The effect of acid rain on ultrastructure and functional parameters of photosynthetic apparatus in pea leaves

The ultrastructure and functional parameters of the photosynthetic apparatus in leaves of 14-day-old pea seedlings were studied in conditions of laboratory simulated acid rain (SAR). Pea seedlings were sprayed with an aqueous solution containing NaNO₃ (0.2 mM) and Na₂SO₄ (0.2 mM) (pH 5.6, a control variant), or with the same solution, which was acidified to pH 2.5 (acid variant). Functional characteristics were determined by chlorophyll fluorescence analysis. There was reduction in the efficiency of the photosynthetic electron transport by 25% accompanied by an increase in the quantum yield of thermal dissipation of excess light quanta by 85% without significant change in maximum quantum yield of PSII photochemistry (F_v/F_m). Ultrastructural changes in chloroplasts were revealed by transmission electron microscopy (TEM) 2 days after the SAR treatment of pea leaves. In this case, changes in the structure of the grana and heterogeneity of the thylakoids packing in the granum, namely, an increase in thylakoid intraspace widths and thickness of granal thylakoids compared to the control, were found. It was shown also that carbonic anhydrase activity was significantly inhibited in chloroplast preparations isolated from SAR-treated pea leaves. We hypothesize possible involvement of chloroplast carbonic anhydrase in thylakoid granal structure maintenance. The structural disturbances and the inhibition of photochemical activity of chloroplasts are possible consequences of the carbonic anhydrase inactivation by SAR treatment leading to violation of HCO₃ ^?–CO₂ equilibrium. The data obtained suggest that acid rains negatively affect the photosynthetic apparatus by disrupting the membrane system of the chloroplast.     

ULTRASTRUCTURAL CHANGES IN SUNFLOWER CHLOROPLASTS FOLLOWING INOCULATION WITH PSEUDOMONAS SYRINGAE PV. TAGETIS

Severe chlorosis and ultrastructural modifications of chloroplasts occur in sunflower in response to infection by Pseudomonas syringae pv. tagetis. Chlorosis became apparent within 2 days after the cotyledons of 10-day-old sunflower seedlings were inoculated with the bacteria. The first symptoms generally appeared in the center of leaves at the second node above the cotyledons. Leaves above the second node lost essentially all of their pigmentation but remained turgid and continued to expand. Grana thylakoids became dilated and separated from the granal stacks. These thylakoid membranes did not chemically breakdown as in the case in chromoplast formation or normal chloroplast senescence. Both grana and stroma thylakoid membranes coalesced to form a large membrane sheet within the plastid. The ultrastructural changes are unlike those reported to be caused by other chlorosis-inducing bacteria or chlorosis associated with normal senescence.     

Chloroplast ultrastructure in leaves of tobacco plants with the introduced gene for the acyl-lipid Δ9-desaturase from <Emphasis Type="Italic">Synechococcus vulcanus</Emphasis> at normal and low temperature

Ultrastructural changes in chloroplasts of tobacco plants (Nicotiana tabacum L.) with the introduced desC gene for the acyl-lipid Δ9-desaturase from the thermophilic cyanobacterium Synechococcus vulcanus were investigated during plant acclimation to cold. Control plants were transformed with an empty pGA482 binary vector. At optimum growth temperature, a decreased number of grana and thylakoids and an increased number of plastoglobules and their larger area were observed in transgenic plants when compared to control ones. In control plants, acclimation to cold (6 days at 10°C) resulted in the larger areas of chloroplasts and grana. These changes indicated starting cold-induced injuries manifested in swelling of the stroma and a slight decrease in the total number of thylakoids in the chloroplast. In contrast, transgenic plants responded to cold by reducing the chloroplast, granal, and plastoglobule areas. Meantime, the number of thylakoids per granum increased noticeably. The total number of thylakoids in the chloroplast increased from 123 to 203. It was concluded that expression of the acyl-lipid Δ9-desaturase gene in tobacco plants provided for the formation of the cell ultrastructure similar to one characteristic of cold-tolerant plants.     

Dynamic flexibility in the structure and function of photosystem II in higher plant thylakoid membranes: the grana enigma

Grana are not essential for photosynthesis, yet they are ubiquitous in higher plants and in the recently evolved Charaphyta algae; hence grana role and its need is still an intriguing enigma. This article discusses how the grana provide integrated and multifaceted functional advantages, by facilitating mechanisms that fine-tune the dynamics of the photosynthetic apparatus, with particular implications for photosystem II (PSII). This dynamic flexibility of photosynthetic membranes is advantageous in plants responding to ever-changing environmental conditions, from darkness or limiting light to saturating light and sustained or intermittent high light. The thylakoid dynamics are brought about by structural and organizational changes at the level of the overall height and number of granal stacks per chloroplast, molecular dynamics within the membrane itself, the partition gap between appressed membranes within stacks, the aqueous lumen encased by the continuous thylakoid membrane network, and even the stroma bathing the thylakoids. The structural and organizational changes of grana stacks in turn are driven by physicochemical forces, including entropy, at work in the chloroplast. In response to light, attractive van der Waals interactions and screening of electrostatic repulsion between appressed grana thylakoids across the partition gap and most probably direct protein interactions across the granal lumen (PSII extrinsic proteins OEEp-OEEp, particularly PsbQ-PsbQ) contribute to the integrity of grana stacks. We propose that both the light-induced contraction of the partition gap and the granal lumen elicit maximisation of entropy in the chloroplast stroma, thereby enhancing carbon fixation and chloroplast protein synthesizing capacity. This spatiotemporal dynamic flexibility in the structure and function of active and inactive PSIIs within grana stacks in higher plant chloroplasts is vital for the optimization of photosynthesis under a wide range of environmental and developmental conditions.     

CO<Subscript>2</Subscript> exchange and structural organization of chloroplasts under hypothermia in potato plants transformed with a gene for yeast invertase

Growth, CO₂ exchange, and the ultrastructure of chloroplasts were investigated in the leaves of potato plants (Solanum tuberosum L., cv. Désirée) of wild type and transformed with a gene for yeast invertase under the control of patatin class I B33 promoter (for apoplastic enzyme) grown in vitro on the Murashige and Skoog medium supplemented with 2% sucrose. At a temperature of 22°C optimal for growth, the transformed plants differed from the plants of wild type in retarded growth and a lower rate of photosynthesis as calculated per plant. On a leaf dry weight basis, photosynthesis of transformed plants was higher than in control plants. Under hypothermia (5°C), dark respiration and especially photosynthesis of transformed plants turned out to be more intense than in control material. After a prolonged exposure to low temperature (6 days at 5°C), in the plants of both genotypes, the ultrastructure of chloroplasts changed. Absolute areas of sections of chloroplasts and starch grains rose, and the area of plastoglobules decreased; in transformed plants, these changes were more pronounced. By some ultrastructural characteristics: a reduction in the cold of relative total area of sections of starch grains and plastoglobules (in percents of the chloroplast section area) and in the number of granal thylakoids (per a chloroplast section area), transformed plants turned out to be more cold resistant than wild-type plants. The obtained results are discussed in connection with changes in source-sink relations in transformed potato plants. These changes modify the balance between photosynthesis and retarded efflux of assimilates, causing an increase in the intracellular level of sugars and a rise in the tolerance to chilling.     

Green tissue within the haustorium of the dwarf mistletoe Korthalsella (Viscaceae). An ultrastructural comparison between chloroplasts of sucker and aerial stem tissues

Summary Korthalsella (Viscaceae) is a dwarf mistletoe attached to its host branch by a single haustorium. Plants are leafless with flattened or cylindrical stems that function in photosynthesis. When a fresh haustorium is cut the sucker within the host appears bright green. Transmission electron microscopy reveals that this greening is due to chloroplasts, but that their organization differs from those of the aerial stem. The three representatives of Korthalsella endemic to New Zealand were the main species investigated. In the stem, chloroplasts have short stacks of cylindrical grana interconnected by stroma thylakoids typical of normal chloroplasts. Sucker chloroplasts have a more variable organization, with most containing extensive granal stacks and poorly differentiated stroma thylakoids. These granal thylakoids exhibit extensive partitions formed by appression of adjacent membranes. Some sucker plastids also approach etioplasts in having a prominent prolamellar body from which radiate thylakoids with short partitions. Sucker chloroplasts usually contain a few large starch grains, plastoglobuli, and sometimes also a stroma centre. The extensive granal thylakoids in sucker chloroplasts of Korthalsella resemble that found in certain shade plants and tissue grown under low light conditions. Sucker chloroplasts probably have a low level of photosynthesis. This activity might provide a local source of osmotically active material used to assist transport between host and parasite.     

Phagocytosis of algal chloroplasts by digestive gland cells in the photosynthesis-capable slug Elysia timida (Mollusca, Opisthobranchia, Sacoglossa)

Parapodia of the sacoglossan slug Elysia timida were preserved by high-pressure cryofixation during feeding experiments and investigated with transmission electron microscopy. This slug has been known for its long-term retention of active chloroplasts and photosynthesis. We observed different stages of phagocytosis of chloroplast components from ingested algal food by slug digestive gland cells. Thylakoid stacks and stroma of chloroplasts were engulfed by the slug cells. In the slug cells thylakoids were surrounded by one membrane only. This membrane is interpreted as having been generated by the mollusk during phagocytosis. It is inferred to be eukaryotic in origin and unlikely, therefore, to be endowed with the translocons system ordinarily regulating import of algal gene-encoded plastid preproteins. Our structural findings suggest that chloroplast components in the slug cells are thylakoid stacks with chloroplast stroma only.     

Effects of drought and waterlogging on ultrastructure of Scots pine and Norway spruce needles

Effects of water stress on needle ultrastructure of 2-year-old Scots pine (Pinus sylvestris L.) and 5-year-old Norway spruce [Picea abies (L.) Karst.] seedlings were studied in greenhouse experiments. Drought stress was induced by leaving seedlings without watering, and waterlogging stress was produced by submerging the seedling containers in water. Needle samples for ultrastructural analyses were collected several times during the experiments, and samples for nutrient analyses at the end of the experiments. In drought stress, plasmolysis of mesophyll and transfusion parenchyma tissues, aggregation of chloroplast stroma and its separation from thylakoids and decreased size and abundance of starch grains in needles of both species were observed. The concentration of lipid bodies around the chloroplasts were detected in pine needles. Calcium and water concentrations in spruce needles were lower by the end of the experiments compared to controls. In waterlogging treatment, swelling of phloem cells in pine needles and large starch grains, slight swelling of thylakoids and increased translucency of plastoglobuli in chloroplasts of both species studied were observed. The phosphorus concentration in pine needles was higher while phosphorus, calcium and magnesium concentrations in spruce needles were lower in the waterlogging treatments compared to controls. Typical symptoms induced by drought stress, e. g. aggregation of chloroplast stroma and its separation from thylakoids, were detected, but, in waterlogging stress, ultrastructural symptoms appeared to be related to the developing nutrient imbalance of needles.     

The response of ultrastructure and function of chloroplasts from cycads to doubled CO2 concentration

This study examines the effects of doubled CO₂ concentration on the ultrastructure and function of chloroplasts from cycads and, for control from two other herbaceous angiosperms. Under a doubled CO₂ concentration condition, the chloroplast ultrastructure of the two cycads (Cycas multipinnata with a shade-type chloroplast andC. panzhihuaensis with a sun-type chloroplast) changed little: The conformation of the thylakoid membrane system kept well, and almost no starch grains accumulated. In contrast, under the same conditions the chloroplast ultrastructure of soybean and foxtail millet changed considerably, with starch grains accumulating in their chloroplasts and some of thylakoids (especially stroma thylakoid) membranes being destroyed to some degree by the more numerous and larger starch grains that accumulated in the chloroplasts. Interestingly, the changes in the ultrastructure of the chloroplasts from the two cycads was correlated with the 77K fluorescence emission spectra of their chlorophyll; i.e., the F685/F734 (PS II / PS I) ratio within the chloroplasts, which were minimal. The absorption spectrum showed decreases in the red and blue peaks. These changes in the absorption spectrum may be related to changes in the structural arrangement of the thylakoid membranes. Preliminarily, this experimental result shows that the cycads may adapt themselves to environmental changes under doubled CO₂ concentration in the coming centuries. However, more studies on this aspect are necessary.     

Insights into the complex 3-D architecture of thylakoid membranes in the unicellular cyanobacterium Cyanothece sp. ATCC 51142

In cyanobacteria and chloroplasts, thylakoids are the complex internal membrane system where the light reactions of oxygenic photosynthesis occur. In plant chloroplasts, thylakoids are differentiated into a highly interconnected system of stacked grana and unstacked stroma membranes. In contrast, in cyanobacteria, the evolutionary progenitors of chloroplasts, thylakoids do not routinely form stacked and unstacked regions, and the architecture of the thylakoid membrane systems is only now being described in detail in these organisms. We used electron tomography to examine the thylakoid membrane systems in one cyanobacterium, Cyanothece sp. ATCC 51142. Our data showed that thylakoids form a complicated branched network with a rudimentary quasi-helical architecture in this organism. A well accepted helical model of grana-stroma architecture of plant thylakoids describes an organization in which stroma thylakoids wind around stacked granum in right-handed spirals. Here we present data showing that the simplified helical architecture in Cyanothece 51142 is lefthanded in nature. We propose a model comparing the thylakoid membranes in plants and this cyanobacterium in which the system in Cyanothece 51142 is composed of non-stacked membranes linked by fret-like connections to other membrane components of the system in a limited left-handed arrangement.Key words: cyanobacteria, Cyanothece 51142, thylakoid membrane, electron tomography, chloroplast     

Effects of Nitrogen Nutrition on Nitrogen Partitioning between Chloroplasts and Mitochondria in Pea and Wheat

Nitrogen partitioning among proteins in chloroplasts and mitochondria was examined in pea (Pisum sativum L.) and wheat (Triticum aestivum L.) grown hydroponically with different nitrogen concentrations. In pea leaves, chloroplast nitrogen accounted for 75 to 80% of total leaf nitrogen. We routinely found that 8% of total ribulose-1,5-bisphosphate carboxylase/oxygenase adhered to thylakoids during preparation and could be removed with Triton X-100. With this precaution, the ratio of stroma nitrogen increased from 53 to 61% of total leaf nitrogen in response to the nitrogen supply, but thylakoid nitrogen remained almost constant around 20% of total. The changes in the activities of the stromal enzymes and electron transport in response to the nitrogen supply reflected the nitrogen partitioning into stroma and thylakoids. On the other hand, nitrogen partitioning into mitochondria was appreciably smaller than that in chloroplasts, and the ratio of nitrogen allocated to mitochondria decreased with increasing leaf-nitrogen content, ranging from 7 to 4% of total leaf nitrogen. The ratio of mitochondrial respiratory enzyme activities to leaf-nitrogen content also decreased with increasing leaf-nitrogen content. These differences in nitrogen partitioning between chloroplasts and mitochondria were reflected in differences in the rates of photosynthesis and dark respiration in wheat leaves measured with an open gas-exchange system. The response of photosynthesis to nitrogen supply was much greater than that of dark respiration, and the CO₂ compensation point decreased with increasing leaf-nitrogen content.     

Entropy-assisted stacking of thylakoid membranes

Chloroplasts in plants and some green algae contain a continuous thylakoid membrane system that is structurally differentiated into stacked granal membranes interconnected by unstacked thylakoids, the stromal lamellae. Experiments were conducted to test the hypothesis that the thermodynamic tendency to increase entropy in chloroplasts contributes to thylakoid stacking to form grana. We show that the addition of bovine serum albumin or dextran, two very different water-soluble macromolecules, to a suspension of envelope-free chloroplasts with initially unstacked thylakoids induced thylakoid stacking. This novel restacking of thylakoids occurred spontaneously, accompanied by lateral segregation of PSII from PSI, thereby mimicking the natural situation. We suggest that such granal formation, induced by the macromolecules, is partly explained as a means of generating more volume for the diffusion of macromolecules in a crowded stromal environment, i.e., greater entropy overall. This mechanism may be relevant in vivo where the stroma has a very high concentration of enzymes of carbon metabolism, and where high metabolic fluxes are required.     

Changes in structure of isolated chloroplasts ofCodium fragile andCaulerpa filiformis in response to osmotic shock and detergent treatment

Summary The ultrastructure of chloroplasts from two genera of coenocytic green algae,Codium andCaulerpa, were examined after suspension in hypotonic solution and in detergent at various concentrations. The capacity of the suspensions to carry out CO₂-dependent and ferricyanide-dependent O₂ evolution was measured under the same conditions of osmotic strength and detergent concentration.The chloroplasts in the preparations were in the form of cytoplasts and gave rates of O₂ evolution comparable with those expected from undamaged chloroplasts. Suspension in hypotonic solution depressed the rate of CO₂-dependent O₂ evolution in both species, but this was partially restored in theCodium chloroplasts when these were re-suspended in iso-osmotic solutions. Major structural changes were observed only after suspension in buffer when theCodium chloroplasts lost their outer envelope, most of their stroma, and the thylakoids became swollen.Caulerpa chloroplasts were more variable in their response and, even when suspended in buffer only, the proportion of the plastids which had lost all of their stroma and thylakoid swelling was never as common as inCodium chloroplasts. However, once suspended in hyper-osmotic medium below 700 mosmolar,Caulerpa chloroplasts could not regain their capacity for CO₂-dependent O₂ evolution.Detergent treatment removed the cytoplast membrane but not the cytoplasmic material adhering to the chloroplast envelope. High concentrations of detergent were needed to cause loss of the chloroplast envelope, loss of stromal contents and unstacking of the thylakoids.Caulerpa chloroplasts were less sensitive to detergent than those ofCodium. There was no indication that specific structures such as the thylakoid organizing body were resistant to detergent action. The results show that exposure to hypotonic solutions and to detergent results in less damage to these chloroplasts than it would to those of higher plants. It is proposed that the basis of this unusual resistance is not due to the properties of the chloroplast membranes but to the presence of material which coats the organelles during isolation. This material is likely to be identical with the sulphated xylo-mannogalactan isolated from the vacuole contents of these algae and which has the visco-elastic properties essential to allow the organelles to resist disruption by osmotic forces and disintegration by detergents.     

Further characterization of ribosome binding to thylakoid membranes

Previous work indicated more polysomes bound to pea (Pisum sativum cv Progress No. 9) thylakoids in light than in the dark, in vivo (LE Fish, AT Jagendorf 1982 Plant Physiol 69: 814-825). With isolated intact chloroplasts incubated in darkness, addition of MgATP had no effect but 24 to 74% more RNA was thylakoid-bound at pH 8.3 than at pH 7. Thus, the major effect of light on ribosome-binding in vivo may be due to higher stroma pH. In isolated pea chloroplasts, initiation inhibitors (pactamycin and kanamycin) decreased the extent of RNA binding, and elongation inhibitors (lincomycin and streptomycin) increased it. Thus, cycling of ribosomes is controlled by translation, initiation, and termination. Bound RNA accounted for 19 to 24% of the total chloroplast RNA and the incorporation of [³H]leucine into thylakoids was proportional to the amount of this bound RNA. These data support the concept that stroma ribosomes are recruited into thylakoid polysomes, which are active in synthesizing thylakoid proteins.     

New insights in thylakoid membrane organization

High-pressure freezing (HPF) in combination with freeze substitution (FS) was used to analyse changes in the structure of barley chloroplasts during the daily change of light and darkness. In contrast to conventional treatment of samples, HPF-FS revealed substantial differences in chloroplast shape, volume and ultrastructure in the light period and during darkness. While chloroplasts have an ellipsoidal shape in the light, they have an enlarged and round form during the dark period. Samples collected in the light show the typical differentiation of stroma and grana thylakoids as observed by conventional ultrastructural analyses. In chloroplasts of samples collected during the dark period, thylakoids were swollen and grana stacks to a large extent were disintegrated. Similar changes occurred when leaves in the light were treated with the uncoupler gramicidin. The results suggest that the light-dependent changes in thylakoid membrane organization are related to the light-dependent changes in the ionic milieu of the thylakoid lumen and the stroma.     

Entropy-assisted stacking of thylakoid membranes

Chloroplasts in plants and some green algae contain a continuous thylakoid membrane system that is structurally differentiated into stacked granal membranes interconnected by unstacked thylakoids, the stromal lamellae. Experiments were conducted to test the hypothesis that the thermodynamic tendency to increase entropy in chloroplasts contributes to thylakoid stacking to form grana. We show that the addition of bovine serum albumin or dextran, two very different water-soluble macromolecules, to a suspension of envelope-free chloroplasts with initially unstacked thylakoids induced thylakoid stacking. This novel restacking of thylakoids occurred spontaneously, accompanied by lateral segregation of PSII from PSI, thereby mimicking the natural situation. We suggest that such granal formation, induced by the macromolecules, is partly explained as a means of generating more volume for the diffusion of macromolecules in a crowded stromal environment, i.e., greater entropy overall. This mechanism may be relevant in vivo where the stroma has a very high concentration of enzymes of carbon metabolism, and where high metabolic fluxes are required.     

Dimorphic chloroplasts in the epidermis of Podostemoideae,a subfamily of the unique aquatic angiosperm family Podostemaceae

Plants of the Podostemoideae, a subfamily of the unique aquatic angiosperm family Podostemaceae, which are found in rapids and waterfalls of the tropics and subtropics, have two different sizes of chloroplasts in their epidermis. These small and large chloroplasts are located separately in each epidermal cell along its upper and inner tangential walls, respectively. This is the first case of the chloroplast dimorphism in a single epidermal cell of angiosperms. While the large chloroplasts have well developed starch grains, the small chloroplasts have a normal granal ultrastructure but very few starch grains. This suggests that the small chloroplasts mainly function in CO₂ uptake for photosynthesis from torrential water.     

Morphometric Analysis of Chloroplasts of Cotton Leaf and Fruiting Organs

We examined morphological and ultrastructural differences in chloroplasts of cotton leaves and the fruiting organs, bract, and capsule wall to advance our understanding of their commonly observed differences in photosynthetic efficiency. Chloroplasts from leaves were large (7.1 μm long in cross section), lens shaped with a well developed membrane system differentiated into grana and stroma lamellae that occupied the large cross-sectional area (12.3 μm²) of the organelle. A few small plastoglobuli and starch grains were scattered in the stroma region. The bract chloroplasts were correlative of leaf chloroplasts in size (6.8 μm in length) and shape with the exception that the bract chloroplasts exhibited greater thylakoid number per granum (15.8) than the leaf chloroplasts (10.5). In contrast to leaf and bract, the capsule wall chloroplasts were smaller in size (4.3 μm) and cross sectional area (6.8 μm²) than either the leaf or bract. The most intriguing feature of the capsule wall chloroplasts was its domination by large starch granules (5.3 μm²) in the stroma which filled the whole chloroplast coercing the membrane system to move towards the periphery of the organelle. Grana number and thylakoids per granum were lowest in the capsule wall chloroplasts. This revised version was published online in July 2006 with corrections to the Cover Date.     

A mechanism for the formation of inside-out membrane vesicles. Preparation of inside-out vesicles from membrane-paired randomized chloroplast lamellae

Inside-out thylakoid membrane vesicles can be isolated by aqueous polymer two-phase partition of Yeda press-fragmented spinach chloroplasts (Andersson, B. and Åkerlund, H.-E. (1978) Biochim. Biophys. Acta 503, 462–472). The mechanism for their formation has been investigated by studying the yield of inside-out vesicles after various treatments of the chloroplasts prior to fragmentation. No inside-out vesicles were isolated during phase partitioning if the chloroplasts had been destacked in a low-salt medium prior to the fragmentation. Only in those cases where the chloroplast lamellae had been stacked by cations or membrane-paired by acidic treatment did we get any yield of inside-out vesicles. Thus, the intrinsic properties of chloroplast thylakoids seem to be such that they seal into right-side out vesicles after disruption unless they are in an appressed state. This favours the following mechanism for the formation of inside-out thylakoids. After press treatment, a ruptured membrane still remains appressed with an adjacent membrane. Resealing of such an appressed membrane pair would result in an inside-out vesicle.If the compartmentation of chloroplast lamellae into appressed grana and unappressed stroma lamellae is preserved by cations before fragmentation, the inside-out vesicles are highly enriched in photosystem II. This indicates a granal origin which is consistent with the proposed model outlined. Inside-out vesicles possessing photosystem I and II properties in approximately equal proportions could be obtained by acid-induced membrane-pairing of chloroplasts which had been destacked and randomized prior to fragmentation. Since this new preparation of inside-out thylakoid vesicles also exposes components derived from the stroma lamellae it complements the previous preparation.It is suggested that fragmentation of paired membranes followed by phase partitioning should be a general method of obtaining inside-out vesicles from membranes of various biological sources.     

Remodeling of tobacco thylakoids by over-expression of maize plastidial transglutaminase

Transglutaminases (TGases, EC 2.3.2.13) are intra- and extra-cellular enzymes that catalyze post-translational modification of proteins by establishing ?-(γ-glutamyl) links and covalent conjugation of polyamines. In chloroplast it is well established that TGases specifically polyaminylate the light-harvesting antenna of Photosystem (PS) II (LHCII, CP29, CP26, CP24) and therefore a role in photosynthesis has been hypothesised (Della Mea et al. [23] and refs therein). However, the role of TGases in chloroplast is not yet fully understood. Here we report the effect of the over-expression of maize (Zea mays) chloroplast TGase in tobacco (Nicotiana tabacum var. Petit Havana) chloroplasts. The transglutaminase activity in over-expressers was increased 4 times in comparison to the wild-type tobacco plants, which in turn increased the thylakoid associated polyamines about 90%. Functional comparison between Wt tobacco and tgz over-expressers is shown in terms of fast fluorescence induction kinetics, non-photochemical quenching of the singlet excited state of chlorophyll a and antenna heterogeneity of PSII. Both in vivo probing and electron microscopy studies verified thylakoid remodeling. PSII antenna heterogeneity in vivo changes in the over-expressers to a great extent, with an increase of the centers located in grana-appressed regions (PSIIα) at the expense of centers located mainly in stroma thylakoids (PSIIβ). A major increase in the granum size (i.e. increase of the number of stacked layers) with a concomitant decrease of stroma thylakoids is reported for the TGase over-expressers.