Photoprotective mechanisms in Elysia species hosting Acetabularia chloroplasts shed light on host-donor compatibility in photosynthetic sea slugs.

Sacoglossa sea slugs have garnered attention due to their ability to retain intracellular functional chloroplasts from algae, while degrading other algal cell components. While protective mechanisms that limit oxidative damage under excessive light are well documented in plants and algae, the photoprotective strategies employed by these photosynthetic sea slugs remain unresolved. Species within the genus Elysia are known to retain chloroplasts from various algal sources, but the extent to which the metabolic processes from the donor algae can be sustained by the sea slugs is unclear. By comparing responses to high-light conditions through kinetic analyses, molecular techniques, and biochemical assays, this study shows significant differences between two photosynthetic Elysia species with chloroplasts derived from the green alga Acetabularia acetabulum. Notably, Elysia timida displayed remarkable tolerance to high-light stress and sophisticated photoprotective mechanisms such as an active xanthophyll cycle, efficient D1 protein recycling, accumulation of heat-shock proteins and α-tocopherol. In contrast, Elysia crispata exhibited absence or limitations in these photoprotective strategies. Our findings emphasize the intricate relationship between the host animal and the stolen chloroplasts, highlighting different capacities to protect the photosynthetic organelle from oxidative damage.

Prey species and abundance affect growth and photosynthetic performance of the polyphagous sea slug Elysia crispata.

Some sacoglossan sea slugs steal functional macroalgal chloroplasts (kleptoplasts). In this study, we investigated the effects of algal prey species and abundance on the growth and photosynthetic capacity of the tropical polyphagous sea slug Elysia crispata. Recently hatched sea slugs fed and acquired chloroplasts from the macroalga Bryopsis plumosa, but not from Acetabularia acetabulum. However, adult sea slugs were able to switch diet to A. acetabulum, rapidly replacing the great majority of the original kleptoplasts. When fed with B. plumosa, higher feeding frequency resulted in significantly higher growth and kleptoplast photosynthetic yield, as well as a slower relative decrease in these parameters upon starvation. Longevity of A. acetabulum-derived chloroplasts in E. crispata was over twofold that of B. plumosa. Furthermore, significantly lower relative weight loss under starvation was observed in sea slugs previously fed on A. acetabulum than on B. plumosa. This study shows that functionality and longevity of kleptoplasts in photosynthetic sea slugs depend on the origin of the plastids. Furthermore, we have identified A. acetabulum as a donor of photosynthetically efficient chloroplasts common to highly specialized monophagous and polyphagous sea slugs capable of long-term retention, which opens new experimental routes to unravel the unsolved mysteries of kleptoplasty.

Ultraviolet screening by slug tissue and tight packing of plastids protect photosynthetic sea slugs from photoinhibition.

One of the main mysteries regarding photosynthetic sea slugs is how the slug plastids handle photoinhibition, the constant light-induced damage to Photosystem II of photosynthesis. Recovery from photoinhibition involves proteins encoded by both the nuclear and plastid genomes, and slugs with plastids isolated from the algal nucleus are therefore expected to be incapable of constantly repairing the damage as the plastids inside the slugs grow old. We studied photoinhibition-related properties of the sea slug Elysia timida that ingests its plastids from the green alga Acetabularia acetabulum. Spectral analysis of both the slugs and the algae revealed that there are two ways the slugs use to avoid major photoinhibition of their plastids. Firstly, highly photoinhibitory UV radiation is screened by the slug tissue or mucus before it reaches the plastids. Secondly, the slugs pack the plastids tightly in their thick bodies, and therefore plastids in the outer layers protect the inner ones from photoinhibition. Both properties are expected to greatly improve the longevity of the plastids inside the slugs, as the plastids do not need to repair excessive amounts of damage.

Genetic autonomy and low singlet oxygen yield support kleptoplast functionality in photosynthetic sea slugs.

The kleptoplastic sea slug Elysia chlorotica consumes Vaucheria litorea, stealing its plastids, which then photosynthesize inside the animal cells for months. We investigated the properties of V. litorea plastids to understand how they withstand the rigors of photosynthesis in isolation. Transcription of specific genes in laboratory-isolated V. litorea plastids was monitored for 7 days. The involvement of plastid-encoded FtsH, a key plastid maintenance protease, in recovery from photoinhibition in V. litorea was estimated in cycloheximide-treated cells. In vitro comparison of V. litorea and spinach thylakoids was applied to investigate reactive oxygen species formation in V. litorea. In comparison to other tested genes, the transcripts of ftsH and translation elongation factor EF-Tu (tufA) decreased slowly in isolated V. litorea plastids. Higher levels of FtsH were also evident in cycloheximide-treated cells during recovery from photoinhibition. Charge recombination in PSII of V. litorea was found to be fine-tuned to produce only small quantities of singlet oxygen, and the plastids also contained reactive oxygen species-protective compounds. Our results support the view that the genetic characteristics of the plastids are crucial in creating a photosynthetic sea slug. The plastid's autonomous repair machinery is likely enhanced by low singlet oxygen production and elevated expression of FtsH.

Photosynthetic sea slugs induce protective changes to the light reactions of the chloroplasts they steal from algae.

Sacoglossan sea slugs are able to maintain functional chloroplasts inside their own cells, and mechanisms that allow preservation of the chloroplasts are unknown. We found that the slug Elysia timida induces changes to the photosynthetic light reactions of the chloroplasts it steals from the alga Acetabularia acetabulum. Working with a large continuous laboratory culture of both the slugs (>500 individuals) and their prey algae, we show that the plastoquinone pool of slug chloroplasts remains oxidized, which can suppress reactive oxygen species formation. Slug chloroplasts also rapidly build up a strong proton-motive force upon a dark-to-light transition, which helps them to rapidly switch on photoprotective non-photochemical quenching of excitation energy. Finally, our results suggest that chloroplasts inside E. timida rely on oxygen-dependent electron sinks during rapid changes in light intensity. These photoprotective mechanisms are expected to contribute to the long-term functionality of the chloroplasts inside the slugs.

Plants, algae and a few other organisms rely on a process known as photosynthesis to fuel themselves, as they can harness cellular structures called chloroplasts to convert light into usable energy. Animals typically lack chloroplasts, making them unable to use photosynthesis to power themselves. The sea slug Elysia timida, however, can steal whole chloroplasts from the cells of the algae it consumes: the stolen structures then become part of the cells in the gut of the slug, allowing the animal to gain energy from sunlight. Once they are in the digestive system of the slug, the chloroplasts survive and keep working for longer than expected. Indeed, these structures are often harmed as a side effect of photosynthesis, but the sea slug does not have the right genes to help repair this damage. In addition, conditions inside animal cells are widely different to the ones found inside algae and plants. It is not clear then how the sea slug extends the lifespan of its chloroplasts by preventing damage caused by sunlight. To investigate this question, Havurinne and Tyystjärvi compared photosynthesis in sea slugs and the algae they eat. A range of methods, including measuring fluorescence from the chloroplasts, was used: this revealed that the slug changes the inside of the stolen chloroplasts, making them more resistant to damage. First, when exposed to light the stolen chloroplasts can quickly switch on a mechanism that dissipates light energy to heat, which is less damaging. Second, a molecule that serves as an intermediate during photosynthesis is kept in a 'safe' state which prevents it from creating harmful compounds. And finally, additional safeguard molecules 'deactivate' compounds that could otherwise mediate damaging reactions. Overall, these measures may reduce the efficiency of the chloroplasts but allow them to keep working for much longer. Early chloroplasts were probably independent bacteria that were captured and 'domesticated' by other cells for their ability to extract energy from the sun. Photosynthesizing sea slugs therefore provide an interesting way to understand some of the challenges of early life. The work by Havurinne and Tyystjärvi may also reveal new ways to harness biological processes such as photosynthesis for energy production in other contexts.

Action spectrum of the redox state of the plastoquinone pool defines its function in plant acclimation.

The plastoquinone (PQ) pool mediates electron flow and regulates photoacclimation in plants. Here we report the action spectrum of the redox state of the PQ pool in Arabidopsis thaliana, showing that 470-500, 560 or 650-660 nm light favors Photosystem II (PSII) and reduces the PQ pool, whereas 420-440, 520 or 690 nm light favors Photosystem I (PSI) and oxidizes PQ. These data were used to construct a model predicting the redox state of PQ from the spectrum of any polychromatic light source. Moderate reduction of the PQ pool induced transition to light state 2, whereas state 1 required highly oxidized PQ. In low-intensity PSI light, PQ was more oxidized than in darkness and became gradually reduced with light intensity, while weak PSII light strongly reduced PQ. Natural sunlight was found to favor PSI, which enables plants to use the redox state of the PQ pool as a measure of light intensity.

Oxygen and ROS in Photosynthesis.

Oxygen is a natural acceptor of electrons in the respiratory pathway of aerobic organisms and in many other biochemical reactions. Aerobic metabolism is always associated with the formation of reactive oxygen species (ROS). ROS may damage biomolecules but are also involved in regulatory functions of photosynthetic organisms. This review presents the main properties of ROS, the formation of ROS in the photosynthetic electron transport chain and in the stroma of chloroplasts, and ROS scavenging systems of thylakoid membrane and stroma. Effects of ROS on the photosynthetic apparatus and their roles in redox signaling are discussed.

Unresolved quenching mechanisms of chlorophyll fluorescence may invalidate MT saturating pulse analyses of photosynthetic electron transfer in microalgae.

Chlorophyll a fluorescence is a powerful tool for estimating photosynthetic efficiency, but there are still unanswered questions that hinder the use of its full potential. The present results describe a caveat in estimation of photosynthetic performance with so-called rapid light curves (RLCs) with pulse amplitude modulation fluorometers. RLCs of microalgae show a severe decrease in photosynthetic performance in high light, although a similar decrease cannot be seen with other methods. We show that this decrease cannot be assigned to energy-dependent non-photochemical quenching or photoinhibition or to the geometry of the algal sample. The measured decrease in electron transfer rate is small in the tested siphonaceuous algae and higher plants, but very notable in all planktonic species, exhibiting species-dependent variation in extent and reversibility. We performed in-depth analysis of the phenomenon in the diatom Phaeodactylum tricornutum, in which the decrease is the most pronounced and reversible among the tested organisms. The results suggest that quenching of fluorescence by oxidized plastoquinone alone cannot explain the phenomenon, and alternative quenching mechanisms within PSII need to be considered.

Photosystem-II D1 protein mutants of Chlamydomonas reinhardtii in relation to metabolic rewiring and remodelling of H-bond network at QB site.

Photosystem II (PSII) reaction centre D1 protein of oxygenic phototrophs is pivotal for sustaining photosynthesis. Also, it is targeted by herbicides and herbicide-resistant weeds harbour single amino acid substitutions in D1. Conservation of D1 primary structure is seminal in the photosynthetic performance in many diverse species. In this study, we analysed built-in and environmentally-induced (high temperature and high photon fluency - HT/HL) phenotypes of two D1 mutants of Chlamydomonas reinhardtii with Ala250Arg (A250R) and Ser264Lys (S264K) substitutions. Both mutations differentially affected efficiency of electron transport and oxygen production. In addition, targeted metabolomics revealed that the mutants undergo specific differences in primary and secondary metabolism, namely, amino acids, organic acids, pigments, NAD, xanthophylls and carotenes. Levels of lutein, ß-carotene and zeaxanthin were in sync with their corresponding gene transcripts in response to HT/HL stress treatment in the parental (IL) and A250R strains. D1 structure analysis indicated that, among other effects, remodelling of H-bond network at the QB site might underpin the observed phenotypes. Thus, the D1 protein, in addition to being pivotal for efficient photosynthesis, may have a moonlighting role in rewiring of specific metabolic pathways, possibly involving retrograde signalling.

Degradation of chlorophyll and synthesis of flavonols during autumn senescence-the story told by individual leaves.

Autumn senescence of deciduous trees is characterized by chlorophyll degradation and flavonoid synthesis. In the present study, chlorophyll and flavonol contents were measured every morning and evening during the whole autumn with a non-destructive method from individual leaves of Sorbus aucuparia, Acer platanoides, Betula pendula and Prunus padus. In most of the studied trees, the chlorophyll content of each individual leaf remained constant until a phase of rapid degradation commenced. The fast phase lasted only ~1 week and ended with abscission. In S. aucuparia, contrary to the other species, the chlorophyll content of leaflets slowly but steadily decreased during the whole autumn, but rapid chlorophyll degradation commenced only prior to leaflet abscission also in this species. An increase in flavonols commonly accompanied the rapid degradation of chlorophyll. The results may suggest that each individual tree leaf retains its photosynthetic activity, reflected by a high chlorophyll content, until a rapid phase of chlorophyll degradation and flavonoid synthesis begins. Therefore, in studies of autumn senescence, leaves whose chlorophyll content is decreasing and leaves with summertime chlorophyll content (i.e. the leaves that have not yet started to degrade chlorophyll) should be treated separately.

Action Spectrum of Photoinhibition in the Diatom Phaeodactylum tricornutum.

Light-dependent electron transfer is necessary for photosynthesis, but light also damages PSII. Light-induced damage to PSII is called photoinhibition, and the damaging reactions of photoinhibition are still under debate. Diatoms possess an exotic combination of light-harvesting pigments, Chls a/c and fucoxanthin, making them an interesting platform for studying the photoreceptors of photoinhibition. We first confirmed the direct proportionality of photoinhibition to the photon flux density of incident light in the diatom Phaeodactylum tricornutum. Phaeodactylum is known for its efficient non-photochemical quenching, and the effect of this photoprotective mechanism on photoinhibition was tested. Photoinhibition proceeded essentially at the same rate in blue-light-grown Phaeodactylum cells that are capable of non-photochemical quenching and in red-light-grown, non-photochemical quenching-deficient cells. To obtain more insight into how the pigment composition of diatoms affects photoinhibition, we measured the action spectrum of photoinhibition in Phaeodactylum. In visible light, the action spectrum resembled the absorption spectrum of Phaeodactylum, and UV radiation caused much more photoinhibition than visible light. Comparison of the action spectrum of photoinhibition with the absorption spectrum and the excitation spectrum of 77 K PSII fluorescence emission confirmed that photosynthetic pigments are involved in photoinhibition, but the photoinhibitory efficiency of red light is weak, suggesting that the role of light-harvesting pigments as light receptors of photoinhibition is secondary. Finally, we compared photoinhibition in Phaeodactylum with that in other photosynthetic organisms, and our data indicate that the PSII reaction centers of Phaeodactylum are not particularly well protected against the primary damage of photoinhibition.

Photoinhibition in marine picocyanobacteria.

Marine Synechococcus and Prochlorococcus cyanobacteria have different antenna compositions although they are genetically near to each other, and different strains thrive in very different illumination conditions. We measured growth and photoinhibition of PSII in two low-light and one high-light Prochlorococcus strains and in one Synechococcus strain. All strains were found to be able to shortly utilize moderate or even high light, but the low-light strains bleached rapidly in moderate light. Measurements of photoinhibition in the presence of the antibiotic lincomycin showed that a low-light Prochlorococcus strain was more sensitive than a high-light strain and both were more sensitive than the marine Synechococcus. The action spectrum of photoinhibition showed an increase from blue to ultraviolet wavelengths in all strains, suggesting contribution of manganese absorption to photoinhibition, but blue light caused less photoinhibition in marine cyanobacteria than expected on the basis of earlier results from plants and cyanobacteria. The visible-light part of the action spectrum resembled the absorption spectrum of the organism, suggesting that photosynthetic antenna pigments, especially divinyl chlorophylls, have a more important role as photoreceptors of visible-light photoinhibition in marine cyanobacteria than in other photoautotrophs.

Reactive oxygen species: Reactions and detection from photosynthetic tissues.

Reactive oxygen species (ROS) have long been recognized as compounds with dual roles. They cause cellular damage by reacting with biomolecules but they also function as agents of cellular signaling. Several different oxygen-containing compounds are classified as ROS because they react, at least with certain partners, more rapidly than ground-state molecular oxygen or because they are known to have biological effects. The present review describes the typical reactions of the most important ROS. The reactions are the basis for both the detection methods and for prediction of reactions between ROS and biomolecules. Chemical and physical methods used for detection, visualization and quantification of ROS from plants, algae and cyanobacteria will be reviewed. The main focus will be on photosynthetic tissues, and limitations of the methods will be discussed.