RESUMO
In addition to linear electron transport from water to NADP(+) , alternative electron transport pathways are believed to regulate photosynthesis. In the two routes of photosystem I (PSI) cyclic electron transport, electrons are recycled from the stromal reducing pool to plastoquinone (PQ), generating additional ΔpH (proton gradient across thylakoid membranes). Plastid terminal oxidase (PTOX) accepts electrons from PQ and transfers them to oxygen to produce water. Although both electron transport pathways share the PQ pool, it is unclear whether they interact in vivo. To investigate the physiological link between PSI cyclic electron transport-dependent PQ reduction and PTOX-dependent PQ oxidation, we characterized mutants defective in both functions. Impairment of PSI cyclic electron transport suppressed leaf variegation in the Arabidopsis immutans (im) mutant, which is defective in PTOX. The im variegation was more effectively suppressed in the pgr5 mutant, which is defective in the main pathway of PSI cyclic electron transport, than in the crr2-2 mutant, which is defective in the minor pathway. In contrast to this chloroplast development phenotype, the im defect alleviated the growth phenotype of the crr2-2 pgr5 double mutant. This was accompanied by partial suppression of stromal over-reduction and restricted linear electron transport. We discuss the function of the alternative electron transport pathways in both chloroplast development and photosynthesis in mature leaves.
Assuntos
Arabidopsis/fisiologia , Plastoquinona/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Transporte de Elétrons , Mutação , Oxirredução , Fotossíntese , Folhas de Planta/metabolismo , Folhas de Planta/fisiologiaRESUMO
The Arabidopsis thaliana kas3 mutant was isolated based on the hypersensitivity of PSII to low temperature using a Chl fluorescence imaging technique. Chl content was lower in kas3 seedlings cultured at 23 degrees C than in the wild type, but PSII activity was only mildly affected. However, after the chilling treatment at 4 degrees C for 7 d, PSII activity was severely impaired in kas3. PSII was more sensitive to light at 4 degrees C in the presence of lincomycin, suggesting that the kas3 mutation accelerates at least the PSII photodamage. The kas3 mutation causes an amino acid alteration in 3-ketoacyl-ACP synthase III (KasIII), leading to the partial loss of the de novo synthesis pathway for fatty acids in plastids. Consequently, the total fatty acid level was reduced to 75% of the wild-type level in kas3 at 23 degrees C and was further reduced to 60% at 4 degrees C. The composition of fatty acids was also slightly affected in kas3 at both 4 and 23 degrees C. Consistent with the results of the electron transport analysis, the chilling treatment also destabilized PsaA and cytochrome (Cyt) f and D1 in kas3. An analysis of double mutants with pgr1 conditionally defective in Cyt b(6)f activity and with var2 defective in FtsH protease suggested that the kas3 mutation has pleiotropic effects on chloroplast function, probably impacting both the Cyt b(6)f activity and translation in chloroplasts at 23 degrees C. The full activity of KasIII is required for the biogenesis of the intact electron transport machinery in thylakoid membranes and is especially important for the process of responding to low temperature.
Assuntos
Arabidopsis/fisiologia , Temperatura Baixa , Ácidos Graxos/biossíntese , Fotossíntese , 3-Oxoacil-(Proteína de Transporte de Acila) Sintase/genética , 3-Oxoacil-(Proteína de Transporte de Acila) Sintase/metabolismo , Sequência de Aminoácidos , Arabidopsis/enzimologia , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Clonagem Molecular , Transporte de Elétrons , Dados de Sequência Molecular , MutaçãoRESUMO
A transient in chlorophyll fluorescence after cessation of actinic light illumination, which has been ascribed to electron donation from stromal reductants to plastoquinone (PQ) by the NAD(P)H-dehydrogenase (NDH) complex, was investigated in Arabidopsis thaliana. The transient was absent in air in a mutant lacking the NDH complex (ndhM). However, in ndhM, the transient was detected in CO(2)-free air containing 2% O(2). To investigate the reason, ndhM was crossed with a pgr5 mutant impaired in ferredoxin (Fd)-dependent electron donation from NADPH to PQ, which is known to be redundant for NDH-dependent PQ reduction in the cyclic electron flow around photosystem I (PSI). In ndhM pgr5, the transient was absent even in CO(2)-free air with 2% O(2), demonstrating that the post-illumination transient can also be induced by the Fd- (or PGR5)-dependent PQ reduction. On the other hand, the transient increase in chlorophyll fluorescence was found to be enhanced in normal air in a mutant impaired in plastid fructose-1,6-bisphosphate aldolase (FBA) activity. The mutant, termed fba3-1, offers unique opportunities to examine the relative contribution of the two paths, i.e., the NDH- and Fd- (or PGR5)-dependent paths, on the PSI cyclic electron flow. Crossing fba3-1 with either ndhM or pgr5 and assessing the transient suggested that the main route for the PSI cyclic electron flow shifts from the NDH-dependent path to the Fd-dependent path in response to sink limitation of linear electron flow.
Assuntos
Arabidopsis/metabolismo , Arabidopsis/efeitos da radiação , Clorofila/metabolismo , Cloroplastos/metabolismo , Cloroplastos/efeitos da radiação , Fluorescência , Complexo de Proteína do Fotossistema I/metabolismo , Arabidopsis/enzimologia , Transporte de Elétrons/efeitos da radiação , Ferredoxinas/metabolismo , Frutose-Bifosfato Aldolase/antagonistas & inibidores , Frutose-Bifosfato Aldolase/metabolismo , Immunoblotting , Luz , Mutação/genética , NADPH Desidrogenase/metabolismo , Oxirredução/efeitos da radiação , Plastoquinona/metabolismoRESUMO
In flowering plants, RNA editing is a posttranscriptional process that converts specific C to U in organelle mRNAs. Nicotiana tabacum is an allotetraploid species derived from the progenitors of Nicotiana sylvestris and Nicotiana tomentosiformis. These Nicotiana species have been used as a model for understanding the mechanism and evolution of RNA editing in plastids. In Nicotiana species, the ndhD-1 site is edited to create the translational initiation codon of ndhD that encodes a subunit of the NAD(P)H dehydrogenease (NDH) complex. An analysis of this RNA editing revealed that editing efficiency in N. tomentosiformis is lower (15%) than that in N. tabacum (42%) and N. sylvestris (37%). However, this level of editing is sufficient for accumulating the NDH complex and its activity. The heterogous complementation of Arabidopsis crr4-3 mutant, in which RNA editing of ndhD-1 is completely impaired, with CRR4 orthologous genes derived from Nicotiana species suggested that the reduction in editing efficiency in N. tomentosiformis is caused by amino acid variations accumulating in CRR4.
Assuntos
Cloroplastos/genética , NADPH Desidrogenase/genética , Nicotiana/genética , Proteínas de Plantas/genética , Edição de RNA , Proteínas de Ligação a RNA/genética , Sequência de Aminoácidos , Cloroplastos/enzimologia , Genes de Plantas , Teste de Complementação Genética , Variação Genética , Dados de Sequência Molecular , NADPH Desidrogenase/metabolismo , Proteínas de Plantas/química , Proteínas de Ligação a RNA/química , Especificidade da Espécie , Nicotiana/enzimologiaRESUMO
PSI cyclic electron transport is essential for photosynthesis and photoprotection. In higher plants, the antimycin A-sensitive pathway is the main route of electrons in PSI cyclic electron transport. Although a small thylakoid protein, PGR5 (PROTON GRADIENT REGULATION 5), is essential for this pathway, its function is still unclear, and there are numerous debates on the rate of electron transport in vivo and its regulation. To assess how PGR5-dependent PSI cyclic electron transport is regulated in vivo, we characterized its activity in ruptured chloroplasts isolated from Arabidopsis thaliana. The activity of ferredoxin (Fd)-dependent plastoquinone (PQ) reduction in the dark is impaired in the pgr5 mutant. Alkalinization of the reaction medium enhanced the activity of Fd-dependent PQ reduction in the wild type. Even weak actinic light (AL) illumination also markedly activated PGR5-dependent PSI cyclic electron transport in ruptured chloroplasts. Even in the presence of linear electron transport [11 mumol O2 (mg Chl)(-1) h(-1)], PGR5-dependent PSI electron transport was detected as a difference in Chl fluorescence levels in ruptured chloroplasts. In the wild type, PGR5-dependent PSI cyclic electron transport competed with NADP+ photoreduction. These results suggest that the rate of PGR5-dependent PSI cyclic electron transport is high enough to balance the production ratio of ATP and NADPH during steady-state photosynthesis, consistently with the pgr5 mutant phenotype. Our results also suggest that the activity of PGR5-dependent PSI cyclic electron transport is regulated by the redox state of the NADPH pool.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Cloroplastos/metabolismo , Complexo de Proteínas do Centro de Reação Fotossintética/metabolismo , Complexo de Proteína do Fotossistema I/metabolismo , Cloroplastos/efeitos da radiação , Transporte de Elétrons/efeitos da radiação , Concentração de Íons de Hidrogênio/efeitos da radiação , Luz , NADP/metabolismo , Oxigênio/metabolismoRESUMO
The low-wave phenomenon, i.e., the transient drop of yield of modulated chlorophyll fluorescence shortly after application of a pulse of saturating light, was investigated in intact leaves of tobacco and Camellia by measuring fluorescence, CO(2) assimilation and absorption at 830 nm simultaneously. Limitations on linear electron flow, due to low electron acceptor levels that were induced by low CO(2), induced the low waves of chlorophyll fluorescence. Low-wave amplitudes obtained under different CO(2) concentrations and photon-flux densities yielded single-peak curves when plotted as functions of fluorescence parameters such as PhiPS II (quantum yield of Photosystem II) and qN (coefficient of non-photochemical quenching), suggesting that low-wave formation depends on the redox state of the electron transport chain. Low waves paralleled redox changes of P700, the reaction center of Photosystem I (PS I), and an additional electron flow through PS I was detected during the application of saturating pulses that induced low-waves. It is suggested that low waves of chlorophyll fluorescence are induced by increased non-photochemical quenching, as a result of the formation of a trans-thylakoid proton gradient due to cyclic electron flow around PS I.
RESUMO
CO2 uptake and chlorophyll fluorescence were measured under non-photorespiratory conditions in leaves from 14 plant species. The rate of CO2-dependent electron transport (JCO2) was calculated as four times rate of gross photosynthesis. The quantum yield of electron transport in photosystem II was estimated from the ratio delta F/Fm', where delta F is the difference between steady-state and maximal fluorescence in the light. As photon flux density (PFD) increased, JCO2 increased linearly first, and then reached saturation. The product (delta F/Fm')PFD, which is a function of electron transport rate, showed a similar response. Therefore, the relationship between (delta F/Fm') PFD versus JCO2 was proportional. However, under high light, a linear correlation was not always maintained. Factors affecting the linear correlation were analyzed by measuring CO2 uptake and chlorophyll fluorescence under illumination from either the upper (adaxial) or lower (abaxial) leaf surface, and by using plants with anatomically symmetric leaves having palisade tissues on both sides. Consequently, it was shown that the parameter delta F/Fm' is based on chlorophyll fluorescence emitted from chloroplasts present near the illuminated surface. Further, it was suggested that this restriction of the origin of fluorescence actually measured is significant in a leaf with high chlorophyll content, resulting in the deviation from linearity in the relationship between JCO2 and (delta F/Fm')PFD.
Assuntos
Dióxido de Carbono/metabolismo , Clorofila/metabolismo , Folhas de Planta/metabolismo , Transporte de Elétrons , Fluorescência , Cinética , Fotossíntese , Plantas/metabolismo , Especificidade da EspécieRESUMO
The mechanism of post-illumination chlorophyll fluorescence transient (PIFT) was investigated in Arabidopsis. PIFT was detected in the wild type after illumination with low light. In the fba3-2 (fructose-1,6-bisphosphate aldolase) mutant, in which PIFT is enhanced, strong light also induced PIFT. PIFT was suppressed not only in the triose phosphate/phosphate translocator (tpt-2) mutant, but also in tpt-2 fba3-2, suggesting that triose phosphates, such as dihydroxyacetone phosphate (DHAP), are involved in the PIFT mechanism. We concluded that PIFT is associated with ribulose-1,5-bisphosphate (RuBP)-regeneration limitation of photosynthesis in low light.
Assuntos
Arabidopsis/metabolismo , Arabidopsis/genética , Clorofila/genética , Clorofila/metabolismo , Fluorescência , Frutose/genética , Luz , Iluminação , Pentoses , Fosfatos , Estimulação Luminosa , Fotossíntese/genética , Ribulosefosfatos/metabolismoRESUMO
During steady-state photosynthesis in low-light, 830-nm absorption (A(830)) by leaves was close to that in darkness in Arabidopsis, indicating that the primary donor P700 in the reaction center of photosystem I (PSI) was in reduced form. However, P700 was not fully oxidized by a saturating light pulse, suggesting the presence of a population of PSI centers with reduced P700 that remains thermodynamically stable during the application of the saturating light pulse (i.e., reduced-inactive P700). To substantiate this, the effects of methyl viologen (MV) and far-red light on P700 oxidation by the saturating light pulse were analyzed, and the cumulative effects of repetitive application of the saturating light pulse on photosynthesis were analyzed using a mutant crr2-2 with impaired PSI cyclic electron flow. We concluded that the reduced-inactive P700 in low-light as revealed by saturating light pulse indicates limitations of electron flow at the PSI acceptor side.
Assuntos
Arabidopsis/metabolismo , Arabidopsis/efeitos da radiação , Clorofila/metabolismo , Luz , Fotossíntese/efeitos da radiação , Complexo de Proteína do Fotossistema I/metabolismo , Arabidopsis/efeitos dos fármacos , Elétrons , Oxirredução/efeitos dos fármacos , Oxirredução/efeitos da radiação , Oxigênio , Paraquat/farmacologia , Fotossíntese/efeitos dos fármacosRESUMO
Expression of miR398 is induced in response to copper deficiency and is involved in the degradation of mRNAs encoding copper/zinc superoxide dismutase in Arabidopsis thaliana. We found that SPL7 (for SQUAMOSA promoter binding protein-like7) is essential for this response of miR398. SPL7 is homologous to Copper response regulator1, the transcription factor that is required for switching between plastocyanin and cytochrome c(6) in response to copper deficiency in Chlamydomonas reinhardtii. SPL7 bound directly to GTAC motifs in the miR398 promoter in vitro, and these motifs were essential and sufficient for the response to copper deficiency in vivo. SPL7 is also required for the expression of multiple microRNAs, miR397, miR408, and miR857, involved in copper homeostasis and of genes encoding several copper transporters and a copper chaperone, indicating its central role in response to copper deficiency. Consistent with this idea, the growth of spl7 plants was severely impaired under low-copper conditions.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Cobre/metabolismo , Fatores de Transcrição/metabolismo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Regulação da Expressão Gênica de Plantas , MicroRNAs/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , Regiões Promotoras Genéticas , Ligação Proteica , RNA Mensageiro/metabolismo , RNA de Plantas/genética , Fatores de Transcrição/genética , Transcrição GênicaRESUMO
Major copper proteins in the cytoplasm of plant cells are plastocyanin, copper/zinc superoxide dismutase, and cytochrome c oxidase. Under copper limited conditions, expression of copper/zinc superoxide dismutase is down-regulated and the protein is replaced by iron superoxide dismutase in chloroplasts. We present evidence that a micro-RNA, miR398, mediates this regulation in Arabidopsis thaliana, by directing the degradation of copper/zinc superoxide dismutase mRNA when copper is limited. Sequence analysis indicated that the transcripts encoding cytosolic copper/zinc superoxide dismutase and COX5b-1, a subunit of the mitochondrial cytochrome c oxidase, are also targeted by miR398. This regulation via miR398 takes place in response to changes in a low range of copper levels (0.2-0.5 microM), indicating that miR398 is involved in a response to copper limitation. On the other hand, another major copper protein, plastocyanin, which is involved in photosynthetic electron flow and is essential in higher plants, was not regulated via miR398. We propose that miR398 is a key factor in copper homeostasis in plants and regulates the stability of mRNAs of major copper proteins under copper-limited conditions.
Assuntos
Arabidopsis/metabolismo , Cobre/metabolismo , Regulação da Expressão Gênica de Plantas/fisiologia , Homeostase/fisiologia , MicroRNAs/metabolismo , Estabilidade de RNA/fisiologia , RNA de Plantas/biossíntese , Arabidopsis/citologia , Arabidopsis/genética , Cloroplastos/genética , Cloroplastos/metabolismo , Cobre/farmacologia , Relação Dose-Resposta a Droga , Transporte de Elétrons/efeitos dos fármacos , Transporte de Elétrons/fisiologia , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Homeostase/efeitos dos fármacos , Ferro/metabolismo , Metaloproteínas/biossíntese , Metaloproteínas/genética , MicroRNAs/genética , Proteínas Mitocondriais/biossíntese , Proteínas Mitocondriais/genética , Proteínas de Plantas/biossíntese , Proteínas de Plantas/genética , Estabilidade de RNA/efeitos dos fármacos , RNA de Plantas/genéticaRESUMO
PSI cyclic electron transport contributes markedly to photosynthesis and photoprotection in flowering plants. Although the thylakoid protein PGR5 (Proton Gradient Regulation 5) has been shown to be essential for the main route of PSI cyclic electron transport, its exact function remains unclear. In transgenic Arabidopsis plants overaccumulating PGR5 in the thylakoid membrane, chloroplast development was delayed, especially in the cotyledons. Although photosynthetic electron transport was not affected during steady-state photosynthesis, a high level of non-photochemical quenching (NPQ) was transiently induced after a shift of light conditions. This phenotype was explained by elevated activity of PSI cyclic electron transport, which was monitored in an in vitro system using ruptured chloroplasts, and also in leaves. The effect of overaccumulation of PGR5 was specific to the antimycin A-sensitive pathway of PSI cyclic electron transport but not to the NAD(P)H dehydrogenase (NDH) pathway. We propose that a balanced PGR5 level is required for efficient regulation of the rate of antimycin A-sensitive PSI cyclic electron transport, although the rate of PSI cyclic electron transport is probably also regulated by other factors during steady-state photosynthesis.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Cloroplastos/metabolismo , Transporte de Elétrons/fisiologia , Complexo de Proteínas do Centro de Reação Fotossintética/metabolismo , Complexo de Proteína do Fotossistema I/metabolismo , Antimicina A/farmacologia , Arabidopsis/efeitos dos fármacos , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Cotilédone/metabolismo , Regulação da Expressão Gênica de Plantas , Luz , Oxirredução , Complexo de Proteínas do Centro de Reação Fotossintética/genética , Fatores de TempoRESUMO
The chloroplast NAD(P)H dehydrogenase (NDH) complex is involved in photosystem I cyclic electron transport and chlororespiration in higher plants. An Arabidopsis (Arabidopsis thaliana) chlororespiratory reduction 6 (crr6) mutant lacking NDH activity was identified by means of chlorophyll fluorescence imaging. Accumulation of the NDH complex was impaired in crr6. Physiological characterization of photosynthetic electron transport indicated the specific defect of the NDH complex in crr6. In contrast to the CRR7 protein that was recently identified as a potential novel subunit of the NDH complex by means of the same screening, the CRR6 protein was stable under the crr2 mutant background in which the NDH complex does not accumulate. The CRR6 gene (At2g47910) encodes a novel protein without any known motif. Although CRR6 does not have any transmembrane domains, it is localized in the thylakoid membrane fraction of the chloroplast. CRR6 is conserved in phototrophs, including cyanobacteria, from which the chloroplast NDH complex has evolutionally originated, but not in Chlamydomonas reinhardtii, in which the NDH complex is absent. We believe that CRR6 is a novel specific factor for the assembly or stabilization of the NDH complex.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimologia , Cloroplastos/enzimologia , NADPH Desidrogenase/metabolismo , Sequência de Aminoácidos , Proteínas de Arabidopsis/química , Sequência de Bases , Clonagem Molecular , Primers do DNA , Fluorescência , Dados de Sequência Molecular , Homologia de Sequência de AminoácidosRESUMO
The NAD(P)H dehydrogenase (NDH) complex in chloroplasts mediates photosystem I cyclic and chlororespiratory electron transport. Eleven chloroplast genes and three nuclear genes have been identified as encoding Ndh subunits, but the entire subunit composition is still unknown. An Arabidopsis (Arabidopsis thaliana) chlororespiratory reduction (crr3) mutant was isolated based on its lack of transient increase in chlorophyll fluorescence after actinic light illumination; this was due to a specific defect in accumulation of the NDH complex. The CRR3 gene (At2g01590) encodes a novel protein containing a putative plastid-targeting signal and a transmembrane domain. Consistent with the gene structure, CRR3 localized to the membrane fraction of chloroplasts. In addition to the essential function of CRR3 in stabilizing the NDH complex, the NDH complex is also required for the accumulation of CRR3. These results suggest that CRR3 interacts with the NDH complex in the thylakoid membrane. In contrast to other subunits in the chloroplast NDH complex, CRR3 is not conserved in cyanobacteria from which the chloroplast NDH complex is believed to have originated. We propose that CRR3 is a subunit of the NDH complex, which is specific to the chloroplast.
Assuntos
Proteínas de Arabidopsis/fisiologia , Arabidopsis/metabolismo , Cloroplastos/metabolismo , Proteínas de Membrana/fisiologia , NADPH Desidrogenase/metabolismo , Sequência de Aminoácidos , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Proteínas de Membrana/química , Proteínas de Membrana/genética , Dados de Sequência Molecular , Mutação , Alinhamento de Sequência , Tilacoides/metabolismoRESUMO
Although photosystem I (PSI) cyclic electron transport is essential for plants, our knowledge of the route taken by electrons is very limited. To assess whether ferredoxin (Fd) donates electrons directly to plastoquinone (PQ) or via a Q-cycle in the cytochrome (cyt) b(6)f complex in PSI cyclic electron transport, we characterized the activity of PSI cyclic electron transport in an Arabidopsis mutant, pgr1 (proton gradient regulation). In pgr1, Q-cycle activity was hypersensitive to acidification of the thylakoid lumen because of an amino acid alteration in the Rieske subunit of the cyt b(6)f complex, resulting in a conditional defect in Q-cycle activity. In vitro assays using ruptured chloroplasts did not show any difference in the activity of PGR5-dependent PQ reduction by Fd, which functions in PSI cyclic electron transport in vivo. In contrast to the pgr5 defect, the pgr1 defect did not show any synergistic effect on the quantum yield of photosystem II in crr2-2, a mutant in which NDH (NAD(P)H dehydrogenase) activity was impaired. Furthermore, the simultaneous determination of the quantum yields of both photosystems indicated that the ratio of linear and PSI cyclic electron transport was not significantly affected in pgr1. All the results indicated that the pgr1 mutation did not affect PGR5-dependent PQ reduction by Fd. The phenotypic differences between pgr1 and pgr5 indicate that maintenance of the proper balance of linear and PSI cyclic electron transport is essential for preventing over-reduction of the stroma.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Complexo Citocromos b6f/genética , Proteínas Ferro-Enxofre/metabolismo , Complexo de Proteínas do Centro de Reação Fotossintética/metabolismo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Clorofila/metabolismo , Complexo Citocromos b6f/metabolismo , Transporte de Elétrons , Proteínas Ferro-Enxofre/genética , Cinética , Modelos Biológicos , Complexo de Proteínas do Centro de Reação Fotossintética/genéticaRESUMO
An Arabidopsis thaliana mutant, crr7 (chlororespiratory reduction), was isolated using chlorophyll fluorescence imaging to detect reduced activity in NAD(P)H dehydrogenase (NDH). The chloroplast NDH complex is considered to have originated from cyanobacteria in which the NDH complex is involved in respiration, photosystem I (PSI) cyclic electron transport and CO2 uptake. In higher plants the NDH complex functions in PSI cyclic electron transport within the chloroplast. Despite exhaustive biochemical approaches, the entire subunit composition of the NDH complex is unclear in both cyanobacteria and chloroplasts. In crr7 accumulation of the NDH complex was specifically impaired. In vivo analysis of electron transport supported the specific loss of the NDH complex in crr7. CRR7 (At5g39210) encodes a protein of 156 amino acids, including a putative plastid target signal, and does not contain any known motifs. In contrast to CRR2 and CRR4, involved in the expression of chloroplast ndh genes, CRR7 is conserved in cyanobacterial genomes. Although CRR7 did not contain any transmembrane domains, it localized to the membrane fraction of the chloroplast. CRR7 was unstable in the crr2-2 mutant background, in which the expression of ndhB was impaired. These results strongly suggest that CRR7 is a novel subunit of the chloroplast NDH complex.
Assuntos
Proteínas de Arabidopsis/fisiologia , Arabidopsis/enzimologia , Cloroplastos/enzimologia , Proteínas de Membrana/fisiologia , NADPH Desidrogenase/metabolismo , Sequência de Aminoácidos , Arabidopsis/genética , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/metabolismo , Clonagem Molecular , Sequência Conservada , Transporte de Elétrons/fisiologia , Estabilidade Enzimática , Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Dados de Sequência Molecular , Estrutura Terciária de Proteína , Proteínas Recombinantes de Fusão/metabolismo , Alinhamento de Sequência , Homologia de Sequência de AminoácidosRESUMO
Using thylakoid membranes, we previously demonstrated that accumulated electrons in the photosynthetic electron transport system induces the electron flow from the acceptor side of PSII to its donor side only in the presence of a pH gradient ((Delta)pH) across the thylakoid membranes. This electron flow has been referred to as cyclic electron flow within PSII (CEF-PSII) [Miyake and Yokota (2001) Plant Cell Physiol. 42: 508]. In the present study, we examined whether CEF-PSII operates in isolated intact chloroplasts from spinach leaves, by correlating the quantum yield of PSII [Phi(PSII)] with the activity of the linear electron flow [V(O(2))]. The addition of the protonophore nigericin to the intact chloroplasts decreased Phi(PSII), but increased V(O(2)), and relative electron flux in PSII [Phi(PSII) x PFD] and V(O(2)) were proportional to one another. Phi(PSII) x PFD at a given V(O(2)) was much higher in the presence of (Delta)pH than that in its absence. These effects of nigericin on the relationship between Phi(PSII) x PFD and V(O(2)) are consistent with those previously observed in thylakoid membranes, indicating the occurrence of CEF-PSII also in intact chloroplasts. In the presence of (Delta)pH, CEF-PSII accounted for the excess electron flux in PSII that could not be attributed to photosynthetic linear electron flow. The activity of CEF-PSII increased with increased light intensity and almost corresponded to that of the water-water cycle (WWC), implying that CEF-PSII can dissipate excess photon energy in cooperation with WWC to protect PSII from photoinhibition under limited photosynthesis conditions.
Assuntos
Cloroplastos/metabolismo , Oxigênio/metabolismo , Complexo de Proteínas do Centro de Reação Fotossintética/metabolismo , Folhas de Planta/metabolismo , Spinacia oleracea/metabolismo , Clorofila/metabolismo , Cloroplastos/efeitos dos fármacos , Cloroplastos/efeitos da radiação , Transporte de Elétrons/efeitos dos fármacos , Transporte de Elétrons/efeitos da radiação , Concentração de Íons de Hidrogênio , Ionóforos/farmacologia , Luz , Complexos de Proteínas Captadores de Luz , Nigericina/farmacologia , Fotossíntese/efeitos dos fármacos , Fotossíntese/fisiologia , Fotossíntese/efeitos da radiação , Complexo de Proteínas do Centro de Reação Fotossintética/efeitos dos fármacos , Complexo de Proteínas do Centro de Reação Fotossintética/efeitos da radiação , Folhas de Planta/efeitos dos fármacos , Folhas de Planta/efeitos da radiação , Spinacia oleracea/efeitos dos fármacos , Spinacia oleracea/efeitos da radiação , Água/farmacologia , Água/fisiologiaRESUMO
The rice mutants M249 and M134 accumulate chlorophyllides a and b which are esterified with incompletely reduced alcohols such as geranylgeraniol, dihydrogeranylgeraniol, and tetrahydrogeranylgeraniol. Quantities of alpha-tocopherol, phylloquinone, and menaquinones in leaves of these mutants were determined by high performance liquid chromatography (HPLC) with a fluorescence detector after post-column chemical reduction to convert quinones to fluorescent quinols. Methylnaphthoquinones, varying in the reduction state of the side chain (menaquinones), were detected in leaf segments of the rice mutants on HPLC analyses with both high selectivity and sensitivity to plant quinones. Mutant M249 preferentially accumulated menaquinone, which contains tetrahydrogeranylgeraniol as its side chain. However, mutant M134 exhibited preferential accumulation of menaquinone with a geranylgeraniol side chain. In both mutants, the accumulation patterns of menaquinones with different prenyl side chains were similar to those of chlorophyll with the corresponding prenyl side chains. The content of P700, the photosystem I primary electron donor, in the wild type was greater than that of either mutant, on both a chlorophyll and a fresh weight basis. However, the ratios of total methylnaphthoquinones to P700 were similar in both the wild type and the mutants. Since no comparative large differences in photosynthetic activity exist between the wild type and the mutants, these results suggest that the hydrogenation of the methylnaphthoquinone side chain to phytol is not an essential requirement for it to function as an electron acceptor in photosystem I. On the other hand, alpha-tocopherol was detected in fully developed leaves of the wild type, but not in those of the mutants. Accumulation of menaquinones and the loss of alpha-tocopherol in mutant leaves suggest that the reduction of chlorophyll-geranylgeraniol to phytol and that of geranylgeranyl pyrophosphate to phytyl pyrophosphate are catalysed by the same enzyme.