Etioplasts Research Articles

De-etiolation is Almost Colour Blind: the Study of Photosynthesis Awakening Under Blue and Red Light.

The synthesis and assembly of functioning photosynthetic complexes in chloroplasts developing from etioplasts during the de-etiolation of angiosperm seedlings are imperative for the plant's autotrophic lifestyle. This study compared de-etiolation process under monochromatic red or blue light of equal photon flux density during a 24-hour illumination period of etiolated Arabidopsis seedlings. The aim was to elucidate the impact of these light wavelength on the etioplast-to-chloroplast transformation and the initiation of light-dependent photosynthetic reactions. Both treatments lead to the formation of functional young chloroplasts; however, the etioplast-to-chloroplast transition and the assembly of photosynthetic complexes occurred unevenly, with individual steps tuned by red or blue light. Ultrastructural analysis suggested faster prolamellar bodies disassembly under blue light, while low temperature fluorescence studies indicated a slower transformation of protochlorophyllide to chlorophyllide, and chlorophyll a, under these conditions. Red light further promoted the synthesis of chlorophyll b and LHCII antenna proteins. However, the efficiency of antennae in dissipating excess absorbed energy was higher for seedlings de-etiolated under blue light; the maximum quantum yield of the photosystem II reached 0.81 after 24-hour de-etiolation, equivalent to mature plants. Blue light seemed to enhance the development of well-functioning photosystems (I and II) and antennae. These findings are important for gaining a deeper understanding of photoreceptor regulation of de-etiolation and for utilizing selected light regimes to improve crop yield.

Membrane protein provision controls prothylakoid biogenesis in tobacco etioplasts.

The cytochrome b559 heterodimer is a conserved component of photosystem II whose physiological role in photosynthetic electron transfer is enigmatic. A particularly puzzling aspect of cytochrome b559 has been its presence in etiolated seedlings, where photosystem II is absent. Whether or not the cytochrome has a specific function in etioplasts is unknown. Here, we have attempted to address the function of cytochrome b559 by generating transplastomic tobacco (Nicotiana tabacum) plants that overexpress psbE and psbF, the plastid genes encoding the two cytochrome b559 apoproteins. We show that strong overaccumulation of the PsbE apoprotein can be achieved in etioplasts by suitable manipulations of the promoter and the translation signals, while the cytochrome b559 level is only moderately elevated. The surplus PsbE protein causes striking ultrastructural alterations in etioplasts; most notably, it causes a condensed prolamellar body and a massive proliferation of prothylakoids, with multiple membrane layers coiled into spiral-like structures. Analysis of plastid lipids revealed that increased PsbE biosynthesis strongly stimulated plastid lipid biosynthesis, suggesting that membrane protein abundance controls prothylakoid membrane biogenesis. Our data provide evidence for a structural role of PsbE in prolamellar body formation and prothylakoid biogenesis, and indicate that thylakoid membrane protein abundance regulates lipid biosynthesis in etioplasts.

Molecular insights into the differences between cyanobacterial and plant LPORs and prolamellar body formation: In vitro studies

Light-dependent protochlorophyllide oxidoreductase (LPOR) has captivated the interest of the research community for decades. One reason is the photocatalytic nature of the reaction catalyzed by the enzyme, and the other is the involvement of LPOR in the formation of a paracrystalline lattice called a prolamellar body (PLB) that disintegrates upon illumination, initiating a process of photosynthetic membrane formation.In this paper, we have integrated three traditional methods previously employed to study the properties of the enzyme: molecular biology, spectroscopy, and electron microscopy. We found that for cyanobacterial LPOR, substrates binding appears to be independent of lipids, with membrane interaction primarily affecting the enzyme post-reaction, with MGDG and PG having opposite effects on SynPOR. In contrast, plant isoforms exhibit sequence alterations, rendering the enzyme effective in substrate binding mainly in the presence of anionic lipids, depending on residues at positions 122, 312, and 318. Moreover, we demonstrated that the interaction with MGDG could initially serve as enhancement of the substrate specificity towards monovinyl-protochlorophyllide (Pchlide). We have shown that the second LPOR isoforms of eudicots and monocots accumulated mutations that made these variants less and more dependent on anionic lipids, respectively. Finally, we have shown that in the presence of Pchlide, NADP+, and the lipids, plant but not cyanobacterial LPOR homolog remodel membranes into the cubic phase. The cubic phase is preserved if samples supplemented with NADP + are enriched with NADPH.The results are discussed in the evolutionary context, and the model of PLB formation is presented.

Dynamic adaptation of the extremophilic red microalga Cyanidioschyzon merolae to high nickel stress

The order of Cyanidiales comprises seven acido-thermophilic red microalgal species thriving in hot springs of volcanic origin characterized by extremely low pH, moderately high temperatures and the presence of high concentrations of sulphites and heavy metals that are prohibitive for most other organisms. Little is known about the physiological processes underlying the long-term adaptation of these extremophiles to such hostile environments. Here, we investigated the long-term adaptive responses of a red microalga Cyanidioschyzon merolae, a representative of Cyanidiales, to extremely high nickel concentrations. By the comprehensive physiological, microscopic and elemental analyses we dissected the key physiological processes underlying the long-term adaptation of this model extremophile to high Ni exposure. These include: (i) prevention of significant Ni accumulation inside the cells; (ii) activation of the photoprotective response of non-photochemical quenching; (iii) significant changes of the chloroplast ultrastructure associated with the formation of prolamellar bodies and plastoglobuli together with loosening of the thylakoid membranes; (iv) activation of ROS amelioration machinery; and (v) maintaining the efficient respiratory chain functionality. The dynamically regulated processes identified in this study are discussed in the context of the mechanisms driving the remarkable adaptability of C. merolae to extremely high Ni levels exceeding by several orders of magnitude those found in the natural environment of the microalga. The processes identified in this study provide a solid basis for the future investigation of the specific molecular components and pathways involved in the adaptation of Cyanidiales to the extremely high Ni concentrations.

Anionic lipids facilitate membrane development and protochlorophyllide biosynthesis in etioplasts.

Dark-germinated angiosperm seedlings develop chloroplast precursors called etioplasts in cotyledon cells. Etioplasts develop lattice membrane structures called prolamellar bodies (PLBs), where the chlorophyll intermediate protochlorophyllide (Pchlide) forms a ternary complex with NADPH and light-dependent NADPH:Pchlide oxidoreductase (LPOR). The lipid bilayers of etioplast membranes are mainly composed of galactolipids, which play important roles in membrane-associated processes in etioplasts. Although etioplast membranes also contain 2 anionic lipids, phosphatidylglycerol (PG) and sulfoquinovosyldiacylglycerol (SQDG), their roles are unknown. To determine the roles of PG and SQDG in etioplast development, we characterized etiolated Arabidopsis (Arabidopsis thaliana) mutants deficient in PG and SQDG biosynthesis. A partial deficiency in PG biosynthesis loosened the lattice structure of PLBs and impaired the insertion of Mg2+ into protoporphyrin IX, leading to a substantial decrease in Pchlide content. Although a complete lack of SQDG biosynthesis did not notably affect PLB formation and Pchlide biosynthesis, lack of SQDG in addition to partial PG deficiency strongly impaired these processes. These results suggested that PG is required for PLB formation and Pchlide biosynthesis, whereas SQDG plays an auxiliary role in these processes. Notably, PG deficiency and lack of SQDG oppositely affected the dynamics of LPOR complexes after photoconversion, suggesting different involvements of PG and SQDG in LPOR complex organization. Our data demonstrate pleiotropic roles of anionic lipids in etioplast development.

Heterologous codA Gene Expression Leads to Mitigation of Salt Stress Effects and Modulates Developmental Processes.

Transgenic tobacco plants overexpressing the choline oxidase gene from A. globiformis showed an increase in resistance at the level of primary and secondary biosynthesis of metabolites, removing the damage characteristic of salinity and stabilizing the condition of plants. We used 200 mM NaCl, which inhibits the growth of tobacco plants at all stages of development. Leaves of transgenic and wild-type (WT) plants Nicotiána tabácum were used for biochemical, cytological and molecular biological analysis. However, for transgenic lines cultivated under normal conditions (without salinity), we noted juvenile characteristics, delay in flowering, and slowing down of development, including the photosynthetic apparatus. This caused changes in the amount of chlorophyll, a delay in the plastid grana development with the preservation of prolamellar bodies. It also caused changes in the amount of sugars and indirectly downstream processes. A significant change in the activity of antioxidant enzymes and a change in metabolism is probably compensated by the regulation of a number of genes, the expression level of which was also changed. Thus, the tolerance of transgenic tobacco plants to salinity, which manifested itself as a result of the constitutive expression of codA, demonstrates an advantage over WT plants, but in the absence of salinity, transgenic plants did not have such advantages due to juvenilization.

Electron tomography of prolamellar bodies and their transformation into grana thylakoids in cryofixed Arabidopsis cotyledons.

The para-crystalline structures of prolamellar bodies (PLBs) and light-induced etioplast-to-chloroplast transformation have been investigated via electron microscopy. However, such studies suffer from chemical fixation artifacts and limited volumes of 3D reconstruction. Here, we examined Arabidopsis thaliana cotyledon cells by electron tomography (ET) to visualize etioplasts and their conversion into chloroplasts. We employed scanning transmission ET to image large volumes and high-pressure freezing to improve sample preservation. PLB tubules were arranged in a zinc blende-type lattice-like carbon atoms in diamonds. Within 2 h after illumination, the lattice collapsed from the PLB exterior and the disorganized tubules merged to form thylakoid sheets (pre-granal thylakoids), which folded and overlapped with each other to create grana stacks. Since the nascent pre-granal thylakoids contained curved membranes in their tips, we examined the expression and localization of CURT1 (CURVATURE THYLAKOID1) proteins. CURT1A transcripts were most abundant in de-etiolating cotyledon samples, and CURT1A was concentrated at the PLB periphery. In curt1a etioplasts, PLB-associated thylakoids were swollen and failed to form grana stacks. In contrast, PLBs had cracks in their lattices in curt1c etioplasts. Our data provide evidence that CURT1A is required for pre-granal thylakoid assembly from PLB tubules during de-etiolation, while CURT1C contributes to cubic crystal growth in the dark.

SPIRE-a software tool for bicontinuous phase recognition: application for plastid cubic membranes.

Bicontinuous membranes in cell organelles epitomize nature’s ability to create complex functional nanostructures. Like their synthetic counterparts, these membranes are characterized by continuous membrane sheets draped onto topologically complex saddle-shaped surfaces with a periodic network-like structure. Their structure sizes, (around 50–500 nm), and fluid nature make transmission electron microscopy (TEM) the analysis method of choice to decipher their nanostructural features. Here we present a tool, Surface Projection Image Recognition Environment (SPIRE), to identify bicontinuous structures from TEM sections through interactive identification by comparison to mathematical “nodal surface” models. The prolamellar body (PLB) of plant etioplasts is a bicontinuous membrane structure with a key physiological role in chloroplast biogenesis. However, the determination of its spatial structural features has been held back by the lack of tools enabling the identification and quantitative analysis of symmetric membrane conformations. Using our SPIRE tool, we achieved a robust identification of the bicontinuous diamond surface as the dominant PLB geometry in angiosperm etioplasts in contrast to earlier long-standing assertions in the literature. Our data also provide insights into membrane storage capacities of PLBs with different volume proportions and hint at the limited role of a plastid ribosome localization directly inside the PLB grid for its proper functioning. This represents an important step in understanding their as yet elusive structure–function relationship.

Curvature thylakoid 1 proteins modulate prolamellar body morphology and promote organized thylakoid biogenesis in Arabidopsis thaliana

The term "de-etiolation" refers to the light-dependent differentiation of etioplasts to chloroplasts in angiosperms. The underlying process involves reorganization of prolamellar bodies (PLBs) and prothylakoids into thylakoids, with concurrent changes in protein, lipid, and pigment composition, which together lead to the assembly of active photosynthetic complexes. Despite the highly conserved structure of PLBs among land plants, the processes that mediate PLB maintenance and their disassembly during de-etiolation are poorly understood. Among chloroplast thylakoid membrane-localized proteins, to date, only Curvature thylakoid 1 (CURT1) proteins were shown to exhibit intrinsic membrane-bending capacity. Here, we show that CURT1 proteins, which play a critical role in grana margin architecture and thylakoid plasticity, also participate in de-etiolation and modulate PLB geometry and density. Lack of CURT1 proteins severely perturbs PLB organization and vesicle fusion, leading to reduced accumulation of the light-dependent enzyme protochlorophyllide oxidoreductase (LPOR) and a delay in the onset of photosynthesis. In contrast, overexpression of CURT1A induces excessive bending of PLB membranes, which upon illumination show retarded disassembly and concomitant overaccumulation of LPOR, though without affecting greening or the establishment of photosynthesis. We conclude that CURT1 proteins contribute to the maintenance of the paracrystalline PLB morphology and are necessary for efficient and organized thylakoid membrane maturation during de-etiolation.

Glucose Induces Thylakoid Formation and Upregulates Green Pigment Contents in Complete Dark Culture of the Angiosperm Pachiramacrocarpa

In addition to angiosperms, most plants are able to synthesize chlorophyll (Chl)-generating green tissues in total darkness. In this study, 140 plants of the angiosperm Pachira macrocarpa were divided into five groups. Among them, one group was grown for 2 weeks under natural light conditions, whereas the others were grown in complete darkness (0 μmol m−2 s−1). Dark-grown plants were then treated with 0~6% glucose for another 8 weeks. The budding and greening ratios, ultrastructure of chloroplasts (ChlPs) of newly developed leaves, and green pigment contents of pre-illuminated mature and young leaves, and totally dark-grown newly developed leaves were measured. Results showed that glucose inhibited the budding and promoted the greening of newly developed leaves. Pre-illuminated mature and young leaves were able to synthesize green pigments during the 2 weeks of dark adaption. Dark-grown newly developed leaves contained high levels of green pigments at 2 and 3 weeks after budding. Green pigments of glucose-fed newly developed leaves had increased, whereas they had decreased in control leaves. In addition, ChlPs of dark-grown glucose-fed newly developed leaves contained both giant grana and prolamellar bodies (PLBs), usually found in shade plants and etiolated seedlings, respectively. The higher the glucose concentration was, the greater the numbers of grana, thylakoids, and PLBs. Glucose increased the green pigment contents and grana formation in newly developed leaves in a dark condition, and the mechanisms are discussed.

The Role of Membranes and Lipid-Protein Interactions in the Mg-Branch of Tetrapyrrole Biosynthesis.

Chlorophyll (Chl) is essential for photosynthesis and needs to be produced throughout the whole plant life, especially under changing light intensity and stress conditions which may result in the destruction and elimination of these pigments. All steps of the Mg-branch of tetrapyrrole biosynthesis leading to Chl formation are carried out by enzymes associated with plastid membranes. Still the significance of these protein-membrane and protein-lipid interactions in Chl synthesis and chloroplast differentiation are not very well-understood. In this review, we provide an overview on Chl biosynthesis in angiosperms with emphasis on its association with membranes and lipids. Moreover, the last steps of the pathway including the reduction of protochlorophyllide (Pchlide) to chlorophyllide (Chlide), the biosynthesis of the isoprenoid phytyl moiety and the esterification of Chlide are also summarized. The unique biochemical and photophysical properties of the light-dependent NADPH:protochlorophyllide oxidoreductase (LPOR) enzyme catalyzing Pchlide photoreduction and located to peculiar tubuloreticular prolamellar body (PLB) membranes of light-deprived tissues of angiosperms and to envelope membranes, as well as to thylakoids (especially grana margins) are also reviewed. Data about the factors influencing tubuloreticular membrane formation within cells, the spectroscopic properties and the in vitro reconstitution of the native LPOR enzyme complexes are also critically discussed.

Molecular landscape of etioplast inner membranes in higher plants

Etioplasts are photosynthetically inactive plastids that accumulate when light levels are too low for chloroplast maturation. The etioplast inner membrane consists of a paracrystalline tubular lattice and peripheral, disk-shaped membranes, respectively known as the prolamellar body and prothylakoids. These distinct membrane regions are connected into one continuous compartment. To date, no structures of protein complexes in or at etioplast membranes have been reported. Here, we used electron cryo-tomography to explore the molecular membrane landscape of pea and maize etioplasts. Our tomographic reconstructions show that ATP synthase monomers are enriched in the prothylakoids, and plastid ribosomes in the tubular lattice. The entire tubular lattice is covered by regular helical arrays of a membrane-associated protein, which we identified as the 37-kDa enzyme, light-dependent protochlorophyllide oxidoreductase (LPOR). LPOR is the most abundant protein in the etioplast, where it is responsible for chlorophyll biosynthesis, photoprotection and defining the membrane geometry of the prolamellar body. Based on the 9-Å-resolution volume of the subtomogram average, we propose a structural model of membrane-associated LPOR.

Photocatalytic LPOR forms helical lattices that shape membranes for chlorophyll synthesis.

Chlorophyll biosynthesis, crucial to life on Earth, is tightly regulated because its precursors are phototoxic1. In flowering plants, the enzyme light-dependent protochlorophyllide oxidoreductase (LPOR) captures photons to catalyse the penultimate reaction: the reduction of a double bond within protochlorophyllide (Pchlide) to generate chlorophyllide (Chlide)2,3. In darkness, LPOR oligomerizes to facilitate photon energy transfer and catalysis4,5. However, the complete three-dimensional structure of LPOR, the higher-order architecture of LPOR oligomers and the implications of these self-assembled states for catalysis, including how LPOR positions Pchlide and the co-factor NADPH, remain unknown. Here, we report the atomic structure of LPOR assemblies by electron cryo-microscopy. LPOR polymerizes with its substrates into helical filaments around constricted lipid bilayer tubes. Portions of LPOR and Pchlide insert into the outer membrane leaflet, targeting the product, Chlide, to the membrane for the final reaction site of chlorophyll biosynthesis. In addition to its crucial photocatalytic role, we show that in darkness LPOR filaments directly shape membranes into high-curvature tubules with the spectral properties of the prolamellar body, whose light-triggered disassembly provides lipids for thylakoid assembly. Moreover, our structure of the catalytic site challenges previously proposed reaction mechanisms6. Together, our results reveal a new and unexpected synergy between photosynthetic membrane biogenesis and chlorophyll synthesis in plants, orchestrated by LPOR.

Spatial Nano-Morphology of the Prolamellar Body in Etiolated Arabidopsis thaliana Plants With Disturbed Pigment and Polyprenol Composition.

The prolamellar body (PLB) is a periodic bicontinuous membrane structure based on tubular tetrahedral units. PLBs are present in plant etioplasts and, upon illumination, directly transform into the lamellar thylakoid networks within chloroplasts. Efficient tubular-lamellar rearrangement and later formation of the photosynthetically active thylakoid membranes are crucial steps in the development of plant autotrophy. PLB membranes are mainly composed of galactolipids, carotenoids, and protochlorophyllide (Pchlide), the chlorophyll precursor, bound in a complex with NADPH and Pchlide oxidoreductase. Although the PLB structure has been studied for over 50 years, the direct role of particular membrane components in the formation of the PLB paracrystalline network remains elusive. Moreover, despite the numerous literature data regarding the PLB geometry, their reliable comparative analysis is complicated due to variable experimental conditions. Therefore, we performed comprehensive ultrastructural and low-temperature fluorescence analysis of wild type Arabidopsis thaliana (Arabidopsis) seedlings grown in different conditions typical for studies on etiolated seedlings. We established that the addition of sucrose to the growing media significantly affected the size and compactness of the PLB. The etiolation period was also an important factor influencing the PLB structural parameters and the ratio of free to complex-bound Pchlide. Thus, a reliable PLB structural and spectral analysis requires particular attention to the applied experimental conditions. We investigated the influence of the pigment and polyprenol components of the etioplast membranes on the formation of the PLB spatial structure. The PLB 3D structure in several Arabidopsis mutants (ccr1-1, lut5-1, szl1-1npq1-2, aba1-6, pif1, cpt7) with disturbed levels of particular pigments and polyprenols using electron tomography technique was studied. We found that the PLB nano-morphology was mainly affected in the pif1 and aba1-6 mutants. An increased level of Pchlide (pif1) resulted in the substantial shift of the structural balance between outer and inner PLB water channels and overall PLB compactness compared to wild type plants. The decrease in the relative content of β-branch xanthophylls in aba1-6 plants was manifested by local disturbances in the paracrystalline structure of the PLB network. Therefore, proper levels of particular etioplast pigments are essential for the formation of stable and regular PLB structure.

Dual role of the active site 'lid' regions of protochlorophyllide oxidoreductase in photocatalysis and plant development.

Protochlorophyllide oxidoreductase (POR) catalyses reduction of protochlorophyllide (Pchlide) to chlorophyllide, a light-dependent reaction of chlorophyll biosynthesis. POR is also important in plant development as it is the main constituent of prolamellar bodies in etioplast membranes. Prolamellar bodies are highly organised, paracrystalline structures comprising aggregated oligomeric structures of POR-Pchlide-NADPH complexes. How these oligomeric structures are formed and the role of Pchlide in oligomerisation remains unclear. POR crystal structures highlight two peptide regions that form a 'lid' to the active site, and undergo conformational change on binding Pchlide. Here, we show that Pchlide binding triggers formation of large oligomers of POR using size exclusion chromatography. A POR 'octamer' has been isolated and its structure investigated by cryo-electron microscopy at 7.7Å resolution. This structure shows that oligomer formation is most likely driven by the interaction of amino acid residues in the highly conserved lid regions. Computational modelling indicates that Pchlide binding stabilises exposure of hydrophobic surfaces formed by the lid regions, which supports POR dimerisation and ultimately oligomer formation. Studies with variant PORs demonstrate that lid residues are involved in substrate binding and photocatalysis. These highly conserved lid regions therefore have a dual function. The lid residues position Pchlide optimally to enable photocatalysis. Following Pchlide binding, they also enable POR oligomerisation - a process that is reversed through subsequent photocatalysis in the early stages of chloroplast development.

The Effect of Light on Plastid Differentiation, Chlorophyll Biosynthesis, and Essential Oil Composition in Rosemary (Rosmarinus officinalis) Leaves and Cotyledons.

It is unclear whether light affects the structure and activity of exogenous secretory tissues like glandular hairs. Therefore, transmission electron microscopy was first used to study plastid differentiation in glandular hairs and leaves of light-grown rosemary (Rosmarinus officinalis “Arp”) plants kept for 2 weeks under ambient light conditions. During our detailed analyses, among others, we found leucoplasts with tubuloreticular membrane structures resembling prolamellar bodies in stalk cell plastids of peltate glandular hairs. To study the effect of darkness on plastid differentiation, we then dark-forced adult, light-grown rosemary plants for 2 weeks and observed occasionally the development of new shoots with elongated internodes and pale leaves on them. Absorption and fluorescence spectroscopic analyses of the chlorophyllous pigment contents, the native arrangement of the pigment-protein complexes and photosynthetic activity confirmed that the first and second pairs of leaf primordia of dark-forced shoots were partially etiolated (contained low amounts of protochlorophyll/ide and residual chlorophylls, had etio-chloroplasts with prolamellar bodies and low grana, and impaired photosynthesis). Darkness did not influence plastid structure in fifth leaves or secretory tissues (except for head cells of peltate glandular hairs in which rarely tubuloreticular membranes appeared). The mesophyll cells of cotyledons of 2-week-old dark-germinated rosemary seedlings contained etioplasts with highly regular prolamellar bodies similar to those in mesophyll etio-chloroplasts of leaves and clearly differing from tubuloreticular membranes of secretory cells. Analyses of the essential oil composition obtained after solid phase microextraction and gas chromatography-mass spectroscopy showed that in addition to light, the age of the studied organ (i.e., first leaf primordia and leaf tip vs. fifth, fully developed green leaves) and the type of the organ (cotyledon vs. leaves) also strongly influenced the essential oil composition. Therefore, light conditions and developmental stage are both important factors to be considered in case of potential therapeutic, culinary or aromatic uses of rosemary leaves and their essential oils.

A cis-carotene derived apocarotenoid regulates etioplast and chloroplast development.

Carotenoids are a core plastid component and yet their regulatory function during plastid biogenesis remains enigmatic. A unique carotenoid biosynthesis mutant, carotenoid chloroplast regulation 2 (ccr2), that has no prolamellar body (PLB) and normal PROTOCHLOROPHYLLIDE OXIDOREDUCTASE (POR) levels, was used to demonstrate a regulatory function for carotenoids and their derivatives under varied dark-light regimes. A forward genetics approach revealed how an epistatic interaction between a ζ-carotene isomerase mutant (ziso-155) and ccr2 blocked the biosynthesis of specific cis-carotenes and restored PLB formation in etioplasts. We attributed this to a novel apocarotenoid retrograde signal, as chemical inhibition of carotenoid cleavage dioxygenase activity restored PLB formation in ccr2 etioplasts during skotomorphogenesis. The apocarotenoid acted in parallel to the repressor of photomorphogenesis, DEETIOLATED1 (DET1), to transcriptionally regulate PROTOCHLOROPHYLLIDE OXIDOREDUCTASE (POR), PHYTOCHROME INTERACTING FACTOR3 (PIF3) and ELONGATED HYPOCOTYL5 (HY5). The unknown apocarotenoid signal restored POR protein levels and PLB formation in det1, thereby controlling plastid development.

Formation of prolamellar-body-like ultrastructures in etiolated cyanobacterial cells overexpressing light-dependent protochlorophyllide oxidoreductase in Leptolyngbya boryana.

Protochlorophyllide (Pchlide) reduction is the penultimate step of chlorophyll (Chl) biosynthesis, and is catalyzed by two evolutionarily unrelated enzymes: dark-operative Pchlide oxidoreductase (DPOR) and light-dependent Pchlide oxidoreductase (LPOR). Because LPOR is the sole Pchlide reductase in angiosperms, dark-grown seedlings of angiosperms become etiolated. LPOR exists as a ternary complex of Pchlide-NADPH-LPOR to form paracrystalline prolamellar bodies (PLBs) in etioplasts. Because LPOR is distributed ubiquitously across oxygenic phototrophs including cyanobacteria, it would be important to determine whether cyanobacterial LPOR has the ability to form PLBs. We isolated a DPOR-less transformant ΔchlL/LPORox, carrying a plasmid to overexpress cyanobacterial LPOR in the cyanobacterium Leptolyngbya boryana. The transformant did not produce Chl in the dark and became etiolated with an accumulation of Pchlide and LPOR. Novel PLB-like ultrastructures were observed in etiolated cells, which disappeared during the early stage of the light-dependent greening process. However, the rate of Chl production in the greening process of ΔchlL/LPORox was almost the same as that observed in the control cells, which carried an empty vector. An in vitro LPOR assay of extracts of dark-grown ΔchlL/LPORox cells suggested that the PLB-like structures are deficient in NADPH. Low-temperature fluorescence emission spectra of membrane fractions of the etiolated cells indicated the absence of the photoactive form of Pchlide, which was consistent with the inefficiency of the greening process. Cyanobacterial LPOR exhibited an intrinsic ability to form PLB-like ultrastructures in the presence of the co-accumulation of Pchlide; however, the PLB-like structure differed from the authentic PLB regarding NADPH deficiency.

Role of Galactolipids in Plastid Differentiation Before and After Light Exposure.

Galactolipids, monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG), are the predominant lipid classes in the thylakoid membrane of chloroplasts. These lipids are also major constituents of internal membrane structures called prolamellar bodies (PLBs) and prothylakoids (PTs) in etioplasts, which develop in the cotyledon cells of dark-grown angiosperms. Analysis of Arabidopsis mutants defective in the major galactolipid biosynthesis pathway revealed that MGDG and DGDG are similarly and, in part, differently required for membrane-associated processes such as the organization of PLBs and PTs and the formation of pigment–protein complexes in etioplasts. After light exposure, PLBs and PTs in etioplasts are transformed into the thylakoid membrane, resulting in chloroplast biogenesis. During the etioplast-to-chloroplast differentiation, galactolipids facilitate thylakoid membrane biogenesis from PLBs and PTs and play crucial roles in chlorophyll biosynthesis and accumulation of light-harvesting proteins. These recent findings shed light on the roles of galactolipids as key facilitators of several membrane-associated processes during the development of the internal membrane systems in plant plastids.

Cytological attributes of storage tissues in nematode and eriophyid galls: pectin and hemicellulose functional insights.

Cell walls and protoplast may work together or distinctly in the establishment of the functional profiles of gall tissue compartments. This presumption is herein evaluated in three gall systems by immunocytochemical and ultrastructural analyses. The common storage tissues (CSTs) of leaf galls induced by Eriophyidae on Miconia ibaguensis leaves and by Ditylenchus gallaeformans on M. ibaguensis and M. albicans have rigid and porous cell walls due to their composition of pectins. Hemicelluloses in CST cell walls are scarcer when compared to the cell walls of the control leaves, being functionally compensated by rigid pectate gels. The typical nutritive tissues (TNTs) in galls induced by Ditylenchus gallaeformans are similar to promeristematic and secretory cells regarding their enriched cytoplasm, several mitochondria, and proplastids, as well as multivesicular and prolamellar bodies in cell membranes. The cytological features of the feeding cells of Eriophyidae galls indicate that they are not as metabolically active as the cells of the TNT in nematode galls. However, their cell wall composition suggests more plasticity and porosity than the cells of the TNT, which can compensate the less production of nutrients with more transport. The ultrastructural and immunocytochemical profiles of CST cells reveal functional similarities, which are independent of the taxa of the gall inducer or of the host plant. Despite their analogous functionalities, the protoplast and cell wall features of TNT cells of nematode galls and of the feeding cells of the Eriophyidae galls are distinct, and work out through different strategies toward keeping gall developmental site active.