Mobility of the IsiA Chlorophyll-binding Protein in Cyanobacterial Thylakoid Membranes

We are using fluorescence recovery after photobleaching (FRAP) to probe the dynamics of thylakoid membranes in vivo in cells of the cyanobacterium Synechococcus sp. PCC7942. We have shown previously that the light-harvesting phycobilisomes diffuse quite rapidly on the thylakoid membrane surface. However, the photosystem II core complexes appear completely immobile. This raises the possibility that all of the membrane integral protein complexes in the thylakoid membrane are locked into a rather rigid array. Alternatively, it is possible that photosystem II is specifically anchored in the membrane, with other membrane proteins able to diffuse around it. We have now resolved this question by studying the diffusion of a second integral membrane protein, the IsiA chlorophyll-binding protein. IsiA is induced under iron starvation and some other stress conditions. In iron-stressed cyanobacterial cells, a high proportion of chlorophyll fluorescence comes from IsiA. This makes it straightforward to examine the diffusion of IsiA by FRAP. We find that the complex is mobile with a mean diffusion coefficient of ∼3 × 10–11 cm2 s–1. Thus it is clear that some thylakoid membrane proteins are mobile and that there must be a specific anchor that prevents photosystem II diffusion. We discuss the implications for the structure and function of the cyanobacterial thylakoid membrane. We are using fluorescence recovery after photobleaching (FRAP) to probe the dynamics of thylakoid membranes in vivo in cells of the cyanobacterium Synechococcus sp. PCC7942. We have shown previously that the light-harvesting phycobilisomes diffuse quite rapidly on the thylakoid membrane surface. However, the photosystem II core complexes appear completely immobile. This raises the possibility that all of the membrane integral protein complexes in the thylakoid membrane are locked into a rather rigid array. Alternatively, it is possible that photosystem II is specifically anchored in the membrane, with other membrane proteins able to diffuse around it. We have now resolved this question by studying the diffusion of a second integral membrane protein, the IsiA chlorophyll-binding protein. IsiA is induced under iron starvation and some other stress conditions. In iron-stressed cyanobacterial cells, a high proportion of chlorophyll fluorescence comes from IsiA. This makes it straightforward to examine the diffusion of IsiA by FRAP. We find that the complex is mobile with a mean diffusion coefficient of ∼3 × 10–11 cm2 s–1. Thus it is clear that some thylakoid membrane proteins are mobile and that there must be a specific anchor that prevents photosystem II diffusion. We discuss the implications for the structure and function of the cyanobacterial thylakoid membrane. In plants and cyanobacteria the light reactions of photosynthesis are mediated by protein complexes in the thylakoid membranes. Considerable information is now available on the structure and function of these complexes (1Jordan P. Fromme P. Witt H.T. Klukas O. Saenger W. Krauss N. Nature. 2001; 411: 909-917Crossref PubMed Scopus (2082) Google Scholar, 2Kurisu G. Zhang H. Smith J.L. Cramer W.A. Science. 2003; 302: 1009-1014Crossref PubMed Scopus (607) Google Scholar, 3Ferreira K.N. Iverson T.M. Maghlaoui K. Barber J. Iwata S. Science. 2004; 303: 1831-1838Crossref PubMed Scopus (2872) Google Scholar). A full understanding of thylakoid membrane function will also require knowledge of the dynamics of the membrane in vivo. The diffusion of proteins in the thylakoid membrane is likely to play a crucial role in processes including photosynthetic electron transport, the regulation of photosynthesis, and the assembly and turnover of the photosynthetic complexes (4Kirchhoff H. Mukherjee U. Galla H.-J. Biochemistry. 2002; 41: 4872-4882Crossref PubMed Scopus (163) Google Scholar, 5Allen J.F. Forsberg J. Trends Plant Sci. 2001; 6: 317-326Abstract Full Text Full Text PDF PubMed Scopus (356) Google Scholar, 6Baena-Gonzalez E. Barbato R. Aro E.-M. Planta. 1999; 208: 196-204Crossref Scopus (99) Google Scholar). We have been using the cyanobacterium Synechococcus sp. PCC7942 (Synechococcus 7942) as a model system for investigating thylakoid membrane dynamics in vivo (7Sarcina M. Tobin M.J. Mullineaux C.W. J. Biol. Chem. 2001; 276: 46830-46834Abstract Full Text Full Text PDF PubMed Scopus (109) Google Scholar, 8Sarcina M. Murata N. Tobin M.J. Mullineaux C.W. FEBS Lett. 2003; 553: 295-298Crossref PubMed Scopus (35) Google Scholar, 9Aspinwall C.L. Sarcina M. Mullineaux C.W. Photosynth. Res. 2004; 79: 179-187Crossref PubMed Scopus (36) Google Scholar). Our technique of choice is fluorescence recovery after photobleaching (FRAP), 1The abbreviations used are: FRAP, fluorescence recovery after photobleaching; PSI, photosystem I; PSII, photosystem II. which can be used to observe the diffusion of fluorescently tagged membrane components. A laser-scanning confocal microscope is used to bleach fluorescence in a small region of the cell, and the subsequent changes in the bleaching pattern give quantitative information on the diffusion of the fluorophore (10Kubitscheck U. Wedekind P. Peters R. Biophys. J. 1994; 67: 948-956Abstract Full Text PDF PubMed Scopus (71) Google Scholar, 11Mullineaux C.W. Sarcina M. Trends Plant Sci. 2002; 7: 237-240Abstract Full Text Full Text PDF PubMed Scopus (42) Google Scholar). Synechococcus 7942 has a simple, regular thylakoid membrane organization that makes it ideal for such studies (11Mullineaux C.W. Sarcina M. Trends Plant Sci. 2002; 7: 237-240Abstract Full Text Full Text PDF PubMed Scopus (42) Google Scholar), and we have developed a one-dimensional variant of FRAP that has enabled us to measure the diffusion of several thylakoid membrane components in vivo (11Mullineaux C.W. Sarcina M. Trends Plant Sci. 2002; 7: 237-240Abstract Full Text Full Text PDF PubMed Scopus (42) Google Scholar, 12Mullineaux C.W. Tobin M.J. Jones G.R. Nature. 1997; 390: 421-424Crossref Scopus (194) Google Scholar, 13Mullineaux C.W. J. Exp. Bot. 2004; 55: 1207-1211Crossref PubMed Scopus (24) Google Scholar). Our previous FRAP studies have shown the mobility of some thylakoid membrane components, including a fluorescently tagged lipid analogue (8Sarcina M. Murata N. Tobin M.J. Mullineaux C.W. FEBS Lett. 2003; 553: 295-298Crossref PubMed Scopus (35) Google Scholar) and the light-harvesting phycobilisomes, which diffuse rather rapidly on the membrane surface (7Sarcina M. Tobin M.J. Mullineaux C.W. J. Biol. Chem. 2001; 276: 46830-46834Abstract Full Text Full Text PDF PubMed Scopus (109) Google Scholar, 9Aspinwall C.L. Sarcina M. Mullineaux C.W. Photosynth. Res. 2004; 79: 179-187Crossref PubMed Scopus (36) Google Scholar, 12Mullineaux C.W. Tobin M.J. Jones G.R. Nature. 1997; 390: 421-424Crossref Scopus (194) Google Scholar). We have looked previously at the diffusion of only one integral membrane protein complex. This is the photosystem II reaction center, which is amenable to FRAP measurements because its chlorophylls fluoresce significantly at room temperature. Surprisingly, photosystem II appears completely immobile even over long time scales (7Sarcina M. Tobin M.J. Mullineaux C.W. J. Biol. Chem. 2001; 276: 46830-46834Abstract Full Text Full Text PDF PubMed Scopus (109) Google Scholar), a situation that is very unusual for a membrane protein (14Zhang F. Lee G.M. Jacobson K. BioEssays. 1993; 15: 579-588Crossref PubMed Scopus (84) Google Scholar). Two explanations, outlined below, are possible. The first possibility is that photosystem II is specifically anchored in the membrane. In eukaryotic plasma membranes this situation can arise because of the binding of membrane proteins to components of the cytoskeleton (14Zhang F. Lee G.M. Jacobson K. BioEssays. 1993; 15: 579-588Crossref PubMed Scopus (84) Google Scholar). In cyanobacterial thylakoid membranes it is more plausible that photosystem II is rendered immobile by oligomerization into rows of PSII dimers (15Olive J. Ajlani G. Astier C. Recouvreur M. Vernotte C. Biochim. Biophys. Acta. 1997; 1319: 275-282Crossref Scopus (63) Google Scholar) and/or because of the presence of a large domain in the thylakoid lumen (16Zouni A. Witt H.-T. Kern J. Fromme P. Krauss N. Saenger W. Orth P. Nature. 2001; 409: 739-743Crossref PubMed Scopus (1776) Google Scholar). The lumen appears to be an extremely crowded environment (17Albertsson P.-A.̊. Trends Plant Sci. 2001; 6: 349-354Abstract Full Text Full Text PDF PubMed Scopus (289) Google Scholar), and, thus, it is possible that protein diffusion in the lumen is greatly restricted. A second possibility would be that all of the integral membrane protein complexes in the thylakoid membrane are locked into a rather rigid array. Such a situation would be very different from that normally found in eukaryotic plasma membranes (10Kubitscheck U. Wedekind P. Peters R. Biophys. J. 1994; 67: 948-956Abstract Full Text PDF PubMed Scopus (71) Google Scholar, 14Zhang F. Lee G.M. Jacobson K. BioEssays. 1993; 15: 579-588Crossref PubMed Scopus (84) Google Scholar). However, thylakoid membranes have a different lipid composition and an exceptionally high ratio of protein to lipid (18Szalontai B. Nishiyama Y. Gombos Z. Murata N. Biochim. Biophys. Acta. 2000; 1509: 409-419Crossref PubMed Scopus (121) Google Scholar). Thylakoid membrane lipids are mobile, although their diffusion is slower than that typically found in eukaryotic plasma membranes (8Sarcina M. Murata N. Tobin M.J. Mullineaux C.W. FEBS Lett. 2003; 553: 295-298Crossref PubMed Scopus (35) Google Scholar). The lipids could, however, be percolating through a rigid matrix of proteins. In the present study we resolve this question by observing the diffusion of a second integral membrane thylakoid protein. This is the IsiA protein, which is induced under iron starvation or under oxidative stress conditions (19Michel K.-P. Pistorius E.K. Physiol. Plant. 2004; 120: 36-50Crossref PubMed Scopus (130) Google Scholar). IsiA (also called CP43′) is a chlorophyll-binding protein whose sequence shows some similarity with that of the CP43 protein of photosystem II (20Burnap R.L. Troyan T. Sherman L.A. Plant Physiol. 1993; 103: 893-902Crossref PubMed Scopus (164) Google Scholar). During iron deficiency this protein is expressed at a high level and is present in the cells in at least two forms. Some IsiA appears to be free in the membrane, where it may act as a chlorophyll reservoir (19Michel K.-P. Pistorius E.K. Physiol. Plant. 2004; 120: 36-50Crossref PubMed Scopus (130) Google Scholar, 20Burnap R.L. Troyan T. Sherman L.A. Plant Physiol. 1993; 103: 893-902Crossref PubMed Scopus (164) Google Scholar). Some IsiA is bound to photosystem I as a ring of IsiA subunits surrounding the photosystem I trimer (21Boekema E.J. Hifney A. Yakushevska A.E. Piotrowski M. Keegstra W. Berry S. Michel K.P. Pistorius E.K. Kruip J. Nature. 2001; 412: 745-748Crossref PubMed Scopus (278) Google Scholar, 22Bibby T.S. Nield J. Barber J. Nature. 2001; 412: 743-745Crossref PubMed Scopus (298) Google Scholar). The association of IsiA with photosystem II is also a possibility, because there is evidence that IsiA can quench excess PSII excitation (23Sandström S. Park Y.I. Öquist G. Gustafsson P. Photochem. Photobiol. 2001; 74: 431-437Crossref PubMed Scopus (79) Google Scholar). IsiA is characterized by strong chlorophyll fluorescence at ∼680–685 nm (20Burnap R.L. Troyan T. Sherman L.A. Plant Physiol. 1993; 103: 893-902Crossref PubMed Scopus (164) Google Scholar, 21Boekema E.J. Hifney A. Yakushevska A.E. Piotrowski M. Keegstra W. Berry S. Michel K.P. Pistorius E.K. Kruip J. Nature. 2001; 412: 745-748Crossref PubMed Scopus (278) Google Scholar, 22Bibby T.S. Nield J. Barber J. Nature. 2001; 412: 743-745Crossref PubMed Scopus (298) Google Scholar, 23Sandström S. Park Y.I. Öquist G. Gustafsson P. Photochem. Photobiol. 2001; 74: 431-437Crossref PubMed Scopus (79) Google Scholar), most of which is likely to come from the IsiA that is free in the membrane. The fluorescence of the IsiA that is bound to photosystem I is strongly quenched by efficient energy transfer to the chlorophylls of the photosystem I core complex (22Bibby T.S. Nield J. Barber J. Nature. 2001; 412: 743-745Crossref PubMed Scopus (298) Google Scholar, 24Andrizhiyevskaya E.G. Schwabe T.M.E. Germano M. D'Haene S. Kruip J. van Grondelle R. Dekker J.P. Biochim. Biophys. Acta. 2002; 1556: 265-272Crossref PubMed Scopus (86) Google Scholar). The high fluorescence from IsiA makes it amenable to FRAP studies. Here we show that at least the highly fluorescent pool of IsiA is freely mobile in the membrane. We discuss the implications for thylakoid membrane structure and function. Growth of Cells—Wild-type cells of Synechococcus sp. PCC 7942 were grown in BG11 medium (25Castenholz R.W. Methods Enzymol. 1988; 167: 68-93Crossref Scopus (350) Google Scholar) supplemented with 10 mm NaHCO3. Liquid cultures were grown in an orbital shaking incubator at 30 °C with white illumination at ∼10 microeinsteins m–2 s–1. To induce IsiA expression, cells were harvested by centrifugation and washed and inoculated in BG11 Liquid cultures were grown for at least under iron S. Park Y.I. Öquist G. Gustafsson P. Physiol. Plant. 2002; PubMed Scopus Google Scholar). were at room with an were with a with a excitation at and the excitation and were fluorescence were using at a chlorophyll of in were to the of the at fluorescence were at using a at a chlorophyll of were into and by into were to the because their are at 30 °C in a from with illumination from an of were from at nm in a The using a a were from using the coefficient of W.A. Biochim. Biophys. Acta. Scopus Google Scholar) FRAP were using a laser-scanning confocal microscope with a The using a A used with a confocal with a light of than were by in of were in a with a the surface. The at 30 °C using a were by the confocal in the in a of × cells in the were for FRAP a the microscope to the and the confocal in the for to bleach a the A of at were using A one-dimensional bleaching the long of the the in the for on the (11Mullineaux C.W. Sarcina M. Trends Plant Sci. 2002; 7: 237-240Abstract Full Text Full Text PDF PubMed Scopus (42) Google Scholar, 12Mullineaux C.W. Tobin M.J. Jones G.R. Nature. 1997; 390: 421-424Crossref Scopus (194) Google Scholar, 13Mullineaux C.W. J. Exp. Bot. 2004; 55: 1207-1211Crossref PubMed Scopus (24) Google Scholar). The fluorescence from the and the bleaching to a using to a of the bleach were by bleach time using a one-dimensional diffusion as previously (11Mullineaux C.W. Sarcina M. Trends Plant Sci. 2002; 7: 237-240Abstract Full Text Full Text PDF PubMed Scopus (42) Google Scholar, 12Mullineaux C.W. Tobin M.J. Jones G.R. Nature. 1997; 390: 421-424Crossref Scopus (194) Google Scholar, 13Mullineaux C.W. J. Exp. Bot. 2004; 55: 1207-1211Crossref PubMed Scopus (24) Google Scholar). of can be induced by iron starvation and typically in a of the chlorophyll by greatly fluorescence at nm (20Burnap R.L. Troyan T. Sherman L.A. Plant Physiol. 1993; 103: 893-902Crossref PubMed Scopus (164) Google Scholar, S. Park Y.I. Öquist G. Gustafsson P. Physiol. Plant. 2002; PubMed Scopus Google Scholar). of iron starvation we a in the chlorophyll from to nm room fluorescence from and The excitation is and are on the of In cells, the comes from the chlorophylls of PSII at starvation in greatly chlorophyll fluorescence and a in the from to nm fluorescence for iron-stressed and In cells, the comes from PSII and In the iron-stressed cells, is by the at nm are of the of IsiA in Synechococcus 7942 (20Burnap R.L. Troyan T. Sherman L.A. Plant Physiol. 1993; 103: 893-902Crossref PubMed Scopus (164) Google Scholar, S. Park Y.I. Öquist G. Gustafsson P. Physiol. Plant. 2002; PubMed Scopus Google Scholar). is clear from at room with chlorophyll most fluorescence comes from PSII in cells from IsiA in iron-stressed iron starvation to of the phycobilisomes and to bleaching of the cells (19Michel K.-P. Pistorius E.K. Physiol. Plant. 2004; 120: 36-50Crossref PubMed Scopus (130) Google Scholar, 20Burnap R.L. Troyan T. Sherman L.A. Plant Physiol. 1993; 103: 893-902Crossref PubMed Scopus (164) Google Scholar). However, at the we used iron-stressed cells of chlorophylls and The of phycobilisomes by the at after iron the cells of phycobilisomes for of Synechococcus 7942 at room with chlorophyll excitation have been to for of Synechococcus 7942 at with chlorophyll excitation have been to from at of the iron-stressed cells photosynthetic we the of on a the were of for iron-stressed cells and of for the iron-stressed cells of photosynthetic electron transport, although their a than that of the FRAP on cyanobacterial cells, most fluorescence at room comes from photosystem II the excitation light is at a by chlorophyll rather than by the FRAP measurements with chlorophyll excitation and in the on the mobility of photosystem II C.W. Tobin M.J. Jones G.R. Nature. 1997; 390: 421-424Crossref Scopus (194) Google Scholar). We have shown previously that photosystem II appears completely immobile even on a time of in the cyanobacterium C.W. Tobin M.J. Jones G.R. Nature. 1997; 390: 421-424Crossref Scopus (194) Google Scholar) and that photosystem II diffusion can be on time scales in Synechococcus 7942 (7Sarcina M. Tobin M.J. Mullineaux C.W. J. Biol. Chem. 2001; 276: 46830-46834Abstract Full Text Full Text PDF PubMed Scopus (109) Google Scholar). shows a long FRAP on a of Synechococcus 7942 at a by chlorophyll We a the by the confocal in the the of the (11Mullineaux C.W. Sarcina M. Trends Plant Sci. 2002; 7: 237-240Abstract Full Text Full Text PDF PubMed Scopus (42) Google Scholar, 12Mullineaux C.W. Tobin M.J. Jones G.R. Nature. 1997; 390: 421-424Crossref Scopus (194) Google Scholar). This a whose is C.W. Tobin M.J. Jones G.R. Nature. 1997; 390: 421-424Crossref Scopus (194) Google Scholar, 13Mullineaux C.W. J. Exp. Bot. 2004; 55: 1207-1211Crossref PubMed Scopus (24) Google Scholar) with a of of the fluorescence at the of the to bleaching in the of the The show recovery of the bleach the time of the the light-harvesting phycobilisomes are there is fluorescence recovery on a time of a (7Sarcina M. Tobin M.J. Mullineaux C.W. J. Biol. Chem. 2001; 276: 46830-46834Abstract Full Text Full Text PDF PubMed Scopus (109) Google Scholar, 9Aspinwall C.L. Sarcina M. Mullineaux C.W. Photosynth. Res. 2004; 79: 179-187Crossref PubMed Scopus (36) Google Scholar, 12Mullineaux C.W. Tobin M.J. Jones G.R. Nature. 1997; 390: 421-424Crossref Scopus (194) Google Scholar). The bleach in by fluorescence and as under The bleach as the of the fluorescence and after 30 it there recovery of the bleach even after 30 and, diffusion of photosystem II The fluorescence recovery that would be for a of in the FRAP shown in can be by using the one-dimensional diffusion C.W. Tobin M.J. Jones G.R. Nature. 1997; 390: 421-424Crossref Scopus (194) Google Scholar) and the of the A diffusion coefficient of cm2 would to fluorescence recovery in the of the bleach in 30 a diffusion coefficient of × cm2 would to recovery in 30 We can be that fluorescence recovery after 30 and, that the diffusion coefficient for PSII is × s–1. FRAP on IsiA is strongly most of the fluorescence at room comes from IsiA chlorophyll is to a first the confocal fluorescence with chlorophyll excitation under these conditions show the of IsiA. This us to FRAP to probe the mobility of IsiA. We cells of Synechococcus 7942 in for FRAP measurements because this to of photosynthetic function C.L. Sarcina M. Mullineaux C.W. Photosynth. Res. 2004; 79: 179-187Crossref PubMed Scopus (36) Google Scholar). of the diffusion coefficient are more in cells C.W. Tobin M.J. Jones G.R. Nature. 1997; 390: 421-424Crossref Scopus (194) Google Scholar). In these however, we to because a proportion of the iron-stressed cells were shows from a FRAP sequence for an iron-stressed with excitation at A and recovery of the bleach can be on a time of diffusion of IsiA. IsiA diffusion were from such as previously C.L. Sarcina M. Mullineaux C.W. Photosynth. Res. 2004; 79: 179-187Crossref PubMed Scopus (36) Google Scholar, 12Mullineaux C.W. Tobin M.J. Jones G.R. Nature. 1997; 390: 421-424Crossref Scopus (194) Google Scholar, 13Mullineaux C.W. J. Exp. Bot. 2004; 55: 1207-1211Crossref PubMed Scopus (24) Google Scholar). IsiA diffusion were quite in all of the cells The mean diffusion coefficient × 10–11 cm2 of an immobile fluorescent and fluorescence recovery appears after ∼3 we that all of the highly fluorescent IsiA is The FRAP in this were by the cells with light by chlorophyll a the from chlorophyll a in the region of the In cells of Synechococcus fluorescence under these conditions is by PSII In iron-stressed cells fluorescence is and is by fluorescence from IsiA we can FRAP to the mobility of PSII in cells and IsiA in iron-stressed The IsiA present in the thylakoid membranes of iron-stressed cells is likely to be present in several free IsiA with (21Boekema E.J. Hifney A. Yakushevska A.E. Piotrowski M. Keegstra W. Berry S. Michel K.P. Pistorius E.K. Kruip J. Nature. 2001; 412: 745-748Crossref PubMed Scopus (278) Google Scholar, 22Bibby T.S. Nield J. Barber J. Nature. 2001; 412: 743-745Crossref PubMed Scopus (298) Google Scholar), and also IsiA with PSII, where it may act as a of excess excitation (23Sandström S. Park Y.I. Öquist G. Gustafsson P. Photochem. Photobiol. 2001; 74: 431-437Crossref PubMed Scopus (79) Google Scholar). IsiA is with reaction its fluorescence is strongly quenched because of energy transfer (22Bibby T.S. Nield J. Barber J. Nature. 2001; 412: 743-745Crossref PubMed Scopus (298) Google Scholar, 24Andrizhiyevskaya E.G. Schwabe T.M.E. Germano M. D'Haene S. Kruip J. van Grondelle R. Dekker J.P. Biochim. Biophys. Acta. 2002; 1556: 265-272Crossref PubMed Scopus (86) Google Scholar). fluorescence measurements in iron-stressed cells on the pool of IsiA that is free in the membrane rather than specifically to reaction this pool of free IsiA a fluorescent probe that us to the to which an integral membrane protein is free to diffuse in the thylakoid membrane. on the cyanobacterium that PSII is completely immobile with diffusion even on long time scales C.W. Tobin M.J. Jones G.R. Nature. 1997; 390: 421-424Crossref Scopus (194) Google Scholar). The present study shows that the situation is in cells of Synechococcus diffusion is even after 30 that the diffusion coefficient for PSII must be × cm2 This situation is quite unusual for a membrane protein. quantitative for the mobility of membrane proteins in vivo come from eukaryotic plasma membranes. In these proteins typically have diffusion in the of to cm2 (14Zhang F. Lee G.M. Jacobson K. BioEssays. 1993; 15: 579-588Crossref PubMed Scopus (84) Google Scholar). Some proteins have diffusion slower than because are anchored or by the cytoskeleton (14Zhang F. Lee G.M. Jacobson K. BioEssays. 1993; 15: 579-588Crossref PubMed Scopus (84) Google Scholar). proteins with diffusion × cm2 are as immobile (14Zhang F. Lee G.M. Jacobson K. BioEssays. 1993; 15: 579-588Crossref PubMed Scopus (84) Google Scholar), and PSII into this This raises the question as to the of photosystem II is a specific of this complex or all of the integral membrane proteins in the thylakoid membrane are locked into a rigid array. FRAP measurements on iron-stressed cells show and recovery of the bleach on a time of a chlorophyll fluorescence in these cells is by IsiA we that IsiA is mobile with a diffusion coefficient of ∼3 × 10–11 cm2 s–1. PSII fluorescence in iron-stressed cells is with IsiA we PSII immobile under these and we have information on the mobility of other complexes such as with a eukaryotic plasma membrane protein, IsiA diffusion is (14Zhang F. Lee G.M. Jacobson K. BioEssays. 1993; 15: 579-588Crossref PubMed Scopus (84) Google Scholar). This may be a of the very crowded environment of the thylakoid membrane with its of protein complexes C.W. J. Plant Physiol. 1999; Scopus Google Scholar) and very high ratio of protein to lipid (18Szalontai B. Nishiyama Y. Gombos Z. Murata N. Biochim. Biophys. Acta. 2000; 1509: 409-419Crossref PubMed Scopus (121) Google Scholar). it is clear that under these conditions the integral thylakoid membrane proteins are all locked into a rigid matrix diffusion. function in iron-stressed cells that there is of photosynthetic the thylakoid membranes of the iron-stressed cells are likely to be as with protein as the thylakoid membranes of The mobility of IsiA that the of PSII has a specific rather than a of all thylakoid membrane proteins. The PSII complex is than this for such an in diffusion possibility is that PSII may be by large oligomerization because it is found as rows of dimers in cyanobacterial thylakoid membranes (15Olive J. Ajlani G. Astier C. Recouvreur M. Vernotte C. Biochim. Biophys. Acta. 1997; 1319: 275-282Crossref Scopus (63) Google Scholar). Alternatively, it is possible that PSII is by and/or specific in the thylakoid PSII has a large domain on the of the membrane of the complex and proteins (16Zouni A. Witt H.-T. Kern J. Fromme P. Krauss N. Saenger W. Orth P. Nature. 2001; 409: 739-743Crossref PubMed Scopus (1776) Google Scholar). IsiA has only in the lumen (20Burnap R.L. Troyan T. Sherman L.A. Plant Physiol. 1993; 103: 893-902Crossref PubMed Scopus (164) Google Scholar). The lumen is crowded with protein (17Albertsson P.-A.̊. Trends Plant Sci. 2001; 6: 349-354Abstract Full Text Full Text PDF PubMed Scopus (289) Google Scholar), it is possible that PSII may be by its In the most IsiA and the CP43 protein of PSII is that the of CP43 is greatly in IsiA (20Burnap R.L. Troyan T. Sherman L.A. Plant Physiol. 1993; 103: 893-902Crossref PubMed Scopus (164) Google Scholar). that the of this of IsiA the thylakoid membrane system and that this be for a function as a chlorophyll the assembly of reaction (20Burnap R.L. Troyan T. Sherman L.A. Plant Physiol. 1993; 103: 893-902Crossref PubMed Scopus (164) Google Scholar). Our that thylakoid membrane proteins can be mobile, at least have This is possible for the assembly and turnover of the A immobile of thylakoid proteins would that all and of the integral membrane protein complexes would have to in and be mediated only by proteins. is evidence to that the of of the reaction may in the plasma membrane rather than the thylakoid membrane E. B. R. F. B. Sci. U. S. A. 2001; PubMed Scopus Google Scholar). The by which be to the thylakoid membranes is because the of the plasma and thylakoid membranes is clear C.W. J. Plant Physiol. 1999; Scopus Google Scholar). However, that it is plausible that or reaction diffuse the thylakoid membrane system to their Alternatively, proteins such as which is to be in the PSII P. S. O. Mullineaux C.W. C. Plant 2003; 15: PubMed Scopus Google Scholar), diffuse to turnover and in We for including the of the used for the FRAP We also for