Anna V. Komarova

Long-distance signal transmission and regulation of photosynthesis in characean cells

Photosynthetic electron transport in an intact cell is finely regulated by the structural flexibility of thylakoid membranes, existence of alternative electron-transport pathways, generation of electrochemical proton gradient, and continuous exchange of ions and metabolites between cell organelles and the cytoplasm. Long-distance interactions underlying reversible transitions of photosynthetic activity between uniform and spatially heterogeneous distributions are of particular interest. Microfluorometric studies of characean cells with the use of saturating light pulses and in combination with electrode micromethods revealed three mechanisms of distant regulation ensuring functional coordination of cell domains and signal transmission over long distances. These include: (1) circulation of electric currents between functionally distinct cell domains, (2) propagation of action potential along the cell length, and (3) continuous cyclical cytoplasmic streaming. This review considers how photosynthetic activity depends on membrane transport of protons and cytoplasmic pH, on ion fluxes associated with the electrical excitation of the plasmalemma, and on the transmission of photoinduced signals with streaming cytoplasm. Because of signal transmission with cytoplasmic flow, dynamic changes in photosynthetic activity can develop far from the point of photostimulus application and with a long delay (up to 100 s) after a light pulse stimulus is extinguished.

Proton flows across the plasma membrane in microperforated characean internodes: tonoplast injury and involvement of cytoplasmic streaming

Microperforation of characean cell wall with a glass micropipette in the absence of the tonoplast impalement was found to cause rapid alkalinization of the apoplast by 2–3 pH units, which may rigidify the cell wall structure, thus protecting the cell from further injury. A similar but a deeper insertion of a microneedle, associated with piercing the tonoplast and with an action potential generation, led to a considerable delay in the apoplast alkalinization without affecting the amplitude of the eventual increase in pH. The retardation by the mechanically elicited action potential of the incision-mediated pH transients in the apoplast contrasted sharply to the enhancement of these pH transients by the action potential triggered electrically before the microperforation. Hence, the delay of the apoplast alkalinization was not related to basic ionic mechanisms of plant action potentials. Measurements of the vacuolar pH after mechanical elicitation of an action potential indicate that the tonoplast piercing was accompanied by leakage of protons from the vacuole into the cytoplasm, which may strongly acidify the cytoplasm around the wounded area, thus collapsing the driving force for H+ influx from the medium into the cytoplasm. The lag period preceding the onset of external alkalinization was found linearly related to the duration of temporal cessation of cytoplasmic streaming. The results suggest that the delayed alkalinization of the apoplast in the region of tonoplast wounding reflects the localized recovery of the proton motive force across the plasmalemma during replacement of the acidic cytoplasm with fresh portions of unimpaired cytoplasm upon restoration of cytoplasmic streaming.

Photoregulation of photosystem II activity mediated by cytoplasmic streaming in Chara and its relation to pH bands

Chloroplasts in vivo exposed to strong light export assimilates and excess reducing power to the cytoplasm for metabolic conversions and allocation to neighboring and distant organelles. The cytoplasmic streaming, being particularly fast in characean internodes, distributes the exported metabolites from brightly illuminated cell spots to light-limited regions, which is evident from the transient increase in chlorophyll fluorescence of shaded areas in response to illumination of distant cell regions situated upstream the liquid flow. It is not yet known whether long-distance communications between anchored chloroplasts are interfered by pH banding that commonly arises in characean internodes under the action of continuous or fluctuating light. In this study, microfluorometry, pH-microsensors, and local illumination were combined to examine long-distance transport and subsequent reentry of photosynthetic metabolites, including triose phosphates, into chloroplasts of cell regions producing external alkaline and acid bands. The lateral transmission of metabolic signals between distant chloroplasts was found to operate effectively in cell areas underlying acid zones but was almost fully blocked under alkaline zones. The rates of linear electron flow in chloroplasts of these regions were nearly equal under dim background light, but differed substantially at high light when availability of CO2, rather than irradiance, was the rate-limiting factor. Different productions of assimilates by chloroplasts underlying CO2-sufficient acid and CO2-deficient alkaline zones were a cause for contrasting manifestations of long-distance transport of photosynthetic metabolites. Nonuniform cytoplasmic pH in cells exhibiting pH bands might contribute to different activities of metabolic translocators under high and low pH zones.

Influence of light on the apoplastic pH in microwounded cells of Chara corallina

Microscopic wounding of plant cell walls by pathogens or by feeding insects triggers the defense responses, including a sharp rise in pH at the cell surface (pHo). Using internodal cells of Chara corallina Klein ex Willd., we show here that the elevated pHo in the area of cell wall microincision decreases in darkness and increases on illumination. These pHo changes occurred specifically in cell areas affected by microincision and were lacking in intact areas with active pHotosynthesis (acid zones). Localized illumination of a remote cell region located upstream the cytoplasmic flow at a 1.5-mm distance from the analyzed area also caused a transient increase in pHo in the area of microwounding but had no such effect in unwounded cell regions having weakly acidic pHo. Apparently, the increase in pHo after wounding is mediated by a metabolite released from illuminated chloroplasts, which is transported with the cytoplasmic flow for long distances. The transient pHo increase in the area of cell wall incision after illumination of a distant cell region coincided with a temporal increase in chlorophyll fluorescence F’. This implies the concurrent influence of the transported reductant (presumably NADH) on light emission of chloroplasts and on the H+ flow across the plasmalemma. We suppose that the alkalinization of cell surface in the area of microincision arises from H+ consumption in the apoplast in association with the transmembrane electron transport from cytoplasmic reducing equivalents to molecular oxygen.

Lateral transport of photosynthetically active intermediate at rest and after excitation of Chara cells

Cytoplasmic streaming is vital for plant cells; however, its relation to cell functions remains largely undisclosed. Microfluorometry of chloroplasts in vivo and measurements of cell surface pH under localized illumination of cell regions located upstream the cytoplasmic flow, at a distance of few millimeters from the analyzed area, is a new means to reveal the role of liquid flow for signal transmission in large cells, such as internodes of characean algae. Properties of photoinduced signals transmitted along the cell can be clarified by comparing the effects of pointed illumination under conditions of continuous and briefly arrested cytoplasmic flow. Chlorophyll fluorescence measurements with the use of saturation pulse method showed that excitation-induced cessation of cytoplasmic streaming, concomitant with the period of localized illumination, caused a significant delay and deceleration of the lateral transmission of the photoinduced signal and, in addition, diminished the peak of maximal fluorescence Fm′ in the cell response to propagated signals. The relative extent of the peak suppression was small in cell regions producing light-dependent external alkaline zones and increased substantially for cell regions with slightly acidic external pH. These and other results indicate the possible role of cytoplasmic pH in controlling chlorophyll fluorescence and photosynthetic activity in vivo. When the period of streaming cessation coincided with localized illumination, the velocity of cytoplasmic flow recovered slower than after arrest of the flow without additional illumination. The results are promising for further analysis of regulatory and protective functions of cytoplasmic streaming in photosynthesizing plant cells.