Reaction Center Core Research Articles

Elevated Inorganic Carbon Concentrating Mechanism Confers Tolerance to High Light in an Arctic Chlorella sp. ArM0029B.

Microalgae and higher plants employ an inorganic carbon (Ci) concentrating mechanism (CCM) to increase CO2 availability to Rubisco. Operation of the CCM should enhance the activity of the Calvin cycle, which could act as an electron sink for electrons generated by photosynthesis, and lower the redox status of photosynthetic electron transport chains. In this study, a hypothesis that microalgal cells with fully operating CCM are less likely to be photodamaged was tested by comparing a Chlorella mutant with its wild type (WT). The mutant acquired by screening gamma-ray-induced mutant libraries of Chlorella sp. ArM0029B exhibited constitutively active CCM (CAC) even in the presence of additional Ci sources under mixotrophic growth conditions. In comparison to the WT alga, the mutant named to constitutively active CCM1 (CAC1) showed more transcript levels for genes coding proteins related to CCM such as Ci transporters and carbonic anhydrases (CA), and greater levels of intracellular Ci content and CA activity regardless of whether growth is limited by light or not. Under photoinhibitory conditions, CAC1 mutant showed faster growth than WT cells with more PSII reaction center core component D1 protein (encoded by psbA), higher photochemical efficiency as estimated by the chlorophyll fluorescence parameter (Fv/Fm), and fewer reactive oxygen species (ROS). Interestingly, high light (HL)-induced increase in ROS contents in WT cells was significantly inhibited by bicarbonate supplementation. It is concluded that constitutive operation of CCM endows Chlorella cells with resistance to HL partly by reducing the endogenous generation of ROS. These results will provide useful information on the interaction between CCM expression, ROS production, and photodamage in Chlorella and related microalgae.

Characterization of thermotolerant phototrophic bacteria, Rhodoplanes tepidicaeni sp. nov. and Rhodoplanes azumiensis sp. nov., isolated from a geothermal spring.

Two strains of thermotolerant phototrophic alphaproteobacteria, designated strains TUT3542T and TUT3581T, were isolated from sediment mud and cyanobacterial mats in a geothermal spring in Japan, respectively, and taxonomically characterized. Both the strains were budding motile rods and were able to grow at 45 °C. Phototrophically grown cells of strains TUT3542T and TUT3581T produced pink and brownish red cultures, respectively, and showed in vivo absorption maxima at 800, 858-859 and 892-895 nm in the near infrared region, indicating the presence of a core reaction centre and peripheral pigment complexes with bacteriochlorophyll a. The intracytoplasmic membrane system was of the lamellar type parallel to the cytoplasmic membrane. 16S rRNA gene sequence comparisons showed that strains TUT3542T and TUT3581T had the highest similarity level to Rhodoplanes oryazae NBRC 109406T (99.6 %) and Rhodoplaneselegans AS130T (99.3 %), respectively. Genomic DNA-DNA reassociation studies revealed that strains TUT3542T and TUT3581T had hybridization levels of less than 62 and 56 % to the type strains of all established species of the genus Rhodoplanes, respectively. The G+C contents of genomic DNA were 67.7 mol% for strain TUT3542T and 70.4 mol% for strain TUT3581T. Results of phenotypic studies showed that the two novel strains could be differentiated from any of the previously described Rhodoplanes species. Thus, the author proposes the names Rhodoplanes tepidicaeni sp. nov. for strain TUT3542T and Rhodoplanes azumiensis sp. nov. for strain TUT3581T. The type strain of Rhodoplanes tepidicaeni is TUT3542T (=KCTC 15602T=NBRC 112815T) and the type strain of Rhodoplanes azumiensis is TUT3581T (=KCTC 15603T=NBRC 112816T).

The Alleviation of Heat Damage to Photosystem II and Enzymatic Antioxidants by Exogenous Spermidine in Tall Fescue.

Tall fescue (Festuca arundinacea Schreb) is a typical cool-season grass that is widely used in turf and pasture. However, high temperature as an abiotic stress seriously affects its utilization. The objective of this study was to explore the effect of spermidine (Spd) on heat stress response of tall fescue. The samples were exposed to 22°C (normal condition) or 44°C (heat stress) for 4 h. The results showed that exogenous Spd partially improved the quality of tall fescue leaves under normal temperature conditions. Nevertheless, after heat stress treatment, exogenous Spd significantly decreased the electrolyte leakage of tall fescue leaves. Spd also profoundly reduced the H2O2 and O2⋅- content and increased antioxidant enzymes activities. In addition, PAs can also regulate antioxidant enzymes activities including SOD, POD, and APX which could help to scavenge ROS. Moreover, application of Spd could also remarkably increase the chlorophyll content and had a positive effect on the chlorophyll α fluorescence transients under high temperature. The Spd reagent enhanced the performance of photosystem II (PSII) as observed by the JIP-test. Under heat stress, the Spd profoundly improved the partial potentials at the steps of energy bifurcations (PIABS and PItotal) and the quantum yields and efficiencies (φP0, δR0, φR0, and γRC). Exogenous Spd could also reduce the specific energy fluxes per QA- reducing PSII reaction center (RC) (TP0/RC and ET0/RC). Additionally, exogenous Spd improved the expression level of psbA and psbB, which encoded the proteins of PSII core reaction center complex. We infer that PAs can stabilize the structure of nucleic acids and protect RNA from the degradation of ribonuclease. In brief, our study indicates that exogenous Spd enhances the heat tolerance of tall fescue by maintaining cell membrane stability, increasing antioxidant enzymes activities, improving PSII, and relevant gene expression.

(Invited) Nanophotonic Structures for Solar Light Harvesting and the Generation of Hot Plasmonic Electrons

Future nanophotonic architectures for optical energy conversion and photocatalysis will require nanostructures with roles that work together cooperatively, such as structures with a large absorption cross section combined with features that enable guiding, propagating, and converting energy. Ultrafast optical responses in nanostructures are potentially important for modulating energy flow and also for characterizing and minimizing energy dissipation pathways in nanophotonic structures. Also, ultrafast extraction of hot plasmonic electrons from metal nanostructures is an important opportunity for increasing efficiency of solar energy conversion and photocatalysis. Efforts to greatly improve the efficiency of hot electron generation through plasmonic metamaterials, and characterization of hot electron dissipation pathways are described. I further describe efforts to induced long-range energy propagation in nanostructures. Nanostructured materials inspired by natural photosynthetic membranes represent an opportunity for efficient and directional energy transport. In natural photosystems, light harvesting complexes can transport excitons to the reaction center core with near unity conversion of absorbed photons to separated charge. A key to this process is the high rate of exciton hopping and the directionality of exciton flow due to an energy level waterfall effect of excitons in complexes as the reaction center core is approached. Efforts to induce similar behavior in biomimetically inspired nanostructures are described. Finally efforts to image exciton flow in light harvesting nanoscale materials are introduced. Use of the Center for Nanoscale Materials was supported by the U. S. Department of Energy, Office of Science, Office of Basic Energy Sciences, under Contract No. DE-AC02-06CH11357.

Novel insights into the origin and diversification of photosynthesis based on analyses of conserved indels in the core reaction center proteins.

The evolution and diversification of different types of photosynthetic reaction centers (RCs) remains an important unresolved problem. We report here novel sequence features of the core proteins from Type I RCs (RC-I) and Type II RCs (RC-II) whose analyses provide important insights into the evolution of the RCs. The sequence alignments of the RC-I core proteins contain two conserved inserts or deletions (indels), a 3 amino acid (aa) indel that is uniquely found in all RC-I homologs from Cyanobacteria (both PsaA and PsaB) and a 1 aa indel that is specifically shared by the Chlorobi and Acidobacteria homologs. Ancestral sequence reconstruction provides evidence that the RC-I core protein from Heliobacteriaceae (PshA), lacking these indels, is most closely related to the ancestral RC-I protein. Thus, the identified 3 aa and 1 aa indels in the RC-I protein sequences must have been deletions, which occurred, respectively, in an ancestor of the modern Cyanobacteria containing a homodimeric form of RC-I and in a common ancestor of the RC-I core protein from Chlorobi and Acidobacteria. We also report a conserved 1 aa indel in the RC-II protein sequences that is commonly shared by all homologs from Cyanobacteria but not found in the homologs from Chloroflexi, Proteobacteria and Gemmatimonadetes. Ancestral sequence reconstruction provides evidence that the RC-II subunits lacking this indel are more similar to the ancestral RC-II protein. The results of flexible structural alignments of the indel-containing region of the RC-II protein with the homologous region in the RC-I core protein, which shares structural similarity with the RC-II homologs, support the view that the 1 aa indel present in the RC-II homologs from Cyanobacteria is a deletion, which was not present in the ancestral form of the RC-II protein. Our analyses of the conserved indels found in the RC-I and RC-II proteins, thus, support the view that the earliest photosynthetic lineages with living descendants likely contained only a single RC (RC-I or RC-II), and the presence of both RC-I and RC-II in a linked state, as found in the modern Cyanobacteria, is a derivation from these earlier phototrophs.

The plastoquinol-plastoquinone exchange mechanism in photosystem II: insight from molecular dynamics simulations.

In the photosystem II (PSII) of oxygenic photosynthetic organisms, the reaction center (RC) core mediates the light-induced electron transfer leading to water splitting and production of reduced plastoquinone molecules. The reduction of plastoquinone to plastoquinol lowers PSII affinity for the latter and leads to its release. However, little is known about the role of protein dynamics in this process. Here, molecular dynamics simulations of the complete PSII complex embedded in a lipid bilayer have been used to investigate the plastoquinol release mechanism. A distinct dynamic behavior of PSII in the presence of plastoquinol is observed which, coupled to changes in charge distribution and electrostatic interactions, causes disruption of the interactions seen in the PSII-plastoquinone complex and leads to the "squeezing out" of plastoquinol from the binding pocket. Displacement of plastoquinol closes the second water channel, recently described in a 2.9Å resolution PSII structure (Guskov et al. in Nat Struct Mol Biol 16:334-342, 2009), allowing to rule out the proposed "alternating" mechanism of plastoquinol-plastoquinone exchange, while giving support to the "single-channel" one. The performed simulations indicated a pivotal role of D1-Ser264 in modulating the dynamics of the plastoquinone binding pocket and plastoquinol-plastoquinone exchange via its interaction with D1-His252 residue. The effects of the disruption of this hydrogen bond network on the PSII redox reactions were experimentally assessed in the D1 site-directed mutant Ser264Lys.

Carotenoid-to-Bacteriochlorophyll Energy Transfer in the LH1-RC Core Complex of a Bacteriochlorophyll b Containing Purple Photosynthetic Bacterium Blastochloris viridis.

Carotenoid-to-bacteriochlorophyll energy transfer has been widely investigated in bacteriochlorophyll (BChl) a-containing light harvesting complexes. Blastochloris viridis utilizes BChl b, whose absorption spectrum is more red-shifted than that of BChl a. This has implications on the efficiency and pathways of carotenoid-to-BChl energy transfer in this organism. The carotenoids that comprise the light-harvesting reaction center core complex (LH1-RC) of B. viridis are 1,2-dihydroneurosporene and 1,2-dihydrolycopene, which are derivatives of carotenoids found in the light harvesting complexes of several BChl a-containing purple photosynthetic bacteria. Steady-state and ultrafast time-resolved optical spectroscopic measurements were performed on the LH1-RC complex of B. viridis at room and cryogenic temperatures. The overall efficiency of carotenoid-to-bacteriochlorophyll energy transfer obtained from steady-state absorption and fluorescence measurements were determined to be ∼27% and ∼36% for 1,2-dihydroneurosporene and 1,2-dihydrolycopene, respectively. These results were combined with global fitting and target analyses of the transient absorption data to elucidate the energetic pathways by which the carotenoids decay and transfer excitation energy to BChl b. 1,2-Dihydrolycopene transfers energy to BChl b via the S2 → Qx channel with kET2 = (500 fs)(-1) while 1,2-dihydroneurosporene transfers energy via S1→ Qy (kET1 = (84 ps)(-1)) and S2 → Qx (kET2 = (2.2 ps)(-1)) channels.

The variability of light-harvesting complexes in aerobic anoxygenic phototrophs

Light-harvesting capacity was investigated in six species of aerobic anoxygenic phototrophic (AAP) bacteria using absorption spectroscopy, fluorescence emission spectroscopy, and pigment analyses. Aerobically grown AAP cells contained approx. 140-1800 photosynthetic reaction centers per cell, an order of magnitude less than purple non-sulfur bacteria grown semiaerobically. Three of the studied AAP species did not contain outer light-harvesting complexes, and the size of their reaction center core complexes (RC-LH1 core complexes) varied between 29 and 36 bacteriochlorophyll molecules. In AAP species containing accessory antennae, the size was frequently reduced, providing between 5 and 60 additional bacteriochlorophyll molecules. In Roseobacter litoralis, it was found that cells grown at a higher light intensity contained more reaction centers per cell, while the size of the light-harvesting complexes was reduced. The presented results document that AAP species have both the reduced number and size of light-harvesting complexes which is consistent with the auxiliary role of phototrophy in this bacterial group.

Dynamics of Energy and Electron Transfer in the FMO-Reaction Center Core Complex from the Phototrophic Green Sulfur Bacterium Chlorobaculum tepidum

The reaction center core (RCC) complex and the RCC with associated Fenna-Matthews-Olson protein (FMO-RCC) complex from the green sulfur bacterium Chlorobaculum tepidum were studied comparatively by steady-state and time-resolved fluorescence (TRF) and femtosecond time-resolved transient absorption (TA) spectroscopies. The energy transfer efficiency from the FMO to the RCC complex was calculated to be ∼40% based on the steady-state fluorescence. TRF showed that most of the FMO complexes (66%), regardless of the fact that they were physically attached to the RCC, were not able to transfer excitation energy to the reaction center. The TA spectra of the RCC complex showed a 30-38 ps lifetime component regardless of the excitation wavelengths, which is attributed to charge separation. Excitonic equilibration was shown in TA spectra of the RCC complex when excited into the BChl a Qx band at 590 nm and the Chl a Qy band at 670 nm, while excitation at 840 nm directly populated the low-energy excited state and equilibration within the excitonic BChl a manifold was not observed. The TA spectra for the FMO-RCC complex excited into the BChl a Qx band could be interpreted by a combination of the excited FMO protein and RCC complex. The FMO-RCC complex showed an additional fast kinetic component compared with the FMO protein and the RCC complex, which may be due to FMO-to-RCC energy transfer.

Characterization of the quinones in purple sulfur bacterium Thermochromatium tepidum

Quinone distributions in the thermophilic purple sulfur bacterium Thermochromatium tepidum have been investigated at different levels of the photosynthetic apparatus. Here we show that, on average, the intracytoplasmic membrane contains 18 ubiquinones (UQ) and 4 menaquinones (MQ) per reaction center (RC). About one-third of the quinones are retained in the light-harvesting–reaction center core complex (LH1–RC) with a similar ratio of UQ to MQ. The numbers of quinones essentially remains unchanged during crystallization of the LH1–RC. There are 1–2 UQ and 1 MQ associated with the RC-only complex in the purified solution sample. Our results suggest that a large proportion of the quinones are confined to the core complex and at least five UQs remain invisible in the current LH1–RC crystal structure.

Chemically modified reaction centers of photosystem II: Exchange of pheophytin a with 7-deformyl-7-hydroxymethyl-pheophytin b

The native pheophytin a (Pheo a) in isolated reaction centers of photosystem II (PSII RCs) has been chemically exchanged with extraneous 7-deformyl-7-hydroxymethyl-Pheo b (71-OH-Pheo b) which differs from Pheo a by the C-7 substituent (hydroxymethyl instead of methyl). The two pigments have similar reduction potentials in vitro [M. Meyer, Dissertation, Universität München, 1997], while their absorption spectra show small but distinct differences in the visible region. The resulting 71-OH-Pheo b-modified reaction center preparations were characterized by high-performance liquid chromatography, electronic absorption and light-induced Fourier transform infra red absorption difference spectroscopies, together with photoaccumulation of the reduced pheophytin electron acceptor and NaBH4-treatment. About 70% of the total Pheo a molecules are found to be replaced by 71-OH-Pheo b molecules in modified preparations, indicating that both the photochemically active (PheoD1) and inactive (PheoD2) binding sites were subjected to pigment exchange. The 71-OH-Pheo b molecule located at the PheoD1 site is able to functionally replace the native Pheo a, participating in primary charge separation as an electron acceptor. The Qx absorption band of this modified pheophytin molecule is localized at ~546nm; its Qy band is blue-shifted with respect to the absorption of other reaction center core pigments, being located at ~665nm. The Qy and Qx optical transitions of the 71-OH-Pheo b molecule exchanged into the PheoD2 site are identified at 677 and 543.5nm, respectively. The photochemically active double-modified PSII RCs additionally containing 7-deformyl-7-hydroxymethyl-131-deoxo-131-hydroxy-Pheo b at the PheoD2 site were obtained by treatment of the 71-OH-Pheo b-modified RCs with NaBH4.

Defining the Far-red Limit of Photosystem I: The Primary Charge Separation is Functional to 840 nm

The far-red limit of photosystem I (PS I) photochemistry was studied by EPR spectroscopy using laser flashes between 730 and 850 nm. In manganese-depleted spinach thylakoid membranes, the primary donor in PS I, P700, was oxidized simultaneously with tyrosine Z, the secondary donor in PS II. It was found that at 295 K PS I photochemistry, observed as P700 (+) formation, was functional up to 840 nm. This is 30 nm further to the red region than was reported for PS II photochemistry (Thapper, A., Mamedov, F., Mokvist, F., Hammarström, L., and Styring, S. (2009) Plant Cell 21, 2391-2401). The same far-red limit for the P700 (+) formation was observed in a PS I reaction center core preparation from Nostoc punctiforme. The reduction of the acceptor side of PS I, observed as reduction of the iron-sulfur centers FA and FB by low temperature EPR measurements, was also functional at 15 K with light up to >830 nm. Taken together, these results, obtained from both plants and cyanobacteria, most likely rule out involvement of the red-absorbing antenna chlorophylls in this reaction. Instead we propose the existence of weak charge transfer bands absorbing in the far-red region in the ensemble of excitonically coupled chlorophyll a molecules around P700 similar to what has been found in the reaction center of PS II. These charge transfer bands could be responsible for the far-red light absorption leading to PS I photochemistry at wavelengths up to 840 nm.

Exchange and complementation of genes coding for photosynthetic reaction center core subunits among purple bacteria.

A mutant of the phototrophic species belonging to the β-proteobacteria, Rubrivivax gelatinosus, lacking the photosynthetic growth ability was constructed by the removal of genes coding for the L, M, and cytochrome subunits of the photosynthetic reaction center complex. The L, M, and cytochrome genes derived from five other species of proteobacteria, Acidiphilium rubrum, Allochromatium vinosum, Blastochloris viridis, Pheospirillum molischianum, and Roseateles depolymerans, and the L and M subunits from two other species, Rhodobacter sphaeroides and Rhodopseudomonas palustris, respectively, have been introduced into this mutant. Introduction of the genes from three of these seven species, Rte. depolymerans, Ach. vinosum, and Psp. molischianum, restored the photosynthetic growth ability of the mutant of Rvi. gelatinosus, although the growth rates were 1.5, 9.4, and 10.7 times slower, respectively, than that of the parent strain. Flash-induced kinetic measurements for the intact cells of these three mutants showed that the photo-oxidized cytochrome c bound to the introduced reaction center complex could be rereduced by electron donor proteins of Rvi. gelatinosus with a t1/2 of less than 10 ms. The reaction center core subunits of photosynthetic proteobacteria appear to be exchangeable if the sequence identities of the LM core subunits between donor and acceptor species are high enough, i.e., 70% or more.

Structural analysis of the homodimeric reaction center complex from the photosynthetic green sulfur bacterium Chlorobaculum tepidum.

The reaction center (RC) complex of the green sulfur bacterium Chlorobaculum tepidum is composed of the Fenna–Matthews–Olson antenna protein (FMO) and the reaction center core (RCC) complex. The RCC complex has four subunits: PscA, PscB, PscC, and PscD. We studied the FMO/RCC complex by chemically cross-linking the purified sample followed by biochemical and spectroscopic analysis. Blue-native gels showed that there were two types of FMO/RCC complexes, which are consistent with complexes with one copy of FMO per RCC and two copies of FMO per RCC. Sodium dodecyl sulfate–polyacrylamide gel electrophoresis analysis of the samples after cross-linking showed that all five subunits of the RC can be linked by three different cross-linkers: bissulfosuccinimidyl suberate, disuccinimidyl suberate, and 3,3-dithiobis-sulfosuccinimidyl propionate. The interaction sites of the cross-linked complex were also studied using liquid chromatography coupled to tandem mass spectrometry. The results indicated that FMO, PscB, PscD, and part of PscA are exposed on the cytoplasmic side of the membrane. PscD helps stabilize FMO to the reaction center and may facilitate transfer of the electron from the RC to ferredoxin. The soluble domain of the heme-containing cytochrome subunit PscC and part of the core subunit PscA are located on the periplasmic side of the membrane. There is a close relationship between the periplasmic portions of PscA and PscC, which is needed for the efficient transfer of the electron between PscC and P840.

The architecture of Rhodobacter sphaeroides chromatophores

The chromatophores of Rhodobacter (Rb.) sphaeroides represent a minimal bio-energetic system, which efficiently converts light energy into usable chemical energy. Despite extensive studies, several issues pertaining to the morphology and molecular architecture of this elemental energy conversion system remain controversial or unknown. To tackle these issues, we combined electron microscope tomography, immuno-electron microscopy and atomic force microscopy. We found that the intracellular Rb. sphaeroides chromatophores form a continuous reticulum rather than existing as discrete vesicles. We also found that the cytochrome bc1 complex localizes to fragile chromatophore regions, which most likely constitute the tubular structures that interconnect the vesicles in the reticulum. In contrast, the peripheral light-harvesting complex 2 (LH2) is preferentially hexagonally packed within the convex vesicular regions of the membrane network. Based on these observations, we propose that the bc1 complexes are in the inter-vesicular regions and surrounded by reaction center (RC) core complexes, which in turn are bounded by arrays of peripheral antenna complexes. This arrangement affords rapid cycling of electrons between the core and bc1 complexes while maintaining efficient excitation energy transfer from LH2 domains to the RCs.

Primary radical ion pairs in photosystem II core complexes.

Ultrafast absorption spectroscopy with 20-fs resolution was applied to study primary charge separation in spinach photosystem II (PSII) reaction center (RC) and PSII core complex (RC complex with integral antenna) upon excitation at maximum wavelength 700-710 nm at 278 K. It was found that the initial charge separation between P680* and ChlD1 (Chl-670) takes place with a time constant of ~1 ps with the formation of the primary charge-separated state P680* with an admixture of: P680*((1-δ)) (P680(δ+)ChlD1(δ-)), where δ ~ 0.5. The subsequent electron transfer from P680(δ+)ChlD1(δ-) to pheophytin (Pheo) occurs within 13 ps and is accompanied by a relaxation of the absorption band at 670 nm (ChlD1(δ-)) and bleaching of the PheoD1 bands at 420, 545, and 680 nm with development of the Pheo(-) band at 460 nm. Further electron transfer to QA occurs within 250 ps in accordance with earlier data. The spectra of P680(+) and Pheo(-) formation include a bleaching band at 670 nm; this indicates that Chl-670 is an intermediate between P680 and Pheo. Stimulated emission kinetics at 685 nm demonstrate the existence of two decaying components with time constants of ~1 and ~13 ps due to the formation of P680(δ+)ChlD1(δ-) and P680(+)PheoD1(-), respectively.

Loss‐of‐function of OsSTN8 suppresses the photosystem II core protein phosphorylation and interferes with the photosystem II repair mechanism in rice (Oryza sativa)

STN8 kinase is involved in photosystem II (PSII) core protein phosphorylation (PCPP). To examine the role of PCPP in PSII repair during high light (HL) illumination, we characterized a T-DNA insertional knockout mutant of the rice (Oryza sativa) STN8 gene. In this osstn8 mutant, PCPP was significantly suppressed, and the grana were thin and elongated. Upon HL illumination, PSII was strongly inactivated in the mutants, but the D1 protein was degraded more slowly than in wild-type, and mobilization of the PSII supercomplexes from the grana to the stromal lamellae for repair was also suppressed. In addition, higher accumulation of reactive oxygen species and preferential oxidation of PSII reaction center core proteins in thylakoid membranes were observed in the mutants during HL illumination. Taken together, our current data show that the absence of STN8 is sufficient to abolish PCPP in osstn8 mutants and to produce all of the phenotypes observed in the double mutant of Arabidopsis, indicating the essential role of STN8-mediated PCPP in PSII repair.

Alleviation of heat damage to photosystem II by nitric oxide in tall fescue

Nitric oxide (NO) has been found to mediate plant responses to heat stress. The objective of this study was to investigate the protective role of NO in the recovery process of photosystem II (PSII) in tall fescue (Festuca arundinacea) against heat stress. Treatment of tall fescue leaves with NO donor sodium nitroprusside significantly improved the overall behavior of PSII probed by the chlorophyll a fluorescence transients, while the inhibition of NO accumulation by 2-phenyl-4,4,5,5-tetramethyl-imidazoline-1-oxyl-3-oxide (PTIO, a NO scavenger) plus N (G)-nitro-L-arginine-methyl ester (L-NAME, NO synthase inhibitor) dramatically disrupted the operation of PSII. Specifically, under heat stress, the exogenous NO reduced the initial fluorescence (F 0), increased the maximal quantum yield (F V/F M), and disappeared the K-step of 0.3ms. By the analysis of the JIP-test, the exogenous NO improved the quantum yield of the electron transport flux from Q A to Q B (ET0/ABS), and decreased the trapped excitation flux per reaction center (RC) (TR0/RC), electron transport flux per RC (ET0/RC), and electron flux reducing end electron acceptors per RC (RE0/RC). In addition, the exogenous NO reduced the content of H2O2, O 2 (•-) , and malondialdehyde and electrolyte leakage of tall fescue leaves. These data suggest that exogenous NO could protect plants, increase the amount of activated RC and improve the electron transport from oxygen evolving complex to D1 protein. Moreover, quantitative RT-PCR revealed that, in the presence of hydrogen peroxide, NO induced the gene expression of psbA, psbB, and psbC, which encode proteins belonging to subunits of PSII core reaction center (Psb) complex. These findings indicate that, as an important strategy to protect plants against heat stress, NO could improve the recovery process of PSII by the up regulation of the transcriptions of genes encoding PSII core proteins.

Evaluation of nuclear ribosomal RNA and chloroplast gene markers for the DNA taxonomy of centric diatoms

Abstract Diatoms are highly diversified microeukaryotes with a taxonomy that has been under perpetual revision, particularly by means of molecular analyses. In this study, we evaluated typical genes to find the best molecular marker for the DNA taxonomy of centric diatoms, including Chaetoceros Ehrenberg , Cyclotella Kutzing ex de Brebisson, Discostella Houk & Klee , Stephanodiscus Ehrenberg, and Thalassiosira Cleve. Our test genes included nuclear ribosomal RNA (e.g. small subunit, 5.8S, and large subunit [LSU]), and chloroplast genes (e.g. ribulose-1, 5-biphoshate carboxylase oxygenase and D1 protein of the photosystem II reaction centre core complex [ psb A]). Calculated genetic divergence was highest for LSU ribosomal RNA D1-D5 ( p -distance of 12.3), followed by 5.8S (7.7), ribulose-1, 5-biphoshate carboxylase oxygenase (7.4), small subunit (6.6), and psb A (3.7). The phylogenetic trees for individual genes effectively separated taxonomically tested centric diatoms with different phylogenetic resolutions; however, psb A was incongruent with others. These taxonomic resolving powers were in agreement with genetic divergences. Parsimony analysis showed that LSU evolved 1.97 times more rapidly than psb A and 1.07 times more rapidly than 5.8S. These results suggest that all of the tested genes except psb A can be used as taxonomic markers for centric diatoms and that LSU is the best molecular marker.

Photochemical trapping heterogeneity as a function of wavelength, in plant photosystem I (PSI–LHCI)

In the present paper the marked changes in photochemical trapping time over the absorption/fluorescence band of isolated PSI–LHCI are studied by means of time resolved fluorescence decay measurements. For emission at 680–690nm the effective trapping time is close to 17–18ps, and represents the effective trapping time from the bulk antenna. At wavelengths above 700nm the effective trapping time increases in a monotonic way, over the entire emission band, to attain values in the range of 70–80ps near 760nm. This is argued to be caused by “uphill” energy transfer from the low energy states to the core antenna and reaction centre. These data, together with the steady state emission spectrum, permit calculation of the overall trapping time for maize PSI–LHCI, which is estimated to be approximately 40ps. The wavelength dependence of the trapping time indicates, that in PSI–LHCI there exists at least one red form which emits at lower energies than the 735nm state. These data indicate that Photosystem I is about 55% diffusion limited.