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T Pfannschmidt

Publications and source records attributed to T Pfannschmidt.

11 recordsLinked to original sources
PubMed2001

A novel mechanism of nuclear photosynthesis gene regulation by redox signals from the chloroplast during photosystem stoichiometry adjustment.

Photosynthetic organisms acclimate to long term changes in the environmental light quality by an adjustment of their photosystem stoichiometry to maintain photosynthetic efficiency. By using light sources that predominantly excite either photosystem I (PSI) or photosystem II (PSII), we studied the effects of excitation imbalances between both photosystems on nuclear PSI gene transcription in transgenic tobacco seedlings with promoter::beta-glucuronidase gene fusions. Shifts from PSI to PSII light sources (and vice versa) induced changes in the reduction/oxidation state of intersystem redox components, and acclimation of tobacco seedlings to such changes were monitored by changes in chlorophyll a/b ratios and in vivo chlorophyll a fluorescence. The ferredoxin-NADP(+)-oxidoreductase gene promoter did not respond to these treatments, those from the genes for subunits PsaD and PsaF of PSI are activated by a reduction signal, and the plastocyanin promoter responded to both reduction and oxidation signals. Additional experiments with photosynthetic electron transport inhibitors 3-(3',4'-dichlorophenyl)-1,1'-dimethyl urea and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone demonstrated that the redox state of the plastoquinone pool controls the activity of the plastocyanin promoter, whereas subunit PsaD and PsaF gene transcription is regulated by other photosynthesis-derived signals. Thus, the expression of nuclear-encoded PSI genes is controlled by diverse light quality-dependent redox signals from the plastids during photosystem stoichiometry adjustment.

Binding Sites↗
PubMed2001

A repressor with similarities to prokaryotic and eukaryotic DNA helicases controls the assembly of the CAAT box binding complex at a photosynthesis gene promoter.

A single nucleotide exchange in a promoter region located immediately upstream of the CAAT box of the spinach photosynthesis gene AtpC (gene product is subunit gamma of the chloroplast ATP synthase) prevents the formation of a secondary structure and causes an unregulated, constitutive high level of expression (Kusnetsov, V., Landsberger, M., Oelmüller, R. (1999) J. Biol. Chem. 274, 36009-36014). We have isolated cDNAs for ATPC-2, a new polypeptide with homologies to pro- and eukaryotic helicases, which specifically binds to this promoter region. Binding of ATPC-2 competes strongly with that of a CAAT box binding factor (CBF), consistent with the idea that both complexes cannot be formed simultaneously because of sterical reasons. In gel mobility shift assays, high binding activities of ATPC-2 and low binding activities of CBF were observed with nuclear extracts from tissue with low AtpC expression levels, and the opposite was observed with extracts from tissues with high AtpC expression levels. Binding of ATPC-2 to the mutant sequence, which directs a constitutively high level expression in vivo and prevents the formation of a secondary structure in vitro, is significantly weaker than binding to the wild-type sequence. Again, the opposite results were obtained for the CBF. Thus, we conclude that the assembly of the CBF.DNA complex stimulates transcription of AtpC and that CBF binding is prevented if ATPC-2 is bound to the promoter region. The novel mechanism of gene regulation and the role of the helicase-like protein ATPC-2 as a potential transcriptional repressor is discussed in relation to its modular structure.

Amino Acid Sequence↗
PubMed2000

Balancing the two photosystems: photosynthetic electron transfer governs transcription of reaction centre genes in chloroplasts.

Chloroplasts are cytoplasmic organelles whose primary function is photosynthesis, but which also contain small, specialized and quasi-autonomous genetic systems. In photosynthesis, two energy converting photosystems are connected, electrochemically, in series. The connecting electron carriers are oxidized by photosystem I (PS I) and reduced by photosystem II (PS II). It has recently been shown that the oxidation reduction state of one connecting electron carrier, plastoquinone, controls transcription of chloroplast genes for reaction centre proteins of the two photosystems. The control counteracts the imbalance in electron transport that causes it: oxidized plastoquinone induces PS II and represses PS I; reduced plastoquinone induces PS I and represses PS II. This complementarity is observed both in vivo, using light favouring one or other photosystem, and in vitro, when site-specific electron transport inhibitors are added to transcriptionally and photosynthetically active chloroplasts. There is thus a transcriptional level of control that has a regulatory function similar to that of purely post-translational 'state transitions' in which the redistribution of absorbed excitation energy between photosystems is mediated by thylakoid membrane protein phosphorylation. The changes in rates of transcription that are induced by spectral changes in vivo can be detected even before the corresponding state transitions are complete, suggesting the operation of a branched pathway of redox signal transduction. These findings suggest a mechanism for adjustment of photosystem stoichiometry in which initial events involve a sensor of the redox state of plastoquinone, and may thus be the same as the initial events of state transitions. Redox control of chloroplast transcription is also consistent with the proposal that a direct regulatory coupling between electron transport and gene expression determines the function and composition of the chloroplast's extra-nuclear genetic system.

Animals↗
PubMed2000

Transcripts and sequence elements suggest differential promoter usage within the ycf3-psaAB gene cluster on mustard (Sinapis alba L.) chloroplast DNA.

The mustard chloroplast DNA region spanning the ycf3 gene and part of the psaAB operon was investigated. The ycf3 gene reveals two class-II introns that are removed during processing to give a mature 0.7-kb transcript, but no RNA editing seems to be involved. RNase protection and RT-PCR experiments suggest cotranscription of ycf3 with the downstream psaA gene, possibly from a NEP promoter upstream of ycf3, whereas distinct ycf3 and psaA transcripts are each initiated from PEP promoters. This situation is reminiscent of that for the trnK-psbA gene region. The implications for light-regulated versus light-independent expression of photosystem core-protein genes are discussed.

Amino Acid Sequence↗
PubMed2000

The multisubunit chloroplast RNA polymerase A from mustard (Sinapis alba L.). Integration of a prokaryotic core into a larger complex with organelle-specific functions.

We previously identified two multisubunit plastid RNA polymerases termed A and B. The B enzyme has a bacterial-type polypeptide composition and is sensitive to the prokaryotic transcription inhibitor rifampicin (Rif); the A enzyme has a more complex subunit structure and is Rif-resistant. Here we report results of N-terminal sequencing and MS carried out with the A enzyme, which establish that the latter contains rpo gene products and is structurally related to the B enzyme. Furthermore, evidence is provided that the A enzyme can be converted into a Rif-sensitive enzyme form in a phosphorylation-dependent manner in vitro by a treatment that results in depletion of a beta-like subunit. Database searches using sequence information derived from additional polypeptides that are present in purified A preparations revealed sequence similarity with chloroplast proteins involved in RNA processing and redox control. This proteomics approach thus points to the complexity of the chloroplast transcription apparatus and its interconnections with post-transcriptional and signalling mechanisms.

Alkaline Phosphatase↗
PubMed2000

Photosynthetic electron flow regulates transcription of the psaB gene in pea (Pisum sativum L.) chloroplasts through the redox state of the plastoquinone pool.

Plants respond to changing light conditions by altering the stoichiometry between components of the photosynthetic electron transport chain of chloroplast thylakoids. We measured specific run-on transcription of the chloroplast genes psaB, psbA and rbcL in pea (Pisum sativum L.) seedlings grown under three different conditions of illumination: light selective for photosystem I (PSI-light); light selective for photosystem II (PSII-light); and a combination of PSI- and PSII-light (mixed light, ML). The transcriptional rate of the psaB gene increased under PSII-light and decreased under PSI-light, while the transcriptional rates of the psbA and rbcL genes were affected only in a non-specific way. Similar effects also occurred in plants grown under ML and switched to either PSI- or PSII-light for 4 h. Addition of the inhibitors of photosynthetic electron transport 3-(3,4 dichlorophenyl)-1,1-dimethylurea (DCMU) and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB) influenced psaB transcription in isolated, illuminated chloroplasts: DCMU addition resulted in oxidation of the plastoquinone pool and decreased transcription of psaB; DBMIB addition resulted in reduction of the plastoquinone pool and increased transcription of psaB. The experimental results obtained in vivo and in vitro provide evidence for coupling between the redox state of plastoquinone and the rate of transcription of the psaB gene in pea.

Chloroplasts↗
PubMed1999

PTK, the chloroplast RNA polymerase-associated protein kinase from mustard (Sinapis alba), mediates redox control of plastid in vitro transcription.

The major RNA polymerase from mustard chloroplasts is a multi-subunit enzyme consisting of core components and associated factors. Among the latter is a heterotrimeric factor named PTK (plastid transcription kinase) because of its serine/threonine-type protein kinase activity. PTK activity itself depends on its phosphorylation state. In addition, we show that it responds to glutathione but not to other redox-reactive reagents that were tested, and both glutathione and phosphorylation act antagonistically. Using a homologous in vitro system, we find that PTK selectively phosphorylates subunit(s) of plastid RNA polymerase and is involved in determining the level of faithful transcription from the chloroplast psbA promoter. Together, these results establish a role for phosphorylation and redox state in the regulation of plastid gene expression.

Chloroplasts↗
PubMed1999

Direct transcriptional control of the chloroplast genes psbA and psaAB adjusts photosynthesis to light energy distribution in plants.

Two photosystems, I and II, absorb and convert light energy in photosynthesis in chloroplasts of green plants. The genes psbA and psaAB of the cytoplasmic chloroplast genome encode core components of photosystem II and photosystem I, respectively. Here we show that the absolute amounts of photosystem I and photosystem II respond, in a complementary manner, to changes in light quality that preferentially excite each photosystem in mustard seedlings. We also show that the initial response to altered energy distribution is a change in the rates of transcription of psbA and psaAB. Changes in chlorophyll fluorescence emission in vivo suggest that the signal initiating this change is the oxidation-reduction state of plastoquinone, a component of the photosynthetic electron transport chain that connects photosystem I and photosystem II. The results are consistent with transcriptional effects observed previously with chloroplasts isolated in vitro and demonstrate that redox control of chloroplast transcription initiates long-term adjustments that compensate for imbalance in energy distribution and adapt the whole plant to altered light environments.

Chloroplasts↗
PubMed1997

The A and B forms of plastid DNA-dependent RNA polymerase from mustard (Sinapis alba L.) transcribe the same genes in a different developmental context.

Two RNA polymerases, termed A (cp-pol A) and B (cp-pol B), are known to be present in mustard plastids. In vitro, the two enzymes have different requirements for DNA binding, but both bind to, and transcribe from, the same set of chloroplast promoters. The B enzyme is sensitive to rifampicin (Rif), whereas the A enzyme is not. When seedlings were grown in the presence of Rif, RNA pool sizes of the photosynthesis-related plastid genes rbcL and psbA were smaller than in untreated controls, whereas transcripts of the non-photosynthetic genes rps16, trnG, rrn and rpoB remained virtually unaffected by the drug. The Rif inhibition patterns of rbcL and psbA transcripts reflect the relative abundance of the A and B enzymes at different stages and light/dark conditions. These genes can thus be transcribed by either of the two enzymes in vivo, whereas the non-photosynthetic genes are transcribed mostly or exclusively by the A enzyme, or by another Rif-resistant plastid polymerase. Among several nuclear gene transcripts that were tested for Rif inhibition, only those of the RbcS gene family for the plastid-bound small subunit of Rubisco revealed a decrease in pool size, which may imply that mechanisms exist that serve to coordinate patterns of gene expression in the different cellular compartments.

DNA, Plant↗
PubMed1994

Separation of two classes of plastid DNA-dependent RNA polymerases that are differentially expressed in mustard (Sinapis alba L.) seedlings.

Chloroplast and etioplast in vitro transcription systems from mustard have different functional properties, which is reflected in differences in phosphorylation status. Here we report another transcription control mechanism, which involves two plastid DNA-dependent RNA polymerases designated as peak A and peak B enzymes. Both are large multi-subunit complexes, but differ in their native molecular mass (> 700 kDa for peak A and ca. 420 kDa for peak B) and in their polypeptide composition. The A enzyme is composed of at least 13 polypeptides, while the B enzyme contains only four putative subunits. Peak B activity is inhibited by rifampicin, whereas that of peak A is resistant. RNA polymerase activity was compared for plastids from cotyledons of 4-day-old seedlings that were grown either under continuous light (chloroplasts) or in darkness (etioplasts), or were first dark-grown and then transferred to light for 16 h ('intermediate-type' plastids). While the total activity was approximately the same in all three cases, enzyme B was the predominant activity obtained from etioplasts and enzyme A that obtained from chloroplasts. Both had equal activity in preparations from the 'intermediate-type' plastid form. Both activation/inactivation and differential gene expression seem to play a role in the regulation of the plastid transcription machinery.

DNA-Directed RNA Polymerases↗