醛脱氢酶活性降低为出发菌株的30%;敲除CtAld1基因尽管会使细胞损失一部分醛脱氢酶活性,但能够一定程度地提升细胞在十二烷中的生长性能。敲除CtAld1或CtAld2会降低菌株二元酸产量,组合敲除CtAld1和CtAld2严重削弱菌株十二碳二元酸的合成能力。【结论】CtAld2对热带假丝酵母细胞的生长和十二碳二元酸的合成具有重要作用,缺失CtAld1或CtAld2基因降低细胞的二元酸合成能力。CtAld1和CtAld2可作为热带假丝酵母ω-氧化途径代谢工程改造的潜在靶点。     

芥菜BjuWRKY75基因表达及其与开花整合子BjuFT互作

芥菜(Brassica juncea)是十字花科芸薹属一年或二年生蔬菜,其产品器官的产量和品质会受到开花时间的影响。WRKY家族成员具有响应生物和非生物胁迫、发育调控和信号转导等作用。WRKY75是WRKY家族中能够调节开花的重要成员,但在芥菜中的开花调控机制还未见报道。本研究克隆了芥菜BjuWRKY75基因,发现其编码蛋白具有高度保守的WRKY结构域,属于Ⅱ类WRKY蛋白,与黑芥BniWRKY75同源性最高。BjuWRKY75在花中表达丰度显著高于叶和茎,并且在叶中表达较为稳定。BjuWRKY75定位于细胞核,能够与含有W-box应答元件的开花整合子BjuFT的启动子相互作用,且能转录激活下游基因表达。BjuWRKY75转入拟南芥可显著提早开花。综上说明,BjuWRKY75能够直接靶向BjuFT从而促进开花。这为深入研究BjuWRKY75开花分子调控奠定了基础。     

马尾松PmPGK1和PmGPIC基因的克隆和表达分析

为了解马尾松（Pinus massoniana）磷酸甘油酸激酶1（PGK1）与胞质溶胶葡萄糖磷酸异构酶（GPIC）的功能，采用RACE技术克隆了PmPGK1和PmGPIC基因，并进行了生物信息学分析与亚细胞定位，采用实时荧光定量PCR技术分析PmPGK1和PmGPIC的表达特性。结果表明，PmPGK1和PmGPIC全长为2 106和1 848 bp，分别编码507和566个氨基酸。PmPGK1和PmGPIC分别定位于叶绿体和胞质溶胶。PmPGK1表达量为新叶 > 老叶 > 新茎 > 根 > 花；而PmGPIC为老叶 > 花 > 新叶 > 新茎 > 根。低温胁迫24 h，PmPGK1和PmGPIC的表达量均随时间延长先降低后升高，且PmGPIC的表达量在处理2 h后即降至较低水平；高浓度CO₂胁迫24 h，PmPGK1的表达量随时间延长呈降低-升高-再降低的变化趋势，PmGPIC的表达下调但变化较不显著。因此，推测PmPGK1主要参与卡尔文循环及叶绿体/质体糖酵解，PmGPIC主要参与细胞质基质糖酵解；PmPGK1、PmGPIC活性在低温胁迫下均受抑制；PmPGK1活性在CO₂胁迫下受到显著抑制，而PmGPIC活性的影响不大。     

ΓОРИЗОНТ С Polydesmia И ИХ РОДСТВЕННЫЕ СВЯЗИ С Ordosoceras

1.BBe及eH“e B几aHHo仑eTaTbe onHeHBa幻Te只3K3eMn几兄PH,eo6PaHHNe B oeHoBHoM nPo中.几y只Hb一xao,np帅.My 3Hb一q、H H aBTopoM B 19,9 ro八y BoBpeM只pa6oTHB玖30以3。-山aHbeKoM Pa益oHe BHyTPeHHe益MOHrO涯HH H qacTHqHO 3Kcne八H从H分MH从633,从20,CeBe-Po一3ana八Horo reo几orllqecKoroy且paB几eHH只H OP八oCKo盆3Kene及H玖He丘Feo几orHqeeKoroynpaB几eHH只BHyTpeHHe直MOHro月HHM对HHcTepcTBa几。几or“H KHP Bo BpeM分o6e几e八。-BaTe几beKo益reo卫JHqeeK诚Pa6oT曰B双aHHoM’…     

Der Bienenfresser(Merops apiaster L.) in ?sterreich

Ohne Zusammenfassung     

Production of fertile somatic hybrids of Gossypium hirsutum?+?G. bickii and G. hirsutum?+?G. stockii via protoplast fusion

Fertile somatic hybrids were obtained via symmetric electrofusion of protoplasts from two combinations of tetraploid cotton (G. hirsutum cv. Coker 201, AD genome) and diploid wild cottons G. bickii (G genome) and G. stockii (E genome), respectively. Observation by morphological, flow cytometric analysis, chromosome counting and RAPD analysis of the tested hybrids of Coker 201 + G. bickii and Coker 201 + G. stockii confirmed the regenerated plants as hybrid status. Cytological investigation of the metaphase root-tip cells revealed there were 78 chromosomes in the hybrids. Flow cytometric analysis showed the tested plants had a relative DNA contents close to the total DNA contents of the two parents. RAPD analysis revealed the hybrids contained specific genomic fragments from both fusion partners, further confirmed their hybridity. The morphology of the hybrids was intermediate between the two fusion partners. The hybrid plants were successfully transferred to the soil, and they bloomed and set bolls. It is sure that the new hexaploids developed by cell fusion would contribute to cotton breeding through backcrossing with the elite genotypes of G. hirsutum.     

Transformation of androgenic-derived Festulolium plants (Lolium?perenne L.?×?Festuca?pratensis Huds.) by Agrobacterium?tumefaciens

Genetic transformation of androgenic-derived amphidiploid Festulolium plants (Lolium perenne L. × Festuca pratensis Huds., 2n = 4x = 28) by Agrobacterium tumefaciens has been achieved. Anther culture-induced calli of Festulolium “Bx351” were inoculated with Agrobacterium tumefaciens strain LBA4404 carrying pIG121-Hm encoding the hygromycin resistance (hph) and β-glucuronidase (uidA) genes under the control of a CaMV 35S promoter. Twenty-three putative transformants were obtained from the hygromycin selection, 19 of which (82.6%) showed GUS activity. The integration of transgene was detected by using genomic DNA PCR analysis, RT-PCR analysis and Southern blot hybridization, respectively, which revealed that foreign gene was integrated into the genomes of dihaploid transformants (2n = 2x = 14). The haploid embryogenic system offers a stable means of transformation, as the introduced trait can be readily fixed through chromosome doubling. An erratum to this article can be found at