Expression patterns of melanin-related genes are linked to crypsis and conspicuousness in a pumpkin toadlet.

Colour signals play pivotal roles in different communication systems, and the evolution of these characters has been associated with behavioural ecology, integumentary production processes and perceptual mechanisms of the species involved. Here, we present the first insight into the molecular and histological basis of skin colour polymorphism within a miniaturized species of pumpkin toadlet, potentially representing the lowest size threshold for colour polytypism in tetrapods. Brachycephalus actaeus exhibits a coloration ranging from cryptic green to conspicuous orange skin, and our findings suggest that colour morphs differ in their capability to be detected by potential predators. We also found that the distribution and abundance of chromatophores are variable in the different colour morphs. The expression pattern of coloration related genes was predominantly associated with melanin synthesis (including dct, edn1, mlana, oca2, pmel, slc24a5, tyrp1 and wnt9a). Up-regulation of melanin genes in grey, green and brown skin was associated with higher melanophore abundance than in orange skin, where xanthophores predominate. Our findings provide a significant foundation for comparing and understanding the diverse pathways that contribute to the evolution of pigment production in the skin of amphibians.

The dynamic behavior of chromatophores marks the transition from bands to spots in leopard geckos.

Reptilian skin coloration is spectacular and diverse, yet little is known about the ontogenetic processes that govern its establishment and the molecular signaling pathways that determine it. Here, we focus on the development of the banded pattern of leopard gecko hatchlings and the transition to black spots in the adult. With our histological analyses, we show that iridophores are present in the white and yellow bands of the hatchling and they gradually perish in the adult skin. Furthermore, we demonstrate that melanophores can autonomously form spots in the absence of the other chromatophores both on the regenerated skin of the tail and on the dorsal skin of the Mack Super Snow (MSS) leopard geckos. This color morph is characterized by uniform black coloration in hatchlings and black spots in adulthood; we establish that their skin is devoid of xanthophores and iridophores at both stages. Our genetic analyses identified a 13-nucleotide deletion in the PAX7 transcription factor of MSS geckos, affecting its protein coding sequence. With our single-cell transcriptomics analysis of embryonic skin, we confirm that PAX7 is expressed in iridophores and xanthophores, suggesting that it plays a key role in the differentiation of both chromatophores. Our in situ hybridizations on whole-mount embryos document the dynamics of the skin pattern formation and how it is impacted in the PAX7 mutants. We hypothesize that the melanophores-iridophores interactions give rise to the banded pattern of the hatchlings and black spot formation is an intrinsic capacity of melanophores in the postembryonic skin.

The influence of cationic antiseptics on the processes of light energy conversion in various photosynthetic pigment-protein complexes.

The widespread use of disinfectants and antiseptics, and consequently their release into the environment, determines the relevance of studying their potential impact on the main producers of organic matter on the planet-photosynthetic organisms. The review examines the effects of some biguanides and quaternary ammonium compounds, octenidine, miramistin, chlorhexidine, and picloxidine, on the functioning of the photosynthetic apparatus of various organisms (Strakhovskaya et al. in Photosynth Res 147:197-209, 2021; Knox et al. in Photosynth Res 153:103, 2022; Paschenko et al. in Photosynth Res 155:93-105, 2023a, Photosynth Res 2023b). A common feature of these antiseptics is the combination of hydrophobic and hydrophilic regions in the molecules, the latter carrying a positive charge(s). The comparison of the results obtained with intact bacterial membrane vesicles (chromatophores) and purified pigment-protein complexes (photosystem II and I) of oxygenic organisms allows us to draw conclusions about the mechanisms of the cationic antiseptic action on the functional properties of the components of the photosynthetic apparatus.

On the role of TFEC in reptilian coloration.

Reptilian species, particularly snakes and lizards, are emerging models of animal coloration. Here, I focus on the role of the TFEC transcription factor in snake and lizard coloration based on a study on wild-type and piebald ball pythons. Genomic mapping previously identified a TFEC mutation linked to the piebald ball python phenotype. The association of TFEC with skin coloration was further supported by gene-editing experiments in the brown anole lizard. However, novel histological analyses presented here reveal discrepancies between the ball python and the anole TFEC mutants phenotype, cautioning against broad generalizations. Indeed, both wild-type and piebald ball pythons completely lack iridophores, whereas the TFEC anole lizard mutants lose their iridophores compared to the wild-type anole. Based on these findings, I discuss the potential role of the MiT/TFE family in skin pigmentation across vertebrate lineages and advocate the need for developmental analyses and additional gene-editing experiments to explore the reptilian coloration diversity.

Use of bacterial photosynthetic vesicles to evaluate the effect of ionic liquids on the permeability of biological membranes.

Ionic liquids (ILs) are salts composed of a combination of organic or inorganic cations and anions characterized by a low melting point, often below 100 °C. This property, together with an extremely low vapor pressure, low flammability and high thermal stability, makes them suitable for replacing canonical organic solvents, with a reduction of industrial activities impact on the environment. Although in the last decades the eco-compatibility of ILs has been extensively verified through toxicological tests performed on model organisms, a detailed understanding of the interaction of these compounds with biological membranes is far from being exhaustive. In this context, we have chosen to evaluate the effect of some ILs on native membranes by using chromatophores, photosynthetic vesicles that can be isolated from Rhodobacter capsulatus, a member of the purple non­sulfur bacteria. Here, carotenoids associated with the light-harvesting complex II, act as endogenous spectral probes of the transmembrane electrical potential (ΔΨ). By measuring through time-resolved absorption spectroscopy the evolution of the carotenoid band shift induced by a single excitation of the photosynthetic reaction center, information on the ΔΨ dissipation due to ionic currents across the membrane can be obtained. We found that some ILs cause a rather fast dissipation of the transmembrane ΔΨ even at low concentrations, and that this behavior is dose-dependent. By using two different models to analyze the decay of the carotenoid signals, we attempted to interpret at a mechanistic level the marked increase of ionic permeability caused by specific ILs.

Lel A. Drachev and the Direct Electrometric Method.

In the bioenergetics studies, the direct electrometric method played an important role. This method is based on measuring the electrical potential difference (Δψ) between two compartments of the experimental cell generated by some membrane proteins. These proteins are incorporated into closed lipid-protein membrane vesicles associated with an artificial lipid membrane that separates the compartments. The very existence of such proteins able to generate Δψ was one of the consequences of Peter Mitchell's chemiosmotic concept. The discovery and investigation of their functioning contributed to the recognition of this concept and, eventually the well-deserved awarding of the Nobel Prize to P. Mitchell. Lel A. Drachev (1926-2022) was one of the main authors of the direct electrometrical method. With his participation, key studies were carried out on the electrogenesis of photosynthetic and respiratory membrane proteins, including bacteriorhodopsin, visual rhodopsin, photosynthetic bacterial reaction centers, cytochrome oxidase and others.

Generation of Electric Potential Difference by Chromatophores from Photosynthetic Bacteria in the Presence of Trehalose under Continuous Illumination.

Measurement of electrical potential difference (Δψ) in membrane vesicles (chromatophores) from the purple bacterium Rhodobacter sphaeroides associated with the surface of a nitrocellulose membrane filter (MF) impregnated with a phospholipid solution in decane or immersed into it in the presence of exogenous mediators and disaccharide trehalose demonstrated an increase in the amplitude and stabilization of the signal under continuous illumination. The mediators were the ascorbate/N,N,N'N'-tetramethyl-p-phenylenediamine pair and ubiquinone-0 (electron donor and acceptor, respectively). Although stabilization of photoelectric responses upon long-term continuous illumination was observed for both variants of chromatophore immobilization, only the samples immersed into the MF retained the functional activity of reaction centers (RCs) for a month when stored in the dark at room temperature, which might be due to the preservation of integrity of chromatophore proteins inside the MF pores. The stabilizing effect of the bioprotector trehalose could be related to its effect on both the RC proteins and the phospholipid bilayer membrane. The results obtained will expand current ideas on the use of semi-synthetic structures based on various intact photosynthetic systems capable of converting solar energy into its electrochemical form.

Exploring the mechanisms and impacts of melatonin on fish colouration.

The pineal hormone melatonin is a multi-functional molecule with a recognized role in pigment aggregation in chromatophores, mediating its actions through binding to subtypes of its specific receptors. Since its discovery, melatonin has been known to be responsible for pigment aggregation towards the cell centre in fishes, including their embryos, as an adaptation to reduced light and thus results in pale body colouration. Diversity exists in the sensitivity of melanophores towards melatonin at interspecies, intraspecific levels, seasons, and amongst chromatophores at different regions of the animal body. In most of the fishes, melatonin leads to their skin paling at night. It is indicated that the melatonin receptors have characteristically maintained to show the same aggregating effects in fishes and other vertebrates in the evolutionary hierarchy. However, besides this aggregatory effect, melatonin is also responsible for pigment dispersion in certain fishes. Here is the demand in our review to explore further the nature of the dispersive behaviour of melatonin through the so-called ß-melatonin receptors. It is clear that the pigment translocations in lower vertebrates under the effect of melatonin are mediated through the melatonin receptors coupled with other hormonal receptors as well. Therefore, being richly supplied with a variety of receptors, chromatophores and melanocytes can be used as in vitro test models for pharmacological applications of known and novel drugs. In this review, we present diverse effects of melatonin on chromatophores of fishes in particular with appropriate implications on most of the recent findings.

Investigation of the Efficacy of Lanthanoid Heavy Metal Acetates as Electron Staining Reagents for Biomembrane Vesicles.

Transmission electron microscopy (TEM) has revolutionized our understanding of protein structures by enabling atomic-resolution visualization without the need for crystallography, thanks to advancements in cryo-TEM and single particle analysis methods. However, conventional electron microscopy remains relevant for studying stained samples, as it allows the practical determination of optimal conditions through extensive experimentation. TEM also facilitates the examination of supramolecular complexes encompassing proteins, lipids, and nucleic acids. In this study, we investigated the applicability of lanthanoid reagents as electron-staining alternatives to uranyl acetate, which is globally regulated as a nuclear fuel material. We focus on a model biomembrane vesicle system, the chromatophores from the purple photosynthetic eubacterium Rhodospirillum rubrum, which integrate proteins and lipids. Through density distribution analysis of electron micrographs, we evaluated the efficacy of various lanthanoid acetates and found that triacetates of neodymium, samarium, and gadolinium exhibited similar staining effectiveness to uranyl acetate. Additionally, triacetates of praseodymium, erbium, and lutetium, followed by europium show promising results as secondary candidates. Our findings suggest that lanthanoid transition heavy metal acetates could serve as viable alternatives for electron staining in TEM, offering potential advantages over uranyl acetate.

Melatonin modulates the hypothalamic-pituitary neuroendocrine axis to regulate physiological color change in teleost fish.

Melatonin (MT) is a crucial neuroendocrine regulator of various physiological activities in vertebrates, especially in circadian or seasonal rhythm control. In the present study, the large yellow croaker (Larimichthys crocea), a marine bony fish with circadian body color change behavior, is chosen for functional investigation on teleost MT signaling systems that remain uncharacterized. All five melatonin receptors (LcMtnr1a1, LcMtnr1a2, LcMtnr1b1, LcMtnr1b2, and LcMtnr1c) were significantly activated by MT, triggering extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation through different G protein coupling signaling pathways, with exclusive Gαi-dependency for LcMtnr1a2 and LcMtnr1c, and Gαq-dependency for two LcMtnr1b paralogs, whereas LcMtnr1a1 activated Gαi and Gαs dual-dependent signaling pathways. A comprehensive model of the MT signaling system in the hypothalamic-pituitary neuroendocrine axis was further constructed based on ligand-receptor interaction analysis using single-cell RNA-seq data, as well as spatial expression patterns of Mtnrs and related neuropeptides in central neuroendocrine tissues. A novel regulatory pathway of MT/melanin-concentrating hormone (MCH) and MT/(tachykinin precursor 1 (TAC1)+corticotropin-releasing hormone (CRH))/melanocyte-stimulating hormone (MSH) was discovered that functions in chromatophore mobilization and physiological color change and was further validated by pharmacological experiments. Together, our findings define multiple intracellular signaling pathways mediated by L. crocea melatonin receptors and provide the first in-depth evidence that uncover the upstream modulating roles of the MT signaling system in the hypothalamic-pituitary neuroendocrine axis of a marine teleost species, particularly in chromatophore mobilization and physiological color change.

FOF1-ATPase Motor-Embedded Chromatophore as Drug Delivery System: Extraction, Cargo Loading Ability and Mucus Penetration Ability.

Mucosal drug delivery permits direct and prompt drug absorption, which is capable of reducing undesirable decomposition that occurs before absorption. However, mucus clearance of those mucosal drug delivery systems strongly retards their actual application. Herein, we propose chromatophore nanoparticles embedded with FOF1-ATPase motors to promote mucus penetration. The FOF1-ATPase motor-embedded chromatophores were firstly extracted from Thermus thermophilus by using a gradient centrifugation method. Then, the model drug (curcumin) was loaded onto the chromatophores. The drug loading efficiency and entrapment efficiency were optimized by using different loading approaches. The activity, motility, stability and mucus permeation of the drug-loaded chromatophore nanoparticles were thoroughly investigated. Both the in vitro and in vivo studies revealed that the FOF1-ATPase motor-embedded chromatophore successfully enhanced mucus penetration glioma therapy. This study indicates that the FOF1-ATPase motor-embedded chromatophore is a promising alternative as a mucosal drug delivery system.

Can canthaxanthin intensify the colour of the blood swordtail Xiphophorus helleri?

Colouring has a great influence on the commercialization of ornamental fish. The aim of this study was to evaluate different concentrations of canthaxanthin in the diet of the blood swordtail Xiphophorus helleri in an effort to obtain a more intense red colour. Six concentrations of canthaxanthin (0, 50, 100, 250, 400 and 600 mg kg-1 diet) were used. The experiment lasted 60 days. Fish were evaluated for increased red pigmentation through the use of photographs (performed by smartphone) and imaging applications considering the Hunter method and the CMYK and productive performance. No significant differences were observed for productive performance. The use of photographs by means of a smartphone and the use of imaging applications proved to be adequate to assess differences in colour in the species. Swordtail red pigmentation did not show significant increase regardless of canthaxanthin dosages. Varieties of ornamental fish bred for red coloration may have limits for increased colour due to the storage capacity of carotenoids by chromatophores.

Identification of a Ubiquinone-Ubiquinol Quinhydrone Complex in Bacterial Photosynthetic Membranes and Isolated Reaction Centers by Time-Resolved Infrared Spectroscopy.

Ubiquinone redox chemistry is of fundamental importance in biochemistry, notably in bioenergetics. The bi-electronic reduction of ubiquinone to ubiquinol has been widely studied, including by Fourier transform infrared (FTIR) difference spectroscopy, in several systems. In this paper, we have recorded static and time-resolved FTIR difference spectra reflecting light-induced ubiquinone reduction to ubiquinol in bacterial photosynthetic membranes and in detergent-isolated photosynthetic bacterial reaction centers. We found compelling evidence that in both systems under strong light illumination-and also in detergent-isolated reaction centers after two saturating flashes-a ubiquinone-ubiquinol charge-transfer quinhydrone complex, characterized by a characteristic band at ~1565 cm-1, can be formed. Quantum chemistry calculations confirmed that such a band is due to formation of a quinhydrone complex. We propose that the formation of such a complex takes place when Q and QH2 are forced, by spatial constraints, to share a common limited space as, for instance, in detergent micelles, or when an incoming quinone from the pool meets, in the channel for quinone/quinol exchange at the QB site, a quinol coming out. This latter situation can take place both in isolated and membrane bound reaction centers Possible consequences of the formation of this charge-transfer complex under physiological conditions are discussed.

Conversion of light into electricity in a semi-synthetic system based on photosynthetic bacterial chromatophores.

Chromatophores (Chr) from photosynthetic nonsulfur purple bacterium Rhodobacter sphaeroides immobilized onto a Millipore membrane filter (MF) and sandwiched between two semiconductor indium tin oxide (ITO) electrodes (termed ITO|Chr - MF|ITO) have been used to measure voltage (ΔV) induced by continuous illumination. The maximum ΔV was detected in the presence of ascorbate / N,N,N'N'-tetramethyl-p-phenylenediamine couple, coenzyme UQ0, disaccaride trehalose and antimycin A, an inhibitor of cytochrome bc1 complex. In doing so, the light-induced electron transfer in the reaction centers was the major source of photovoltages. The stability of the voltage signal upon prolonged irradiation (>1 h) may be due to the maintenance of a conformation that is optimal for the functioning of integral protein complexes and stabilization of lipid bilayer membranes in the presence of trehalose. Retaining ∼70 % of the original photovoltage performance on the 30th day of storage at 23 °C in the dark under air was achieved after re-injection of fresh buffer (∼40 µL) containing redox mediators into the ITO|Chr - MF|ITO system. The approach we use is easy and can be extended to other biological intact systems (cells, thylakoid membranes) capable of converting energy of light.

Intracytoplasmic-membrane development in alphaproteobacteria involves the homolog of the mitochondrial crista-developing protein Mic60.

Mitochondrial cristae expand the surface area of respiratory membranes and ultimately allow for the evolutionary scaling of respiration with cell volume across eukaryotes. The discovery of Mic60 homologs among alphaproteobacteria, the closest extant relatives of mitochondria, suggested that cristae might have evolved from bacterial intracytoplasmic membranes (ICMs). Here, we investigated the predicted structure and function of alphaproteobacterial Mic60, and a protein encoded by an adjacent gene Orf52, in two distantly related purple alphaproteobacteria, Rhodobacter sphaeroides and Rhodopseudomonas palustris. In addition, we assessed the potential physical interactors of Mic60 and Orf52 in R. sphaeroides. We show that the three α helices of mitochondrial Mic60's mitofilin domain, as well as its adjacent membrane-binding amphipathic helix, are present in alphaproteobacterial Mic60. The disruption of Mic60 and Orf52 caused photoheterotrophic growth defects, which are most severe under low light conditions, and both their disruption and overexpression led to enlarged ICMs in both studied alphaproteobacteria. We also found that alphaproteobacterial Mic60 physically interacts with BamA, the homolog of Sam50, one of the main physical interactors of eukaryotic Mic60. This interaction, responsible for making contact sites at mitochondrial envelopes, has been conserved in modern alphaproteobacteria despite more than a billion years of evolutionary divergence. Our results suggest a role for Mic60 in photosynthetic ICM development and contact site formation at alphaproteobacterial envelopes. Overall, we provide support for the hypothesis that mitochondrial cristae evolved from alphaproteobacterial ICMs and have therefore improved our understanding of the nature of the mitochondrial ancestor.

Tumor microenvironment responded naturally extracted FOF1-ATPase loaded chromatophores for antitumor therapy.

Tumor microenvironment (TME) plays an important role in the growth, invasion, and metastasis of tumor cells. The pH of TME is more acidic in solid tumors than in normal tissues. Although targeted delivery in TME has progressed, the complex and expensive construction of delivery systems has limited their application. FOF1-ATP synthase (FOF1-ATPase) is a rotation molecular motor found in bacteria, chloroplasts, and mitochondria. Here, FOF1-ATPase loaded chromatophores (chroma) isolated from thermophilic bacteria were extracted and utilized as a new delivery system targeting TME for the first time. Curcumin as model drug was successfully loaded by a filming-rehydration ultrasonic dispersion method to prepare a curcumin-loaded chroma delivery system (Cur-Chroma). The mobility and propensity distributions of Cur-Chroma reveal its specific pH-sensitive targeting driven by the transmembrane proton kinetic potential, demonstrating its distinct distribution in the TME and more favorable targeting delivery. Cellular uptake experiments indicated that Cur-Chroma entered cells through grid pathway-mediated endocytosis. In vivo studies have shown that Cur-Chroma can specifically target tumor tissue and effectively inhibit tumor growth with good safety. Curcumin's bioavailability and anti-tumor effects were significantly improved. These studies demonstrate that ATPase-loaded chromatophores are potentially ideal vehicles for anti-tumor drug delivery and have promising applications.

Mechanisms of alkali pH-shifted colour changes in squid (Uroteuthis edulis) subjected to frozen storage.

The skin discoloration of squid subjected to frozen storage negatively affects market price. In this study, various alkali treatments were investigated for effects on red granules and yellow pigments of squid skin and corresponding mechanisms were investigated at the tissue, cellular and molecular level. A significant colour improvement was observed when subjected to a pH 12 treatment, supported by decreased Δb* and increased Δa* values. Neither lower nor harsher alkali treatments than pH 12 can not obtain such results. HE staining and the UV-vis spectrum suggest that the improved red colour in skin was ascribed to the release of red pigment granules from damaged chromatophores by alkaline treatment and the release of red pigments in alkaline aqueous solutions from granules. However, based on TEM and particle size analysis, an excessive alkali treatment of pH 13 would degrade granules into smaller particles. The degradation of yellowness pigments indicated high sensitivity to alkali environments according to HPLC results. This study provides a valuable reference for improving the colour appearance of squid skin subjected to frozen storage.

Effect of Dipyridamole on Membrane Energization and Energy Transfer in Chromatophores of Rba. sphaeroides.

Effect of dipyridamole (DIP) at concentrations up to 1 mM on fluorescent characteristics of light-harvesting complexes LH2 and LH1, as well as on conditions of photosynthetic electron transport chain in the bacterial chromatophores of Rba. sphaeroides was investigated. DIP was found to affect efficiency of energy transfer from the light-harvesting complex LH2 to the LH1-reaction center core complex and to produce the long-wavelength ("red") shift of the absorption band of light-harvesting bacteriochlorophyll molecules in the IR spectral region at 840-900 nm. This shift is associated with the membrane transition to the energized state. It was shown that DIP is able to reduce the photooxidized bacteriochlorophyll of the reaction center, which accelerated electron flow along the electron transport chain, thereby stimulating generation of the transmembrane potential on the chromatophore membrane. The results are important for clarifying possible mechanisms of DIP influence on the activity of membrane-bound functional proteins. In particular, they might be significant for interpreting numerous therapeutic effects of DIP.

The physiological cost of colour change: evidence, implications and mitigations.

Animals benefit from phenotypic plasticity in changing environments, but this can come at a cost. Colour change, used for camouflage, communication, thermoregulation and UV protection, represents one of the most common plastic traits in nature and is categorised as morphological or physiological depending on the mechanism and speed of the change. Colour change has been assumed to carry physiological costs, but current knowledge has not advanced beyond this basic assumption. The costs of changing colour will shape the evolution of colour change in animals, yet no coherent research has been conducted in this area, leaving a gap in our understanding. Therefore, in this Review, we examine the direct and indirect evidence of the physiological cost of colour change from the cellular to the population level, in animals that utilise chromatophores in colour change. Our Review concludes that the physiological costs result from either one or a combination of the processes of (i) production, (ii) translocation and (iii) maintenance of pigments within the colour-containing cells (chromatophores). In addition, both types of colour change (morphological and physiological) pose costs as they require energy for hormone production and neural signalling. Moreover, our Review upholds the hypothesis that, if repetitively used, rapid colour change (i.e. seconds-minutes) is more costly than slow colour change (days-weeks) given that rapidly colour-changing animals show mitigations, such as avoiding colour change when possible. We discuss the potential implications of this cost on colour change, behaviour and evolution of colour-changing animals, generating testable hypotheses and emphasising the need for future work to address this gap.

Disproportionate effect of cationic antiseptics on the quantum yield and fluorescence lifetime of bacteriochlorophyll molecules in the LH1-RC complex of R. rubrum chromatophores.

Photosynthetic membrane complexes of purple bacteria are convenient and informative macromolecular systems for studying the mechanisms of action of various physicochemical factors on the functioning of catalytic proteins both in an isolated state and as part of functional membranes. In this work, we studied the effect of cationic antiseptics (chlorhexidine, picloxydine, miramistin, and octenidine) on the fluorescence intensity and the efficiency of energy transfer from the light-harvesting LH1 complex to the reaction center (RC) of Rhodospirillum rubrum chromatophores. The effect of antiseptics on the fluorescence intensity and the energy transfer increased in the following order: chlorhexidine, picloxydine, miramistin, octenidine. The most pronounced changes in the intensity and lifetime of fluorescence were observed with the addition of miramistin and octenidine. At the same concentration of antiseptics, the increase in fluorescence intensity was 2-3 times higher than the increase in its lifetime. It is concluded that the addition of antiseptics decreases the efficiency of the energy migration LH1 → RC and increases the fluorescence rate constant kfl. We associate the latter with a change in the polarization of the microenvironment of bacteriochlorophyll molecules upon the addition of charged antiseptic molecules. A possible mechanism of antiseptic action on R. rubrum chromatophores is considered. This work is a continuation of the study of the effect of antiseptics on the energy transfer and fluorescence intensity in chromatophores of purple bacteria published earlier in Photosynthesis Research (Strakhovskaya et al. in Photosyn Res 147:197-209, 2021).