Light-Dependent Redistribution of Ions in Suspensions of Chloroplast Thylakoid Membranes

Abstract

Ion movements associated with the pH rise that is observed upon illumination of thylakoid suspensions at low pH have been studied by a multiparameter technique. Light-dependent, dark-reversible fluxes of H+, Cl-, Na +, K + and divalent cations were monitored, together with simultaneous changes in the optical density of the suspension. Extensive uptake of Cl-and efflux of Mg2 + accompany the apparent inward movement of H + in the light. Only minor efflux of K+ is seen and Na+ appears immobile. The Cl-and Mg2+ fluxes together compensate for most of the charge transferred as H+, contributing respectively about 49% and 43% on an equivalent basis. The ratio of Cl-influx to Mg2 + efflux is variable, but usually >1.0., The Mg2 + flux can be supplanted by (1) K +flux, if the K+/Mg2+ activity ratio in the suspension is high, and (2) Ca2+ flux, if the thylakoids are equilibrated with suspending media containing Ca2+. The affinity of the divalentcation-binding sites, or carrier mechanism, is greater for Ca2 + than for Mg2 +. Schemes can be drawn up to account for the observed ion movements on the basis of either a chemical or a chemiosmotic mechanism for energy transduction in chloroplasts. In intact chloroplasts, lightdependent control of Mg2 + distribution between thylakoid and stroma could serve to regulate enzyme activities in the carbon fixation pathway, and hence photosynthesis.Ten years ago, the first description appeared (1) of the lightdriven apparent uptake of hydrogen ions by isolated thylakoids (chloroplast inner membranes). The pH rise in the suspending medium was associated with a reversible increase in sample turbidity, and with the ability of energized thylakoids to phosphorylate ADP, upon relaxation in the dark under conditions conducive to photophosphorylation.These observations have been extensively elaborated, and reviewed in detail (2, 3). The pH rise phenomenon attracts continued attention, since it has seemed to provide the most tangible evidence in favor of the chemiosmotic mechanism (4) of energy transduction. Estimates of up to 3.0 pH units have been made (5-7) for the magnitude of the H+ gradient across illuminated thylakoid membranes, and this has been construed as establishing the thermodynamic feasibility of chemiosmotic coupling.Such massive pH shifts cannot occur without concomitant flux of some ion other than H+, as required by the law of electroneutrality. It is commonly assumed that Cl-serves as a counterion, and Cl-uptake has been observed (8-10). That reported by Deamer and Packer (10) was seen under conditions favoring uptake of the phenazine methosulfate cation (11), in a manner presumably analogous to the uptake of amine cations and Cl-(12). A simultaneous demonstration of stoichiometric H+ and Cl-uptake in the absence of uncoupler has yet to be given. Indeed, Dilley and Vernon (13) Abbreviations: HEPES, N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid; PIPES, piperazine-N,N'-bis(2-ethanesulfonic acid).

Assessment of the requirement for aquaporins in the thylakoid membrane of plant chloroplasts to sustain photosynthetic water oxidation

“…Even though information about concentration on other osmolytes in the stroma (e.g., sugars) is not considered here, it is obvious that in dark-adapted chloroplasts the direction of the osmotic gradient would not favor net flux of water movement from the stroma into the thylakoid lumen. Since increase in thylakoid volume has been observed in the light, we suggest that this is due to massive water flow into its lumen following influx of Cl À ions in response to lumen acidification [53], and possibly also influx of other yet unknown osmolytes. This would mean that the direction of the osmotic gradient in the light has been reversed as compared to the dark state.…”

“…The mechanism behind the observed regulatory changes in the volume of the lumen is largely unknown. Nevertheless, there is experimental evidence for extensive light-dependent and dark-reversible ion fluxes across the thylakoid membrane, specifically Cl À uptake into the lumen, which was proposed to maintain electroneutrality during photosynthetic H + uptake [53]. Large influx or eflux of Cl À ions and other solutes may happen in very short time scales (seconds to minutes).…”

Direct Measurement of K+ Channels in Thylakoid Membranes by Incorporation of Vesicles into Planar Lipid Bilayers

“…Historically, it was assumed that the main ion counterbalancing H+ accumulation was K+, largely because of the abundance of this ion in chloroplasts (33). However, various measurements showed a clear increase upon illumination of Mg2" concentration in the stroma, the aqueous solution outside the thylakoid, with the Mg2+ ions coming from the intrathylakoidal lumen (1,4,6,12,16,23). Stromal Mg2+ concentrations rose by I to 3 mm in the light, roughly doubling dark levels of free Mg2+ ( 16,23).…”

“…' (6,12,21). Significant movements of Cl-have also been reported (12,30). Overall, it is conceivable that fluxes of other ions could be many times greater than the Mg2`movement.…”

Involvement of Na+ in Active Uptake of Pyruvate in Mesophyll Chloroplasts of Some C4 Plants

“…Cations, especially Mg2+, are reported to be transported out from thylakoid lumen to compensate for the H+ accumulation in the lumen in C3 chloroplasts (10). Although K+ and Na+ were shown to be rather immobile in C3 thylakoid preparations (10), a possibility still remains that Na+/H+ exchange occurs on the thylakoid of C4 chloroplasts.…”