Alternative electron pathways of photosynthesis power green algal CO2 capture.

Microalgae contribute to about half of global net photosynthesis, which converts sunlight into the chemical energy (ATP and NADPH) used to transform CO2 into biomass. Alternative electron pathways of photosynthesis have been proposed to generate additional ATP that is required to sustain CO2 fixation. However, the relative importance of each alternative pathway remains elusive. Here, we dissect and quantify the contribution of cyclic, pseudo-cyclic and chloroplast-to-mitochondria electron flows for their ability to sustain net photosynthesis in the microalga Chlamydomonas reinhardtii. We show that (i) each alternative pathway can provide sufficient additional energy to sustain high CO2 fixation rates, (ii) the alternative pathways exhibit cross-compensation, and (iii) the activity of at least one of the three alternative pathways is necessary to sustain photosynthesis. We further show that all pathways have very different efficiencies at energizing CO2 fixation, with the chloroplast-mitochondria interaction being the most efficient. Overall, our data lay bioenergetic foundations for biotechnological strategies to improve CO2 capture and fixation.

Exploring LHCSR3 expression and its role in Chlamydomonas reinhardtii under osmotic stress: Implications for non-photochemical quenching mechanism.

Plants have a protective mechanism called non-photochemical quenching to prevent damage caused by excessive sunlight. A critical component of this mechanism is energy-dependent quenching (qE). In Chlamydomonas reinhardtii, the protein expression called light-harvesting complex stress-related protein 3 (LHCSR3) is crucial for the qE mechanism. LHCSR3 expression is observed in various conditions that result in photooxidation, such as exposure to high light or nutrient deprivation, where the amount of captured light surpasses the maximum photosynthetic capacity. Although the role of LHCSR3 has been extensively studied under high light (HL) conditions, its function during nutrient starvation remains unclear. In this study, we demonstrate that LHCSR3 expression can occur under light intensities below saturation without triggering qE, particularly when nutrients are limited. To investigate this, we cultivated C. reinhardtii cells under osmotic stress, which replicates conditions of nutrient scarcity. Furthermore, we examined the photosynthetic membrane complexes of wild-type (WT) and npq4 mutant strains grown under osmotic stress. Our analysis revealed that LHCSR3 expression might modify the interaction between the photosystem II core and its peripheral light-harvesting complex II antennae. This alteration could potentially impede the transfer of excitation energy from the antenna to the reaction center.

Role of cyclic electron transport mutations pgrl1 and pgr5 in acclimation process to high light in Chlamydomonas reinhardtii.

Light is crucial for photosynthesis, but the amount of light that exceeds an organism's assimilation efficacy can lead to photo-oxidative damage and even cell death. In Chlamydomonas (C). reinhardtii cyclic electron flow (CEF) is very important for the elicitation of non-photochemical quenching (NPQ) by controlling the acidification of thylakoid lumen. This process requires the cooperation of proton gradient regulation (PGR) proteins, PGRL1 and PGR5. Here, we compared the growth pattern and photosynthetic activity between wild type (137c, t222+) and mutants impaired in CEF (pgrl1 and pgr5) under photoautotrophic and photoheterotrophic conditions. We have observed the discriminative expression of NPQ in the mutants impaired in CEF of pgrl1 and pgr5. The results obtained from the mutants showed reduced cell growth and density, Chl a/b ratio, fluorescence, electron transport rate, and yield of photosystem (PS)II. These mutants have reduced capability to develop a strong NPQ indicating that the role of CEF is very crucial for photoprotection. Moreover, the CEF mutant exhibits increased photosensitivity compared with the wild type. Therefore, we suggest that besides NPQ, the fraction of non-regulated non-photochemical energy loss (NO) also plays a crucial role during high light acclimation despite a low growth rate. This low NPQ rate may be due to less influx of protons coming from the CEF in cases of pgrl1 and pgr5 mutants. These results are discussed in terms of the relative photoprotective benefit, related to the thermal dissipation of excess light in photoautotrophic and photoheterotrophic conditions.

Non-photochemical quenching-dependent acclimation and thylakoid organization of Chlamydomonas reinhardtii to high light stress.

Light is essential for all photosynthetic organisms while an excess of it can lead to damage mainly the photosystems of the thylakoid membrane. In this study, we have grown Chlamydomonas reinhardtii cells in different intensities of high light to understand the photosynthetic process with reference to thylakoid membrane organization during its acclimation process. We observed, the cells acclimatized to long-term response to high light intensities of 500 and 1000 µmol m-2 s-1 with faster growth and more biomass production when compared to cells at 50 µmol m-2 s-1 light intensity. The ratio of Chl a/b was marginally decreased from the mid-log phase of growth at the high light intensity. Increased level of zeaxanthin and LHCSR3 expression was also found which is known to play a key role in non-photochemical quenching (NPQ) mechanism for photoprotection. Changes in photosynthetic parameters were observed such as increased levels of NPQ, marginal change in electron transport rate, and many other changes which demonstrate that cells were acclimatized to high light which is an adaptive mechanism. Surprisingly, PSII core protein contents have marginally reduced when compared to peripherally arranged LHCII in high light-grown cells. Further, we also observed alterations in stromal subunits of PSI and low levels of PsaG, probably due to disruption of PSI assembly and also its association with LHCI. During the process of acclimation, changes in thylakoid organization occurred in high light intensities with reduction of PSII supercomplex formation. This change may be attributed to alteration of protein-pigment complexes which are in agreement with circular dichoism spectra of high light-acclimatized cells, where decrease in the magnitude of psi-type bands indicates changes in ordered arrays of PSII-LHCII supercomplexes. These results specify that acclimation to high light stress through NPQ mechanism by expression of LHCSR3 and also observed changes in thylakoid protein profile/supercomplex formation lead to low photochemical yield and more biomass production in high light condition.

Thylakoid membrane dynamics and state transitions in Chlamydomonas reinhardtii under elevated temperature.

Moderately elevated temperatures can induce state transitions in higher plants by phosphorylation of light-harvesting complex II (LHCII). In this study, we exposed unicellular algae Chlamydomonas reinhardtii to moderately elevated temperatures (38 °C) for short period of time in the dark to understand the thylakoid membrane dynamics and state transition mechanism. Here we report that under elevated temperatures (1) LHCII gets phosphorylated similar to higher plants and (2) there is decreased absorption cross section of photosystem II (PSII), whereas (3) there is no change in absorption cross section of photosystem I (PSI) indicating that LHCII trimers are largely disconnected with both photosystems under moderately elevated temperatures and (4) on return to room temperature after elevated temperature treatment there is a formation of state transition complex comprising of PSII-LHCII-PSI. The temperature-induced state transition mechanism also depends on stt7 kinase-like in light-induced state transition. The protein content was stable at the moderately elevated temperature treatment of 40 °C; however, at 45 °C severe downregulation in photosynthetic performance and protein content was observed. In addition to the known changes to photosynthetic apparatus, elevated temperatures can destabilize the PSII-LHCII complex that can result in decreased photosynthetic efficiency in C. reinhardtii. We concluded that the membrane dynamics of light-induced state transitions differs considerably from temperature-induced state transition mechanisms in C. reinhardtii.

Changes in the photosynthetic apparatus and lipid droplet formation in Chlamydomonas reinhardtii under iron deficiency.

The unicellular photosynthetic alga Chlamydomonas reinhardtii was propagated in iron deficiency medium and patterns of growth, photosynthetic efficiency, lipid accumulation, as well as the expression of lipid biosynthetic and photosynthesis-related proteins were analysed and compared with iron-sufficient growth conditions. As expected, the photosynthetic rate was reduced (maximally after 4 days of growth) as a result of increased non-photochemical quenching (NPQ). Surprisingly, the stress-response protein LHCSR3 was expressed in conditions of iron deficiency that cause NPQ induction. In addition, the protein contents of both the PSI and PSII reaction centres were gradually reduced during growth in iron deficiency medium. Interestingly, the two generations of Fe deficiency cells could be able to recover the photosynthesis but the second generation cells recovered much slower as these cells were severely in shock. Analysis by flow cytometry with fluorescence-activated cell sorting and thin layer chromatography showed that iron deficiency also induced the accumulation of triacylglycerides (TAG), which resulted in the formation of lipid droplets. This was most significant between 48 and 72 h of growth. Dramatic increases in DGAT2A and PDAT1 levels were caused by iron starvation, which indicated that the biosynthesis of TAG had been increased. Analysis using gas chromatography mass spectrometry showed that levels of 16:0, 18:0, 18:2 and 18:3Δ9,12,15 fatty acids were significantly elevated. The results of this study highlight the genes/enzymes of Chlamydomonas that affect lipid synthesis through their influence on photosynthesis, and these represent potential targets of metabolic engineering to develop strains for biofuel production.

Iron deficiency cause changes in photochemistry, thylakoid organization, and accumulation of photosystem II proteins in Chlamydomonas reinhardtii.

A trace element, iron (Fe) plays a pivotal role in photosynthesis process which in turn mediates the plant growth and productivity. Here, we have focused majorly on the photochemistry of photosystem (PS) II, abundance of proteins, and organization of supercomplexes of thylakoids from Fe-depleted cells in Chlamydomonas reinhardtii. Confocal pictures show that the cell's size has been reduced and formed rosette-shaped palmelloids; however, there is no cell death. Further, the PSII photochemistry was reduced remarkably. Further, the photosynthetic efficiency analyzer data revealed that both donor and acceptor side of PSII were equally damaged. Additionally, the room-temperature emission spectra showed the fluorescence emission maxima increased due to impaired energy transfer from PSII to PSI. Furthermore, the protein data reveal that most of the proteins of reaction center and light-harvesting antenna were reduced in Fe-depleted cells. Additionally, the supercomplexes of PSI and PSII were destabilized from thylakoids under Fe-deficient condition showing that Fe is an important element in photosynthesis mechanism.

High light induced changes in organization, protein profile and function of photosynthetic machinery in Chlamydomonas reinhardtii.

The green alga Chlamydomonas (C.) reinhardtii is used as a model organism to understand the efficiency of photosynthesis along with the organization and protein profile of photosynthetic apparatus under various intensities of high light exposure for 1h. Chlorophyll (Chl) a fluorescence induction, OJIPSMT transient was decreased with increase in light intensity indicating the reduction in photochemical efficiency. Further, circular dichroism studies of isolated thylakoids from high light exposed cells showed considerable change in the pigment-pigment interactions and pigment-proteins interactions. Furthermore, the organization of supercomplexes from thylakoids is studied, in which, one of the hetero-trimer of light harvesting complex (LHC) II is affected significantly in comparison to other complexes of LHC's monomers. Also, other supercomplexes, photosystem (PS)II reaction center dimer and PSI complexes are reduced. Additionally, immunoblot analysis of thylakoid proteins revealed that PSII core proteins D1 and D2 were significantly decreased during high light treatment. Similarly, the PSI core proteins PsaC, PsaD and PsaG were drastically changed. Further, the LHC antenna proteins of PSI and PSII were differentially affected. From our results it is clear that LHCs are damaged significantly, consequently the excitation energy is not efficiently transferred to the reaction center. Thus, the photochemical energy transfer from PSII to PSI is reduced. The inference of the study deciphers the structural and functional changes driven by light may therefore provide plants/alga to regulate the light harvesting capacity in excess light conditions.

Photosynthetic membrane organization and role of state transition in cyt, cpII, stt7 and npq mutants of Chlamydomonas reinhardtii.

In Chlamydomonas reinhardtii, cytochrome b6/f and chlorophyll b binding proteins are important in energy distribution between photosystem (PS)II and PSI. In this study, we have used C. reinhardtii mutants deficient in cytochrome b6/f complex (cyt), chlorophyll b binding protein (cpII), non-photochemical quenching (npq) and LHC II kinase (stt7) to study the importance of these proteins in electron transport, phosphorylation, and structural organization of thylakoid supercomplexes under optimum growth conditions. Fast Chl a fluorescence studies have shown that lack of CpII and Cyt b6/f caused reduced photochemical yield (Fv/Fm). The disappearance of I phase in cyt mutant showed that electron transfer from Cyt b6/f to PSI is reduced due to un availability of Q0 site for docking of PQH2 therefore LHC II kinase was unable to phosphorylate LHCII in cyt mutant. Further, blue native gel electrophoresis revealed the differential organization of photosynthetic membrane protein complexes in different mutants. Particularly, LHCII trimerization is more in cyt and stt7 mutants. Also, chemically induced state transition (LHCII phosphorylation) was not observed in cyt mutant, however, all other mutants were similar to that of wild type. Based on our results, we propose that the LHCII trimer accumulation and its organization with other complexes are very important in state transitions.