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Junichi Obokata - One of the best experts on this subject based on the ideXlab platform.

  • early light response of psad psae and psah gene families of photosystem i in Nicotiana sylvestris psi d has an isoform of very quick response
    Plant and Cell Physiology, 1995
    Co-Authors: Yoshiharu Y. Yamamoto, Masayuki Nakamura, Yukiko Kondo, Hideo Tsuji, Junichi Obokata
    Abstract:

    Light response of the psaD, psaE and psaH gene families of photosystem I was studied in etiolated seedlings and dark-adapted mature leaves of Nicotiana sylvestris using RNase protection assays and immunoblot analysis. The results suggest that the initial light-response consists of at least two types; a very quick response found only in the psaDb gene, and a slower response common to all the examined genes.

  • 5′-Leader of a Photosystem I Gene in Nicotiana sylvestris, psaDb, Contains a Translational Enhancer
    The Journal of biological chemistry, 1995
    Co-Authors: Yoshiharu Y. Yamamoto, Hideo Tsuji, Junichi Obokata
    Abstract:

    Messenger RNA primary structures responsible for translational efficiency of a photosystem I gene, psaDb, of Nicotiana sylvestris were studied using a transgenic tobacco system. The entire 5′-leader (23 base pairs) with the first four amino acid codons of the protein coding region was fused in frame with the β-glucuronidase (GUS) gene under the control of the 35 S promoter of cauliflower mosaic virus (CaMV). This construct (CaMV::psaDb-GUS′) was introduced into tobacco. GUS activity and GUS mRNA levels were determined for individual transformants, revealing that the insertion of the psaDb sequence greatly enhanced the GUS activity relative to GUS mRNA abundance. The GUS activity/GUS mRNA was 14 times higher in the CaMV::psaDb-GUS′ transformants than in the control CaMV::GUS′ transformants. The high GUS activity/GUS mRNA of the CaMV::psaDb-GUS′ transformants was reduced 20-fold when 13 bases within the psaDb leader were altered. These 13 bases are common to the leaders of an Arabidopsis ferredoxin gene and the psaDb gene of N. sylvestris. Since GUS proteins encoded by these chimeric GUS genes have identical amino acid sequences, these results indicate that the 5′-leader of the psaDb mRNA contains a translational enhancer element.

  • 5 leader of a photosystem i gene in Nicotiana sylvestris psadb contains a translational enhancer
    Journal of Biological Chemistry, 1995
    Co-Authors: Yoshiharu Y. Yamamoto, Hideo Tsuji, Junichi Obokata
    Abstract:

    Messenger RNA primary structures responsible for translational efficiency of a photosystem I gene, psaDb, of Nicotiana sylvestris were studied using a transgenic tobacco system. The entire 5′-leader (23 base pairs) with the first four amino acid codons of the protein coding region was fused in frame with the β-glucuronidase (GUS) gene under the control of the 35 S promoter of cauliflower mosaic virus (CaMV). This construct (CaMV::psaDb-GUS′) was introduced into tobacco. GUS activity and GUS mRNA levels were determined for individual transformants, revealing that the insertion of the psaDb sequence greatly enhanced the GUS activity relative to GUS mRNA abundance. The GUS activity/GUS mRNA was 14 times higher in the CaMV::psaDb-GUS′ transformants than in the control CaMV::GUS′ transformants. The high GUS activity/GUS mRNA of the CaMV::psaDb-GUS′ transformants was reduced 20-fold when 13 bases within the psaDb leader were altered. These 13 bases are common to the leaders of an Arabidopsis ferredoxin gene and the psaDb gene of N. sylvestris. Since GUS proteins encoded by these chimeric GUS genes have identical amino acid sequences, these results indicate that the 5′-leader of the psaDb mRNA contains a translational enhancer element.

  • Organization of the psaH gene family of photosystem I in Nicotiana sylvestris.
    Plant & cell physiology, 1994
    Co-Authors: Masayuki Nakamura, Junichi Obokata
    Abstract:

    The PSI-H subunit of photosystem I has two isoforms of differing molecular mass in Nicotiana sylvestris [Obokata et al. (1993) Plant Physiol. 102: 1259], and is encoded by a nuclear gene, psaH. We identified three structurally distinct psaH genes in the nuclear genome of N. sylvestris, designated psaHa, psaHb, and psaHc, and all three genes are expressed in young leaves. Each gene has two introns: one between sequences encoding a transit peptide and the N-terminal acidic domain, and one between the N-terminal domain and a central hydrophobic domain. The deduced amino acid sequences are identical in the mature proteins and differ only in the transit peptides. Since PSI-H is present in two isoforms in N. sylvestris, the psaH products may be subjected to post-translational modifications.

  • Microheterogeneity of PSI-E subunit of photosystem I in Nicotiana sylvestris.
    Plant & cell physiology, 1994
    Co-Authors: Junichi Obokata, Kohki Mikami, Yoshiharu Y. Yamamoto, Nobuaki Hayashida
    Abstract:

    Microheterogeneity of a photosystem I (PSI) subunit encoded by a nuclear gene psaE was examined in Nicotiana sylvestris, with the aid of cDNA cloning, peptide mapping analysis and protein sequencing. The psaE product of this plant has four isoforms whose mobilities in PAGE are slightly different from each other. We isolated two types of psaE cDNAs from a N. sylvestris cDNA library, and designated the corresponding genes as psaEa and psaEb, respectively. The psaEa and psaEb genes are 77% homologous at DNA level, and their translation products share 80.4% homology for the precursor proteins and 89.1% for the mature forms. Comparative analysis of the four isoproteins and the putative products of the two psaE genes revealed that two isoproteins out of four are derived horn psaEa gene, and the difference between these two isoproteins lies in the respective presence or absence of N-terminal alanine. Likewise, the other two proteins are derived from psaEb with similar N-terminal heterogeneity. These results indicate that multi-gene organization and heterogeneous N-terminal formation at post-translational level are two possible causes for PSI subunit polymorphism in isogenic plant lines.

Fernand Vedel - One of the best experts on this subject based on the ideXlab platform.

  • Organization and expression of the mitochondrial genome in the Nicotiana sylvestris CMSII mutant
    Genetics, 1998
    Co-Authors: Christine Lelandais, R. De Paepe, Fernand Vedel, Béatrice Albert, Sophie Gutierres, Bernard Godelle, P. Chetrit
    Abstract:

    Previous analyses suggested that the Nicotiana sylvestris CMSII mutant carried a large deletion in its mitochondrial genome. Here, we show by cosmid mapping that the deletion is 60 kb in length and contains several mitochondrial genes or ORFs, including the complex I nad7 gene. However, due to the presence of large duplications in the progenitor mitochondrial genome, the only unique gene that appears to be deleted is nad7. RNA gel blot data confirm the absence of nad7 expression, strongly suggesting that the molecular basis for the CMSII abnormal phenotype, poor growth and male sterility, is the altered complex I structure. The CMSII mitochondrial genome appears to consist essentially of one of two subgenomes resulting from recombination between direct short repeats. In the progenitor mitochondrial genome both recombination products are detected by PCR and, reciprocally, the parental fragments are detected at the substoichiometric level in the mutant. The CMSII mtDNA organization has been maintained through six sexual generations.

  • Tissue-specific expression of genes encoding isoforms of the mitochondrial ATPase β subunit in Nicotiana sylvestris
    Plant molecular biology, 1998
    Co-Authors: Eric Lalanne, Fernand Vedel, Chantal Mathieu, R. De Paepe
    Abstract:

    We report here that the catalytic β subunit of the mitochondrial ATPase/ATP synthase is encoded by a small multigenic family in the diploid tobacco Nicotiana sylvestris (nsatp2 genes). cDNAs and genes corresponding to the β1, β2 and β3 (pollen specific) isoforms previously detected by 2D-SDS PAGE were isolated. Nsatp2.1 and nsatp2.2 transcripts were found in all vegetative and reproductive tissues analysed. In contrast, nsatp2.3 transcripts were found exclusively in bicellular pollen. As a whole, steady-state transcript levels of nuclear nsatp2 and mitochondrial atp1 genes were found to be closely correlated.

  • A Promoter Element Active in Run-Off Transcription Controls the Expression of Two Cistrons of Nad and rps Genes in Nicotiana sylvestris Mitochondria
    Nucleic acids research, 1996
    Co-Authors: Christine Lelandais, Chantal Mathieu, Fernand Vedel, Sophie Gutierres, Claire Remacle, Laurence Maréchal-drouard, Axel Brennicke, Stefan Binder, P. Chetrit
    Abstract:

    The expression of two mitochondrial gene clusters (orf87-nad3-nad1/A and orf87-nad3-rps12) was studied in Nicotiana sylvestris. 5' and 3' termini of transcripts were mapped by primer extension and nuclease S1 protection. Processing and transcription initiation sites were differentiated by in vitro phosphorylation and capping experiments. A transcription initiation site, present in both gene clusters, was found 213 nucleotides upstream of orf87. This promoter element matches the consensus motif for dicotyledonous mitochondrial promoters and initiates run-off transcription in a pea mitochondrial purified protein fraction. Processing sites were identified 5' of nad3, nad1/A and rps12 respectively. These results suggest that (i) the expression of the two cistrons is only controlled by one duplicated promoter element, and (ii) multiple processing events are required to produce monocistronic nad3, nad1/A and rps12 transcripts.

  • Cytoplasmic male sterility is associated with large deletions in the mitochondrial DNA of two Nicotiana sylvestris protoclones.
    Current genetics, 1992
    Co-Authors: P. Chetrit, R. De Paepe, R. Rios, V. Vitart, S. Gutierres, Fernand Vedel
    Abstract:

    Two cytoplasmic male-sterile plants (CMSI and CMSII) were obtained by protoplast culture in Nicotiana sylvestris. Both plants showed large deletions (up to 50 kb) in their mitochondrial DNA. Restriction maps of the reorganized regions suggested that the deletions occurred via two homologous recombination events (rec. 1 and rec. 2) in the parental mitochondrial genome. With the exception of nad5, no mitochondrial DNA polymorphism could be detected between parental and CMS lines using different heterologous genes probes. A sequence homologous to the Oenothera nad5 mitochondrial gene was located close to the CMSI-specific rec. 2 region. Moreover, a cDNA probe corresponding to total mitochondrial RNA from the parent line was found to hybridize to mitochondrial DNA fragments involved in the rec. 1 event common to both CMS lines, suggesting that rec. 1 lies in a transcribed region. Cytoplasmic male sterility in the Nicotiana sylvestris CMS mutants could be due either to gene deletion or to a regulatory effect of such a deletion on mitochondrial gene expression, rather than to the presence of specific polypeptides as has been shown in the T cytoplasm of maize, or in CMS Petunia.

  • Oligomycin/venturicidin-resistance is due to a nuclear modification in a somatic hybrid between Nicotiana tabacum and a variant line of Nicotiana sylvestris
    Plant Science, 1991
    Co-Authors: Magali Pla, Fernand Vedel, Chantal Mathieu, J. Durand
    Abstract:

    Abstract Several lines of Nicotiana sylvestris selected for oligomycin-resistance were also found to be resistant to venturicidin, another inhibitor of mitochondrial ATP synthase. One of the lines, previously shown to transmit oligomycin-resistance independently of nuclear and chloroplastic markers through protoplast fusion, was further analysed. Variations of mitochondrial DNA restriction patterns were observed, but the sequence of the mitochondrial gene atp9 , the putative site of mutation, was identical to normal Nicotiana tabacum sequence. In the progeny of one plant regenerated from protoplast fusion, oligomycin-resistance was irregularly transmitted via the pollen. Our conclusions are: no somaclonal variation of mitochondrial DNA affected the sequence of atp9 after protoplast and prolonged callus culture; oligomycin-resistance in this line cannot be related to a mutation in this mitochondrial gene, but is due to a nuclear modification.

David F. Hildebrand - One of the best experts on this subject based on the ideXlab platform.

  • Glycosidically bound volatile components of Nicotiana sylvestris and N. suaveolens flowers
    Phytochemistry, 1992
    Co-Authors: John H. Loughrin, Thomas R. Hamilton-kemp, Roger A. Andersen, Harold R. Burton, David F. Hildebrand
    Abstract:

    Abstract Glycosidically bound volatile components were extracted from flowers of Nicotiana sylvestris and N. suaveolens and isolated using liquid column chromatography with an Amberlite XAD-2 resin. These glycosides, which were putative precursors of fragrance compounds, were hydrolysed enzymatically and a number of the volatiles released were subsequently identified by GC-MS including compounds not previously identified in the floral headspace of these species. Average yields of volatiles released from glycosides were ca 230 and 1050 μg g −1 for N. sylvestris and N. suaveolens flowers, respectively. Compounds in the glycosidically bound fraction were all phenylpropanoid-derived volatiles (e.g. benzyl alcohol, benzaldehyde, E -cinnamyl alcohol, benzyl salicylate) with the exception of the monoterpene α-terpineol. No pronounced diurnal changes in the levels of glycosidically bound volatile compounds could be noted from flowers of either species. However, pronounced differences were obtained in the concentrations of glycosidically bound volatiles at different floral maturity stages.

  • Circadian rhythm of volatile emission from flowers of Nicotiana sylvestris and N. suaveolens
    Physiologia Plantarum, 1991
    Co-Authors: John H. Loughrin, Thomas R. Hamilton-kemp, Roger A. Andersen, David F. Hildebrand
    Abstract:

    The volatile profiles from flowers of Nicotiana sylvestris and N. suaveolens were investigated by means of dynamic headspace sampling and capillary gas chromatography. Under conditions of light/dark entrainment both species emitted phenylpropanoid-derived volatiles (e.g. benzyl alcohol, methyl benzoate) with maximum emission occurring during the dark period. Emission of these compounds was demonstrated to be circadian by continuance of rhythmicity under conditions of constant light and subsequent re-entrainment to a new light/dark cycle. In contrast, emission of the sesquiterpene hydrocarbon, caryophyllene, from N. sylvestris followed no apparent pattern. The emission of monoterpene hydrocarbons from flowers of N. suaveolens showed diurnal differences only under conditions of light/dark entrainment.

Mamoru Sugita - One of the best experts on this subject based on the ideXlab platform.

  • non aug translation initiation of mrna encoding plastid targeted phage type rna polymerase in Nicotiana sylvestris
    Biochemical and Biophysical Research Communications, 2002
    Co-Authors: Yuki Kobayashi, Yuko Dokiya, Yoshinori Kumazawa, Mamoru Sugita
    Abstract:

    A third nuclear gene encoding a bacteriophage T7-type RNA polymerase, NsRpoT-C, was isolated and characterized from Nicotiana sylvestris. The gene, NsRpoT-C, consists of 21 exons and 20 introns and encodes a polypeptide of 977 amino acid residues. The predicted NsRpoT-C protein shows the highest identity (72% amino acid identity) with Arabidopsis thaliana RpoT;3 which is a plastid-targeted protein. Surprisingly, comparison of the deduced amino acid sequence of NsRpoT-C with that of A. thaliana RpoT;3 predicted that the NsRpoT-C starts at a CUG triplet, a rare translation initiation codon. Transient expression assays in protoplasts from tobacco leaves demonstrated that the putative N-terminal transit peptide of NsRpoT-C encodes a targeting signal directing the protein into chloroplasts. This strongly suggests that NsRpoT-C functions as an RNA polymerase transcribing plastid-encoded genes. We have designated this protein NsRpoTp.

  • Organization and transcription of the gene family encoding chlorophyll a/b-binding proteins in Nicotiana sylvestris.
    Gene, 2002
    Co-Authors: Keiko Hasegawa, Mamoru Sugita, Yasushi Yukawa, Masahiro Sugiura
    Abstract:

    Nine Lhcb1 genes encoding the light-harvesting chlorophyll a/b-binding proteins of photosystem II were isolated and characterized from Nicotiana sylvestris. Their nucleotide sequences are highly similar. Lhcb1 transcripts are accumulated in leaves and stems but not in roots and non-green cultured cells. RNase protection assay revealed that no transcripts were detected from the gene, Lhcb1*2, in Nicotiana tabacum. This finding raises the possibility that the amphidiploid tobacco cultivar (N. tabacum) lost one gene from the female progenitor (N. sylvestris) during evolution. Transcriptional initiation sites were mapped and found to be mostly cytidine residues, which is unique to the N. sylvestris Lhcb1 genes. Four of the nine genes have single start sites and the remaining genes possess multiple initiation sites. The TATA-like sequences of nine Lhcb1 genes can be classified into two groups; one that possesses a TTTATA sequence and the other that has a sequence diverged from it. The genes with single initiation sites belong to the first group. A consensus motif for the initiation region is CTC*A (C* for initiation site), which differs from those of other plant genes or mammalian genes.

  • genomic organization and organ specific expression of a nuclear gene encoding phage type rna polymerase in Nicotiana sylvestris
    Gene, 2001
    Co-Authors: Yuki Kobayashi, Masahiro Sugiura, Yuko Dokiya, Yasuo Niwa, Mamoru Sugita
    Abstract:

    We have isolated and sequenced a nuclear gene and cDNA encoding bacteriophage T7-type single subunit RNA polymerase, NsRpoT-A, from Nicotiana sylvestris. NsRpoT-A consists of 19 exons and 18 introns; the first intron is 17 kb, the longest yet identified in a plant gene. Genomic Southern analysis indicated that N. sylvestris contains a small family of NsRpoT genes. Quantitative RT-PCR revealed that steady-state mRNA levels are highest in the leaves and lowest in the cotyledons. Phylogenetic analysis of NsRpoT-A and the RpoT proteins of other plant species suggested that NsRpoT-A is a mitochondrial protein. The TargetP program predicted localization of the NsRpoT-A gene product to the mitochondria. Using a transient expression assay and protoplasts from N. sylvestris mesophyll cells, we clearly demonstrated that the N-terminal sequence of NsRpoT-A targets the protein to the mitochondria. We therefore named this protein NsRpoTm.

William O. Dawson - One of the best experts on this subject based on the ideXlab platform.

  • Structure-function Relationship Between Tobacco Mosaic Virus Coat Protein and Hypersensitivity in Nicotiana sylvestris
    Journal of molecular biology, 1994
    Co-Authors: James N. Culver, Gerald Stubbs, William O. Dawson
    Abstract:

    Alterations in the structure of the tobacco mosaic virus (TMV) coat protein affect the elicitation of the N' gene hypersensitive response (HR) in Nicotiana sylvestris. To investigate this structure-function relationship, amino acid substitutions with predicted structural effects were created throughout the known structure of the TMV coat protein. Substitutions that resulted in the elicitation of the HR resided within and would predictably interfere with interface regions located between adjacent subunits in ordered aggregates of coat protein. Substitutions that did not result in the elicitation of the HR were either conservative or located outside these interface regions. In vitro analysis of coat protein aggregates demonstrated HR-eliciting coat proteins to have reduced aggregate stability in comparison with non-HR-eliciting coat proteins and a correlation existed between the strength of the elicited HR and the ability of a substitution to interfere with ordered aggregate formation. This finding corresponded with the predicted structural effects of HR-eliciting substitutions. Radical substitutions that predictably disrupted coat protein tertiary structure were found to prevent HR elicitation. These findings demonstrate that structural alterations that affect the stability of coat protein quaternary structure but not tertiary structure lead to host cell recognition and HR elicitation. A model for HR elicitation is proposed, in which disassembly of coat protein aggregates exposes a host "receptor" binding site.