Phosphorylation in State Transition Less Cause More Effect - DiVA portal

information om kompetensutvecklin arbetsmöjligheter, senaste nyhetern utbildning och forskning med mera

För Umeå universitet är kontakten m dig och andra alumner oerhört vikti erfarenheter du har spelar stor roll f Phosphorylation in State Transition utveckling. Less fortsatta Cause More Effect Håll kontakten! Jakob Thomas Damkjær

Gå till 0.05 (Not significant), n=27. Df

Sum Sq

Mean Sq

F value

Pr(>F)

Significance

Genotype

1

259

259

30.979

1.8E-07

***

State

1

61.403

61.403

733.737

6.22E-14

***

Sample date

1

34.445

34.445

411.597

3.39E-09

***

Prep. date

1

184

184

2.198

6.401

Genotype:State

1

1.032

1.032

1.233

2.692

Residuals

26

94.564

837

35

Phosphorylation in State Transition Less Cause More Effect

5.2. Short state transition inducing light treatment induces LHCII phosphorylation When plants were exposed to a short (5 min) state 1 or state 2 inducing light treatments an increase of ~55% in wild type and ~45% in koLhcb3 is observed (see figure 19). However in both the state 1 and state 2 induced plants there was no significant difference in the LHCII P-Thr level in between the genotypes. ANOVA analysis of the data revealed a clear significant variation originating from the state of the sample. But neither the genotype nor the interaction between state and genotype showed a significant variation (see table 5).

LHCII Phosphorylation (% Relative to Wildtype St1)

200

150

100

50

0 Wildtype Wild type St1

koLhcb3 St1

Wildtype Wild type St2 St2

koLhcb3 St2

Figure 19: Effect of Lhcb3 depletion on phosphorylation of LHCII trimers in state transition. Plants are treated with a 5 min. light treatment promoting state 1 and state 2. White bars indicate wild type and grey bars indicate koLhcb3. Signal is normalization to wild type state 1 with in-lane normalization to CP43 phosphorylation and chlorophyll content. Error bars indicate SE, n=6. Table 5: ANOVA analysis of the significance of the interaction between genotype (wild type or koLhcb3) and treatment (State 1 or State 2 light) with regard to LHCII phosphorylation. Light treatment duration: 5 min. Significance level indicated is; *** = P < 0.001; ** = P < 0.01; * = P < 0.05; ʼ = P > 0.05 (Not significant) n=6. Df

Sum Sq

Mean Sq

F value

Pr(>F)

Genotype

1

8.8949E+10

8.8949E+10

12.956

2.685

State

1

5.6141E+12

5.6141E+12

817.734

1.67E-08

Genotype:State

1

2.1797E+10

2.1797E+10

3.175

5.794

Residuals

20

1.3731e+12

6.8654E+10

Significance

***

36


III. Discussion The close connection between the state transition and phosphorylation of LHCII trimers have been accepted ever since it was found to be the case in algae. This has been generally accepted to be the case in plants, in particular since the homolog of the Chlamydomonas Lhcb kinase (Stt7) was found in higher plants and the depletion of Stn7 abolished the state transition process entirely (Bellafiore et al., 2005). The target of the kinase was discovered to be the Lhcb1 and Lhcb2 units of LHCII. Depletion of the Lhcb1/2 subunits cause a complete loss of state transition as well (Andersson et al., 2003). However, in the later years investigations have discovered cases in higher plants where it seems that the direct connection between phosphorylation and state transition may not be as tight as in algae. The koPsaH plants exhibit hyperphosphorylation, yet they do not dissociate LHCII from PSII at all. This would suggest that phosphorylation of the LHCII trimers does not directly cause the LHCII trimer to dissociate from the PSII supercomplexes (Lunde et al., 2000). In that paper it was argued that the LHCII required a docking site away from PSII prior to dissociating from PSII. This explanation - absence of state transition even when LHCII was hyperphosphorylated - is not supported by recent findings. These have discovered how the LHCII dissociating from PSII is quenched without being associated to another supercomplex (Ruban and Johnson 2009). Furthermore the LHCII associating to PSI doesnʼt seem to have the same subunit composition as those that dissociate from PSII (Jansson et al., 1997). This makes it unlikely that the inability of some LHCII trimers to associate to PSI would block the M-LHCII trimers' ability to dissociate from PSII and quench as a way to alleviate the imbalanced state photosynthetic apparatus. A lack of change of the PSII antennae size during ST induction is observed in the koPsaL mutant, which shares the HILO docking site depletion of koPsaH (Lunde et al., 2003). Even though the Fm value is unchanged the steady state fluorescence decays during state transition. While the PSII antenna is not decreased the PSII fluorescence is quenched, either photochemically or non-photochemically. If the kinetic decay during state 2 treatment is caused by a increased LHCII to PSI energy (not through HILO) this would enable PSI to reduce the PQ pool when far red light is turned on. This does not happen and the absence of a slightly increased fluorescence (usually interpreted as the return of LHCII to PSII during state 2 to state 1 transition) indicates that the PSII antennae may not be altered in functional size. Unfortunately this leaves the clear and significant state 1-to37


state 2 phenotype in this mutant unexplained. Further investigation of this phenotype is clearly needed as current models do not produce a comprehensive explanation. In high/ low state transition inducing light treatment an increase of LHCII to PSI-LHCII energy transfer can be linked to phosphorylation. This increase is located to the grana margin region supporting the thylakoid macrostructural model. However, the lamella thylakoid region were likewise found to contain more phosphorylated LHCII trimers and although there is a high content of PSI-LHCI in this region no increase in LHCII to PSI-LHCI energy transfer is detected (Tikkanen et al., 2010). In line with these results this work indicate that while phosphorylation of LHCII do indeed increase the rate of state transition, but no change in the extent of the state transition after 15 minutes of induction is observed. If there was a direct connection between the state transition and LHCII phosphorylation it would have been expected that the koLhcb3 mutant, as this is implicit in all the mechanistic models, showed an increase in state 1 to state 2 transition. If instead the rate of the state transition is dependent on the level of LHCII phosphorylation this could be explained by a change in affinity of LHCII to PSII being the determining factor to what rate the LHCII trimer can dissociate from PSII, supporting the molecular recognition mechanistic explanation. While this is the case for the 15 min. treatments it is not possible to detect an increase in the phosphorylation of the koLhcb3 LHCII trimers after 5 minutes even though the koLhcb3 plants show a significant increase in state 1 to state 2 transition. These two ST inducing treatments and the state of the photosynthetic balance would suggest that, while there is no doubt that an imbalance in the photosynthetic apparatus favouring PSII cause the redox state of PQ to become reduced and activate the Stn7 kinase and as a result LHCII is phosphorylated, a direct connection between phosphorylation of LHCII and a state transition response have not been detected in this investigation. Furthermore the level of LHCII phosphorylation does not seem to determine the rate of the state transition. This is revealed when comparing the LHCII phosphorylation and state transition extent and rate in both the 5 and 15 min. ST inducing treatments. A significant difference to wild type in ST extent was observed in koLhcb3 after 5 minutes of state inducing light treatment. But no increase relative to wild type was observed in LHCII phosphorylation. Additionally after 15 minutes of state inducing light treatment a significant 38


difference in LHCII phosphorylation was apparent but no significant change in ST extent was observed. In the 15 min. samples a significant increase in LHCII phosphorylation in koLhcb3 was observed, suggesting that the rate of state transition was increased as a result. However the 5 min. samples show there is a increased rate of state transition with an unchanged LHCII phosphorylation. As there is no relative increase in LHCII phosphorylation after 5 minutes of state transition light treatment it is not possible that an increased rate as well as extent of state transition can be caused by the phosphorylation of the LHCII. An attractive explanation of the increase in LHCII phosphorylation observed in the koLhcb3 plants is that it's simply caused by the increase in available phosphorylation sites since the Lhcb3 subunits are being replaced with Lhcb1 or Lhcb2 subunits. The Stn7 kinase that is induced as a result of the conditions that cause state transition, would phosphorylate available targets (Allen 1992). This explanation then begs the question: what does the LHCII phosphorylation do if not regulate state transition directly? An explanation could be that LHCII phosphorylation enables state transition, but does not determine the extent or the rate of the ST. In such a model unless the LHCII trimers are phosphorylated they are not able to participate in state transition. A tentative explanation of the mechanism could be that the low light intensities state transition is active at, the LHCII, that disconnects from PSII, is not able to quench unless it is phosphorylated. The phosphorylated LHCII trimers might have a decreased ∆pH requirement for activating NPQ. This would make the Stn7 kinase phosphatase the master switch in determining whether the photosynthetic conditions are most optimally acclimated using the NPQ process or the state transition process. Both Lhcb6 and Lhcb3 are involved in the binding site of the LHCII M-trimer and both mutants show an increased rate of state 1 to state 2 transition. Lhcb6 is clearly the most important partner in the binding of LHCII M-trimer as the depletion of Lhcb6 causes the main population of PSII-LHCII to display a C2S2 conformation. The relative Lhcb3 content is decreased in the koLhcb6 mutant suggesting that the LHCII trimers containing Lhcb3 unable to connect to PSII is turned over more frequently than other LHCII trimers. Not being able to bind LHCII M-trimers to PSII koLhcb6 looses almost a full third of the PSII connected antennae. Even so the state transition is not completely depleted. The LHCII Mtrimer seems to be able to bind to PSII to some extent even though the Lhcb6 subunit is 39


missing but the greatly reduced affinity of the LHCII M-trimer causes it to be an easily broken affliation. The koLhcb3 mutant is able to photosynthesize like wild type at low to medium growth light and maintain a wild type like PSII-LHCII supercomplex conformation. Nevertheless the ability to interact with the PSII Lhcb6 binding site (binding the LHCII M-trimer) seems somewhat degraded. The lower affinity of the Lhcb3 less LHCII is a posible mechanistic explanation why the koLhcb3 mutant displays an increased rate of state transition. In addition it could be the reason that koLhcb3 displays a worse ability than wild type to both alleviate a high light shift stress as well as reparing the damage that it causes. This could be explained by an incorrect gene regulation response to the light shifts. Investigations into the gene regulation caused by a state transition inducing light treatment have shown gene expression regulation to be closely linked to redox states known to originate from an inbalanced photosynthesis apparatus. Further more Stn7 have been implicated as a link between short and long time frame photoacclimation, and this might be the primary function of phosphorylating LHCII (Pesaresi et al., 2009). In such a mechanism the redox state would be used to determine if the photoacclimation is in balance. If there iss too much imbalance LHCII would be phosphorylated along with other proteins by the action of the Stn7 kinase and the next level in the regulatory network would be dependent on LHCII phosphorylation. However LHCII phosphorylation would perhaps be an inadequate sensor as the phosphatases activate during the normal daily cycle and erase the records of the stn7 activation had sensed. If such a LHCII phosphorylation sensor system existed it would probably only be useful under extreme conditions. The interdependent Lhcb3 containing LHCII trimers and Lhcb6 evolved during the colonization of the terrestrial environment (Alboresi et al., 2008). As both NPQ and state transition as part of photoacclimation machinery, that enabled the plants to better cope with the new environment, it is obviuos thay Lhcb3, Lhcb6, NPQ and ST have to be studied together to really understand the mechanistic basis of acclimation of photosynthetic light harvesting.

40


IV. Conclusions and outlook In light of the results presented in this thesis along with other evidence, the role of LHCII phosphorylation in determining the extent and rate of the state transition is not as clear cut as it once appeared. In our experimental setup we found no strong evidence that the level of LHCII phosphorylation determined the rate or the extent of the state transition. However, it would be incorrect to suggest that LHCII phosphorylation does not play an important role in state transition. However what that indirect role is I was unable to determine. Furthermore the central role of Lhcb6 in binding/interacting with the M-trimer was investigated and it was revealed how the protein is involved in many other aspects of the photoacclimation process as well. Further research would focus on trying to integrate both aspects of the non photochemical quenching and state transition in a common photoacclimation mechanistic model. Many proteins once thought to be only involved in a limited part of photoacclimation has recently been shown not be be as clear cut. Both state transition processes that was only thought to be active when NPQ was inactive utilize quenching. (Ruban and Johnson 2009) As well as proteins once thought to be primarily involved in highlight photoprotection which also serve functions involved in state transition (Kiss et al., 2008). These developements seem to merit the investigation of a model of photoacclimation that integrates the different processes (ie. NPQ, state transitions, gene regulation ect.) in a unified model. Perhaps the different processes are emergent aspects of a common photoacclimation process, which is able to balance the different mechanisms of photosynthetic light harvesting to meet the current needs of the plant.

41


V. Acknowledgements

I would like to thank my supervisor Stefan Jansson for giving me this opportunity to research photosynthesis with relatively loose reins and backing up most of the good ideas that I brought to his attention. Successes like the digital darkroom camera that enabled steamboating a flotilla of immunoblots through the lab every few days, as well as the less useful immunoblot manifold that to this day is the most expensive paperweight I have had the honor of using. Privilege is something you learn to take for granted every day you come to work at UPSC. The longer I worked there the more I forgot the hoard of privileges had required a lot of work to accumulate. The only thing that topped the privileges bestowed on us all at UPSC was the open doors the INTRO2 network provided us in a host of major laboratories across the european continent. Finally I must give thanks to my stupendously patient copyeditor; my sister Maria Damkjær.

42


VI. References Alboresi, A., Caffarri, S., Nogue, F., Bassi, R., and Morosinotto, T. (2008). In Silico and Biochemical Analysis of Physcomitrella patens Photosynthetic Antenna: Identification of Subunits which Evolved upon Land Adaptation. PLoS ONE 3, e2033. Allen, J., Bennett, J., Steinback, K., and Arntzen, C. (1981). Chloroplast protein phosphorylation couples plastoquinone redox state to distribution of excitation energy between photosystems. Nature 291, 25-29. Allen, J.F. (1992). Protein phosphorylation in regulation of photosynthesis. Biochim Biophys Acta 1098, 275-335. Allen, J.F. (2002). Plastoquinone redox control of chloroplast thylakoid protein phosphorylation and distribution of excitation energy between photosystems: discovery, background, implications. Photosyn Res 73, 139-148. Allen, J.F. (2003). State Transitions--a Question of Balance. Science 299, 1530-1532. Allen, J.F. (2005). Photosynthesis: the processing of redox signals in chloroplasts. Curr Biol 15, R929-932. Allen, J.F., and Pfannschmidt, T. (2000). Balancing the two photosystems: photosynthetic electron transfer governs transcription of reaction centre genes in chloroplasts. Philos Trans R Soc Lond, B, Biol Sci 355, 1351-1359. Allen, J.F., and Forsberg, J. (2001). Molecular recognition in thylakoid structure and function. Trends Plant Sci 6, 317-326. Anderson, J. (1999). Insights into the consequences of grana stacking of thylakoid membranes in vascular plants: a personal perspective. Functional Plant Biology 26, 625-639. Andersson, Wentworth, Walters, Howard, Ruban, Horton, and Jansson. (2003). Absence of the Lhcb1 and Lhcb2 proteins of the light-harvesting complex of photosystem II - effects on photosynthesis, grana stacking and fitness. The Plant journal : for cell and molecular biology 35, 350-361. Bailey, S., Walters, R.G., Jansson, S., and Horton, P. (2001). Acclimation of Arabidopsis thaliana to the light environment: the existence of separate low light and high light responses. Planta 213, 794-801. Barros, T., Royant, A., Standfuss, J., Dreuw, A., and Kühlbrandt, W. (2009). Crystal structure of plant light-harvesting complex shows the active, energy-transmitting state. EMBO J 28, 298-306. Bassi, R., and Dainese, P. (1992). Reorganization of thylakoid membrane lateral heterogeneity following state-I - state-II transition. In Regulation of Chloroplast Biogenesis, J.H. Argyroudiakoyunoglou, ed (New York: Plenum Press Div Plenum Publishing Corp), pp. 511-520. Bellafiore, S., Barneche, F., Peltier, G., and Rochaix, J.-D. (2005). State transitions and light adaptation require chloroplast thylakoid protein kinase STN7. Nature 433, 892-895. Ben-Shem, A., Frolow, F., and Nelson, N. (2003). Crystal structure of plant photosystem I. Nature 426, 630-635. Boekema, E.J., Van Roon, H., Van Breemen, J.F.L., and Dekker, J.P. (1999). Supramolecular organization of photosystem II and its light-harvesting antenna in partially solubilized photosystem II membranes. European Journal of Biochemistry 266, 444-452. Boekema, E.J., Hankamer, B., Bald, D., Kruip, J., Nield, J., Boonstra, A.F., Barber, J., and Rögner, M. (1995). Supramolecular structure of the photosystem II complex from green plants and cyanobacteria. Proceedings of the National Academy of Sciences of the United States of America 92, 175-179. Büchel, C., and Kühlbrandt, W. (2005). Structural differences in the inner part of Photosystem II between higher plants and cyanobacteria. Photosynthesis Research 85, 3-13. 43

Phosphorylation in State Transition Less Cause More Effect Caffarri, S., Kouril, R., Kereiche, S., Boekema, E.J., and Croce, R. (2009). Functional architecture of higher plant photosystem II supercomplexes. EMBO J 28, 3052-3063. Crouchman, S., Ruban, A., and Horton, P. (2006). PsbS enhances nonphotochemical fluorescence quenching in the absence of zeaxanthin. FEBS Lett 580, 2053-2058. Danielsson, R., Albertsson, P., Mamedov, F., and Styring, S. (2004). Quantification, of photosystem I and II in different parts of the thylakoid membrane from spinach. Biochimica et Biophysica Acta-Bioenergetics 1608, 53-61. Dekker, J.P. (2001). Supermolecular organization of photosystem II and its associated light-harvesting antenna in Arabidopsis thaliana. Eur J Biochem 268, 6020-6028. Dekker, J., and Boekema, E. (2005a). Supramolecular organization of thylakoid membrane proteins in green plants. Biochim Biophys Acta 1706, 12-39. Dekker, J.P., and Boekema, E.J. (2005b). Supramolecular organization of thylakoid membrane proteins in green plants. Biochimica et Biophysica Acta 1706, 12-39. Ganeteg, U. (2004). The light-harvesting antenna of higher plant photosystem I. Ganeteg, U., Klimmek, F., and Jansson, S. (2004). Lhca5 – an LHC-Type Protein Associated with Photosystem I. Plant Molecular Biology 54, 641-651. Guskov, A., Kern, J., Gabdulkhakov, A., Broser, M., Zouni, A., and Saenger, W. (2009). Cyanobacterial photosystem II at 2.9-A resolution and the role of quinones, lipids, channels and chloride. Nat Struct Mol Biol 16, 334-342. Haldrup, A., Naver, H., and Scheller, H. (1999). The interaction between plastocyanin and photosystem I is inefficient in transgenic Arabidopsis plants lacking the PSI-N subunit of photosystem I. Plant J 17, 689-698. Haldrup, A., Jensen, P., Lunde, C., and Scheller, H. (2001). Balance of power: a view of the mechanism of photosynthetic state transitions. Trends Plant Sci 6, 301-305. Hankamer, B., Nield, J., Zheleva, D., Boekema, E., Jansson, S., and Barber, J. (1997). Isolation and biochemical characterisation of monomeric and dimeric photosystem II complexes from spinach and their relevance to the organisation of photosystem II in vivo. Eur J Biochem 243, 422-429. Horton, A.V.R. (2000). Allosteric regulation of the light-harvesting system of photosystem II. Philosophical Transactions of the Royal Society B: Biological Sciences 355, 1361-1370. Horton, P., and Ruban, A. (2005). Molecular design of the photosystem II light-harvesting antenna: photosynthesis and photoprotection. J Exp Bot 56, 365-373. Horton, P., Wentworth, M., and Ruban, A. (2005). Control of the light harvesting function of chloroplast membranes: the LHCII-aggregation model for non-photochemical quenching. FEBS Lett 579, 4201-4206. Jansson, S., Stefánsson, H., Nyström, U., and Gustafsson, P. (1997a). Antenna protein composition of PS I and PS II in thylakoid sub-domains. BBA-Bioenergetics. Jansson, S., Stefansson, H., Nystrom, U., Gustafsson, P., and Albertsson, P.-A. (1997b). Antenna protein composition of PS I and PS II in thylakoid sub-domains. Biochimica et Biophysica Acta 1320, 297-309. Jensen, P.E., Haldrup, A., Rosgaard, L., and Scheller, H.V. (2003). Molecular dissection of photosystem I in higher plants: topology, structure and function. Physiologia Plantarum 119, 313-321.

44

Phosphorylation in State Transition Less Cause More Effect Jensen, P., Haldrup, A., Zhang, S., and Scheller, H. (2004). The PSI-O subunit of plant photosystem I is involved in balancing the excitation pressure between the two photosystems. J Biol Chem 279, 24212-24217. Kargul, J., and Barber, J. (2008). Photosynthetic acclimation: structural reorganisation of light harvesting antenna--role of redox-dependent phosphorylation of major and minor chlorophyll a/b binding proteins. FEBS J 275, 1056-1068. Keller, D., and Bustamante, C. (1986). Theory of the interaction of light with large inhomogeneous molecular aggregates. II. Psi-type circular dichroism. The Journal of Chemical Physics 84, 2972-2980. Kiss, A.Z., Ruban, A.V., and Horton, P. (2008). The PsbS protein controls the organization of the photosystem II antenna in higher plant thylakoid membranes. J Biol Chem 283, 3972-3978. Klimmek, F., Sjodin, A., Noutsos, C., Leister, D., and Jansson, S. (2006). Abundantly and Rarely Expressed Lhc Protein Genes Exhibit Distinct Regulation Patterns in Plants. Plant Physiol. 140, 793-804. Kouril, R., Zygadlo, A., Arteni, A., de Wit, C., and al., e. (2005). Structural characterization of a complex of photosystem I and light-harvesting complex II of …. Biochemistry. Kovacs, L., Damkjaer, J., Kereiche, S., Ilioaia, C., Ruban, A., Boekema, E., Jansson, S., and Horton, P. (2006). Lack of the Light-Harvesting Complex CP24 Affects the Structure and Function of the Grana Membranes of Higher Plant Chloroplasts. The Plant Cell 18, 3106. Külheim, C., Ågren, J., and Jansson, S. (2002). Rapid Regulation of Light Harvesting and Plant Fitness in the Field. Science 297, 91-93. Lavergne, J., and Trissl, H.W. (1995). Theory of fluorescence induction in photosystem II: derivation of analytical expressions in a model including exciton-radical-pair equilibrium and restricted energy transfer between photosynthetic units. Biophys J 68, 2474-2492. Liu, Z., Yan, H., Wang, K., Kuang, T., Zhang, J., Gui, L., An, X., and Chang, W. (2004). Crystal structure of spinach major light-harvesting complex at 2.72 A resolution. Nature 428, 287-292. Lunde, C., Jensen, P.E., Haldrup, A., Knoetzel, J., and Scheller, H.V. (2000). The PSI-H subunit of photosystem I is essential for state transitions in plant photosynthesis. Nature 408, 613-615. Lunde, C., Jensen, P., Rosgaard, L., Haldrup, A., Gilpin, M., and Scheller, H. (2003). Plants impaired in state transitions can to a large degree compensate for their defect. Plant Cell Physiol 44, 44-54. Malkin, R., Niyogi, K., and Bob B. Buchanan, W. (2000). Photosynthesis. In Biochemistry and molecular biology of plants, pp. 568-628. Mustardy, L., and Garab, G. (2003). Granum revisited. A three-dimensional model--where things fall into place. Trends Plant Sci 8, 117-122. Niyogi, K.K., Li, X.-P., Rosenberg, V., and Jung, H.-S. (2005). Is PsbS the site of non-photochemical quenching in photosynthesis? J Exp Bot 56, 375-382. Pesaresi, P., Hertle, A., Pribil, M., Kleine, T., Wagner, R., Strissel, H., Ihnatowicz, A., Bonardi, V., Scharfenberg, M., Schneider, A., Pfannschmidt, T., and Leister, D. (2009). Arabidopsis STN7 Kinase Provides a Link between Short- and Long-Term Photosynthetic Acclimation. The Plant Cell 21, 2402. Pascal, A., ZhenFeng, L., Broess, K., Oort, B., and al., e. (2005). Molecular basis of photoprotection and control of photosynthetic light-harvesting. Nature (London). Peter, G.F., and Thornber, J.P. (1991). Biochemical evidence that the higher plant photosystem II core complex is organized as a dimer. Plant and Cell Physiology 32, 1237-1250.

45

Phosphorylation in State Transition Less Cause More Effect Rozak, P., Seiser, R., Wacholts, W., and Wise, R. (2002). Rapid, reversible alterations in spinach thylakoid appression upon changes in light intensity. Plant Cell Environ, 421-429. Ruban, A.V., and Johnson, M.P. (2009). Dynamics of higher plant photosystem cross-section associated with state transitions. Photosynthesis Research 99, 173-183. Ruban, A., Wentworth, M., Yakushevska, A., Andersson, J., Lee, P., Keegstra, W., Dekker, J., Boekema, E., Jansson, S., and Horton, P. (2003). Plants lacking the main light-harvesting complex retain photosystem II macro-organization. Nature 421, 648-652. Scheller, H., Jensen, P., Haldrup, A., Lunde, C., and Knoetzel, J. (2001). Role of subunits in eukaryotic Photosystem I. Biochim Biophys Acta 1507, 41-60. Thidholm, E., Lindström, V., Tissier, C., Robinson, C., P. Schröder, W., and Funk, C. (2002). Novel approach reveals localisation and assembly pathway of the PsbS and PsbW proteins into the photosystem II dimer. FEBS Letters 513, 217-222. Tikkanen, M., Grieco, M., Kangasjarvi, S., and Aro, E.-M. (2010)Thylakoid Protein Phosphorylation in Higher Plant Chloroplasts Optimizes Electron Transfer under Fluctuating Light. Plant Physiol. 152, 723-735. Tikkanen, M., Nurmi, M., Suorsa, M., Danielsson, R., Mamedov, F., Styring, S., and Aro, E.-M. (2008). Phosphorylation-dependent regulation of excitation energy distribution between the two photosystems in higher plants. Biochimica et Biophysica Acta 1777, 425-432. Varotto, C., Pesaresi, P., Jahns, P., Lessnick, A., Tizzano, M., Schiavon, F., Salamini, F., and Leister, D. (2002). Single and double knockouts of the genes for photosystem I subunits G, K, and H of Arabidopsis. Effects on photosystem I composition, photosynthetic electron flow, and state transitions. Plant Physiol 129, 616-624. Wollman, F., Minai, L., and Nechushtai, R. (1999). The biogenesis and assembly of photosynthetic proteins in thylakoid membranes1. Biochim Biophys Acta 1411, 21-85. Yakushevska, A., Keegstra, W., Boekema, E., Dekker, J., Andersson, J., Jansson, S., Ruban, A., and Horton, P. (2003). The structure of photosystem II in Arabidopsis: localization of the CP26 and CP29 antenna complexes. Biochemistry 42, 608-613. Yakushevska, A., Jensen, P., Keegstra, W., van Roon, H., Scheller, H., Boekema, E., and Dekker, J. (2001). Supermolecular organization of photosystem II and its associated light-harvesting antenna in Arabidopsis thaliana. Eur J Biochem 268, 6020-6028.

46