A study of the ultrastructure of the shoot apex and leaf cells in two liverworts,with special reference to the oil bodies

Summary In this investigation attention has been paid to the general ultrastructure of the shoot apical and leaf cells in the liverwortsBazzania trilobata andLophozia ventricosa but especially to the different developmental stages of their oil bodies. These species have been chosen because their oil bodies differ from each other in size and shape.The appearance of the different organelles, nucleus, chloroplasts, mitochondria, ER, and Golgi bodies, are in their main features the same as those of higher plants described in the literature. The dark cytoplasm seen in the leaf cells ofLophozia in the vicinity of the oil bodies but without any surrounding membrane when fixed in double fixative 2, seems to be specific to this species. On the other hand, granular dense bodies were visible in the cells of the shoot apex ofBazzania, which shrank in size as the development of the oil bodies proceeded and were lacking in the mature leaf cells.In both species investigated, the oil bodies have the same component parts: (1) an outer membrane enveloping the whole body, (2) inside this, a granular stroma layer of varying thickness enveloping (3) specific globules of varying size and number, each of which is surrounded by (4) a thin inner membrane (Fig. 28).The oil bodies develop in at least two ways and usually in one way for each species. InBazzania they seem to develop from vacuole-like formations in the shoot apex or in the leaf primordia into which substances have segregated. InLophozia they seem to originate by aggregation and fusion of lipid bodies.     

Development and enzyme activity of protein bodies in proteinoplasts of tobacco root cells

The development of protein bodies in proteinoplasts of tobacco (Nicotiana tabacum L. var. Wis. 38) roots was investigated with TEM, HVEM, and enzyme cytochemistry. These plastids contain a three-dimensional network of fenestrated tubules which originate from invaginations of the inner membrane of the plastid envelope. Elaboration of the network occurs in parallel with cell differentiation: slender tubules common to plastids in meristematic cells undergo dilation as protein accumulates during cell differentiation; proteinoplasts of vacuolate and root cap cells usually contain a large protein body. The contents of the peripheral tubules, originating from the inner membrane, are less electron dense than the tubules making up the central network. Localized dilations within the tubular network result in the formation of dense spheroidal structures, protein bodies, apparently as a result of continued protein accumulation via tubules connecting to the central network. Protein might be imported from segments of rough ER attached to or apposed to the outer membrane of the proteinoplast envelope. The presence of catalase (E.C. 1.11.1.6), peroxidase (E.C. 1.11.1.7), and cytochrome oxidase (E.C. 1.9.3.1) was demonstrated by cytochemistry with diaminobenzidine (DAB) as substrate. Oxidized DAB was found in protein bodies after incubation in each of the specific reaction media. While aminotriazole and sodium azide inhibited oxidation of DAB by catalase and peroxidase, respectively, only potassium cyanide completely inhibited oxidation of DAB in protein bodies. We conclude that protein bodies of proteinoplasts in tobacco roots are not sites for storage of protein, rather protein bodies contain heme protein(s) with strong oxidase activity that may convey a specific function to proteinoplasts.     

Membrane continuity and associations in the fernDryopteris borreri

Summary Occasional direct membrane connections have been observed, inDryopteris borreri gametophyte cells, of the outer membrane of the chloroplast envelope with smooth ER, of the plastid envelope with the plasmalemma, and of the nuclear envelope with the ER. In addition, close spatial associations exist in most cells between ER and both plastids and microbodies.     

Structure of amyloplasts and endoplasmic reticulum in the root caps of Lepidium sativum and Zea mays observed after selective membrane staining and by high-voltage electron microscopy

The structure of plastids in the root cap of cress and maize was studied by low- and high-voltage electron microscopy after staining their membranes with a mixture of zinc iodide and osmium tetroxide. In plastids of both species electron-opaque membranes were found in the plastid interior while membranes of lesser electron-opacity comprised the outer envelope and vesicles and cisternae underlying it. Electron-opaque tubules, often in groups attached to the inner membrane of the amyloplast envelope, were found in cress but not in maize. The internal, less-opaque membranes were often found associated with the starch grains. No specific association could be seen between amyloplasts and endoplasmic reticulum (ER); their surfaces showed no regular contact or connexion, though the amyloplasts clearly indented the underlying ER. The ER in statocytes was predominantly tubular in cress but predominantly cisternal in maize.Abbreviations ER endoplasmic reticulum - ZIO zinc iodideosmium tetroxide     

Lipid trafficking between the endoplasmic reticulum and the plastid in Arabidopsis requires the extraplastidic TGD4 protein

The development of chloroplasts in Arabidopsis thaliana requires extensive lipid trafficking between the endoplasmic reticulum (ER) and the plastid. The biosynthetic enzymes for the final steps of chloroplast lipid assembly are associated with the plastid envelope membranes. For example, during biosynthesis of the galactoglycerolipids predominant in photosynthetic membranes, galactosyltransferases associated with these membranes transfer galactosyl residues from UDP-Gal to diacylglycerol. In Arabidopsis, diacylglycerol can be derived from the ER or the plastid. Here, we describe a mutant of Arabidopsis, trigalactosyldiacylglycerol4 (tgd4), in which ER-derived diacylglycerol is not available for galactoglycerolipid biosynthesis. This mutant accumulates diagnostic oligogalactoglycerolipids, hence its name, and triacylglycerol in its tissues. The TGD4 gene encodes a protein that appears to be associated with the ER membranes. Mutant ER microsomes show a decreased transfer of lipids to isolated plastids consistent with in vivo labeling data, indicating a disruption of ER-to-plastid lipid transfer. The complex lipid phenotype of the mutant is similar to that of the tgd1,2,3 mutants disrupted in components of a lipid transporter of the inner plastid envelope membrane. However, unlike the TGD1,2,3 complex, which is proposed to transfer phosphatidic acid through the inner envelope membrane, TGD4 appears to be part of the machinery mediating lipid transfer between the ER and the outer plastid envelope membrane. The extent of direct ER-to-plastid envelope contact sites is not altered in the tgd4 mutant. However, this does not preclude a possible function of TGD4 in those contact sites as a conduit for lipid transfer between the ER and the plastid.     

Development and enzyme activity of protein bodies in proteinoplasts of tobacco root cells

Summary The development of protein bodies in proteinoplasts of tobacco (Nicotiana tabacum L. var. Wis. 38) roots was investigated with TEM, HVEM, and enzyme cytochemistry. These plastids contain a three-dimensional network of fenestrated tubules which originate from invaginations of the inner membrane of the plastid envelope. Elaboration of the network occurs in parallel with cell differentiation: slender tubules common to plastids in meristematic cells undergo dilation as protein accumulates during cell differentiation; proteinoplasts of vacuolate and root cap cells usually contain a large protein body. The contents of the peripheral tubules, originating from the inner membrane, are less electron dense than the tubules making up the central network. Localized dilations within the tubular network result in the formation of dense spheroidal structures, protein bodies, apparently as a result of continued protein accumulation via tubules connecting to the central network. Protein might be imported from segments of rough ER attached to or apposed to the outer membrane of the proteinoplast envelope.The presence of catalase (E.C. 1.11. 1.6), peroxidase (E.C. 1.11.1.7), and cytochrome oxidase (E.C. 1.9.3.1) was demonstrated by cytochemistry with diaminobenzidine (DAB) as substrate. Oxidized DAB was found in protein bodies after incubation in each of the specific reaction media. While aminotriazole and sodium azide inhibited oxidation of DAB by catalase and peroxidase, respectively, only potassium cyanide completely inhibited oxidation of DAB in protein bodies. We conclude that protein bodies of proteinoplasts in tobacco roots are not sites for storage of protein, rather protein bodies contain heme protein(s) with strong oxidase activity that may convey a specific function to proteinoplasts.Abbreviations used CAT catalase - CYT-OX cytochrome oxidase - DAB diaminobenzidine - ER endoplasmic reticulum - f filaments - HVEM high voltage electron microscopy - M mitochondrion - MT microtubule - P peroxisome - PB protein body - PER peroxidase - Pl plastid - Pg plastoglobuli - RER rough endoplasmic reticulum - RuBPcase ribulose-1,5-bisphosphate carboxylase - S starch - T tubule - V vacuole Scientific Article No. A3997, Contribution No. 6981, of the Maryland Agricultural Experiment StationThe scale bar on each micrograph is 0.1 , unless indicated otherwise     

INTERMEDIATE FEATURES OF CYANELLE DIVISION OF CYANOPHORA PARADOXA (GLAUCOCYSTOPHYTA) BETWEEN CYANOBACTERIAL AND PLASTID DIVISION1

Cyanelles of glaucocystophytes may be the most primitive of the known plastids based on their peptidoglycan content and the sequence phylogeny of cyanelle DNA. In this study, EM observations have been made to characterize the cyanelle division of Cyanophora paradoxa Korshikov and to gain insights into the evolution of plastid division. Constriction of cyanelles involves ingrowth of the septum at the cleavage site with the inner envelope membrane invaginating at the leading edge and the outer envelope membrane invaginating behind the septum. This means the inner and outer envelope membranes do not constrict simultaneously as they do in plastid division in other plants. The septum and the cyanelle envelope became stained after a silver‐methenamine staining was applied for in situ detection of polysaccharides. Septum formation was inhibited by β‐lactams and vancomycin, which are potent inhibitors of bacterial peptidoglycan biosynthesis. These results suggest the presence of peptidoglycan at the septum and the cyanelle envelope. In dividing cyanelles, a single electron‐dense ring (cyanelle ring) was observed on the stromal face of the inner envelope membrane at the isthmus, but no ring‐like structures were detected on the outer envelope membrane. Thus a single, stromal cyanelle ring such as this is quite unique and also distinct from FtsZ rings, which are not detectable by TEM. These features suggest that the cyanelle division of glaucocystophytes represents an intermediate stage between cyanobacterial and plastid division. If monophyly of all plastids is true, the cyanelle ring and the homologous inner plastid dividing ring might have evolved earlier than the outer plastid dividing ring.     

Ontogeny of microbodies (glyoxysomes) in cotyledons of dark-grown watermelon (Citrullus vulgaris Schrad.) seedlings

The development of glyoxysomal marker enzyme activities and concomitant ultrastructural evidence for the ontogeny of glyoxysomes has been studied in cotyledons of dark-grown watermelon seedlings (Citrullus vulgaris Schrad., var. Florida Giant). Catalase (CAT, EC 1.11.1.6) was stained in glyoxysomal structures with the 3,3-diaminobenzidine procedure. Serial sections and high-voltage electron microscopy were used to analyze the three-dimensional structure of the glyoxysomal population. With early germination CAT was localized in three distinct cell structures: spherical microbodies already present in freshly imbibed cotyledons; in appendices on lipid bodies; and in small membrane vesicles between the lipid bodies. Due to their ribosome-binding capacity, both appendices and small vesicles were identified as derivatives of the endoplasmic reticulum (ER). In the following period, glyoxysome formation and lipid body degradation were found to be inseparable processes. The small CAT-containing vesicles attach to a lipid body on a restricted area. Both lipid body appendices and attached cisternae enlarge around and between tightly packed lipid bodies and eventually become pleomorphic glyoxysomes with lipid bodies entrapped into cavities. The close contact between lipid body and glyoxysomes is maintained until the lipid body is digested and the glyoxysomal cavity becomes filled with cytoplasm. During the entire period of increase in glyoxysomal enzyme activities, no evidence was obtained for destruction of glyoxysomes, but small CAT-containing vesicles were observed from day 2 through day 6 after imbibition, indicating a continuous de novo formation of glyoxysomes. This study does not substantiate the hypothesis that glyoxysomes bud directly from the ER. Rather, ER-derivatives, e.g., lipid body appendices or cisternae attached to lipid bodies are interpreted as being glyoxysomal precursors that grow in close contact with lipid bodies both in volume and surface membrane area.Abbreviations CAT catalase - DAB 3,3 diaminobenzidine tetrahydrochloride - ER endoplasmic reticulum - GOX glycolate oxidase - HPR hydroxypyruvate reductase - HVEM high-voltage electron microscopy - ICL isocitrate lyase - MS malate synthase - RER rough endoplasmic reticulum In the figures bars represent 0.1 m (if not stated otherwise)     

SOME SURFACE CHARACTERISTICS OF THE CHLOROPLAST ENVELOPE

Pronase, cationic ferritin, and ferritin-conjugated plant lectins were used to study the chloroplast envelope. Negative charges (binding cationic ferritin) are fairly uniformly distributed over the envelope surfaces in contact with the hyaloplasm and are not appreciably altered by mild pronase treatment of isolated plastids. All surfaces of stroma-free thylakoids previously exposed to the stroma uniformly bind cationic ferritin. Ricin_II-ferritin binding to the membranes of the chloroplast envelope indicates that galactolipids are distributed in the outer membrane in such a way that their galactose moieties are exposed on the envelope surface. In addition, the outer surface of the inner membrane (the intermembrane face) contains uniformly distributed galactose which binds ricin_II when this membrane is exposed to the reaction medium. Isolated vesicles of the chloroplast envelope bind ricin_II, while isolated envelope vesicles as well as the envelopes of intact chloroplasts failed to bind concanavalin A. Thylakoid surfaces showed minor binding of ricin_II as well as concanavalin A.     

Electron Microscopy of the Spherical Body of Oral Spirochetes In Vitro: Further Studies

The surface and inner structure of the spherical bodies (SB) produced by the human oral treponeme strain G7201, similar to Treponema macrodentium, were studied by electron microscopy. Ultrathin sectioning and scanning techniques demonstrated that in the presence of a high concentration of sucrose, the outer envelope of one or both terminal ends of this oral spirochete changed into a swollen structure, the SB. Spirochetal cells adhered firmly to the surface of the resultant body. The membrane of the SB, i.e. the outer envelope, enclosed the coiled protoplasmic cylinder and five axial fibrils which were located between the envelope and the cylinder. Large expanded protoplasmic cylinders were observed, surrounded by a partially disrupted double membrane in some SBs. A number of frizzly fibrous structures, which differed from axial fibrils in number and shape, were also observed within these SBs. Except for abnormal or partially broken cylinders, the protoplasmic cylinders tended to be located close to the inner surface of the SB membrane, resulting in a central vacant space with occasional axial fibrils. These findings suggest that the oral spirochete produces an SB by terminal expansion of the outer envelope in the presence of high concentrations of sucrose. The outer envelope of the SB, which consists of two electron-dense layers, has the property of binding spirochetal cells to its outer layer and the protoplasmic cylinder and axial fibrils to the inner layer. Some protoplasmic cylinders were also observed to be swollen in the presence of high sucrose concentrations.     

Leucoplasts: a distinct kind of organelles lacking typical 70S ribosomes and free thylakoids

Non-pigmented plastids were observed in fully differentiated cells from leaves and stem tissues of various species. Although showing important differences in size and shape, these plastids exhibit permanent structural features which allow to get them together as a distinct kind of organelles: the leucoplasts. Leucoplasts are distinct from the proplastids and every intermediate stage of plastid differentiation, from white chromoplasts and tuber amyloplasts. Mature leucoplasts do not contain an autonomous central system of thylakoids structurally independent from the envelope and, therefore, are never green. However, the envelope inner membrane invaginates within the plastid a cisternal or tubular stroma reticulum connected with the intermembrane space of the envelope. In addition, the leucoplast stroma is often less dense than chloroplasts stroma and contain several nucleoids with DNA fibrils. However, 70S ribosomes either scattered in the stroma or attached to the stroma reticulum or the envelope are not visible in ultrathin sections of leucoplasts stained with uranyl and lead. The existence of more discrete particles as dense as ribosomes is suggested. The relationship between the absence of ribosomes and thylakoids is discussed. Except for their specific role in C10 monoterpene synthesis in glandular cells, the functions of leucoplasts in plant cells remains largely up to now a matter of conjecture.     

Ultrastructural features of the constricted region of dividing plastids

Summary The ultrastructure of the constricted region of dividing plastids of spinach, bean, turnip, tobacco, and wheat has been studied. In these species, an electron-opaque, ring-like structure (RS) girdles the constricted region of plastids in advanced stages of division. The RS is a compound entity composed of two concentric rings of electron-opaque materials; one on the stromal face of the inner membrane and the other on the cytoplasmic face of the outer membrane. It was concluded that the compound nature of the RS is highly conserved in angiosperms being present in some cereal grasses and in plants representing four different orders of dicotyledonous plants. Evidence indicating that the electron-opaque materials of the RS are integrated into the envelope membranes was also provided and it was suggested that the envelope in the region of the RS may have unique properties. For spinach, it was also noted that plastids with deeply constricted necks tend to have RSs with lower volumes than those from wider necks and that endoplasmic reticulum was frequently present in the cytoplasm of the constriction region.Abbreviations RS ring structure - ER endoplasmic reticulum     

Pollen and Tapetum Development in Male Fertile Rosmarinus Officinalis L. (Lamiaceae)

The development of microspores/pollen grains and tapetum was studied in fertile Rosmarinus officinalis L. (Lamiaceae). Most parts of the cell walls of the secretory anther tapetum undergo modifications before and during meiosis: the inner tangential and radial cell walls, and often also the outer tangential and radial wall, acquire a fibrous appearance; these walls become later transformed into a thin poly-saccharidic film, which is finally dissolved after microspore mitosis. Electron opaque granules found within the fibrous/lamellated tapetal walls consist of sporopollenin-like material, but cannot be interpreted as Ubisch bodies. The middle lamella and the primary wall of the outer tangential and radial tapetal walls remain unmodified, but get covered by an electron opaque, sporopollenin-like layer. Pollenkitt is formed only by lipid droplets from the ground plasma and/or ER profiles, the plastids do not form pollenkitt precursor lipids. Tapetum maturation (“degeneration”) does not take place before late vacuolate stage.

The apertures are determined during meiosis by vesicles or membrane stacks on the surface of the plasma membrane. The procolumellae are conical, but at maturity the columellae are more cylindrical in shape. The columellar bases often fuse, but a genuine foot layer is lacking. The formation of the endexine starts with sporopollenin-accumulating white lines adjacent to the columellar bases. Later, the endexine grows more irregularly by the accumulation of sporopollenin globules. In mature pollen the intine is clearly bilayered.

Generative cells (GCs) and sperm cells contain a comparatively large amount of cytoplasm, and organelles like mitochondria, dictyosomes, ER, and multi-vesicular bodies, but no plastids; GCs and sperms are separated from the vegetative cell only by two plasma membranes.     

FINE STRUCTURE OF PROTEIN-STORING PLASTIDS IN BEAN ROOT TIPS

The fine structure of leucoplasts in root tip cells of Phaseolus vulgaris L. has been studied in material fixed in glutaraldehyde followed by osmium tetroxide and poststained in uranyl acetate and lead citrate. Plastid development has been followed from the young stages in and near the meristematic region, through an ameboid stage, to the larger forms with more abundant storage products in the outermost cells. The plastids contain a dense stroma penetrated by tubules and cisternae arising from the inner membrane of the plastid envelope. Also located in the stroma are lamellae, ribosome-like particles, phytoferritin granules, and fine fibrils in less dense regions. In some elongate plastids microfilaments run lengthwise in the stroma near the surface. The same plastids store both starch and protein, but in a strikingly different manner. The starch is deposited in the stroma, while the protein always is accumulated within membrane-bounded sacs. These sacs arise as outgrowths from a complex of interconnected tubules which in turn appears to originate by coalescence and proliferation of tubules and cisternae arising from the inner plastid membrane. This "tubular complex" bears a strong resemblance to the prolamellar body of etiolated chloroplasts, but is smaller and ordinarily less regularly organized, and is apparently light-insensitive. Crystallization of the protein commonly occurs in the sacs and occasionally takes place within the tubules of the complex as well. The fine structure of the leucoplasts is discussed in relation to that of etiolated chloroplasts. Suggestions are made concerning the function of the tubular complex, role of the ameboid plastid forms, and manner of accumulation of the storage protein in the plastids.     

Interactions between the Nucleus and Cytoplasmic Organelles during the Cell Cycle of Euglena gracilis in Synchronized Cultures: I. Associations between the Nucleus and Chloroplasts at an Early Stage in the Cell Cycle under Photoorganotrophic Conditions (Part I)

At an early stage in the cell cycle of Euglena gracilis Z, synchronizedunder 10-h light : 14-h dark alternations in an organic medium,the conjoined chloroplasts that formed made up a single giantbody that came close to the nucleus, covering most of the nuclearperiphery. Three different types of association between thesetwo organelles were observed. In one the outer membrane of thenuclear envelope was in contact, in some narrow regions, withthe chloroplast membrane, the site of contact being filled withdense material. A chromosome in the unfolded, fibrillar structurewas very close to the site of contact, the extreme end of thefibril touching the inner membrane of the nuclear envelope.When cells from the culture used above were stained with DAPIand examined under a fluorescence microscope, chloroplast nucleoidsin the giant body appeared to form, at least in part, a threadwith branchings, and some tips of the branchings came closeto the site of contact with the nucleus. In the second type of association, which was rare, part of thenuclear envelope protruded into the chloroplast, and the siteof contact was filled with dense material. A chromosome wasnear the site of this protrusion. In the third type of chloroplast-nucleusassociation the ER was continuous with the outer membrane ofthe nuclear envelope at one end and in contact with the chloroplastmembrane, at the other end. ¹This work was reported at the 48th Annual Meeting of the BotanicalSociety of Japan in Kyoto, October, 1983. (Received March 14, 1984; Accepted June 27, 1984)     

Electron Microscopic Observations on the Structure of the Envelopes of Mature Elementary Bodies and Developmental Reticulate Forms of Chlamydia psittaci

Purified suspensions of Chlamydia psittaci were prepared from L cells. Thin sections of intact elementary bodies and intact developmental reticulate bodies and of their purified envelopes were observed by electron microscopy. In both intact organisms and partially purified envelopes, two membranous structures, each appearing in electron micrographs as two darkly stained layers, were observed. In the elementary body sections, the outer membrane was round, apparently rigid, and was not soluble in 0.5% sodium dodecyl sulfate. The inner layer was irregular in shape and was completely removed by detergent treatment. We interpret these results to indicate that the outer rigid layer of the envelope is the cell wall and the inner layer is the cytoplasmic membrane. When the fragile reticulate body envelopes were similarly studied, the outer cell wall was clearly visible, and some evidence of an inner membrane was seen. After treatment with nucleases and detergent, all evidence of inner or cytoplasmic membrane was removed, but the outer cell wall remained. Thus, it appears that the cell wall of this organism is continuous throughout the growth cycle and that the fragility and lack of rigidity of the reticulate body cell is due to changes in chemical composition or structure of the cell wall.     

African Swine Fever Virus Is Enveloped by a Two-Membraned Collapsed Cisterna Derived from the Endoplasmic Reticulum

During the cytoplasmic maturation of African swine fever virus (ASFV) within the viral factories, the DNA-containing core becomes wrapped by two shells, an inner lipid envelope and an outer icosahedral capsid. We have previously shown that the inner envelope is derived from precursor membrane-like structures on which the capsid layer is progressively assembled. In the present work, we analyzed the origin of these viral membranes and the mechanism of envelopment of ASFV. Electron microscopy studies on permeabilized infected cells revealed the presence of two tightly apposed membranes within the precursor membranous structures as well as polyhedral assembling particles. Both membranes could be detached after digestion of intracellular virions with proteinase K. Importantly, membrane loop structures were observed at the ends of open intermediates, which suggests that the inner envelope is derived from a membrane cisterna. Ultraestructural and immunocytochemical analyses showed a close association and even direct continuities between the endoplasmic reticulum (ER) and assembling virus particles at the bordering areas of the viral factories. Such interactions become evident with an ASFV recombinant that inducibly expresses the major capsid protein p72. In the absence of the inducer, viral morphogenesis was arrested at a stage at which partially and fully collapsed ER cisternae enwrapped the core material. Together, these results indicate that ASFV, like the poxviruses, becomes engulfed by a two-membraned collapsed cisterna derived from the ER.     

Preparation and characterization of membrane fractions enriched in outer and inner envelope membranes from spinach chloroplasts. II. Biochemical characterization

In the previous paper (Block, M. A., Dorne, A.-J., Joyard, J., and Douce, R. (1983) J. Biol. Chem. 258, 13273-13280), we have described a method for the separation of membrane fractions enriched in outer and inner envelope membranes from spinach chloroplasts. The two envelope membranes have a different weight ratio of acyl lipid to protein (2.5-3 for the outer envelope membrane and 0.8-1 for the inner envelope membrane). The two membranes also differ in their polar lipid composition. However, in order to prevent the functioning of the galactolipid:galactolipid galactosyltransferase during the course of envelope membrane separation, we have analyzed the polar lipid composition of each envelope membrane after thermolysin treatment of the intact chloroplasts. The outer envelope membrane is characterized by the presence of high amounts of phosphatidylcholine and digalactosyldiacylglycerol whereas the inner envelope membrane has a polar lipid composition almost identical with that of the thykaloids. No phosphatidylethanolamine or cardiolipin could be detected in either envelope membranes, thus demonstrating that the envelope membranes, and especially the outer membrane, do not resemble extrachloroplastic membranes. No striking differences were found in the fatty acid composition of the polar lipids from either the outer or the inner envelope membrane. The two envelope membranes also differ in their carotenoid composition. Among the different enzymatic activities associated with the chloroplast envelope, we have shown that the Mg2+-dependent ATPase, the UDP-Gal:diacylglycerol galactosyltransferase, the phosphatidic acid phosphatase, and the acyl-CoA thioesterase are associated with the inner envelope from spinach chloroplasts whereas the acyl-CoA synthetase is located on the outer envelope membrane.     

Conservative sorting in the muroplasts of Cyanophora paradoxa: a reevaluation based on the completed genome sequence

After primary endosymbiosis, massive gene transfer occurred from the genome of the cyanobacterial endosymbiont to the nucleus of the protist host cell. In parallel, a specific protein import apparatus arose for reimport of many, but not all products of the genes moved to the nuclear genome. Presequences evolved to allow recognition of plastid proteins at the envelope and their translocation to the stroma. However, plastids (and cyanobacteria) also comprise five other subcompartments. Protein sorting to the cyanobacterial thylakoid membrane, the thylakoid lumen, the inner envelope membrane, the periplasmic space, and the outer envelope membrane is achieved by prokaryotic protein translocases recognizing, e.g., signal sequences. The “conservative sorting” hypothesis postulates that these translocases remained functional in endosymbiotic organelles and obtained their passengers not only from imported proteins but also from proteins synthesized in organello. For proteins synthesized in the cytosol, a collaboration of the general import apparatus and the former prokaryotic translocase is necessary which is often reflected by the use of bipartite presequences, e.g., stroma targeting peptide and signal peptide. For plants, this concept has been experimentally proven and verified. The muroplasts from Cyanophora paradoxa, that have several features more in common with cyanobacteria than with plastids, were analyzed with the availability of the recently completed nuclear genome sequence. Interesting findings include the absence of the post-translational signal recognition particle pathway, dual Sec translocases in thylakoid and inner envelope membranes that are produced from a single set of genes, and a co-translational signal recognition pathway operating without a 4.5S RNA component.     

The plastid outer membrane localized LPTD1 is important for glycerolipid remodelling under phosphate starvation

Glycerolipid synthesis in plants is coordinated between plastids and the endoplasmic reticulum (ER). A central step within the glycerolipid synthesis is the transport of phosphatidic acid from ER to chloroplasts. The chloroplast outer envelope protein TGD4 belongs to the LptD family conserved in bacteria and plants and selectively binds and may transport phosphatidic acid. We describe a second LptD‐family protein in A. thaliana (atLPTD1; At2g44640) characterized by a barrel domain with an amino‐acid signature typical for cyanobacterial LptDs. It forms a cation selective channel in vitro with a diameter of about 9 Å. atLPTD1 levels are induced under phosphate starvation. Plants expressing an RNAi construct against atLPTD1 show a growth phenotype under normal conditions. Expressing the RNAi against atLPTD1 in the tgd4–1 background renders the plants more sensitive to light stress or phosphate limitation than the individual mutants. Moreover, lipid analysis revealed that digalactosyldiacylglycerol and sulfoquinovosyldiacylglycerol levels remain constant in the RNAi mutants under phosphate starvation, while these two lipids are enhanced in wild‐type. Based on our results, we propose a function of atLPTD1 in the transport of lipids from ER to chloroplast under phosphate starvation, which is combinatory with the function of TGD4.