Storage of light-driven transthylakoid proton motive force as an electric field (Δψ) under steady-state conditions in intact cells of Chlamydomonasreinhardtii

Abstract

Proton motive force (pmf) is physiologically stored as either a DeltapH or a membrane potential (Deltapsi) across bacterial and mitochondrial energetic membranes. In the case of chloroplasts, previous work (Cruz et al. 2001, Biochemistry 40: 1226-1237) indicates that Deltapsi is a significant fraction of pmf, in vivo, and in vitro as long as the activities of counterions are relatively low. Kinetic analysis of light-induced changes in the electrochromic shift (ECS) in intact leaves was consistent with these observations. In this work, we took advantage of the spectroscopic properties of the green alga, Chlamydomonas reinhardtii, to demonstrate that light-driven Deltapsi was stored in vivo over the hours time scale. Analysis of the light-induced ECS kinetics suggested that the steady-state Deltapsi in 400 micromol photons m(-2) s(-1) red light was between 20 and 90 mV and that this represented about 60% of the light-induced increase in pmf. By extrapolation, it was surmised that about half of total (basal and light-induced) pmf is held as Deltapsi. It is hypothesized that Deltapsi is stabilized either by maintaining low chloroplast ionic strength or by active membrane ion transporters. In addition to the strong implications for regulation of photosynthesis by the xanthophyll cycle, these results imply that pmf partitioning is important across a wide range of species.

Acclimation of Chlamydomonas reinhardtii to Different Growth Irradiances

“…When needed, 3 mM n-propyl gallate was added before O 2 evolution measurement to inhibit PTOX enzymatic activity (36). PSI/PSII ratio, trans-thylakoid protonmotive force (PMF), ⌬pH formation, and variation in the electric field (⌬⌿) upon illumination with different light intensities were monitored in the different acclimated cells measuring the carotenoid electro-chromic shift (ECS) with a Joliot-type spectrophotometer (Bio-Logic SAS JTS-10) as described previously (37,38). Maximal state transition induction was measured in the different acclimated cells as described previously (39).…”

The Liverwort, Marchantia, Drives Alternative Electron Flow Using a Flavodiiron Protein to Protect PSI

“…The ECS signal is considered an intrinsic optical voltmeter that responds rapidly to changes in the electrical potential across the thylakoid membrane (Witt, 1979) and can be utilized in a noninvasive spectroscopic measurement (Baker et al, 2007;Bailleul et al, 2010;Klughammer et al, 2013;Johnson and Ruban, 2014). The thylakoid membrane potential during photosynthesis is defined as the total rapid (less than 1 s) change in the ECS signal (ECSt) upon rapidly switching off AL from steady state, which reflects the light-dark difference in proton motive force, and includes two components: transmembrane differences in the concentration of protons (ΔpH) and in the electric field (ΔC; Sacksteder and Kramer, 2000;Cruz et al, 2001Cruz et al, , 2005Baker et al, 2007). In this study, the values of ECSt were normalized by dividing the magnitude of ECS decay in dark-interval relaxation kinetics (DIRK) analysis by the magnitude of ECS induced by a 10-ms single-turnover flash (Klughammer et al, 2013).…”

Nonphotochemical Quenching of Chlorophyll Fluorescence inChlamydomonas reinhardtii

“…Inversion of the membrane potential upon switching the light off is assumed to stem from fast H + flux from the CF 0 /F 1 ATPase complex. Thus the amplitude of the inverted membrane potential can be taken as a growing function of the size of the light-induced ∆pH (52,59). performances (54).…”