LIGHT MICROSCOPE INVESTIGATION OF SIEVE-ELEMENT ONTOGENY AND STRUCTURE IN ULMUS AMERICANA

At maturity the sieve elements of Ulmus americana L. contain a parietal network of very fine strands of slime which is continuous from one sieve element to the next through the sieve-plate pores. Upon injury this parietal network, which is derived from the slime bodies of immature sieve elements, sometimes becomes distorted into longitudinally oriented strands. Some of these strands frequently extend the length of the cells and often are continuous from one sieve element to the next through the sieve-plate pores. At times past such strands have erroneously been interpreted as normal constituents of the mature sieve-element protoplast. Many mature sieve elements of U. americana contain nuclei, which apparently persist for the life of the sieve elements. In addition, some evidence has been found in mature sieve elements for the presence of a membrane which delimits the parietal layer of cytoplasm, including its network of slime strands, from the vacuolar region of the cell.     

ANATOMICAL FEATURES OF METAPHLOEM IN STEMS OF SABAL,COCOS AND TWO OTHER PALMS

Sieve tubes in metaphloem of palm stems function throughout the life of the plant and merit close investigation. A stem of Sabal palmetto estimated to be 50 years old was sampled extensively. Variation in length of sieve-tube elements throughout this stem was measured and is discussed. In the metaphloem of individual vascular bundles companion cells are not sharply differentiated from other phloem parenchyma cells. Definitive callose deposits and slime are normally absent from mature sieve tubes, even in fixed material. Otherwise no conspicuous structural features which might account for the longevity of sieve tubes can be discerned. Occlusion of phloem strands after leaf fall is initially by callose deposition on sieve plates followed immediately by tylosoid formation. Similar sampling of Cocos nucifera, Washingtonia robusta and to a lesser extent Archontophoenix alexandrae confirmed these results except for quantitative differences.     

Observations on Companion Cells and Specialized Phloem Parenchyma Cells of Nelumbo nucifera Gaertn

The phloem of very young petioles of Nelumbo nucifera Gaertn.(Nelumbium speciosum Willd.) was studied with the light microscope.The elongated, mature sieve elements contain slime, plugs, strands,and numerous plastids. Some sieve elements remain nucleatedfor a brief period even after the sieve plates are well developed.The companion cells numbering 8–14 undergo disintegrationbefore the elongation of the ontogenetically related sieve elementis completed. They are uninucleate to begin with but later becomebinucleate and finally degenerate and obliterate. The variousstages in their ontogeny and disintegration are described. Ofthe very few specialized phloem parenchyma cells present, someare associated with sieve elements. They have slime body-likestructures, and plastid-like bodies which group together andeventually disintegrate.     

ULTRASTRUCTURE OF THE SECONDARY PHLOEM OF TILIA AMERICANA

Summer and winter (July and January) samples of secondary phloem of Tilia americana were studied with the electron microscope. Parenchyma cells contain: nuclei, endoplasmic reticulum, ribosomes, plastids, mitochondria and occasional dictyosomes. Well-defined tonoplasts separate vacuoles from cytoplasmic ground substance. Vacuoles often contain tannins. Lipid droplets are common in cytoplasm. Endoplasmic reticulum–connected plasmodesmata are aggregated in primary pit fields. Companion cells differ from parenchyma cells in having numerous sieve-element connections, possibly slime, and in lacking plastids. Mature, enucleate sieve elements possess 1–4 extruded nucleoli. Numerous vesicles occupy a mostly parietal position in association with plasmalemma. The mature sieve element lacks endoplasmic reticulum, organelles (except for few mitochondria) and tonoplast. In OsO_4– and glutaraldehyde-fixed elements, slime has a fine, fibrillar appearance. Normally, these fine fibrils are organized into coarser ones which form strands that traverse the cell and the plasmalemma-lined pores of sieve plates and lateral sieve areas.     

ONTOGENY AND STRUCTURE OF THE SECONDARY PHLOEM IN EPHEDRA

The secondary phloem in Ephedra is atypical of the gymnosperms in general and exhibits several angiosperm-like characteristics. The ray system of the conducting phloem consists of parenchymatous, multiseriate rays. The axial system contains parenchyma cells, sieve cells, and unusual albuminous cells reminiscent of the specialized parenchyma cells found in some angiosperms. These cell types may intergrade with each other. P-protein in the developing sieve element appears early in the form of a single, ovoid slime body. Later, smaller slime bodies appear and quickly disperse. In the mature sieve element the single, ovoid slime body is lost, and P-protein is then evident in the form of a parietal cylinder, thread-like strands, amorphose globules, or a slime plug. Necrotic-appearing nuclei are commonly found in mature sieve cells.     

OBSERVATIONS ON METAPHLOEM IN THE VEGETATIVE PARTS OF PALMS

Metaphloem was studied in available vegetative parts of 374 species in 164 genera of palms. Sieve elements usually have compound sieve plates except in the subfamilies Lepidocaryoideae and Nypoideae. Sieve elements in roots usually have oblique to very oblique end walls, whereas in stems and leaves they have transverse to oblique walls. Within a phloem strand the degree of compounding of a sieve plate is directly correlated with element diameter. Plastids are normally present in functioning, enucleate sieve elements. Small quantities of “slime” substances have been detected in young sieve elements in stems and petioles of a few species. Many sieve plates in functioning sieve elements lacked callose in materials quick-killed in liquid nitrogen or chilled acetic-alcohol. Definitive callose is confined to sieve elements just before their obliteration. Sieve tubes in leaf and stem are usually ensheathed by contiguous parenchyma cells while those in root have very few contiguous parenchyma cells. Two types of contiguous parenchyma cells can be distinguished by difference in cytoplasmic density, especially with the electron microscope. Cells with denser cytoplasm are interpreted as companion cells. Lignified contiguous parenchyma cells are occasionally present in metaphloem of petioles. The possible diagnostic and taxonomic features of metaphloem are discussed.     

Development and Structure of Phloem in the Petiole of Lagenaria siceraria (Mol.) Standl. and Momordica charantia L.

Light microscopic studies of the petioles of Lagenaria sicerariareveal that the external phloem of each bicollateral vascularbundle develops earlier than the internal phloem, and that thesieve elements of the external phloem are arranged in the outerand inner zones. Each sieve element of L. siceraria and Momordicacharantia is vertically associated with a maximum of six andtwo companion cells respectively. Discrete granular bodies seenin the cytoplasm of young sieve elements develop into globular,oval, or elongated slime bodies. Enlargement and fusion of slimebodies, and the subsequent dispersal of slime occur in the parietalcytoplasm. The dispersal of slime coincides with degradationof the nucleus and perforation of the pore sites. Before nucleardisorganization, the sieve-element nucleolus is extruded. Slimeafter its immediate dispersal appears amorphous and uniformlydistributed in the sieve elements. Plugs exhibit varying degreesof condensation of slime near the sieve plates. Certain maturesieve elements in the external phloem of L. siceraria have ovalbodies which we consider reaggregated or undispersed slime.Evidence has been obtained that a central cavity occurs in afew, almost mature, sieve elements wherein the cytoplasm includingthe slime is peripheral.     

The Phloem of Nelumbo nucifera Gaertn

In common with other aquatic angiosperms, Nelumbo nucifera Gaertn.has a relatively strongly developed phloem tissue. The vascularsystem consists of discrete collateral bundles in which no cambiumdevelops and the phloem and xylem are separated by a narrowlayer of parenchyma cells. The phloem consists of sieve elements,companion cells, and phloem parenchyma cells. The sieve elementshave transverse end walls with simple sieve plates. The cellsattain considerable width in the late phloem (metaphloem). Thecompanion cells are in vertical strands. In the early phloem(protophloem) of large bundles the sieve tubes and companioncells are eventually obliterated. The parenchyma cells alsoform vertical strands which may contain tannin cells. Some parenchymacells and companion cells are binucleate. The sieve elementsshow ultrastructural features common for these cells in dicotyledons.At maturity, they lack nuclei, ribosomes, and tonoplasts, butretain a plasmalemma, mitochondria, and plastids. The latterare poorly differentiated and form starch. The endoplasmic reticulumis in part stacked, in part it forms a network next to the plasmalemma.The P-protein occurs in two forms. One consists of tubules notassembled in any specific type of array. The other, possiblycomposed of much extended tubules, is assembled in crystallineaggregates which are retained as such in mature cells. The sieveplate pores are lined with callose and plasmalemma. The lateralwalls are relatively thin and the nacreous layer varies in degreeof distinctness.     

STUDIES ON THE ROOT OF HORDEUM VULGARE L.– ULTRASTRUCTURE OF THE SEMINAL ROOT WITH SPECIAL REFERENCE TO THE PHLOEM

Seminal root tissue of Hordeum vulgare L. var. Barsoy was fixed in glutaraldehyde and osmium tetroxide and studied with the light and electron microscopes. The roots consist of an epidermis, 6–7 layers of cortical cells, a uniseriate endodermis and a central vascular cylinder. Cytologically, the cortical and endodermal cells are similar except for the presence of tubular-like invaginations of the plasmalemma, especially near the plasmodesmata, in the former. The vascular cylinder consists of a uniseriate pericycle surrounding 6–9 phloem strands occurring on alternating radii with an equal number of xylem bundles. The center of the root contains a single, late maturing metaxylem vessel element. Each phloem strand consists of one protophloem sieve element, two companion cells and 1–3 metaphloem sieve elements. The protophloem element and companion cells are contiguous with the pericycle. Metaphloem sieve elements are contiguous with companion cells and are separated from tracheary elements by xylem parenchyma cells. The protoplasts of contiguous cells of the root are joined by various numbers of cytoplasmic connections. With the exception of the pore-plasmodesmata connections between sieve-tube members and parenchymatic elements, the plasmodesmata between various cell types are similar in structure. The distribution of plasmodesmata supports a symplastic pathway for organic solute unloading and transport from the phloem to the cortex. Based on the arrangement of cell types and plasmodesmatal frequencies between various cell types of the root, the major symplastic pathway from sieve elements to cortex appears to be via the companion and xylem parenchyma cells.     

Spatial and temporal regulation of the forisome gene <Emphasis Type="Italic">for1</Emphasis> in the phloem during plant development

Forisomes are protein aggregates found uniquely in the sieve elements of Fabaceaen plants. Upon wounding they undergo a reversible, calcium-dependent conformational switch which enables them to act as cellular stopcocks. Forisomes begin to form in young sieve elements at an early stage of metaphloem differentiation. Genes encoding forisome components could therefore be useful as markers of early sieve element development. Here we present a comprehensive analysis of the developmental expression profile of for1, which encodes such a forisome component. The for1 gene is highly conserved among Fabaceaen species and appears to be unique to this phylogenetic lineage since no orthologous genes have been found in other plants, including Arabidopsis and rice. Even so, transgenic tobacco plants expressing reporter genes under the control of the for1 promoter display reporter activity exclusively in immature sieve elements. This suggests that the regulation of sieve element development is highly conserved even in plants where mature forisomes have not been detected. The promoter system could therefore provide a powerful tool for the detailed analysis of differentiation in metaphloem sieve elements in an unexpectedly broad range of plant species. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Primitive-like metaphloem sieve elements in the aerial stem ofEquisetum hyemale

Summary Internodal metaphloem sieve elements located near the nodes of aerial stems ofEquisetum hyemale contain very oblique end walls. During maturation, the connections, or plasmodesmata, in these walls undergo little or no structural modification. By contrast, the endwall connections uniting the protoplasts of mature sieve elements elsewhere in the aerial stem ofE. hyemale are pores.This work as supported by U.S. National Science Foundation grant GB 31417 to R. F.Evert.     

Preservation of the Tonoplast in Metaphloem Sieve Elements of Embryonic Roots of Zea mays L.

Decortication of embryonic roots of 4- to 5-day-old Zea seedlingsand subsequent chemical fixation permitted comparison of cutand uncut developing sieve elements In a decorticated root wheresieve tubes are not severed, metaphloem sieve elements in latestates of development and some mature sieve elements exhibita highly vacuolate condition When roots are cut or diced inthe course of fixation intact vacuoles are not observed in latestages of sieve-element ontogeny The degree of callose formationat sites of developing sieve-plate pores and in the pores ofmature sieve elements varies greatly with both decorticationand non-decortication treatments Nuclei were not observed insieve elements at the electron microscope level, but they wereseen at the light microscope level in serial sections of sieveelements in the late to mature developmental stages representedAlthough the occurrence and distribution of plastids, mitochondria,endoplasmic reticulum, dictyosomes and nbosomes also vanes insieve elements of decorticated roots, disruption or surgingof sieve-element contents is greater for sieve tubes that aresevered during fixation treatment A discussion is presentedrelating effects of trauma on observed developmental stagesand sieve-element structure Zea mays L, maize, corn, phloem, Sieve elements, tonoplast, ultrastructure     

Structurally,phloem unloading in the maize leaf cannot be symplastic

Developing longitudinal vascular bundles of the leaf blades of maize (Zea mays L. cv. W273) were examined with the transmission electron microscope to determine the frequency of plasmodesmata between the sieve tubes and their neighboring cells. Of particular interest were the protophloem sieve tubes, the first sieve tubes to mature in importing (all large and some intermediate) bundles. The protophloem sieve tubes, most of which lack companion cells, intergrade structurally with the thin-walled metaphloem sieve tubes. Both the protophloem sieve tubes and the thin-walled metaphloem sieve tubes and their companion cells (the sieve tube-companion cell complexes) are virtually isolated symplastically from the rest of the leaf, precluding a symplastic mechanism of phloem unloading in the leaf blade of maize.     

SOLUTE CONCENTRATIONS IN THE PHLOEM AND APEX OF THE ROOT OF ZEA MAYS

Plasmolytic studies utilizing a graded series of mannitol solutions (0.1–1.4 M in 0.1 M increments) were conducted on adventitious roots of Zea mays to determine solute concentrations of cell types at various locations in the root. Results indicated that mature sieve-tube members had the highest solute concentration as determined by their C₅₀ (the estimated mannitol concentration plasmolyzing an average of 50% of a given cell type) of any cell type in the root. In tissue 12 cm from the tip, C₅₀ values calculated for proto- and metaphloem sieve-tube members were 1.15 and 1.19 M, respectively, while in tissue 0.5 cm from the root tip, values for the same cell types were 0.68 and 0.46 M, respectively. The C₅₀ values for sieve elements in tissue 5 cm from the tip were intermediate (1.08 and 1.11 M). Although the companion cells generally plasmolyzed at nearly the same concentrations of mannitol as the sieve elements, their C₅₀ values were slightly lower than adjacent mature sieve elements. The lowest C₅₀ (0.35 M) for any cell type examined was associated with meristematic cells in tissue 0.1 cm from the root tip. Taken collectively, the results indicate that positive concentration gradients exist between mature sieve tubes and meristematic cells of the root apex of maize.     

DEVELOPMENT AND STRUCTURE OF PHLOEM IN THE PETIOLE OF LUFFA CYLINDRICA

Differentiation of external phloem is earlier than that of internal phloem in the young petiole of Luffa cylindrical. For a single sieve-tube element, one to six companion cells are present. The young sieve element shows many globular slime bodies which fuse longitudinally and disperse into the cytoplasm. Simultaneously the nucleus loses its stainable contents and later disorganizes. The contents of the sieve element are in the form of plugs, strands or a granular mass. Undispersed slime in the form of discrete bodies along the lateral walls is also observed. During one stage, at least, the dispersed slime and other contents of a mature sieve element lie at the periphery around a central cavity. A special type of phloem-parenchyma cell shows disorganizing chloroplasts, an extruded nucleolus, and callose on primary pit fields.     

Sieve Element of Impatiens sultanii: 1. Wound Reaction

A Study of wound reaction of the metaphloem of the stem by white-light,fluorescence, and electron microscopy provides evidence forthe structure of mature sieve elements in the intact plant.Starch grains usually are retained in plastids which are locatedagainst the lateral walls of sieve elements and concentratedat both ends of each cell. Slime plugs and dense connectingstrands in the sieve plates seem to result from reactions tocutting or penetration of the killing agent; after appropriatemethods of killing, the contents of a connecting strand maybe only slightly denser, if any, than the milieu on either sideof the sieve plate. A strange accumulation of slime, consistingof streamers directed toward the wound surface from each sieveplate, occurred in tissue boiled immediately after incisionof the phloem. Callose is present in sieve elements of intactplants when the tissue is killed within 4 seconds after injury.Callose is accumulated in response to wounding in added amounts,but only after 5 minutes or more and only within about 15 sieveelements from the wound. Quantities of callose sufficient forplugging the sieve plate accumulate after 30 minutes or more.Sieve-plate callose and deposits on the nearby lateral wallsproduce a cup-shaped mass which is called a cup deposit.     

Expression pattern of cell cycle genes suggests the developmental arrangement of vascular cells in sugarcane strand

Unlike the ordered multiplication of vascular cells deriving from a row of initials in dicotyledons, vascular growth in monocotyledonous vascular strands does not show the procambial pattern but leads to a complex organization of the vascular bundle. Establishment of the bundle should have a specific developmental pattern. The cell cycle conferring cell proliferation represents a active state of growth and development of tissues. Here, we cloned an A-type CDK gene (Sacof;CDKA;1) from sugarcane (Saccharum officinarum cv. ROC16) and confirmed that its encoding protein interacted physically with two sugarcane CYCD4s (Sacof;CYCD4;1 and Sacof;CYCD4;2), which shared only 47% amino acid sequence similarity. The three genes were expressed concurrently in meristems of root tip, stem tip, and young leaf but not in mature leaves. More importantly, they were predominantly expressed in vascular strands of stem tips and young leaves. In stem-tip strands, the expression region extends deep basipetally to where the sieve tube increases in number in the metaphloem and the vessels are produced in the metaxylem showing a pattern of cell division occurring among differentiating or differentiated cells. This pattern suggests a positional determination of vascular cell arrangement in strands during vascular development.     

Ultrastructural indications for coexistence of symplastic and apoplastic phloem loading in Commelina benghalensis leaves

The ultrastructural ontogeny of Commelina benghalensis minor-vein elements was followed. The mature minor vein has a restricted number of elements: a sheath of six to eight mestome cells encloses one xylem vessel, three to five vascular parenchyma cells, a companion cell, a thin-walled protophloem sieve-tube member and a thick-walled metaphloem sieve-tube member. The protophloem sieve-tube member (diameter 4–5 m; wall thickness 0.12 m) and the companion cell originated from a common mother cell. The metaphloem sieve-tube member (diameter 3 m; wall thickness 0.2 m) developed from the same precursor cell as the phloem parenchyma cells. Counting the plasmodesmatal frequencies demonstrated a symplastic continuum from mesophyll to the minor-vein phloem. The metaphloem sievetube member and the phloem parenchyma cells are the termini of this symplast. The protophloem sieve-tube member and companion cell constitute an insulated symplastic domain. The symplastic route, mesophyll to metaphloem sieve tube, appears to offer a path for symplastic loading; the protophloem sieve tube may be capable of accumulation from the apoplast. A similar two-way system of loading may exist in a number of plant families. Plasmodesmograms (a novel way to depict cell elements, plasmodesmatal frequencies and vein architecture) of some other species also displayed the anatomical requirements for two routes from mesophyll to sieve tube and indicate the potential coexistence of symplastic and apoplastic loading.     

Another View of the Ultrastructure of Cucurbita Phloem

The primary phloem of young internodes of Cucurbita maxima wasstudied with the electron microscope. Phloem parenchyma cellsare highly vacuolated and contain nuclei, endoplasmic reticulum,ribosomes, mitochondria, chloro-plasts, and occasional dictyosomes.As compared with parenchyma cells, the most distinctive featuresof companion cells are their extremely dense cytoplasm, lowdegree of vacuolation, lack of chloroplasts, and numerous sieve-elementconnexions. Companion cells contain plastids with few internalmembranes. At maturity the enucleate sieve element is linedby a plasmalemma, one or more cistema-like layers of endoplasmicreticulum, and a membrane which apparently delimits the parietallayer of cytoplasm from a large central cavity. In OsO_4–-andglutaraldehyde-fixed elements, the central cavity is traversedby numerous strands, which run from cell to cell through thepores of sieve plates and lateral sieve areas, and which arederived ontogenetically from the slime bodies of immature cells.Numerous normal-appearing mitochondria are present in the parietallayer of cytoplasm. The pores of sieve plates and lateral sieveareas are lined with cytoplasm. The ultrastructural detailsof young sieve elements differ little from those of other youngnucleate cells. During sieve-element development, the sieveelement increases in vacuolation. At the same time, slime bodiesdevelop in the cytoplasm. With glutaraldehyde fixation, thesebodies often exhibit a double-layered limiting membrane. Asthe sieve element continues to differentiate, the slime bodiesincrease in size and the parietal layer of cytoplasm becomesvery narrow. Presently, the slime bodies begin to disperse andtheir contents fuse. This phenomenon occurs in the parietallayer of cytoplasm, while the latter is still delimited fromthe large central vacuole by a distinct tonoplast. The initiationof slime-body dispersal more or less coincides with perforationof the pore sites, and many pores are traversed by slime earlyin their development. Before slime-body dispersal, all dictyosomesand associated vesicles disappear from the cytoplasm. Eventually,the tonoplast diappears and the slime becomes distributed throughoutthe central cavity in the form of strands. Nuclei and ribosomesdisappear before breakdown of the tonoplast. Sieve elementsare connected with companion cells and parenchyma cells by plasmodesmata.     

Ricerche Sull'infrastruttura del Coleottile di Avena

Abstract

Researches on ultrastructure of Avena coleoptile. 3. The sieve elements. — A study on the ultrastructural organization of the mature sieve elements of Avena coleoptile has been carried out. Data suggest that functional phloem tubes are alive and remain alive until they are working. Judging on morphological basis, the metabolic activity of sieve elements should be of peculiar type and low in comparison to that of the companion cells. In fact the cytoplasm is located in a narrow parietal strand, mitochondria, Golgi apparatus and endoplasmic reticulum are present, but they appear very modified; plastids and nucleus are absent. The cytoplasm is bounded externally by a normal plasmalemma, whilst the vacuole has no visible limits: a tonoplast is, therefore not identifiable.

The strands connecting the superimposed sieve elements with one another through the sieve plate result to be made by a double membrane system very similar to the endoplasmic reticulum, which we believe to realize cytoplasmic continuity between phloem tubes.

The data reported are more favorable to the existence in the sieve tubes of an active mechanism of translocation of organic solutes than a passive mass-flow.

The collaboration of companion cells in the translocation mechanism has been discussed.