Sieve-element plastids ofCyrillaceae,Erythroxylaceae andRhizophoraceae: Description and significance of subtype PV plastids

A new subtype (PV) of protein-containing sieve-element plastids was found to contain a uniquely large number of polygonal protein crystals, sometimes with (PVcf) and sometimes without (PVc) protein filaments. These plastids do not accumulate starch. The PVcf-plastids occur inCyrillaceae only, while the PVc-plastids are limited toErythroxylaceae andRhizophoraceae. The significance of the new P-subtype with respect to the systematic position of the three families is discussed.     

Sieve-element plastids ofGymnospermae: Their ultrastructure in relation to systematics

The distribution of S-type and P-type plastids in the sieve elements of 30 species from 13 families of theConiferophytina andCycadophytina is recorded, of which 21 species were studied for the first time with respect to their sieve-element plastids. While starch storing S-type plastids are the most commonly occurring type throughout both taxa, all thePinaceae examined (11 species of 7 genera) contain P-type plastids characterized by a peripheral, ring-shaped bundle of protein filaments, an additional protein crystalloid, and several starch grains. Starch grains of sieve-element plastids in theConiferophytina andCycadophytina are commonly club-shaped. Taxonomic implications of these ultrastructural findings on sieve-element plastids are discussed.     

Sieve-element plastids,exine sculpturing and the systematic affinities of theBuxaceae

The sieve-element plastids of members of several genera in theBuxaceae (Buxus, Pachysandra andSarcococca) were found to be of the specific subtype PVI, which contains a central globular protein crystal.Simmondsia (Simmondsiaceae) andDaphniphyllum (Daphniphyllaceae), on the other hand, were found to contain S-type sieve-element plastids. The occurrence of the highly restricted PVI plastids in theBuxaceae mitigates against a close relationship between theBuxaceae andSimmondsia, Daphniphyllum andEuphorbiaceae. Exine sculpturing of theBuxaceae andSimmondsiaceae also shows no close similarities. Both of these EM characters are discussed in connection with other available data and with respect to earlier systematic treatment of these families.     

Sieve-element plastids ofCaryophyllales: Additional investigations with special reference to theCaryophyllaceae andMolluginaceae

Dictyonema moorei andD. ligulatum possess a haustorial system quite similar to that of three otherDictyonema species. It is singular among all Asco- and Basidiolichenes and specific for the genusDictyonema. In herbarized specimens it shrinks and becomes nearly indistinguishable with longer time. InDictyonema a sterile thallus may develop which consists of a turf of filaments ofScytonema growing over mosses and each surrounded by a fungal sheath.     

Sieve-element plastids and evolution of monocotyledons, with emphasis on Melanthiaceae sensu lato and Aristolochiaceae-Asaroideae, a putative dicotyledon sister group

Monocotyledons are distinguishable from dicotyledons by their subtype P2 sieve-element plastids containing cuneate protein crystals, a synapomorphic character uniformly present from basal groups through Lilioids to Commelinoids. The dicotyledon generaAsarum andSaruma (Aristolochiaceae-Asaroideae) are the only other taxa with cuneate crystals, but their sieveelement plastids include an additional large polygonal crystal, as is typical of many eumagnoliids. New investigations in Melanthiaceae s.l. revealed the same pattern (polygonal plus cuneate crystals) in the sieve-element plastids ofJaponolirion osense (Japonoliriaceae/Petrosaviaceae), ofHarperocallis flava, Pleea tenuifolia, andTofleldia (all: Tofieldiaceae). InNarthecium ossifragum a large crystal, present in addition to cuneate ones, usually breaks up into several small crystals, whereas inAletris glabra andLophiola americana (Nartheciaceae) and in all of the 15 species studied and belonging to Melanthiaceae s.str. only cuneate crystals are found. Highresolution TEM pictures reveal a crystal substructure that is densely packed in both cuneate and polygonal forms, but in Tofieldiaceae the polygonal crystals stain less densely, probably as a result of the slightly wider spacing of their subunits. The small crystals ofNarthecium are “loose”; that is, much more widely spaced. Such “loose” crystals are commonly found in sieve-element plastids of Velloziaceae, present there in addition to angular crystals, and together with cuneate crystals in a few Lilioids and many taxa of Poales (Commelinoids). Ontogenetic studies of the sieve elements ofSaruma, Aristolochia, and several monocotyledons have shown that in their plastids cuneate crystals develop very early and independent from a polygonal one present in some taxa. Therefore, a conceivable particulation of polygonal into cuneate crystals is excluded. Consequently, mutations of some monocotyledons that contain a lone, large, polygonal crystal in their sieve-element plastids are explained as the result of a complex genetic block. The total result of all studies in sieve-element plastids suggests thatJaponolirion and Tofieldiaceae are the most basal monocotyledons and that Aristolochiaceae are their dicotyledon sister group.     

P-type sieve-element plastids present in members of the tribesTriplareae andCoccolobeae (Polygonaceae) renew the links between thePolygonales and theCaryophyllales

Form-Pfs sieve-element plastids were found inTriplaris, Ruprechtia, andCoccoloba (Polygonaceae) while other genera of the family and those studied from the often associatedPlumbaginaceae contain S-type sieve-element plastids. The rareness of form-Pfs plastids among the angiosperms, their similarity to the peculiar form-P3fs plastids of theChenopodiineae, and the comparatively small plastid diameters measured for all forms present in theCaryophyllales, Polygonales, andPlumbaginales suggest close relationships between these taxa. The restriction inPolygonaceae of form-Pfs plastids to the closely allied tribesTriplareae andCoccolobeae is discussed with regard to both the intrafamilial and ordinal phylogeny, and also considering possible connections to the only magnoliidaean Pfs-taxonCanella. Dedicated to Univ.-Prof. DrF. Ehrendorfer on the occasion of his 70th birthday.     

Functional complementation of the yeast P-type H-ATPase, PMA1, by the Pneumocystis carinii P-type H-ATPase, PCA1

The opportunistic fungus Pneumocystis is the etiologic agent of an interstitial plasma cell pneumonia that primarily afflicts immunocompromised individuals. Like other fungi Pneumocystis maintains a H(+) plasma membrane gradient to drive nutrient uptake and regulates intracellular pH by ATP-dependent proton efflux. Previously, we identified a Pneumocystis gene, PCA1, whose predicted protein product was homologous to fungal proton pumps. In this study, we show by functional complementation in a Saccharomyces strain whose endogenous PMA1 proton pump activity is repressed that the Pneumocystis PCA1 encodes a H(+)-ATPase. The properties of PCA1 characterized in this system closely resemble those of yeast PMA1. Yeast expressing PCA1 grow at low pH and are able to acidify the external media. Maximal enzyme activity (V(max)) and efficiency of substrate utilization (K(m)) in plasma membranes were nearly identical for PCA1 and PMA1. PCA1 contains an inhibitory COOH-terminal domain; removal of the final 40 amino acids significantly increased V(max) and growth at pH 6.5. PCA1 activity was inhibited by proton pump inhibitors omeprazole and lansoprazole, but was unaffected by H(+)/K(+)-ATPase inhibitor SCH28080. Thus, H(+) homeostasis in Pneumocystis is likely regulated as in other fungi. This work also establishes a system for screening PCA1 inhibitors to identify new anti-Pneumocystis agents.     

Protein targeting into plastids: a key to understanding the symbiogenetic acquisitions of plastids

Recent progress in molecular phylogenetics has proven that photosynthetic eukaryotes acquired plastids via primary and secondary endosymbiosis and has given us information about the origin of each plastid. How a photosynthetic endosymbiont became a plastid in each group is, however, poorly understood, especially for the organisms with secondary plastids. Investigating how a nuclear-encoded plastid protein is targeted into a plastid in each photosynthetic group is one of the most important keys to understanding the evolutionary process of symbiogenetic plastid acquisition and its diversity. For organisms which originated through primary endosymbiosis, protein targeting into plastids has been well studied at the molecular level. For organisms which originated through secondary endosymbiosis, molecular-level studies have just started on the plastid-targeted protein-precursor sequences and the targeting pathways of the precursors. However, little information is available about how the proteins get across the inner two or three envelope membranes in organisms with secondary plastids. A good in vitro protein-import system for isolated plastids and a cell transformation system must be established for each group of photosynthetic eukaryotes in order to understand the mechanisms, the evolutionary processes and the diversity of symbiogenetic plastid acquisitions in photosynthetic eukaryotes.     

Protein trafficking to plastids: one theme, many variations

Plastids are a diverse group of essential organelles in plants that include chloroplasts. The biogenesis and maintenance of these organelles relies on the import of thousands of nucleus-encoded proteins. The complexity of plastid structure has resulted in the evolution of at least four general import pathways that target proteins into and across the double membrane of the plastid envelope. Several of these pathways can be further divided into specialty pathways that mediate and regulate the import of specific classes of proteins. The co-ordination of import by these specialized pathways with changes in gene expression is critical for plastid and plant development. Moreover, protein import is acutely regulated in response to physiological and metabolic changes within the cell. In the present review we summarize the current knowledge of the mechanism of import via these pathways and highlight the regulatory mechanisms that integrate the plastid protein-trafficking pathways with the developmental and metabolic state of the plant.     

One,two, three: nature's tool box for building plastids

Summary. Plastids were acquired by different strategies. While in primary endosymbiosis a cyanobacterium was engulfed by a eukaryotic cell and reduced to a plastid, secondarily evolved plastids trace back to an enslaved red or green alga. Nature's recent playground in merging organisms together can be detected in dinoflagellates, which developed additional strategies to acquire their solar-powered factory. Some dinoflagellates possess secondary plastids, other species temporarily use “stolen plastids” of different origin. The highest degree of complexity is reached in dinoflagellates with chloroplasts originating from the uptake of a photosynthetic symbiont with secondary plastids, a process termed tertiary endosymbiosis. Received June 18, 2001 Accepted January 11, 2002     

Specific characters of vessel primary walls during the early stages of wood differentiation

At the peak of its activity, the cambial zone comprises several layers of undifferentiated, apparently identical cells. In order to find criteria indicating the commitment of cambial cells either to phloem or xylem, early changes in primary wall structure and composition were looked for, using sycamore branches as experimental material. Several chemicals were employed to extract cell wall polysaccharides. Treated specimens were studied by electron microscopy after selective staining. Extracted matrix components were analysed through HPLC. Comparison of ultrastructural and biochemical results indicated that in contrast to phloem derivatives cellulose biosynthesis in xylem derivatives was delayed. Among xylem-committed cells, the very young vessels were characterized by a nearly complete lack of a cellulose skeleton and a high amount of xylose-rich hemicelluloses in their primary walls. This organization would cause the wall plasticity necessary for the cell extensive growth in diameter.     

Application of the iterative helical real-space reconstruction method to large membranous tubular crystals of P-type ATPases

Since the development of three-dimensional helical reconstruction methods in the 1960's, advances in Fourier-Bessel methods have facilitated structure determination to near-atomic resolution. A recently developed iterative helical real-space reconstruction (IHRSR) method provides an alternative that uses single-particle analysis in conjunction with the imposition of helical symmetry. In this work, we have adapted the IHRSR algorithm to work with frozen-hydrated tubular crystals of P-type ATPases. In particular, we have implemented layer-line filtering to improve the signal-to-noise ratio, Wiener-filtering to compensate for the contrast transfer function, solvent flattening to improve reference reconstructions, out-of-plane tilt compensation to deal with flexibility in three dimensions, systematic calculation of Fourier shell correlations to track the progress of the refinement, and tools to control parameters as the refinement progresses. We have tested this procedure on datasets from Na(+)/K(+)-ATPase, rabbit skeletal Ca(2+)-ATPase and scallop Ca(2+)-ATPase in order to evaluate the potential for sub-nanometer resolution as well as the robustness in the presence of disorder. We found that Fourier-Bessel methods perform better for well-ordered samples of skeletal Ca(2+)-ATPase and Na(+)/K(+)-ATPase, although improvements to IHRSR are discussed that should reduce this disparity. On the other hand, IHRSR was very effective for scallop Ca(2+)-ATPase, which was too disordered to analyze by Fourier-Bessel methods.     

Diatom plastids: Secondary endocytobiosis, plastid genome and protein import

Plastids of diatoms and other chromophytic algae have four surrounding membranes. In contrast to plastids of green algae, higher plants and red algae chromophytic cells are thought to have evolved by secondary endocytobiosis, i.e. by uptake of a eukaryotic photosynthetic organism by a eukaryotic host cell. This review gives a brief summary of the current views about the origin of diatom plastids and discusses possible mechanisms the cells might employ to transport nucleus-encoded plastid proteins into these organelles.     

The development of specific sieve-element plastids in wound phloem ofColeus blumei (S-type) andPisum sativum (P-type), regenerated from amyloplast-containing parenchyma cells

Protoplasma - In experimentally-induced wound phloem, sieve-element plastids express their genetically determined type in depositing amylopectinrich sieve-tube starch (Coleus, S-type) and polygonal...     

The endosymbiotic origin,diversification and fate of plastids

Plastids and mitochondria each arose from a single endosymbiotic event and share many similarities in how they were reduced and integrated with their host. However, the subsequent evolution of the two organelles could hardly be more different: mitochondria are a stable fixture of eukaryotic cells that are neither lost nor shuffled between lineages, whereas plastid evolution has been a complex mix of movement, loss and replacement. Molecular data from the past decade have substantially untangled this complex history, and we now know that plastids are derived from a single endosymbiotic event in the ancestor of glaucophytes, red algae and green algae (including plants). The plastids of both red algae and green algae were subsequently transferred to other lineages by secondary endosymbiosis. Green algal plastids were taken up by euglenids and chlorarachniophytes, as well as one small group of dinoflagellates. Red algae appear to have been taken up only once, giving rise to a diverse group called chromalveolates. Additional layers of complexity come from plastid loss, which has happened at least once and probably many times, and replacement. Plastid loss is difficult to prove, and cryptic, non-photosynthetic plastids are being found in many non-photosynthetic lineages. In other cases, photosynthetic lineages are now understood to have evolved from ancestors with a plastid of different origin, so an ancestral plastid has been replaced with a new one. Such replacement has taken place in several dinoflagellates (by tertiary endosymbiosis with other chromalveolates or serial secondary endosymbiosis with a green alga), and apparently also in two rhizarian lineages: chlorarachniophytes and Paulinella (which appear to have evolved from chromalveolate ancestors). The many twists and turns of plastid evolution each represent major evolutionary transitions, and each offers a glimpse into how genomes evolve and how cells integrate through gene transfers and protein trafficking.     

Nuclear magnetic resonance of heme protein crystals. General aspects

A new technique capable of determining the static and dynamic structures of heme protein crystals is reported. It is shown that microcrystals of a variety of paramagnetic heme proteins, suspended in approximately 90% saturated (NH4)2SO4, may be perfectly aligned by an intense static external magnetic field, H0, due to the large anisotropy in the magnetic susceptibility of the protein caused by the paramagnetic center. Myoglobin from sperm whale (Physeter catodon) was isotopically enriched at the C epsilon methyl groups of methionine residues 55 and 131 with either 13C or 2H and studied in the crystalline solid state by 2H-quadrupole echo and 13C-Fourier transform nuclear magnetic resonance spectroscopy. It was found that suspensions of both high (S = 5/2) and low (S = 1/2) spin ferric forms of the labeled protein were ordered, the axis of ordering being approximately perpendicular to the low temperature minimum g tensor valve, even though upper Kramers levels are populated at room temperature. The paramagnetic CoII derivative "coboglobin" showed similar ordering behavior, but the diamagnetic carboxymyoglobin was unaffected. The magnetic ordering method permits the recording of "single crystal" NMR spectra from microcrystalline arrays of proteins which cannot be prepared in large enough form (approximately 1 cm3) for single crystal NMR spectroscopy and thereby allows the resolution and assignment of numerous single atom sites in the crystalline solid state. The information from a "single crystal" NMR spectrum combined with that obtained on the crystal powder allows for the direct determination of (i) the spatial orientation of the particular labeled residue within the protein crystal and (ii) the rates and types of side chain motion. Resonances were assigned by spin label broadening experiments and by use of existing x-ray data to predict 2H-NMR spectra. This new technique opens up the possibility of determining directly the dynamic structure of protein crystals and of comparing the structures of proteins in the crystalline solid state with that in solution and is applicable to other heme proteins, e.g. catalase.