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Sequence analysis of mRNA polyadenylation signals of rice genes

Chinese Science Bulletin 2006年第9期51卷 1069-1077页

作者： LU Ying1, 2. GAO Chenxi1,2.3.& HAN Bin1,3.41. National Center for Gene Research, {2. institutes for {2. Sciences, Chinese Academy of Sciences, {2. 2.02.0}, China 2. Graduate School of the Chinese Academy of Sciences, Beijing 10003., China 3. institute of plant physiology and ecology, {2. institutes for {2. Sciences, Chinese Academy of Sciences, {2. 2.003., China National Center for Gene Research Shanghai Institutes for Biological Sciences Chinese Academy of Sciences Shanghai China Graduate School of the Chinese Academy of Sciences Beijing China institute of plant{2. and {3. Shanghai Institutes for Biological Sciences Chinese Academy of Sciences Shanghai China

The formation of eukaryotic mRNAs involves the cleavage and polyadenylation of pre- mRNAs. To investigate the sequence requirement of putative polyadenylation signals (PASs), poly(A) sites and downstream elements (DUEs) in 3.-end-pro- cessing in rice, we compared expressed sequences tags (ESTs) with poly(A) extremity to full-length cDNA sequences and constructed a database of 12.69 pre- mRNA sequences in ?40―+40 nt surrounding the poly(A) sites, which were from 9953.genes. The al- ternative poly(A) sites were revealed in approxi- mately 2.% of mRNAs. Nearly 80% of pre-mRNAs showed stringent requirement of the YA (CA or UA) at poly (A) sites for polyadenylation. About 7.9% had the AAUAAA signals on ?40―?1 nt upstream of the poly(A) sites. Over 60% of mRNAs probably used the one- or two-base variants of AAUAAA hexamers as their PASs in 3. fragments. The single-base variants of AAUGAA revealed the high frequency in 11.5% of 3. fragments. The DUEs were detected in 90% of pre- mRNAs, especially more than half of the pre-mRNAs with multi-base variants of AAUAAA had the DUEs surrounding the poly(A) site. The location of DUE is also important for defining the cleavage site. Al- though most of the rice pre-mRNAs did not contain AAUAAA signal, the existence of downstream ele- ments ensured the efficiency of cleavage-polyade-

关键词：多聚腺苷酸序列分析 mRNA 水稻基因工程

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Anaerobic Induction of Isocitrate Lyase and Malate Synthase in Submerged Rice Seedlings Indicates the Important Metabolic Role of the Glyoxylate Cycle

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Acta Biochimica et Biophysica Sinica 2005年第6期37卷 406-414页

作者： Ying LU~(1.2. Yong-Rui WU~(1.2. Bin HAN~(1.3. 1 National Center for Gene Research. shanghai {2. for {2. Sciences. Chinese Academy of Sciences. shanghai 2.02.0}. China. 2.Graduate School of the Chinese Academy of Sciences. Beijing 10003..China 3.institute of plant physiology and ecology. shanghai {2. for {2. Sciences. Chinese Academy of Sciences. shanghai 2.003.. China National Center for Gene Research Shanghai Institutes for Biological Sciences Chinese Academy of SciencesShanghai 200233 China Graduate School of the Chinese Academy of SciencesBeijing 10003. China institute of plant{2. and {3. Shanghai Institutes for Biological Sciences Chinese Academy of SciencesShanghai 200032 China

The glyoxylate cycle is a modified form of the tricarboxylic acid cycle that converts C2.ompounds into C4 dicarboxylic acids at plant developmental *** studying submerged rice seedlings,we revealed the activation of the glyoxylate cycle by identifying the increased transcripts of mRNAs of thegenes of isocitrate lyase(ICL)and malate synthase(MS),two characteristic enzymes of the *** blot analysis showed that ICL and MS were activated in the prolonged anaerobic *** activity assay of pyruvate decarboxylase and ICL in the submerged seedlings indicated an 8.8-fold and3.5-fold increase over that in the unsubmerged seedlings,*** activity assay of acetyl-coen-zyme A synthetase in the submerged seedlings indicated a 3.fold increase over that in the unsubmergedseedlings,which is important for initiating acetate ***,we concluded that the glyoxylatecycle was involved in acetate metabolism under anaerobic conditions.

关键词： Oryza sativa glyoxylate cycle isocitrate lyase malate synthase acetyl-coenzyme A synthetase anoxia

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Substitutions of the Conserved Gly47 Affect the CF1 Inhibitor and Proton Gate Functions of the Chloroplast ATP Synthase ε Subunit

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Acta Biochimica et Biophysica Sinica 2005年第7期37卷 453-462页

作者： Hui DONG Zhang-Lin NI Jia-Mian WEI institute of plant{2. & {3.,shanghai institutes for biological Sciences,Chinese Academy of Sciences,shanghai 2.003.,China institute of plant{2. & {3. Shanghai Institutes for Biological Sciences Chinese Academy of SciencesShanghai 200032 China

The conserved residue Gly47 of the chloroplast ATP synthase ε subunit was substituted withLeu,Arg,Ala and Glu by site-directed *** process generated the mutants εG47L,εG47R,εG47A and εG47E,*** the ε variants showed lower inhibitory effects on the soluble CF1（-ε）Ca2.-ATPase compared with wild-type ε.In reduced conditions,εG47E and εG47R had a lower inhibitoryeffect on the oxidized CF1（-ε）Ca2.-ATPase compared with wild-type ε.In contrast,εG47L and εG47Aincreased the Ca2.-ATPase activity of soluble oxidized CF1（-ε）.The replacement of Gly47 significantlyimpaired the interaction between the subunit ε and γ in an in vitro binding *** study showed thatall ε variants were more effective in blocking proton leakage from the thylakoid *** enhancedATP synthesis of the chloroplast and restored ATP synthesis activity of the reconstituted membranes to alevel that was more efficient than that achieved by wild-type ε.These results indicate that the conservedGly47 residue of the ε subunit is very important for maintaining the structure and function of the ε subunitand may affect the interaction between the ε subunit,β subunit of CF1and subunit Ⅲ of CF_o,therebyregulating the ATP hydrolysis and synthesis,as well as the proton translocation role of the subunit Ⅲ of CF_o.

关键词： chloroplast ATP synthase ε subunit GST pull-down assay site-directed mutagenesis

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Expressional Analysis of an EREBP Transcription Factor Gene OsEBP-89 in Rice

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Acta Biochimica et Biophysica Sinica 2004年第1期36卷 21-26页

作者： Hui SHEN and Zong-Yang WANG( The State Key Laboratory of plant Molecular Genetics, institute of plant {3. & ecology, shanghai institutes for biological Sciences,the Chinese Academy of Sciences, shanghai 2.003., China ) The State Key Laboratory of plant Molecular Genetics Institute of Plant Physiology & Ecology Shanghai Institutes for Biological Sciences the Chinese Academy of Sciences Shanghai 200032 China

OsEBP-89 gene encodes an ethylene responsive element binding protein (EREBP) transcription factor from rice (Oryza sativa). Northern blot analysis revealed that OsEBP-89 was expressed in root, stem, seeds, flowers and leaves. Histochemical assay showed that β-glucuronidase (GUS) was expressed mainly in phloem of vascular tissues of the root and stem transition region (RST), basal part of sheath roots, stem node and basal part of adventitious roots, also in endosperm of seeds in transgenic rice harboring OsEBP-89/GUS construct (pNSG). A sequence region from -2.9 to -97 was found to play an important role for OsEBP-89 gene expression through promoter deletion assay. The possible function of OsEBP-89 gene was discussed.

关键词： OsEBP-89 gene rice EREBP subfamily transcription factor tissue expression

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Transiently Expressed Short Hairpin RNA Targeting 12. kDa Protein of Tobacco Mosaic Virus Interferes with Virus Infection

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Acta Biochimica et Biophysica Sinica 2006年第1期38卷 22-28页

作者： Ming-Min ZHAO~(1,2.,De-Rong AN~(1*),Jian ZHAO~2.Guang-Hua HUANG~3.Zu-Hua HE~4,and Jiang-Ye CHEN~3. ~1 College of {2. Protection,Northwest Science and Technology University of Agriculture and Forestry,Yangling 712.00,China ~2.College of Agriculture,Yangtze University,Jingzhou 43.02.,China ~3.State Key Laboratory of Molecular Biology,institute of Biochemistry and Cell Biology,shanghai {3. for biological Sciences, Chinese Academy of Sciences,shanghai 2.003.,China ~4 State Key Laboratory of {2. Molecular Genetics,institute of {2. physiology and ecology,shanghai {3. for biological Sciences, Chinese Academy of Sciences,shanghai 2.003.,China College of plant Protection Northwest Science and Technology University of Agriculture and Forestry Yang ling 712100 China College of Agriculture Yangtze University Jingzhou 434025 China State Key Laboratory of Molecular Biology Institute of Biochemistry and Cell Biology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences Shanghai 200031 China State Key Laboratory of plant Molecular Genetics Institute of Plant Physiology and Ecology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences Shanghai 200032 China

RNA-interference (RNAi) silences gene expression by'guiding mRNA degradation in asequence-specific *** interfering RNA (siRNA),an intermediate of the RNAi pathway,has beenshown to be very effective in inhibiting virus infection in mammalian cells and cultured plant ***,wereport that Agrobacterium tumefaciens-mediated transient expression of short hairpin RNA (shRNA) couldinhibit tobacco mosaic virus (TMV) RNA accumulation by targeting the gene encoding the replication-asso-ciated 12. kDa protein in intact plant *** results indicate that transiently expressed shRNA efficientlyinterfered with TMV *** interference observed is sequence-specific,and time-and *** expressed shRNA corresponding to the TMV 12. kDa protein gene did not inhibit cucumbermosaic virus (CMV),an unrelated *** order to interfere with TMV accumulation in tobaccoleaves,it is essential for the shRNA constructs to be infiltrated into the same leaves as TMV *** support the view that RNAi opens the door for novel therapeutic procedures against virus *** propose that a combination of the RNAi technique and Agrobacterium-mediated transient expressioncould be employed as a potent antiviral treatment in plants.

关键词： tobacco mosaic virus 12. kDa protein RNA interference short hairpin RNA

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cvhA Gene of Streptomyces hygroscopicus 10-2. Encodes a Negative Regulator for Mycelia Development

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Acta Biochimica et Biophysica Sinica 2006年第4期38卷 271-280页

作者： Heng-An WANG~(1,2.) Lei QIN~(2.) Ping LU~(1*) Zhi-Xuan PANG~2.Zi-Xin DENG~(3.) Guo-Ping ZHAO~(2.) 1 Biotechnology institute,School of Agriculture and Biology,{2. Jiaotong University,{2. 2.1101,China 2.Laboratory of Molecular Microbiology,institute of plant physiology and ecology,{2. institute for biological Sciences,Chinese Academy of Sciences,{2. 2.003.,China 3.Bio-X Life Science Research Center,{2. Jiaotong University,{2. 2.003.,China. Biotechnology institute School of Agriculture and Biology Shanghai Jiaotong UniversityShanghai 201101 China Laboratory of Molecular Microbiology Institute of Plant Physiology and Ecology Shanghai Institute for Biological Sciences Chinese Academy of SciencesShanghai 200032 China Bio-X Life Science Research Center Shanghai Jiaotong UniversityShanghai 200030 China.

A five-gene cluster cvhABCDE was identified from Streptomyces hygroscopicus *** thefirst gene of this cluster,cvhA encoded a putative sensor histidine kinase with a predicted sensor domainconsisting of two trans-membrane segments at the N-terminus and a conserved HATPasecdomain at the *** C-terminus polypeptide of CvhA expressed in Escherichia coli was purified and shown to beautophosphorylated with [γ-3.P]ATP in *** phosphoryl group was acid-labile and basic-stable,whichsupported histidine as the phosphorylation *** obvious difference of mycelia development was ob-served between the null mutant of cvhA generated by targeted gene replacement and the wild-type parentalstrain 10-2. grown on solid soya flour medium with 2.-8% glucose or sucrose,but the cvhA mutant couldform much more abundant aerial mycelia and spores than the wild-type strain on solid soya flour mediumsupplemented with 6%-8% mannitol,6%-8% sorbitol,4%-6% mannose,or 4%-6% *** pheno-type was complemented by the cloned wild-type cvhA gene,and no difference was observed for growthcurves of the cvhA mutant and the wild strain in liquid minimal medium with the tested sugars at a concen-tration of 4%,6% and 8%.We thus propose that CvhA is likely a sensor histidine kinase and negativelyregulates the morphological differentiation in a sugar-dependent manner in *** 10-2..

关键词： sensor histidine kinase aerial mycelia sugar metabolism

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Comparative Analysis of Two-component Signal Transduction System in Two Streptomycete Genomes

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Acta Biochimica et Biophysica Sinica 2007年第5期39卷 317-325页

Species of the genus Streptomyces are major bacteria responsible for producing most natural antibiotics. Streptomyces coelicolor A3.2. and Streptomyces avermitilis were sequenced in 2.02.and 2.03. respectively. Two-component signal transduction systems (TCSs), consisting of a histidine sensor kinase (SK) and a cognate response regulator (RR), form the most common mechanism of transmembrane signal transduction in prokaryotes. TCSs in S. coelicolor A3.2. have been analyzed in detail. Here, we identify and classify the SK and RR of S. avermitilis and compare the TCSs with those of S. coelicolor A3.2. by computational approaches. Phylogenetic analysis of the cognate SK-RR pairs of the two species indicated that the cognate SK-RR pairs fall into four classes according to the distribution of their orthologs in other organisms. In addition to the cognate SK-RR pairs, some potential partners of non-cognate SK-RR were found, including those of unpaired SK and orphan RR and the cross-talk between different components in either strain. Our study provides new clues for further exploration of the molecular regulation mechanism of streptomycetes with industrial importance.

关键词： Streptomyces two-component system cross-talk phylogenetic analysis

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Transcriptional, post-transcriptional and post-translational regulations of gene expression during leaf polarity formation

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Cell Research 2007年第6期17卷 512-519页

作者： Lin Xu Li YangHai Huang National Laboratory of plant Molecular Genetics, shanghai{2. of plant physiology and ecology, shanghai institutes for biological Sciences, Chinese Academy of Sciences, 3.0 Fenglin Road, shanghai 2.003., China National Laboratory of plant Molecular Genetics Shanghai Institute of Plant Physiology and Ecology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences Shanghai China Present address: IBMP CNRS Universite Louis Pasteur 67084 Strasbourg France Present address: Department of Biology University of Pennsylvania Philadelphia PA 19104 USA

Leaf morphogenesis requires the establishment of adaxial-abaxial polarity after primordium initiation from the shootapical meristem (SAM). Several families of transcription factors are known to play critical roles in promoting adaxial orabaxial leaf fate. Recently, post-transcriptional gene silencing pathways have been shown to regulate the establishmentof leaf polarity, providing novel and exciting insights into leaf development. For example, microRNAs (miR165/166)and a trans-acting siRNA (TAS3.derived tasiR-ARF) have been shown to repress the expression of several key transcrip-tion factor genes. In addition, yet another level of regulation, post-translational regulation, has been revealed recently bystudies on the role of the 2.S proteasome in leaf polarity. Although our understanding regarding the molecular mecha-nisms underlying establishment of adaxial-abaxial polarity has greatly improved, there is still much that remains *** review aims to discuss recent progress, as well as the remaining questions, regarding the molecular mechanismsunderlying leaf polarity formation.

关键词： Arabidopsis leaf development polarity formation

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Sequencing and Analysis of a Genomic Fragment Provide an Insight into the Dunaliella viridis Genomic Sequence

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Acta Biochimica et Biophysica Sinica 2006年第11期38卷 812-820页

作者： Xiao-Ming SUN~(1#) Yuan-Ping TANG~(1#) Xiang-Zong MENG~1 Wen-Wen ZHANG~1 Shan LI~1 Zhi-Rui DENG~1 Zheng-Kai XU~(1,2. Ren-Tao SONG~1* 1 shanghai Key Laboratory of Bio-energy Crops,School of Life Sciences,shanghai University,shanghai 2.0444,China 2.{3. of plant physiology & ecology,shanghai institutes for biological Sciences,Chinese Academy of Sciences,shanghai 2.003.,China shanghai Key Laboratory of Bio-energy Crops School of Life Sciences Shanghai UniversityShanghai 200444 China institute of plant{2. & {3. Shanghai Institutes for Biological Sciences Chinese Academy of SciencesShanghai 200032 China

Dunaliella is a genus of wall-less unicellular eukaryotic green *** exceptional resistancesto salt and various other stresses have made it an ideal model for stress tolerance ***,very littleis known about its genome and genomic *** this study,we sequenced and analyzed a 2.,2.8 bpgenomic fragment from DunalieIla *** fragment showed low sequence homology to the *** the nucleotide level,only a segment with significant sequence homology to 18S rRNA *** fragment contained six putative genes,but only one gene showed significant homology at theprotein level to GenBank *** average GC content of this sequence was 51.1%,which was muchlower than that of close related green algae Chlamydomonas （65.7%）.Significant segmental duplicationswere found within this *** duplicated sequences accounted for about 3..7% of the *** amounts of simple sequence repeats （microsatellites） were found,with strong bias towards（AC）_n type （76%）.Analysis of other Dunaliella genomic sequences in the GenBank database （total 2.,749bp） was in agreement with these *** sequence features made it difficult to sequence Dunaliellagenomic *** investigation should be made to reveal the biological significance of these uniquesequence features.

关键词： Dunaliella viridis genomic sequence sequence feature sequence duplication simple sequence repeat

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nifH Promoter Activity Is Regulated by DNA Supercoiling in Sinorhizobium meliloti

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Acta Biochimica et Biophysica Sinica 2005年第4期37卷 221-226页

作者： Yan-Jie LIU Biao HU Jia-Bi ZHU Shan-Jiong SHEN Guan-Qiao YU National Laboratory of plant Molecular Genetics,institute of plant {3. and ecology,shanghai institutes for biological Sciences,Chinese Academy of Sciences,shanghai 2.003.,China National Laboratory of plant Molecular Genetics Institute of Plant Physiology and Ecology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences Shanghai 200032 China

In prokaryotes,DNA supercoiling regulates the expression of many genes;for example,theexpression of Klebsiella pneumoniae nifLA operon depends on DNA negative supercoiling in anaerobicallygrown cells,which indicates that DNA supercoiling might play a role in gene regulation of the *** the expression of the nifH promoter in Sinorhizobium meliloti is not repressed by oxygen,itis proposed that the status of DNA supercoiling may not affect the expression of the nifH *** this hypothesis by analyzing nifH promoter activity in wild-type and gyr Escherichia coli in thepresence and absence of DNA gyrase *** results show that gene expression driven by the *** nifH promoter requires the presence of active DNA *** DNA gyrase increases thenumber of negative superhelical turns in DNA in the presence of ATP,our data indicate that negativesupercoiling is also important for nifH promoter *** study also shows that the DNA supercoiling-dependent *** nifH promoter activity is related to the trans-acting factors NtrC and NifA that *** supercoiling appeared to have a stronger effect on NtrC-activated nifH promoter activity than onNifA-activated promoter ***,these results from the *** nifH promoter model systemseem to indicate that,in addition to regulating gene expression during anaerobic signaling,DNA supercoilingmay also provide a favorable topology for trans-acting factor binding and promoter activation regardless ofoxygen status.

关键词： nifH DNA supercoiling gyrase Sinorhizobium meliloti gene expression

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