24 research outputs found
Dynamic regulation of photosynthesis by chloroplast thioredoxin systems
Oxygenic photosynthesis is sunlight-energized conversion of CO2 into
carbohydrates using electrons extracted from water. It occurs in
cyanobacteria and in their endosymbiotic evolutionary descendants, the
chloroplasts of plants and algae, and enables the existence of most
ecosystems on Earth. Electron transfer from water to ferredoxin produces
NADPH and generates an electrochemical proton gradient across the
thylakoid membrane, which is utilized to power the ATP synthase. In the
stroma, the products of the light reactions are then used to assimilate CO2
into sugar phosphates in the Calvin–Benson cycle. In natural growth
conditions, plants experience fast and unpredictable fluctuations in light
intensity and other environmental factors. This has necessitated evolution
of intricate regulatory mechanisms to prevent damage to the photosynthetic
machinery and to avoid energy-expensive futile reactions. An important
way to control these mechanisms is through formation and cleavage of
disulfide bridges in chloroplast proteins by thioredoxins. Indeed, plant
chloroplasts contain a large variety of thioredoxin isoforms, as well as two
distinct thioredoxin systems; one dependent on ferredoxin as reductant, the
other on NADPH.
In this thesis I have investigated the role of the NADPH-dependent
chloroplast thioredoxin system (NTRC) in regulation of photosynthetic
processes, as well as the coordination between the NTRC- and ferredoxindependent
systems. I demonstrate that NTRC forms a crucial regulatory
hub in chloroplasts that allows maintenance of redox balance between the
photosynthetic electron transfer chain and stromal metabolism, particularly
in low light conditions. This is achieved through regulation of the activities
of the ATP synthase and enzymes of the Calvin–Benson cycle, as well as
non-photochemical quenching, cyclic electron transfer around photosystem
I via the NADH dehydrogenase-like complex, and reversible redistribution
of excitation energy between the photosystems. I show that
significant crosstalk exists between the thioredoxin systems, which allows
dynamic control of photosynthetic processes and photoprotective
mechanisms in fluctuating light conditions. Understanding these regulatory
mechanisms of photosynthesis is of utmost importance in bioengineering
projects aiming to maximize crop yields or biofuel production. Moreover,
my results suggest that enhancement of chloroplast thioredoxin activity
may provide a simple but effective tool for those purposesHappea tuottavassa fotosynteesissä hiilidioksidia muunnetaan sokereiksi
auringon valoenergian sekä vedeltä peräisin olevien elektronien avulla.
Syanobakteerit sekä niiden evolutiiviset jälkeläiset eli kasvien ja levien
viherhiukkaset kykenevät fotosynteettiseen sokereiden tuottoon
mahdollistaen lähes kaikkien Maapallon ekosysteemien toiminnan.
Fotosynteesin elektroninsiirto vedeltä ferredoksiinille tuottaa NADPH:ta ja
johtaa elektrokemiallisen protonigradientin muodostumiseen
tylakoidikalvon yli. Protonigradientti toimii ATP-syntaasin
käyttövoimana, ja NADPH:ta ja ATP:tä käytetään energialähteenä Calvin–
Benson–syklissä tapahtuvassa hiilidioksidin sidonnassa.
Luonnonolosuhteissa valon voimakkuus ja muut ympäristötekijät
vaihtelevat nopeasti kasvien kasvupaikoilla. Tämä on luonut
valintapaineen moninaisten säätelymekanismien kehittymiselle
fotosynteesikoneiston energiankeruun turvaamiseksi ja vahingoittumisen
välttämiseksi. Tioredoksiinit kuuluvat säätelyproteiineihin, jotka
katalysoivat proteiinien rikkisiltojen pelkistystä ja ovat keskeisiä
viherhiukkasten toimintaa sääteleviä yhdisteitä. Kasvien viherhiukkasissa
on useita tioredoksiini-proteiineja sekä kaksi erillistä
tioredoksiinijärjestelmää, joista toinen käyttää ferredoksiinia ja toinen
NADPH:ta pelkistyksessä tarvittavien elektronien lähteenä.
Väitöskirjassani olen tutkinut NADPH-riippuvaisen
tioredoksiinijärjestelmän (NTRC:n) roolia fotosynteettisten prosessien
säätelijänä, sekä NTRC- ja ferredoksiini-riippuvaisen järjestelmän välistä
vuorovaikutussuhdetta. Tutkimukseni osoittaa, että NTRC:llä on keskeinen
tehtävä hapetus–pelkistys-tasapainon säilyttämisessä fotosynteesin
valoreaktioiden ja strooman hiilimetabolian välillä, etenkin heikossa
valossa ja valo-olosuhteiden äkillisten muutosten aikana. NTRC säätelee
ATP-syntaasin ja Calvin–Benson-syklin entsyymien aktiivisuutta,
ylimääräisen viritysenergian hajottamista, syklistä elektronikiertoa, sekä
viritysenergian jakautumista valoreaktio II:n ja I:n kesken. Työni osoittaa
myös, että NTRC- ja ferredoksiini-riippuvainen tioredoksiinijärjestelmä
ovat vuorovaikutuksessa keskenään, mikä mahdollistaa fotosynteettisten
reaktioiden ja suojamekanismien dynaamisen säätelyn vaihtelevissa valoolosuhteissa.
Näiden säätelymekanismien ymmärtäminen on hyvin tärkeää
kun pyritään bioteknisin keinoin maksimoimaan viljelykasvien tai
biopolttoaineen tuottoa. Kloroplastin tioredoksiinijärjestelmien toiminnan
tehostaminen yksinkertaisella geenimuokkauksella saattaa olla hyödyllinen
työkalu kasvien kasvun ja tuottavuuden parantamiseksi
Chloroplast thioredoxin systems dynamically regulate photosynthesis in plants
Photosynthesis is a highly regulated process in photoautotrophic cells. The main goal of the regulation is to keep the basic photosynthetic reactions, i.e. capturing light energy, conversion into chemical energy and production of carbohydrates, in balance. The rationale behind the evolution of strong regulation mechanisms is to keep photosynthesis functional under all conditions encountered by sessile plants during their lifetimes. The regulatory mechanisms may, however, also impair photosynthetic efficiency by overriding the photosynthetic reactions in controlled environments like crop fields or bioreactors, where light energy could be used for production of sugars instead of dissipation as heat and down-regulation of carbon fixation. The plant chloroplast has a high number of regulatory proteins called thioredoxins (TRX), which control the function of chloroplasts from biogenesis and assembly of chloroplast machinery to light and carbon fixation reactions as well as photoprotective mechanisms. Here, we review the current knowledge of regulation of photosynthesis by chloroplast TRXs and assess the prospect of improving plant photosynthetic efficiency by modification of chloroplast thioredoxin systems
Cytochrome cM decreases photosynthesis under photomixotrophy in Synechocystis sp. PCC 6803
Photomixotrophy is a metabolic state that enables photosynthetic microorganisms to simultaneously perform photosynthesis and metabolism of imported organic carbon substrates. This process is complicated in cyanobacteria, since many, including Synechocystis sp. PCC 6803, conduct photosynthesis and respiration in an interlinked thylakoid membrane electron transport chain. Under photomixotrophy, the cell must therefore tightly regulate electron fluxes from photosynthetic and respiratory complexes. In this study, we demonstrate, via characterization of photosynthetic apparatus and the proteome, that photomixotrophic growth results in a gradual inhibition of QA- reoxidation in wild-type Synechocystis, which largely decreases photosynthesis over 3 d of growth. This process is circumvented by deleting the gene encoding cytochrome cM (CytM), a cryptic c-type heme protein widespread in cyanobacteria. The ΔCytM strain maintained active photosynthesis over the 3-d period, demonstrated by high photosynthetic O2 and CO2 fluxes and effective yields of PSI and PSII. Overall, this resulted in a higher growth rate compared to that of the wild type, which was maintained by accumulation of proteins involved in phosphate and metal uptake, and cofactor biosynthetic enzymes. While the exact role of CytM has not been determined, a mutant deficient in the thylakoid-localized respiratory terminal oxidases and CytM (ΔCox/Cyd/CytM) displayed a phenotype similar to that of ΔCytM under photomixotrophy. This, in combination with other physiological data, and in contrast to a previous hypothesis, suggests that CytM does not transfer electrons to these complexes. In summary, our data suggest that CytM may have a regulatory role in photomixotrophy by modulating the photosynthetic capacity of cells
Regulatory electron transport pathways of photosynthesis in cyanobacteria and microalgae: Recent advances and biotechnological prospects
Cyanobacteria and microalgae perform oxygenic photosynthesis where light energy is harnessed to split water into oxygen and protons. This process releases electrons that are used by the photosynthetic electron transport chain to form reducing equivalents that provide energy for the cell metabolism. Constant changes in environmental conditions, such as light availability, temperature, and access to nutrients, create the need to balance the photochemical reactions and the metabolic demands of the cell. Thus, cyanobacteria and microalgae evolved several auxiliary electron transport (AET) pathways to disperse the potentially harmful over-supply of absorbed energy. AET pathways are comprised of electron sinks, e.g. flavodiiron proteins (FDPs) or other terminal oxidases, and pathways that recycle electrons around photosystem I, like NADPH-dehydrogenase-like complexes (NDH) or the ferredoxin-plastoquinone reductase (FQR). Under controlled conditions the need for these AET pathways is decreased and AET can even be energetically wasteful. Therefore, redirecting photosynthetic reducing equivalents to biotechnologically useful reactions, catalyzed by i.e. innate hydrogenases or heterologous enzymes, offers novel possibilities to apply photosynthesis research
Overexpression of chloroplast NADPH-dependent thioredoxin reductase in Arabidopsis enhances leaf growth and elucidates in vivo function of reductase and thioredoxin domains
Plant chloroplasts have versatile thioredoxin systems including two thioredoxin reductases and multiple types of thioredoxins. Plastid-localized NADPH-dependent thioredoxin reductase (NTRC) contains both reductase (NTRd) and thioredoxin (TRXd) domains in a single polypeptide and forms homodimers. To study the action of NTRC and NTRC domains in vivo, we have complemented the ntrc knockout line of Arabidopsis with the wild type and full-length NTRC genes, in which 2-Cys motifs either in NTRd, or in TRXd were inactivated. The ntrc line was also transformed either with the truncated NTRd or TRXd alone. Overexpression of wild-type NTRC promoted plant growth by increasing leaf size and biomass yield of the rosettes. Complementation of the ntrc line with the full-length NTRC gene containing an active reductase but an inactive thioredoxin domain, or vice versa, recovered wild-type chloroplast phenotype and, partly, rosette biomass production, indicating that the NTRC domains are capable of interacting with other chloroplast thioredoxin systems. Overexpression of truncated NTRd or TRXd in ntrc background did not restore wild-type phenotype. Modelling of the 3-dimensional structure of the NTRC dimer indicates extensive interactions between the NTR domains and the TRX domains further stabilize the dimeric structure. The long linker region between the NTRd and TRXd, however, allows flexibility for the position of the TRXd in the dimer. Supplementation of the TRXd in the NTRC homodimer model by free chloroplast thioredoxins indicated that TRXf is the most likely partner to interact with NTRC. We propose that overexpression of NTRC promotes plant biomass yield both directly by stimulation of chloroplast biosynthetic and protected pathways controlled by NTRC and indirectly via free chloroplast thioredoxins. Our data indicate that overexpression of chloroplast thiol redox-regulator has a potential to increase biofuel yield in plant and algal species suitable for sustainable bioene
Multilevel regulation of non-photochemical quenching and state transitions by chloroplast NADPH-dependent thioredoxin reductase
In natural growth habitats, plants face constant, unpredictable changes in light conditions. To avoid damage to the photosynthetic apparatus on thylakoid membranes in chloroplasts, and to avoid wasteful reactions, it is crucial to maintain a redox balance both within the components of photosynthetic electron transfer chain and between the light reactions and stromal carbon metabolism under fluctuating light conditions. This requires coordinated function of the photoprotective and regulatory mechanisms, such as non-photochemical quenching (NPQ) and reversible redistribution of excitation energy between photosystem II (PSII) and photosystem I (PSI). In this paper, we show that the NADPH-dependent chloroplast thioredoxin system (NTRC) is involved in the control of the activation of these mechanisms. In plants with altered NTRC content, the strict correlation between lumenal pH and NPQ is partially lost. We propose that NTRC contributes to downregulation of a slow-relaxing constituent of NPQ, whose induction is independent of lumenal acidification. Additionally, overexpression of NTRC enhances the ability to adjust the excitation balance between PSII and PSI, and improves the ability to oxidize the electron transfer chain during changes in light conditions. Thiol regulation allows coupling of the electron transfer chain to the stromal redox state during these changes.</p
Editorial: A Novel Perspective for Photosystem I: An Emerging Hub for the Functional Integration of Photosynthesis and Metabolism
Mapping Nanocellulose- and Alginate-Based Photosynthetic Cell Factory Scaffolds:Interlinking Porosity, Wet Strength, and Gas Exchange
To develop efficient solid-state photosynthetic cell factories for sustainable chemical production, we present an interdisciplinary experimental toolbox to investigate and interlink the structure, operative stability, and gas transfer properties of alginate- and nanocellulose-based hydrogel matrices with entrapped wild-type Synechocystis PCC 6803 cyanobacteria. We created a rheological map based on the mechanical performance of the hydrogel matrices. The results highlighted the importance of Ca2+-cross-linking and showed that nanocellulose matrices possess higher yield properties, and alginate matrices possess higher rest properties. We observed higher porosity for nanocellulose-based matrices in a water-swollen state via calorimetric thermoporosimetry and scanning electron microscopy imaging. Finally, by pioneering a gas flux analysis via membrane-inlet mass spectrometry for entrapped cells, we observed that the porosity and rigidity of the matrices are connected to their gas exchange rates over time. Overall, these findings link the dynamic properties of the life-sustaining matrix to the performance of the immobilized cells in tailored solid-state photosynthetic cell factories.</p
Photosynthetically produced sucrose by immobilized Synechocystis sp. PCC 6803 drives biotransformation in E. coli
Background: Whole-cell biotransformation is a promising emerging technology for the production of chemicals. When using heterotrophic organisms such as E. coli and yeast as biocatalysts, the dependence on organic carbon source impairs the sustainability and economic viability of the process. As a promising alternative, photosynthetic cyanobacteria with low nutrient requirements and versatile metabolism, could ofer a sustainable platform for the heterologous production of organic compounds directly from sunlight and CO2. This strategy has been applied for the photoautotrophic production of sucrose by a genetically engineered cyanobacterium, Synechocystis sp. PCC 6803 strain S02. As the key concept in the current work, this can be further used to generate organic carbon compounds for diferent heterotrophic applications, including for the whole-cell biotransformation by yeast and bacteria. Results: Entrapment of Synechocystis S02 cells in Ca2+-cross-linked alginate hydrogel beads improves the specifc sucrose productivity by 86% compared to suspension cultures during 7 days of cultivation under salt stress. The process was further prolonged by periodically changing the medium in the vials for up to 17 days of efcient production, giving the fnal sucrose yield slightly above 3000 mg l −1 . We successfully demonstrated that the medium enriched with photosynthetically produced sucrose by immobilized Synechocystis S02 cells supports the biotransformation of cyclohexanone to ε-caprolactone by the E. coli WΔcscR Inv:Parvi strain engineered to (i) utilize low concentrations of sucrose and (ii) perform biotransformation of cyclohexanone to ε-caprolactone. Conclusion: We conclude that cell entrapment in Ca2+-alginate beads is an efective method to prolong sucrose production by the engineered cyanobacteria, while allowing efcient separation of the cells from the medium. This advantage opens up novel possibilities to create advanced autotroph–heterotroph coupled cultivation systems for solar-driven production of chemicals via biotransformation, as demonstrated in this work by utilizing the photosynthetically produced sucrose to drive the conversion of cyclohexanone to ε-caprolactone by engineered E. coli.</p
Flv3A facilitates O2 photoreduction and affects H2 photoproduction independently of Flv1A in diazotrophic Anabaena filaments
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