The Arabidopsis Thylakoid Chloride Channel AtCLCe Functions in Chloride Homeostasis and Regulation of Photosynthetic Electron Transport

Abstract

Chloride ions can be translocated across cell membranes through Cl− channels or Cl−/H+ exchangers. The thylakoid-located member of the Cl− channel CLC family in Arabidopsis thaliana (AtCLCe) was hypothesized to play a role in photosynthetic regulation based on the initial photosynthetic characterization of clce mutant lines. The reduced nitrate content of Arabidopsis clce mutants suggested a role in regulation of plant nitrate homeostasis. In this study, we aimed to further investigate the role of AtCLCe in the regulation of ion homeostasis and photosynthetic processes in the thylakoid membrane. We report that the size and composition of proton motive force were mildly altered in two independent Arabidopsis clce mutant lines. Most pronounced effects in the clce mutants were observed on the photosynthetic electron transport of dark-adapted plants, based on the altered shape and associated parameters of the polyphasic OJIP kinetics of chlorophyll a fluorescence induction. Other alterations were found in the kinetics of state transition and in the macro-organization of photosystem II supercomplexes, as indicated by circular dichroism measurements. Pre-treatment with KCl but not with KNO3 restored the wild-type photosynthetic phenotype. Analyses by transmission electron microscopy revealed a bow-like arrangement of the thylakoid network and a large thylakoid-free stromal region in chloroplast sections from the dark-adapted clce plants. Based on these data, we propose that AtCLCe functions in Cl− homeostasis after transition from light to dark, which affects chloroplast ultrastructure and regulation of photosynthetic electron transport.

An update on the regulation of photosynthesis by thylakoid ion channels and transporters in Arabidopsis

“…Nevertheless, the mechanism is probably similar to that of nigericin, as this eliminates the ΔpH without affecting ΔΨ by exchanging K + with H + . Interestingly, the PMF partitioning phenotype of kea3 mutants is observed only in low light conditions (90–160 µmol photons m −2 s −1 ; Armbruster et al ), whereas in the case of clce and vccn1 mutants, this is observed regardless of light intensity (Duan et al , Herdean et al , Herdean et al ). Conversely, the phenotype of tpk3 ‐silenced lines was only seen in plants grown in light at 90 µmol photons m −2 s −1 (Carraretto et al ).…”

“…The clce mutants displayed higher steady state of NPQ than the wild‐type at all tested light intensities without altering the induction phase, and despite a reduced PMF partitioning to the ΔpH (Herdean et al ). Slower NPQ relaxation and lower ETR in the clce mutants than in the wild‐type were observed upon transition from light to dark, suggesting involvement of this transport protein in achieving a ‘resting state’ of the light‐dependent photosynthetic reactions.…”

“…Duan et al () did not analyze the ultrastructure of thylakoids from the atbest1 mutants, but reported a larger thylakoid lumen in the atbest ( atbest1atbest2 ) double mutant. The clce chloroplasts and thylakoid network had a peculiar bow‐like shape leaving a larger thylakoid‐free stromal space in dark‐adapted plants, but were indistinguishable from the wild‐type in light‐adapted plants (Herdean et al ). The kea3 chloroplasts did not exhibit morphological defects, but those in the kea1kea2kea3 triple mutant were fewer and had reduced number of thylakoids as compared with the double mutants of envelope‐localized homologous K + /H + antiporters KEA1 and KEA2 (Kunz et al ).…”

Hacking the thylakoid proton motive force for improved photosynthesis: modulating ion flux rates that control proton motive force partitioning into Δ ψ and ΔpH

“…In any case, the balancing of Δψ/ΔpH, and its kinetics, are dependent on the regulation of counter-ion homeostasis in the chloroplast (34), and recent work from several labs has identified putative components of these ion homeostatic machineries, including a thylakoid potassium channel (35), a K + /H + antiporter (36), as well as transporters for other charged species (37)(38)(39)(40)(41). There are also indications that the Δψ/ΔpH balance is controlled by the synthesis of membrane permeable weak bases, such as putrescene, that effectively increase the proton buffering capacity of the lumen (42).…”

Oolong tea extract as a substitute for uranyl acetate in staining of ultrathin sections based on examples of animal tissues for transmission electron microscopy

“…Uranyl acetate (UA) was first introduced by Watson () as an electron stain for transmission electron microscopy (TEM) observations. The method involves staining the ultrathin sections with UA followed by lead salts (Pb) and has since been conventionally applied in TEM of a wide variety of samples (Reynolds, ; Sato et al ., ; Miller et al ., ; Nakakoshi et al ., ; Carpentier et al ., ; He et al ., ), due to the unrivalled image quality (Bai et al ., ; Akita et al ., ; Bardelcikova et al ., ; Fernandez‐Avila et al ., ; Herdean et al ., ; Ota et al ., ; Sobala et al ., ). However, because of the instability, high toxicity and radioactivity of UA, it has become increasingly difficult to obtain it due to strict regulations on internationally controlled nuclear materials even for scientific research purposes (Miller et al ., ).…”