Pathways for external alkalinization in intact and in microwounded Chara cells are differentially sensitive to wortmannin

Abstract

Proton flows across the plant cell membranes play a major role in electrogenesis and regulation of photosynthesis and ion balance. The profiles of external pH along the illuminated internodal cells of characean algae consist of alternating high- and low-pH zones that are spatially coordinated with the distribution of photosynthetic activity of chloroplasts underlying these zones. The results based on confocal laser scanning fluorescence microscopy, pH microsensors, and pulse-amplitude-modulated chlorophyll microfluorometry revealed that the coordination of H+ transport and photosynthesis is disrupted by the 2 different environmental cues (low light and wounding) and by a chemical, wortmannin interfering with the inositol phospholipid metabolism. On the one hand, the transition from moderate to low irradiance diminished the peaks in the profiles of photosystem II (PSII) quantum efficiency but did not remove the pH bands. On the other hand, the microwounding of the internode with a glass micropipette, impacting primarily the cell wall, resulted in a rapid local alkalinization of the external medium (by 2–2.5 pH units) near the cell surface, thus mimicking the appearance of natural pH bands. Despite their seeming similarity, the alkaline bands of intact cells were eliminated by wortmannin, whereas the wound-induced alkalinization was insensitive to this drug. Furthermore, the attenuation of natural pH bands in wortmannin-treated cells was accompanied by the enhancement in spatial heterogeneity of PSII efficiency and electron transport rates, which indicates the complexity of chloroplast–plasma membrane interactions. The results suggest that the light- and wound-induced alkaline areas on the cell surface are associated with different ion-transport systems.

PhenoChip: A single-cell phenomic platform for high-throughput photophysiological analyses of microalgae

“…However, cells of Symbiodinium are also adapted to very low pH; recent studies have shown that the coral host acidifies the compartment (the "symbiosome") around endosymbionts to a pH of approximately 4, which provides the most photosynthetically accessible form of C i (CO 2 ) to Symbiodinium (29). Within PhenoChip, our expectation was thus that Symbiodinium cells exposed to pH < 6.0 would show increased carbon fixation and thereby increased effective quantum yields ( II ), as previously reported in filamentous green algae (30). Instead, in both strains, we found the highest  II to occur at pH 6.8.…”

Inhibition of endosomal trafficking by brefeldin A interferes with long‐distance interaction between chloroplasts and plasma membrane transporters

“…). Consistently, wortmannin also inhibits both pH banding and endocytosis in Chara , although to a lesser extend (Foissner et al , Bulychev and Foissner ). An argument against the hypothesis that continuous renewal of transporters is required to maintain ion transport is, that ikarugamycin, another inhibitor of endocytosis, has no effect on pH banding in Chara (own unpublished data).…”

“…In previous studies, we applied wortmannin, an inhibitor of PIP3 and PIP4 kinases which induces the formation of large aggregates consisting of TGN and MVBs (Foissner et al ). Wortmannin reduced the amplitude of pH peaks (Bulychev and Foissner ) but, in contrast to BFA, had only minor effects on long‐distance transmission of photometabolites (Bulychev and Foissner , own unpublished data). This may indicate that chloroplast‐released substances bind (preferentially) to the membrane of specific organelles.…”

“…There is evidence that the intracellular turnover of membrane material might be important for the circulation of electric currents between functionally distinct regions in illuminated characean internodes and for long‐distance signaling within the cell. For example, an inhibitor of cytoplasmic vesicle trafficking, wortmannin suppressed the pH banding in Chara and eliminated a minor component in the cyclosis‐mediated changes of chlorophyll (Chl) fluorescence (Bulychev and Foissner ). The action of other inhibitors of vesicular transport, e.g.…”

“…Boot et al () observed the polar movement of indole‐3‐acetic acid (IAA) along Chara internodes at velocities comparable to or lower than the rate of cytoplasmic streaming. However, IAA transport was insensitive to inhibition of cyclosis, unlike metabolite signaling detected with pulse amplitude‐modulated microscopy (PAM) fluorometry (Bulychev and Foissner ).…”