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Title: The Role of Light–Dark Regulation of the Chloroplast ATP Synthase

Abstract

The chloroplast ATP synthase catalyzes the light-driven synthesis of ATP and is activated in the light and inactivated in the dark by redox-modulation through the thioredoxin system. It has been proposed that this down-regulation is important for preventing wasteful hydrolysis of ATP in the dark. To test this proposal, we compared the effects of extended dark exposure in Arabidopsis lines expressing the wild-type and mutant forms of ATP synthase that are redox regulated or constitutively active. In contrast to the predictions of the model, we observed that plants with wild-type redox regulation lost photosynthetic capacity rapidly in darkness, whereas those expressing redox-insensitive form were far more stable. To explain these results, we propose that in wild-type plants, down-regulation of ATP synthase inhibits ATP hydrolysis, leading to dissipation of thylakoid proton motive force (pmf) and subsequent inhibition of protein transport across the thylakoid through the twin arginine transporter (Tat)-dependent and Secdependent import pathways, resulting in the selective loss of specific protein complexes. By contrast, in mutants with a redox-insensitive ATP synthase, pmf is maintained by ATP hydrolysis, thus allowing protein transport to maintain photosynthetic activities for extended periods in the dark. Hence, a basal level of Tat-dependent, as well as, Sec-dependentmore » import activity, in the dark helps replenishes certain components of the photosynthetic complexes and thereby aids in maintaining overall complex activity. But, the influence of a dark pmf on thylakoid protein import, by itself, could not explain all the effects we observed in this study. For example, we also observed in wild type plants a large transient buildup of thylakoid pmf and nonphotochemical exciton quenching upon sudden illumination of dark adapted plants. Thus, we conclude that down-regulation of the ATP synthase is probably not related to preventing loss of ATP per se. Instead, ATP synthase redox regulation may be impacting a number of cellular processes such as (1) the accumulation of chloroplast proteins and/or ions or (2) the responses of photosynthesis to rapid changes in light intensity. A model highlighting the complex interplay between ATP synthase regulation and pmf in maintaining various chloroplast functions in the dark is presented.« less

Authors:
 [1];  [2];  [3];  [1];  [4];  [4];  [2];  [2]
  1. Michigan State Univ., East Lansing, MI (United States). Dept. of Energy Plant Research Lab.
  2. Michigan State Univ., East Lansing, MI (United States). Dept. of Energy Plant Research Lab., Dept. of Biochemistry and Molecular Biology
  3. Michigan State Univ., East Lansing, MI (United States). Dept. of Energy Plant Research Lab., Dept. of Cell and Molecular Biology
  4. Washington State Univ., Pullman, WA (United States). Dept. of Horticulture and Landscape Architecture
Publication Date:
Research Org.:
Washington State Univ., Pullman, WA (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Basic Energy Sciences (BES) (SC-22)
OSTI Identifier:
1392983
Grant/Contract Number:
FG02-04ER15559
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
Frontiers in Plant Science
Additional Journal Information:
Journal Volume: 8; Journal ID: ISSN 1664-462X
Publisher:
Frontiers Research Foundation
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 60 APPLIED LIFE SCIENCES; NPQ; ATP synthase; pmf; protein transport; twin arginine transporter; coupling factor; energy sensing; thioredoxin

Citation Formats

Kohzuma, Kaori, Froehlich, John E., Davis, Geoffry A., Temple, Joshua A., Minhas, Deepika, Dhingra, Amit, Cruz, Jeffrey A., and Kramer, David M.. The Role of Light–Dark Regulation of the Chloroplast ATP Synthase. United States: N. p., 2017. Web. doi:10.3389/fpls.2017.01248.
Kohzuma, Kaori, Froehlich, John E., Davis, Geoffry A., Temple, Joshua A., Minhas, Deepika, Dhingra, Amit, Cruz, Jeffrey A., & Kramer, David M.. The Role of Light–Dark Regulation of the Chloroplast ATP Synthase. United States. doi:10.3389/fpls.2017.01248.
Kohzuma, Kaori, Froehlich, John E., Davis, Geoffry A., Temple, Joshua A., Minhas, Deepika, Dhingra, Amit, Cruz, Jeffrey A., and Kramer, David M.. 2017. "The Role of Light–Dark Regulation of the Chloroplast ATP Synthase". United States. doi:10.3389/fpls.2017.01248. https://www.osti.gov/servlets/purl/1392983.
@article{osti_1392983,
title = {The Role of Light–Dark Regulation of the Chloroplast ATP Synthase},
author = {Kohzuma, Kaori and Froehlich, John E. and Davis, Geoffry A. and Temple, Joshua A. and Minhas, Deepika and Dhingra, Amit and Cruz, Jeffrey A. and Kramer, David M.},
abstractNote = {The chloroplast ATP synthase catalyzes the light-driven synthesis of ATP and is activated in the light and inactivated in the dark by redox-modulation through the thioredoxin system. It has been proposed that this down-regulation is important for preventing wasteful hydrolysis of ATP in the dark. To test this proposal, we compared the effects of extended dark exposure in Arabidopsis lines expressing the wild-type and mutant forms of ATP synthase that are redox regulated or constitutively active. In contrast to the predictions of the model, we observed that plants with wild-type redox regulation lost photosynthetic capacity rapidly in darkness, whereas those expressing redox-insensitive form were far more stable. To explain these results, we propose that in wild-type plants, down-regulation of ATP synthase inhibits ATP hydrolysis, leading to dissipation of thylakoid proton motive force (pmf) and subsequent inhibition of protein transport across the thylakoid through the twin arginine transporter (Tat)-dependent and Secdependent import pathways, resulting in the selective loss of specific protein complexes. By contrast, in mutants with a redox-insensitive ATP synthase, pmf is maintained by ATP hydrolysis, thus allowing protein transport to maintain photosynthetic activities for extended periods in the dark. Hence, a basal level of Tat-dependent, as well as, Sec-dependent import activity, in the dark helps replenishes certain components of the photosynthetic complexes and thereby aids in maintaining overall complex activity. But, the influence of a dark pmf on thylakoid protein import, by itself, could not explain all the effects we observed in this study. For example, we also observed in wild type plants a large transient buildup of thylakoid pmf and nonphotochemical exciton quenching upon sudden illumination of dark adapted plants. Thus, we conclude that down-regulation of the ATP synthase is probably not related to preventing loss of ATP per se. Instead, ATP synthase redox regulation may be impacting a number of cellular processes such as (1) the accumulation of chloroplast proteins and/or ions or (2) the responses of photosynthesis to rapid changes in light intensity. A model highlighting the complex interplay between ATP synthase regulation and pmf in maintaining various chloroplast functions in the dark is presented.},
doi = {10.3389/fpls.2017.01248},
journal = {Frontiers in Plant Science},
number = ,
volume = 8,
place = {United States},
year = 2017,
month = 7
}

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  • The rate of ADP-glucose formation from (/sup 14/C)glucose 6-phosphate and ATP by the soluble fraction of lysed chloroplasts is studied as a function of the levels of metabolites (3-phosphoglycerate, orthophosphate, hexose monophosphate, and ATP) as determined in whole chloroplasts of Spinacia oleracea in light and dark. A change in 3-phosphoglycerate concentration (from 4 to 1.4 millimolar, as in whole chloroplasts during light-dark transition) decreases the rate of ADP-glucose formation 6- to 7-fold. An increase in hexose monophosphate concentration from 2 to 6 millimolar, which occurs at the same time in whole chloroplasts, stimulates ADP-glucose formation only slightly. At constant levelsmore » of orthophosphate (4 millimolar) and 3-phosphoglycerate (4 millimolar), a change in ATP concentration from 0.2 to 1 millimolar causes an immediate 4- to 5-fold increase in the rate of ADP-glucose formation. Another significant stimulation of ADP-glucose formation (about 4- to 6-fold) is obtained after addition of dithiothreitol at high concentrations (50 millimolar). A simultaneous increase in the concentrations of 3-phosphoglycerate, ATP, and dithiothreitol, with orthophosphate and Mg/sup 2 +/ being constant at 4 and 5 millimolar, respectively, causes a 130-fold increase in the rate of ADP-glucose formation (from 0.042 to 5.49 microgram atoms carbon per milligram chlorophyll per hour). The role of these and other factors is discussed with respect to light-dark regulation of starch formation in intact chloroplasts.« less
  • The rate of ADP-glucose formation from (/sup 14/C)glucose 6-phosphate and ATP by the soluble fraction of lysed chloroplasts is studied as a function of the levels of metabolites (3-phosphoglycerate, orthophosphate, hexose monophosphate, and ATP) as determined in whole chloroplasts of Spinacia oleracea in light and dark. A change in 3-phosphoglycerate concentration (from 4 to 1.4 millimolar, as in whole chloroplasts during light-dark transition) decreases the rate of ADP-glucose formation 6- to 7-fold. An increase in hexose monophosphate concentration from 2 to 6 millimolar, which occurs at the same time in whole chloroplasts, stimulates ADP-glucose formation only slightly. At constant levelsmore » of orthophosphate (4 millimolar) and 3-phosphoglycerate (4 millimolar), a change in ATP concentration from 0.2 to 1 millimolar causes an immediate 4- to 5-fold increase in the rate of ADP-glucose formation. Another significant stimulation of ADP-glucose formation (about 4- to 6-fold) is obtained after addition of dithiothreitol at high concentrations (50 millimolar). A simultaneous increase in the concentrations of 3-phosphoglycerate, ATP, and dithiothreitol, with orthophosphate and Mg/sup 2 +/ being constant at 4 and 5 millimolar, respectively, causes a 130-fold increase in the rate of ADP-glucose formation (from 0.042 to 5.49 microgram atoms carbon per milligram chlorophyll per hour). The role of these and other factors is discussed with respect to light-dark regulation of starch formation in intact chloroplasts.« less
  • The incorporation of water oxygens into ATP made by photophosphorylation is known to be increased markedly when either Pi or ADP concentration is lowered. The present studies show a similar increase in oxygen exchange when light intensity is lowered even with ample ADP and Pi present. The number of reversals of bound ATP formation prior to release increases about 1 to about 27 in the presence of dithiothreitol and to 5 in its absence. The equilibrium of the bound reactants still favors ATP at low light intensity, as shown by measurement of the amount of bound ATP rapidly labeled frommore » (/sup 32/P)Pi during steady-state photophosphorylation. Changes observed in the interconversion rate in the absence of added thiol are likely involved in the regulation of the dark ATPase activity in the chloroplast. The interconversion rate of bound ATP to bound ADP and Pi in the presence of thiol is about the same at low and high light intensities. This rate of bound ATP formation is not sufficient, however, to account for the maximum rate of photophosphorylation. Thus, when adequate protonmotive force is present, the rate of conversion of bound ADP and Pi to bound ATP, and possibly that of bound ATP to bound ADP and Pi, must be increased, with proton translocation being completed only when bound ATP is present to be released. These observations are consistent with the predictions of the binding change mechanism with sequential participation of catalytic sites and are accommodated by a simplified general scheme for the binding change mechanism that is presented here.(ABSTRACT TRUNCATED AT 250 WORDS)« less
  • In Spinacia oleracea the kinetics of CO/sub 2/ fixation, of starch formation, and of changes in the levels of metabolites in chloroplasts and the surrounding medium has been investigated during light-dark and dark-light transitions with isolated intact chloroplasts. The internal level of orthophosphate stays constant throughout a light-dark-light cycle. The concentration of 3-phosphoglycerate in the chloroplasts is about 4 millimolar in the light and decreases in the dark within 3 minutes to about 1.6 millimolar. The level of the hexose monophosphates shows a reverse trend, increasing from about 2.2 millimolar in the light to 6 millimolar in darkness. In themore » subsequent light period both compounds reach their original levels within 2 minutes. The chloroplastic concentrations of dihydroxyacetone phosphate, of the pentose monophosphates, and of the hexose- and heptose bisphosphates remain constant at about 0.4 millimolar throughout the light-dark-light cycle. In the medium, the concentration of 3-phosphoglycerate increases and dihydroxyacetone phosphate decreases in the dark phase: this is due to an exchange of internal 3-phosphoglycerate for external dihydroxyacetone phosphate. Part of the reimported dihydroxyacetone phosphate is converted into hexose monophosphates via aldolase and fructose bisphosphatase during the first minutes of darkness. Due to the observed exchange transport reactions, the large difference between the transenvelope concentration gradients of 3-phosphoglycerate, dihydroxyacetone phosphate, and orthophosphate which exist in the light, is completely abolished after 2 to 3 minutes in the dark. The kinetics and the magnitudes of the changes of metabolite concentrations during the light-dark-light cycle are compared to the kinetics of starch formation, and their relevance for a possible light-dark regulation of starch synthesis is discussed.« less