The occurrence of amoeboid plastids in the actinorhizal root nodules of Alnus glutinosa (L.) Gaertn.

Abstract. The invasion of the actinomycete Frankia into the root cells of Alnus glutinosa with subsequent nodule formation effects a number of ultrastructural changes in the host cell cytoplasm. Among other changes the amyloplasts rapidly lose their starch and acquire an amoeboid or pleomorphic form. Such plastids occur predominantly in the mature vesicle-containing, nitrogen-fixing cells of the nodule. They lack starch, have an electron dense stroma and a complex lamellar system. This last would appear to be associated with a distinct membranous reticulum which can be extensive. The flexible form of these plastids is mirrored in their ability to enclose portions of host cytoplasm together with organelles and even other plastids. Their close association with cristate mitochondria suggests an active metabolic role in the nodule symbiosis.     

Ultrastructure of organelles during microsporogenesis inTillandsia pallidoflavens (Bromeliaceae)

InTillandsia pallidoflavens none of the organelles undergoes fundamental de- and redifferentiation during microsporogenesis. The plastids are amoeboid, exhibit complex internal structures and gradually start accumulating polysaccharides from meiotic prophase I onwards. These observations contradict reports for other taxa. The ultrastructure of mitochondria and dictyosomes, respectively, is more or less orthodox. The extensive ER, which is only poorly stained by standard methods was identified by image intensifiying techniques. The ribosomes are not only associated with the ER or occur as polyribosomes free in the cytoplasm, but can also form more or less dense clusters.     

Evidence for intracellular spatial separation of hexokinases and fructokinases in tomato plants

Four hexokinase (LeHXK1–4) and four fructokinase (LeFRK1–4) genes were identified in tomato plants. Previous GFP fusion studies indicate that the gene product of LeHXK3 is associated with the mitochondria while that of LeHXK4 is located within plastids. In this study we found that the enzyme encoded by the fructokinase gene LeFRK3 is also located within plastids. The presence of LeFrk3 enzyme in plastids raises the question of the origin of fructose in these organelles. The other three FRKs enzymes, LeFrk1&2&4, are located in the cytosol. Unlike LeFrk1&2&4, the two additional HXKs, LeHxk1&2, share a common membrane anchor domain and are associated with the mitochondria similar to LeHxk3. The difference in the locations of the cytoplasmic FRK and HXK isozymes suggests that glucose phosphorylation is confined to defined special intracellular localizations while fructose phosphorylation is less confined.Contribution from the Agriculture Research Organization, The Volcani Center, Bet Dagan, Israel, No. 126/2006 series.     

The biology ofChlamydomyxa montana: Ultrastructure of the cyst

Summary Cells ofChlamydomyxa montana Lankester photosynthesize within a cyst under bright light and ingest diatoms in an excysted amoeboid form during periods of low light intensity. Cyst cell walls are partially comprised of cellulose and vary in thickness. The cells are multinucleate and intracellular bacteria are closely associated with the nuclei. A pair of centrioles is also present adjacent to individual nuclei. Chloroplasts are numerous and thylakoids are generally organized into bands of three. No endoplasmic reticulum was found surrounding the plastids, nor was a complete girdling lamella ever observed within.C. montana chloroplasts appear to be its own but this protist does not precisely fit into an algal class.     

Amyloplast development in etiolated and ethylene-treated pea epicotyls

The amyloplasts found in the apical hook cells of etiolated pea (Pisum sativum L.) epicotyls were randomly distributed. Sedimentation of endodermal amyloplasts in the direction of gravity became apparent in the transition from the hook to the top of the main axis of the epicotyl. Cortical amyloplasts in this region were not, however, sedimented. These patterns of sedimentation could not be related to changes in amyloplast size, and it is proposed that cytoplasmic properties determine amyloplast behaviour.The differentiation of plastids in the hook differed between the amyloplast-containing endodermal cells and the cortical cells, in which amoeboid plastids predominated over amyloplasts. Amyloplasts disappeared from the cortical cells in the main axis of the epicotyl, but in the endodermal cells sedimented amyloplasts were found throughout the upper epicotyl.Etiolated epicotyls induced to grow horizontally by treatment with ethylene had a normal content of amyloplasts, sedimented in the direction of gravity.     

Ultrastructure and autoradiography of dormant and activated parenchyma ofHelianthus tuberosus

Summary Parenchyma cells of dormant tubers ofHelianthus tuberosus L. cv. OB1 (Jerusalem artichoke) contain a very low amount of hormones, therefore they respond to 2,4-D or IAA treatment by dividing and synthesizing RNA, DNA, and polyamines.In particular the activation of the dormant tissues induces an early synthesis of DNA, which reaches the maximum at 3 hours, much before the beginning of the S phase (12 hours). By supplying [6-³H] thymidine and carrying out electron microscopic autoradiography, we were able to determine that plastids and mitochondria were the organelles responsible for this early synthesis while the DNA in the nucleus first appeared labeled at 15 hours.In addition, ultrastructural observations carried out to compare the dormant cells with activated ones, showed an increase in the nucleolar volume, a different organization of the tubular complex of the plastids and several other ultrastructural changes which indicate that at 3 hours some fundamental metabolic processes are already active; they become even more evident later on.The implications of these results in the physiology of the tuber cells during activation are discussed.     

Rapid separation of the plastid,mitochondrial, and cytoplasmic fractions from intact leaf protoplasts of Avena

Purified intact protoplasts were isolated from etiolated and greening leaves of Avena sativa. They were ruptured by forcing them through a 20-m aperture nylon net and immediately thereafter fractionated into a pure pellet of plastids (well above 70% of total plastids), a layer of mitochondria only slightly contaminated by other cellular constituents (about 50% of total mitochondria), and a cytoplasmic supernatant. This was achieved within 60 s by an integrated method of homogenation of protoplasts and centrifugal filtration of the homogenate on a gradient of silicone oils, contained together with the nylon net in 450 l microtubes, and verified by comparing the levels of activity of specific markers within the three fractions obtained. With appropriate modifications to immediately quench metabolic reactions within the fractions, this method allows the determination of metabolite levels within plastids, mitochondria, and the cytoplasmic compartment of intact protoplasts. The applicability of this technique is demonstrated by the determination of ATP in the plastids, mitochondria, and the cytoplasm of protoplasts obtained from etiolated and greening primary leaves of Avena. The levels of ATP, corrected for contamination of the fractions by each other, exhibit a pronounced transient increase during greening, especially within the cytoplasm.Abbreviations BSA bovine serum albumin - Cyt c cytochrome c - EDTA ethylenediamine tetraacetic acid - HEPES N-2-hydroxyethyl-piperazine-N-2-ethane sulphonic acid - MES 2(N-morpholino) ethane sulphonic acid - PGA 3-phosphoglyceric acid - PEP phosphoenol pyruvic acid - RuBP ribulose-1.5-bis-phosphate     

Microgametogenesis in Plumbago zeylanica (Plumbaginaceae). 2. Quantitative cell and organelle dynamics of the male reproductive cell lineage

Quantitative cell and organelle dynamics of the male gamete-producing lineage of Plumbago zeylanica were examined using serial transmission electron microscopic reconstruction at five stages of development from generative cell inception to sperm cell maturity. The founder population of generative cell organelles includes an average of 3.88 plastids, 54.9 mitochondria, and 3.7 vacuoles. During development the volume of the pollen grain increases from 6,200 μm³ in early microspores to 115,000 μm³ at anthesis, cell volume of the male germ lineage decreases more than 67% from 362.3 μm³ to 118.4 μm³. By the time the generative cell separates from the intine, plastid numbers increase by >600%, mitochondria by 250%, and vesicles by 43 times. A cellular projection elongates toward and establishes an association with the vegetative nucleus; this leading edge contains plastids and numerous mitochondria. When the generative cell completes its separation from the intine, organellar polarity is reversed and plastids migrate to the opposite pole of the cell. Cytoplasmic microtubules are common in association with cellular organelles. Plastids accumulate at the distal end of the cell as a linked mass, apparently adhered by lateral electron dense regions. Before division of the highly polarized generative cell, plastids decrease in number by 16%, whereas mitochondria increase by ∼90% and vacuoles increase by ∼140% from the prior stage. After mitosis, the resultant sperm cells differ in size and organelle content. The sperm cell associated with the vegetative nucleus (S_vn) contains 62.7% of the cytoplasm volume, 87% of the mitochondria, 280.4 vesicles (79% of those in the generative cell), and 0.6% of the plastids. At maturity, the S_vn mitochondria increase by 31% and the cell contains an average of 0.4 plastids, 158.9 vesicles, and 0.36 microbodies. The mature unassociated sperm (S_ua) contains 39.8 mitochondria (up 3.3%), 24.3 plastids (down 31%), 91.1 vesicles (up 54.9%), and 3.18 microbodies. The small number of organelles initially in the generative cell, followed by their rapid multiplication in a shrinking cytoplasm suggests a highly competitive cytoplasmic environment that would tend to eliminate residual organellar heterogeneity. Cell and cytoplasmic volumes vary as a consequence of fluctuations in the number and size of large vesicles or vacuoles, as well as loss of cytoplasmic volume by (1) formation of “false cells” involving amitotic cytokinesis, (2) “pinching off” of cytoplasm, and (3) dehydration of pollen contents prior to anthesis.     

Acid phosphatase activity in plastids (plastolysomes) of senescing embryo-suspensor cells

Autolysis of the suspensor, an embryonal haustorium, starts in the basal cells and proceeds in the direction of the embryo. InPhaseolus vulgaris, acid phosphatase activity is first found in transforming plastids, similar to the acid phosphatase activity inPh. coccineus [Nagl (1977) Z. Pflanzenphysiol.85, 45–51], although the ultrastructural details are different. InTropaeolum majus, autolysis begins in the most distal part of the suspensor, i.e., the chalazal or carpel haustorium. First the endoplasmic reticulum shows acid phosphatase activity, but neither the mitochondria, which undergo transformation similar to that observed in plastids ofPhaseolus, nor the leucoplasts show such activity. Later, however, the plastids exhibit low activity. Contrarily, the plastids in the suspensor cells adjacent to the embryo show increasing activity during senescence of the suspensor. During final autolysis, activity is found in all cytoplasmic membranes, while it is reduced in plastids. The visible ultrastructural transformations of various organelles into cytolysomes does not necessarily coincide with acid phosphatase activity. Our findings are a further indication of the high diversification and specialization of plastids during plant embryogenesis.     

Plastid degeneration induced by vanadium

In the cells of primary cortex in the roots of horse bean and of root cap of maize, vanadium induced striking changes in the shape of plastids,i.e. the origin of amoeboid, ribbonlike plastids and plastid protrusions.     

Cytological features of developing embryogenic callus and somatic embryos of Iris pumila L. and Iris setosa Pall

Abstract

Cytological examination during somatic embryogenesis in Iris pumila L. and Iris setosa Pall, were performed using light and electron microscopy. The first sign of the cellular differentiation in the initial embryogenic callus (EC; stage 1) of both Iris species was the formation of short and elongated cell types. After the onset of embryogenesis, short cells divided producing a mass of densely packed meristematic cells, closely connected with numerous plasmodesmata. Further differentiation into globular embryos (GE) led to a loss of plasmodesmata and cell separation. In vacuolated elongated cells, cytoplasm was located near the wall and around the nucleus. In both cell types amyloplasts and small mitochondria with poorly developed crystae were abundant.

Cell of GE (stage 2) contained an increased number of mitochondria and plastids comparing to those from stage 1, indicating further differentiation. Thylakoids and starch grains were observed within the plastids, while the number of cristae within the mitochondria was increasing.

In cells of embryos with coleoptile (ECl) (stage 3), plastids differentiated into chloroplasts with thylakoids. In all stages of cell differentiation, short and long cisternae of endoplasmic reticulum with ribosomes were seen. Activity of dictyosomes was increased in stages 1 and 2, then reduced in stage 3.

Ultrastructure of EC cells was identical to that of proembryogenic cells, i.e. of early GE. Ultrastructural appearance of GE cells was identical in both Iris species, but evident, and increasing, differences in mitochondria and plastids were observed between GE and ECl embryos.The presence of bi-, three- and eight- cell proembryos demonstrates that they originate from a single cell in both Iris species.     

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.     

NaCl胁迫对宁夏枸杞叶和幼根显微及超微结构的影响

以宁夏枸杞为材料,采用超薄切片技术制备样品,应用光学显微镜和透射电镜分析了不同浓度NaCl胁迫条件下宁夏枸杞叶和幼根显微及超微结构的变化。结果表明：随着NaCl胁迫的加重,(1)叶片上表皮细胞增厚,栅栏组织细胞出现缩短现象,排列疏松且紊乱;幼根的初生结构无明显变化。(2)叶片栅栏组织中叶绿体不再紧靠在细胞膜上,叶绿体双层膜破坏,基粒片层松散排列,杂乱无章,出现膨胀和空泡现象,淀粉粒和嗜锇颗粒增多,叶肉细胞中线粒体发生轻微变化;幼根中皮层薄壁细胞线粒体形状发生改变,结构破坏,内膜和外膜模糊甚至破裂,大多数嵴模糊,出现空泡现象;细胞核解体,基质外溢。研究表明, 不同浓度的NaCl胁迫对宁夏枸杞叶片和幼根细胞的显微及超微结构影响不同,NaCl浓度大于200 mmol/L时,宁夏枸杞叶片和幼根细胞的显微及超微结构发生了明显变化,且叶肉细胞中线粒体的变化没有叶绿体的变化显著,推测叶肉细胞中线粒体的耐盐性比叶绿体强。     

Chloroplast development in low light-grown barley seedlings

Segments of 7-d low light-grown barley laminae cut at 0.5 cm intervals up from the intercalary meristem were examined ultrastructurally and biochemically. The different regions upwards showed the succession of plastid development in light-grown tissues of eoplasts, amyloplasts, amoeboid, immature and mature plastids as described by Whatley (1977). Semi-crystalline bodies were detected in all of them. The eoplast-amyloplast regions are characterised by a greater proportion of mitochondria and high levels of ATP and 3-phosphoglyceric acid, together with low levels of inorganic phosphate conducive to the activation of ADP glucose pyrophosphorylase. The amoeboid and immature plastid regions have higher levels of inhibitory phosphate and starch breakdown may be responsible for the release of metabolites and energy for development. Segments containing amoeboid and immature plastids also have reduced levels of ATP (and 3-phosphoglyceric acid) as photosynthetic components are synthesised. Using ultrastructural assessments of areas of thylakoids, first -carotene and violaxanthin, followed by chlorophyll a and lutein and, lastly, chlorophyll b are concentrated in the developing lamellar systems of the immature and mature chloroplasts. The formation of additional membraneous material which spreads these pigment systems over a greater thylakoid area within the plastids is the final stage of plastid morphogenesis in low light-grown seedlings.Abbreviations Chl chlorophyll - 3-PGA 3 phosphoglyceric acid     

Paternal cytoplasmic transmission in Chinese pine (Pinus tabulaeformis)

Summary. In this paper, the stages of normal sexual reproduction between pollen tube penetration of the archegonium and early embryo formation in Pinus tabulaeformis are described, emphasizing the transmission of parental cytoplasm, especially the DNA-containing organelles – plastids and mitochondria. The pollen tube growing in the nucellus contained an irregular tube nucleus followed by a pair of sperm cells. The tube cytoplasm contained abundant organelles, including starch-containing plastids and mitochondria. The two sperm cells differed in their volume of cytoplasm. The leading sperm, with more cytoplasm, contained abundant plastids and mitochondria, while the trailing one, with a thin layer of cytoplasm, had very few organelles. The mature egg cell contained a great number of mitochondria, whereas it lacked normal plastids. At fertilization, the pollen tube penetrated into the egg cell at the micropylar end and released all of its contents, including the two sperms. One of the sperm nuclei fused with the egg nucleus, whereas the other one was retained by the receptive vacuole. Very few plastids and mitochondria of male origin were observed around the fusing sperm and egg nuclei, while the retained sperm nucleus was surrounded by a large amount of male cytoplasm. The discharged tube cytoplasm occupied a large micropylar area in the egg cell. In the free nuclear proembryo, organelles of maternal and paternal origins intermingled in the neocytoplasm around the free nuclei. Most of the mitochondria had the same features as those of the egg cell, but some appeared to be from sperm cells and tube cytoplasm. Plastids were obviously of male origin, with an appearance similar to those of the sperm or tube cells. After cellularization of the proembryo, maternal mitochondria became more abundant than the paternal ones and the plastids enlarged and began to accumulate starch. The results reveal the cytological mechanism for paternal inheritance of plastids and biparental inheritance of mitochondria in Chinese pine. Correspondence and reprints: State Key Laboratory of Plant Physiology and Biochemistry, College of Biological Science, China Agricultural University, Beijing 100094, People’s Republic of China.     

Cytological evidence for preservation of mitochondrial and plastid DNA in the mature generative cells of Chlorophytum spp. (Liliaceae)

Summary.  Following 4′,6-diamidino-2-phenylindole staining of mature pollen grains of Chlorophytum comosum, fluorescence microscopy confirmed that cytoplasmic nucleoids (DNA aggregates) were present in the generative cells, which indicated the possibility of biparental cytoplasmic inheritance. Electron and immuno-electron microscopy showed that both plastids and mitochondria were present in the generative cells, and both organelles contained DNA. These results indicate that mitochondria and plastids of C. comosum have the potential for biparental inheritance. Similar results were obtained with mature pollen grains of C. chinense. Therefore, we conclude the coincident biparental inheritance for mitochondria and plastids in the members of the genus Chlorophytum. Received June 28, 2002; accepted September 26, 2002; published online April 2, 2003 RID="*" ID="*" Correspondence and reprints: College of Life Science, Peking University, Bejing 100871, People's Republic of China.     

Quantitative analysis of ultrastructural changes during zygotic and somatic embryogenesis in pearl millet (Pennisetum glaucum [L.] R. Br.)

Ultrastructural changes during zygotic and somatic embryogenesis in pearl millet (Pennisetum glaucum [L.] R. Br.) were quantified using morphometric techniques. The total area per cell profile and the cell volume percentage of the whole cell, endoplasmic reticulum (ER), Golgi bodies, mitochondria, nuclei, lipids, plastids, starch grains and vacuoles were measured and comparisons made between three zygotic and three somatic embryo developmental stages. All measurements were taken from scutellar or scutellar-derived cells. Zygotic embryogenesis was characterized by increases in cell size, lipids, plastids, starch, Golgi bodies, mitochondria and ER. Somatic embryogenesis was characterized by two phases of cell development: (1) the dedifferentiation of scutellar cells involving a reduction in cell and vacuole size and an increase in cell activity during somatic proembryoid formation and (2) the development of somatic embryos in which most cell organelle quantities returned to values found in late coleoptile or mature predesiccation zygotic stages. In summary, although their developmental pathways differed, the scutella of somatic embryos displayed cellular variations which were within the ranges observed for later stages of zygotic embryogenesis.     

Ultramorphometric study of the plastids in apical tuber cells of original and transgenic potato plants treated with ambiol

Ultramorphometric characteristics of plastids in cells of apical tuber meristems of original and defensin gene-transfected potato (Solanum tuberosum L.) plants, either maintained under normal conditions or subjected to treatment with the antioxidant ambiol, were compared. Under normal conditions, the tuber cells of the original and transgenic potato plants differed in neither the number nor size of the plastids. Only certain quantitative distinctions in the development of individual ultrastructural characteristics of plastids were detected. Treatment with ambiol enhanced the differentiation of the internal membrane system of plastids in the cells of original and transgenic plants, especially the tubular membrane systems. Certain differences in the responses to ambiol of cell plastids of original and transgenic plants were related to plastid sizes and development of individual intraplastid structures. The results comply with earlier data on varying responses of mitochondria of original and transgenic plants to ambiol treatment.__________Translated from Prikladnaya Biokhimiya i Mikrobiologiya, Vol. 41, No. 3, 2005, pp. 330–339.Original Russian Text Copyright © 2005 by Platonova, Evsyunina, Belikov, Korableva.     

Presence of plastid and absence of mitochondrial DNA in male reproductive cells as evidence for cytoplasmic inheritance in Turnera ulmifolia and Zantedeschia aethiopica

Summary. Epifluorescence microscopy of mature pollen grains of Turnera ulmifolia and Zantedeschia aethiopica stained with 4,6-diamidino-2-phenylindole demonstrated the presence of fluorescent cytoplasmic DNA aggregates in the male reproductive cells of both species. Double staining of the cells with 4,6-diamidino-2-phenylindole and 3,3-dihexyloxacarbocyanine iodide in Technovit resin sections showed that the mitochondria of these cells did not correspond to the fluorescent cytoplasmic DNA aggregates. Electron microscopy studies revealed both plastids and mitochondria in the cells of these species. In addition, immunoelectron microscopy using an anti-DNA monoclonal antibody showed clear labeling of plastids but not mitochondria. These data provide cytological evidence for biparental plastid inheritance and maternal mitochondrial inheritance in these species.Correspondence and reprints: College of Life Sciences, Peking University, Beijing 100871, Peoples Republic of China.     

Subcellular location of lincomycin resistance in Nicotiana mutants

Summary Lincomycin-resistant Nicotiana plumbaginifolia plastid mutants were considered also to carry mitochondrial mutations on the basis of their ability to grow in the dark under selective conditions. To clarify the role of mitochondria, individual protoplasts of the green, lincomycin-resistant N. plumbaginifolia mutant LR400 were microfused with protoplasts of the N. tabacum plastid albino line 92V37, which possesses N. undulata cytoplasm. The production of lincomycin-resistant albino cybrid lines, with N. undulata plastids and recombinant mitochondria, strongly indicated a determining role for mitochondria in the lincomycin resistance. Sequence analysis of the region encompassing putative mutation sites in the 26S rRNA genes from the LR400 and several other lincomycin-resistant N. plumbaginifolia mutants revelaed, however, no differences from the wild-type sequence. As an alternative source of the resistance of the fusion products, the N. tabacum fusion partner was also taken into account. Surprisingly, a natural lincomycin resistance of tobacco was detected, which was inherited as a dominant nuclear trait. This result compromises the interpretation of the fusion data suggested above. Thus, to answer the original question definitively, the mutant LR400 was crossed as a female parent with a N. plumbaginifolia line carrying streptomycin-resistant N. tabacum plastids. Calli were then induced from the seedlings. Occasional paternal plastid transmissions were selected as streptomycin-resistant calli on selective medium. These cell lines were shown by restriction enzyme analysis to contain paternal plastids and maternal mitochondria. They were tested for greening and growing ability in the presence of lincomycin. These resistance traits proved to be genetically linked and exclusively located in the plastids.EMBL accession number X68710