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At least 73 records · Page 4Linked to original sources
PubMed2006

Arabidopsis SAMT1 defines a plastid transporter regulating plastid biogenesis and plant development.

S-Adenosylmethionine (SAM) is formed exclusively in the cytosol but plays a major role in plastids; SAM can either act as a methyl donor for the biogenesis of small molecules such as prenyllipids and macromolecules or as a regulator of the synthesis of aspartate-derived amino acids. Because the biosynthesis of SAM is restricted to the cytosol, plastids require a SAM importer. However, this transporter has not yet been identified. Here, we report the molecular and functional characterization of an Arabidopsis thaliana gene designated SAM TRANSPORTER1 (SAMT1), which encodes a plastid metabolite transporter required for the import of SAM from the cytosol. Recombinant SAMT1 produced in yeast cells, when reconstituted into liposomes, mediated the counter-exchange of SAM with SAM and with S-adenosylhomocysteine, the by-product and inhibitor of transmethylation reactions using SAM. Insertional mutation in SAMT1 and virus-induced gene silencing of SAMT1 in Nicotiana benthamiana caused severe growth retardation in mutant plants. Impaired function of SAMT1 led to decreased accumulation of prenyllipids and mainly affected the chlorophyll pathway. Biochemical analysis suggests that the latter effect represents one prominent example of the multiple events triggered by undermethylation, when there is decreased SAM flux into plastids.

Anion Transport Proteins↗
PubMed1984

DNA sequence requirements for the accurate transcription of a protein-coding plastid gene in a plastid in vitro system from mustard (Sinapis alba L.).

A nuclease-treated plastid extract from mustard (Sinapis alba L.) allows efficient transcription of cloned plastid DNA templates. In this in vitro system, the major runoff transcript of the truncated gene for the 32 000 mol. wt. photosystem II protein was accurately initiated from a site close to or identical with the in vivo start site. By using plasmids with deletions in the 5'-flanking region of this gene as templates, a DNA region required for efficient and selective initiation was detected 28-35 nucleotides upstream of the transcription start site. This region contains the sequence element TTGACA, which matches the consensus sequence for prokaryotic ;-35' promoter elements. In the absence of this region, a region 13-27 nucleotides upstream of the start site still enables a basic level of specific transcription. This second region contains the sequence element TATATAA, which matches the consensus sequence for the ;TATA' box of genes transcribed by RNA polymerase II (or B). The region between the ;TATA'-like element and the transcription start site is not sufficient but may be required for specific transcription of the plastid gene. This latter region contains the sequence element TATACT, which resembles the prokaryotic ;-10' (Pribnow) box. Based on the structural and transcriptional features of the 5' upstream region, a ;promoter switch' mechanism is proposed, which may account for the developmentally regulated expression of this plastid gene.

Journal Article↗
PubMed1993

SecA is plastid-encoded in a red alga: implications for the evolution of plastid genomes and the thylakoid protein import apparatus.

Partial sequence analysis of the plastid DNA (ptDNA) from a red alga, Antithamnion sp., revealed the presence of a homologue to the Escherichia coli secA gene as well as two open reading frames (ORF 510, ORF 179). In addition a sec Y homologue has been detected on the plastid genome by heterologous hybridization. None of these genes has been found in completely sequenced chlorophytic plastid genomes. SecA and secY gene copies were also detected in the ptDNA of a chromophytic alga, indicating that secA Y may be ubiquitous in rhodophytes and chromophytes. The significance of these findings for the evolution of plastid genomes and the thylakoid protein import mechanism is discussed.

Adenosine Triphosphatases↗
PubMed1970

In vitro protein synthesis by plastids of Phaseolus vulgaris. IV. Amino acid incorporation by etioplasts and effect of illumination of leaves on incorporation by plastids.

Protein synthesis in vitro by etioplasts and chloroplasts from Phaseolus vulgaris was examined to study the factors regulating the development of etioplasts into chloroplasts. The properties of incorporation of (14)C-leucine into protein by etioplasts from plants grown 6.5 days in darkness are similar to those of chloroplasts from plants of the same age that were illuminated for 12 hours. However, the rate of incorporation per plastid by chloroplasts is 4 times higher than the rate of amino acid incorporation by etioplasts. When 6-day-old plants are placed in light, this 4-fold increase occurs within 6 hours and is maintained up to 36 hours. The difference in rate of amino acid incorporation into protein between etioplasts and chloroplasts represents a real difference in the ability of etioplasts and chloroplasts to synthesize protein. A difference in pool size of leucine between etioplasts and chloroplasts does not account for the difference in amino acid incorporation between etioplasts and chloroplasts. Also the difference in photosynthetic capabilities of etioplasts and chloroplasts does not account for the difference in the ability to incorporate amino acid into protein. Furthermore, there are no factors in homogenates of etiolated leaves which inactivate amino acid incorporation into protein by chloroplasts. The difference in rates of amino acid incorporation between etioplasts and chloroplasts is correlated with the state of development of the plastids. The plastids have increased ability to incorporate amino acid into protein when the plastids are undergoing growth and differentiation.

Amino Acids↗
PubMed1987

Plastid run-on transcription. Application to determine the transcriptional regulation of spinach plastid genes.

We have developed a spinach plastid run-on transcription system to determine the extent of transcriptional regulation of chloroplast genes during morphogenetic changes of the organelle. In contrast to transcription in a spinach chloroplast extract, which requires initiation of exogenously added genes (Gruissem, W., Greenberg, B. M., Zurawski, G., and Hallick, R. B. (1983) Cell 35, 815-828), RNA synthesis in the run-on system is not affected by heparin or different salt concentrations. Transcription is asymmetric, and the size of the run-on transcripts varies between 75 nucleotides and 8 kilobases. Quantitative filter hybridization studies included gene-specific probes for the ribosomal RNA genes and nine protein-coding genes. Based on the amounts of hybridizable run-on transcripts, these genes can be ordered according to their respective transcriptional activities. The relative transcriptional activities of psbA, rbcL, and atpB in the run-on assay correlate closely with their reported promoter strengths in vitro. The plastid run-on transcription assay has been applied to determine the transcriptional regulation of plastid genes. Hybridization of run-on transcripts to regions of the spinach chloroplast genome containing at least nine tRNA genes indicates that most or all loci are highly transcribed. No significant qualitative and quantitative differences are detected when run-on transcripts from plastids of etiolated and greening cotyledons are hybridized to total, restriction enzyme-digested chloroplast DNA, demonstrating limited transcriptional regulation during chloroplast development.

Chloroplasts↗
PubMed1997

Analysis of the cluster of ribosomal protein genes in the plastid genome of a unicellular red alga Cyanidioschyzon merolae: translocation of the str cluster as an early event in the rhodophyte-chromophyte lineage of plastid evolution.

The nucleotide sequence of a cluster of ribosomal protein genes in the plastid genome of a unicellular red alga, Cyanidioschyzon merolae, which has been supposed to be the most primitive alga, was determined. The phylogenetic tree inferred from the amino acid sequence of ribosomal proteins of two rhodophytes, a chromophyte, a glaucophyte, two chlorophytes (land plants), a cyanobacterium, and three eubacteria suggested a close relationship between the cyanobacterium Synechocystis PCC6803 and the plastids of various species in the kingdom Plantae, which is consistent with the hypothesis of the endosymbiotic origin of plastids. In this tree, the two species of rhodophytes were grouped with the chromophyte, and the glaucophyte was grouped with the chlorophytes. Analysis of the organization of the genes encoding the ribosomal proteins suggested that the translocation of the str cluster occurred early in the lineage of rhodophytes and chromophytes after these groups had been separated from chlorophytes and glaucophytes.

Cyanobacteria↗
PubMed2001

Plastids undifferentiated, a nuclear mutation that disrupts plastid differentiation in Zea mays L.

Photosynthetic development in any plant requires the intracellular co-ordination of chloroplast and nuclear gene expression programs. In this report, we investigate the role of a nuclear gene in photosynthetic development by examining C4 photosynthetic differentiation in a yellow mutant of maize (Zea mays L.). The plastids undifferentiated (pun) mutation disrupts plastid biogenesis in both bundle sheath and mesophyll cells, at an early developmental stage and in a light-independent manner. Chloroplast thylakoids are disrupted in the mutant and both membrane-associated and soluble chloroplast-encoded proteins accumulate at much reduced levels. The observed plastid morphology is consistent with a general defect in chloroplast biogenesis that is most likely exerted at the post-translational level. Despite aberrant chloroplast development, nuclear photosynthetic genes are expressed normally in pun mutants. Thus, neither functional chloroplasts nor the Pun gene product are required to establish nuclear photosynthetic gene expression patterns in maize.

Cell Nucleus↗
PubMed2000

Molecular evidence of a unique lipoamide dehydrogenase in plastids: analysis of plastidic lipoamide dehydrogenase from Arabidopsis thaliana.

Lipoamide dehydrogenase is a subunit of the alpha-ketoacid dehydrogenases and the glycine decarboxylase complex in mitochondria, and the pyruvate dehydrogenase complex in plastids. We report here the unexpected finding of two plastidic isoforms of lipoamide dehydrogenase from Arabidopsis thaliana that are different from the mitochondrial form of the enzyme. The cDNA clones were confirmed by sequence alignment analysis and their location verified by chloroplast import assay. They are single copy genes that appear to be expressed in parallel in different tissues with highest level in developing siliques. Phylogenetic analysis gives further exemplary evidence for the plastidic evolution derived from cyanobacteria.

Amino Acid Sequence↗
PubMed1994

Controlled expression of plastid transgenes in plants based on a nuclear DNA-encoded and plastid-targeted T7 RNA polymerase.

Phage T7 RNA polymerase has been used extensively in Escherichia coli for high-level expression of selected genes placed under the control of the phage T7 gene 10 promoter. We have constructed an analogous system for use in plastids of higher plants. A T7 RNA polymerase chimeric gene containing a cauliflower mosaic virus 35S promoter and a tobacco ribulose-bisphosphate carboxylase/oxygenase small-subunit chloroplast transit-peptide sequence was introduced into tobacco by nuclear transformation. Stable plastid transformation of tobacco expressing the T7 RNA polymerase activity with a T7 promoter/beta-glucuronidase (GUS) reporter gene construct resulted in expression of GUS mRNA and enzyme activity in all tissues examined. Expression of GUS activity was extremely high in mature leaves, moderate in young leaves and petals, and low in stems, roots, and developing seeds. Plastid transformation of wild-type tobacco with the same chimeric GUS gene resulted in undetectable levels of GUS mRNA and enzyme activity. Genetic crosses demonstrated that a silent T7/GUS reporter gene could be activated in the F1 generation by transmission of an active nuclear T7 RNA polymerase gene from the male parent.

Base Sequence↗
PubMed2006

Nucleus-encoded plastid sigma factor SIG3 transcribes specifically the psbN gene in plastids.

We have investigated the function of one of the six plastid sigma-like transcription factors, sigma 3 (SIG3), by analysing two different Arabidopsis T-DNA insertion lines having disrupted SIG3 genes. Hybridization of wild-type and sig3 plant RNA to a plastid specific microarray revealed a strong reduction of the plastid psbN mRNA. The microarray result has been confirmed by northern blot analysis. The SIG3-specific promoter region has been localized on the DNA by primer extension and mRNA capping experiments. Results suggest tight regulation of psbN gene expression by a SIG3-PEP holoenzyme. The psbN gene is localized on the opposite strand of the psbB operon, between the psbT and psbH genes, and the SIG3-dependent psbN transcription produces antisense RNA to the psbT-psbH intergenic region. We show that this antisense RNA is not limited to the intergenic region, i.e. it does not terminate at the end of the psbN gene but extends as antisense transcript to cover the whole psbT coding region. Thus, by specific transcription initiation at the psbN gene promoter, SIG3-PEP holoenzyme could also influence the expression of the psbB operon by producing psbT antisense RNA.

Arabidopsis↗
PubMed1999

Identification of two essential sequence elements in the nonconsensus type II PatpB-290 plastid promoter by using plastid transcription extracts from cultured tobacco BY-2 cells.

In higher plants, plastid genes are transcribed by at least two types of DNA-dependent RNA polymerases. One of them is the well-known plastid-encoded prokaryotic type of polymerase that recognizes sigma(70)-type promoters consisting of -35 and -10 consensus elements. The other recently recognized RNA polymerase has been found to be encoded entirely in the nucleus, and it recognizes a completely different set of promoters, designated previously as nonconsensus type II (NCII) promoters. Here, we report the development of an in vitro transcription system using nonphotosynthetic plastids of cultured tobacco BY-2 cells. This system preferentially and accurately initiates transcription from NCII promoters. The conditions for in vitro transcription were optimized by using the tobacco PatpB-290 promoter, which has been found to be the most highly expressed NCII promoter in vivo. Analysis of in vitro transcription initiation in a series of PatpB-290 5' deletion constructs revealed that sequences upstream of nucleotide -41 do not influence the transcriptional activity of this promoter. A 43-bp region (nucleotides -35 to +8) was further analyzed by introducing single or multiple nucleotide substitutions into two regions (box I and box II) of high sequence conservation. We report here that the ATAGAA sequence comprising box II and the -11 to +4 region (relative to transcription initiation) in box I significantly influence the activity of this NCII promoter.

Base Sequence↗
PubMed2006

The function and diversity of plastid protein import pathways: a multilane GTPase highway into plastids.

The photosynthetic chloroplast is the hallmark organelle of green plants. During the endosymbiotic evolution of chloroplasts, the vast majority of genes from the original cyanobacterial endosymbiont were transferred to the host cell nucleus. Chloroplast biogenesis therefore requires the import of nucleus-encoded proteins from their site of synthesis in the cytosol. The majority of proteins are imported by the activity of Toc and Tic complexes located within the chloroplast envelope. In addition to chloroplasts, plants have evolved additional, non-photosynthetic plastid types that are essential components of all cells. Recent studies indicate that the biogenesis of various plastid types relies on distinct but homologous Toc-Tic import pathways that have specialized in the import of specific classes of substrates. These different import pathways appear to be necessary to balance the essential physiological role of plastids in cellular metabolism with the demands of cellular differentiation and plant development.

Forecasting↗
PubMed1993

Detection and characterization of a plastid envelope DNA-binding protein which may anchor plastid nucleoids.

Chloroplast DNA (cpDNA) binds to the envelope membrane of actively dividing chloroplasts (plastids) in young pea leaves. South-western blotting was used to identify and characterize the protein involved in the binding of cpDNA to the envelope membrane. A 130 kDa protein in the inner chloroplast (plastid) envelope membrane binds specific sequences within the cpDNA. These included a 0.41 kbp sequence located upstream of the psaAB gene, a 0.57 kbp sequence located downstream of the petA gene and a 1.2 kbp sequence located within the rpoC2 gene. The protein was detected in the envelope membrane of young pea leaves in which the cpDNA had been located by fluorescence microscopy at the chloroplast periphery, whereas it was undetectable in mature leaves. We therefore propose that the 130 kDa protein is involved in the binding of cpDNA to the envelope membrane, and named it plastid envelope DNA-binding protein.

Amino Acid Sequence↗
PubMed1999

In vivo analysis of plastid psbA, rbcL and rpl32 UTR elements by chloroplast transformation: tobacco plastid gene expression is controlled by modulation of transcript levels and translation efficiency

5' and 3' untranslated regions (UTRs) of plastid RNAs act as regulatory elements for post-transcriptional control of gene expression. Polyethylene glycol-mediated plastid transformation with UTR-GUS reporter gene fusions was used to study the function of the psbA, rbcL and rpl32 UTRs in vivo. All gene fusions were expressed from the same promoter, i.e. the promoter of the 16S-rRNA gene, such that variations in RNA and protein levels would be due to the involved UTR elements alone. Transgenic tobacco lines containing different combinations of UTRs showed fivefold variation in the uidA-mRNA level (RNA stability) and approximately 100-fold differences in GUS activity, a measure of translation activity. The rbcL 5'-UTR conferred greater mRNA stability than the psbA 5'-UTR on uidA transcripts. In contrast, the psbA 5'-UTR enhanced translation of GUS to a much greater extent compared to the rbcL 5'-UTR. The psbA 5'-UTR also mediated light-induced activation of translation which was not observed with other constructs. Deletion mutagenesis of an unanalysed terminal sequence element of the psbA 5'-UTR resulted in a twofold drop in uidA-mRNA level and a fourfold decrease in translation efficiency. Exchange of 3'-UTRs results in up to fivefold changes of mRNA levels and does not significantly influence translation efficiency. The mechanical impacts of these results on plastid translation regulation are discussed.

Journal Article↗
PubMed1987

Plastid Development in Pisum sativum Leaves during Greening : I. A Comparison of Plastid Polypeptide Composition and in Organello Translation Characteristics.

Changes in plastid polypeptide composition during greening of etiolated peas were investigated by two-dimensional gel electrophoresis. One hundred of the more than 250 polypeptides which could be detected upon silver staining were followed during plastid development. Thirty-nine polypeptides decreased in abundance on a per organelle basis. Twentythree of the 46 polypeptides which increased in abundance upon greening could be identified as proteins of the thylakoid membrane. The changes in proteins observed during greening of etiolated leaves corresponded largely to those observed during normal leaf expansion. The origin of some of the polypeptides was traced back by comparing the two-dimensional gels of plastid proteins with in organello translation products and with polypeptides which had been synthesized in vitro from poly(A(+)) mRNA preparations and posttranslationally imported by chloroplasts. Some polypeptides were specifically identified in two-dimensional gels by Western blot analysis.

Journal Article↗
PubMed2002

Plastid transcription in the holoparasitic plant genus Cuscuta: parallel loss of the rrn16 PEP-promoter and of the rpoA and rpoB genes coding for the plastid-encoded RNA polymerase.

The holoparasitic plant genus Cuscuta comprises a range of species whose plastid genomes have different degrees of reductions in their coding capacity. In this study, four Cuscuta species, Cuscuta reflexa, C. gronovii, C. subinclusa and C. odorata, that possess substantial physiological differences, were analysed with respect to the sequence and promoter structure of the rrn16 gene coding for the ribosomal 16S rRNA. Whereas the coding region of this gene is highly conserved among all four Cuscuta species, significant differences were observed in the non-coding region 5' of rrn16 with respect to both the length of the intergenic region between rrn16 and trnV and the promoters used to initiate transcription of the rrn16 gene. In the green species C. reflexa, rrn16 transcription starts from a functional plastid-encoded RNA polymerase (PEP) promoter that is missing in the other three species, C. gronovii, C. odorata and C. subinclusa. Instead, a 15-nucleotide-long conserved sequence immediately upstream of the mapped 5' ends bearing the nuclear-encoded RNA polymerase (NEP) promoter motif could be identified in these three species. The lack of a PEP promoter in these species coincides with the loss of two genes that encode subunits of PEP (rpoA and rpoB).

5' Flanking Region↗
PubMed1997

Targeted deletion of sprA from the tobacco plastid genome indicates that the encoded small RNA is not essential for pre-16S rRNA maturation in plastids.

The small plastid RNA (spRNA) which includes a segment that is complementary to the pre-16S rRNA has been suggested to facilitate maturation of pre-16S rRNA in tobacco. To investigate the function of spRNA, the gene encoding it (sprA) was removed from the plastid genome using targeted gene deletion. We report here that deletion of sprA does not significantly affect pre-16S rRNA maturation, nor does it cause any obvious phenotype. Although the spRNA still may be involved in rRNA maturation, it is non-essential under normal growth conditions.

Gene Deletion↗
PubMed2001

A plastid envelope location of Arabidopsis ent-kaurene oxidase links the plastid and endoplasmic reticulum steps of the gibberellin biosynthesis pathway.

We have used fusions of gibberellin biosynthesis enzymes to green fluorescent protein (GFP) to determine the subcellular localization of the early steps of the pathway. Gibberellin biosynthesis from geranylgeranyl diphosphate is catalysed by enzymes of the terpene cyclase, cytochrome P450 mono-oxygenase and 2-oxoglutarate-dependent dioxygenase classes. We show that the N-terminal pre-sequences of the Arabidopsis thaliana terpene cyclases copalyl diphosphate synthase (AtCPS1) and ent-kaurene synthase (AtKS1) direct GFP to chloroplasts in transient assays following microprojectile bombardment of tobacco leaves. The AtKS1-GFP fusion is also imported by isolated pea chloroplasts. The N-terminal portion of the cytochrome P450 protein ent-kaurene oxidase (AtKO1) directs GFP to chloroplasts in tobacco leaf transient assays. Chloroplast import assays with 35S-labelled AtKO1 protein show that it is targeted to the outer face of the chloroplast envelope. The leader sequences of the two ent-kaurenoic acid oxidases (AtKAO1 and AtKAO2) from Arabidopsis direct GFP to the endoplasmic reticulum. These data suggest that the AtKO1 protein links the plastid- and endoplasmic reticulum-located steps of the gibberellin biosynthesis pathway by association with the outer envelope of the plastid.

Arabidopsis↗