Changes in endoplasmic reticulum during spermiogenesis in the mouse

Summary Changes in the endoplasmic reticulum of mouse spermatids during spermiogenesis were examined by scanning electron microscopy, applying the OsO₄-DMSO-OsO₄ method, which permits 3-dimensional observation of cell organelles. At the same time, the endoplasmic reticulum was stained selectively by the Ur-Pb-Cu method, and 0.5 m-thick sections were prepared for observation by transmission electron microscopy. The results demonstrated stereoscopically the mode of disappearance of the endoplasmic reticulum. In spermatids of the early maturation phase, the endoplasmic reticulum was of uniform diameter, branched and anastomosed, forming a complicated three-dimensional network throughout the cytoplasm. A two-dimensional net was also noted to have formed just beneath the plasma membrane and about Sertoli cell processes invaginating the spermatid cytoplasm. As spermiogenesis progressed, the spread-out endoplasmic reticulum gradually aggregated to form a condensed, glomerulus-like structure consisting of a very thin endoplasmic reticulum connected to the surrounding endoplasmic reticulum. This structure corresponds to the so-called radial body. Thus, the endoplasmic reticulum may aggregate, condense, be transformed into a radial body, and be removed from the cytoplasm. The two-dimensional endoplasmic reticulum-net, just beneath the plasma membrane and surrounding processes of Sertoli cells, disappeared in spaces where the three-dimensional endoplasmic reticulum network was scarce. Both the two-dimensional endoplasmic reticulum-net structure and the three-dimensional endoplasmic reticulum network disappeared at the same time, indicating that they may be closely related.     

Ultrastrukturen der zellulären Autophagie

Zusammenfassung Im Laufe der Cytoplasmareduktion während der Spermiogenese von Eisenia foetida sind zwei Vorgänge zu unterscheiden: 1. Die Bildung autophagischer Vakuolen. Sie entstehen, indem Teile des Grundcytoplasmas in das Kompartiment des ER verlagert werden. Da sie keine Reaktion auf saure Phosphatase geben, sind sie als nicht lysosomale Anfangstadien der zellulären Autophagie zu betrachten. 2. Die Bildung primärer Lysosomen. Sie entstehen in Form von lytische Enzyme enthaltenden Golgivesikeln, die von einer neu im Cytoplasma entstehenden Membran zu größeren Einheiten zusammengefaßt werden: den multivesicular bodies. Autophagische Vakuolen und multivesicular bodies gelangen ins Cytophor das am Ende der Spermiogenese den Charakter eines ausgedehnten Autophagosoms annimmt. Als Struktureigentümlichkeit entstehen in ihm undulierende Tubulikörper. Der coat an den Hüllmembranen junger multivesicular bodies und am Plasmalemm der Spermatidenverbindung zum Cytophor wird in Zusammenhang mit der Membrandifferenzierung diskutiert.

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Ultrastructural equivalents of cellular autophagyElectronmicroscopical observations on spermatids of Eisenia foetida during the cytoplasmic reduction

Summary During the cytoplasmic reduction phase in the spermiogenesis of Eisenia foetida two different processes may be defined: 1. The formation of autophagic vacuoles, which arise by the displacement of cytoplasmic portions into the cisternae of the endoplasmic reticulum. Since they exhibit no acid phosphatase activity they are considered to be early stages in cellular autophagy. 2. The formation of primary lysosomes. They originate in Golgi vesicles and are then enveloped by a membrane, formed in the cytoplasm de novo, which transforms them into multivesicular bodies. Autophagic vacuoles and multivesicular bodies subsequently transfer to the cytophor, which contains at the end of the spermiogenesis the characteristics of a large autophagosom, showing aggregates of undulating tubules. The outer coat of the limiting membranes in the early multivesicular bodies and of the cell membrane of the connecting piece between spermatid and cytophor appear to be associated with the membrane development.

     

The endomembrane system of differentiating carposporangia in the red algaChondria tenuissima: Occurrence and participation in secretion of polysaccharidic and proteinaceous substances

Summary The endomembrane system during carposporogenesis inChondria tenuissima was studied using electron microscopy and histochemistry. Profiles of the nucleus are convoluted, resulting in a highly increased surface area. Stacked cisternae are found within the peripheral part of the nucleus. Vesicles, tubules and membrane bound fibrillar bodies occur within the nucleoplasm. The endoplasmic reticulum surrounds the nuclear envelope.The endoplasmic reticulum and the Golgi apparatus, together with small transition vesicles, represent a functional unit. They form two different secretory substances during carposporogenesis. In young stages, carbohydrates are produced by normal dictyosomes within large, normal exocytotic Golgi vesicles. They do not react positively with PAS or Thiéry method and are believed to represent cell wall material. In later stages, the central area of the Golgi cisternae becomes filled with electron dense material. The individual cisternae are transformed into cored vesicles at the trans-face of the dictyosomes. The dense core of the vesicles is proteinaceous and stains with coomassie brilliant blue R. The peripheral fibrillar material is polysaccharidic and reacts positively using the Thiéry method. The contents of the cored vesicles are believed to participate in carpospore attachment. The ER gives rise to cytolysosomes in which starch grains are sequestrated and digested. Mucilaginous sacs seem to be similarly formed.     

Elektronenmikroskopische Beobachtungen an den Sehzellen des Blutegels,Hirudo medicinalis L.

Summary The visual cell of the leech (Hirudo medicinalis) contains a big vacuole filled with a moderate dense substance (vitreous body). The wall of the vacuole consists of a system of microvilli (brush border) merging into the substance of the vitreous body. The cytoplasmic zone underneath the brush border has a special structure consisting of flattened sacs and vesicles giving a radial striation to this cell region.The cytoplasm is filled with a great number of mitochondria and small vesicles. Bundles of fine filaments are striking features of the cytoplasm; in the periphery of the cell they are in close contact with the cell membrane (half desmosomes). Elements of the endoplasmic reticulum and free ribosomes may also be present. The cells regularly contain membrane-bounded inclusions having a dense granular or cristalline content.The visual fibers (processes of the visual cells) show a certain similarity to the unmyelinated nerve fibers of higher animals. They are ensheathed into sheath cells possibly of glial nature. Processes of these cells often surround the visual cells and are sometimes embedded in their cytoplasm. Both visual and sheath cells are covered with a homogeneous basement membrane.The possible role of the structures and the problems of conduction from the brush border onto the cell surface are discussed.     

The distribution and form of the endoplasmic reticulum during spermatogenesis in the crayfish,Cambaroides japonicus

Summary The form and differentiation of the endoplasmic reticulum has been studied in the developing sperm of the crayfish, Cambaroides japonicus. Throughout development a relationship between the nuclear envelope and cytoplasmic portion of the endoplasmic reticulum has been shown to exist. Furthermore, large contributions of material from the nuclear envelope to extranuclear cytoplasmic systems has been noted in the development of early spermatids and nearly mature sperm.A sequential predominance of several types of endoplasmic reticulum has been described in the differentiating sperm. An agranular vesicular reticulum is the most common in the early stages although annulate lamellar stacks and rough surfaced stacks are scattered randomly throughout the cytoplasm. Blebs of the nuclear envelope appear to contribute rough surfaced reticulum to the cytoplasmic system in the early spermatid. A fusion of vesicular elements results in the formation of the dense filamentous reticulum which is typical of the nearly mature sperm. Densely packed lamellae develop on the nuclear envelope in the maturing sperm and are connected to both the nuclear envelope and filamentous endoplasmic reticulum. The possible relationships of these lamellar groups to mitochondria or Golgi is discussed.Supported in part by Grant No B-2314 of the National Institute of Neurological Diseases and Blindness, U.S. Public Health Service.Predoctoral Research Fellow of the National Institute of Neurological Diseases and Blindness, U.S. Public Health Service.     

Ultrastructural localization of cellular compartments involved in secretion of the low molecular weight,alkaline xylanase by Trichoderma reesei

The intracellular location of the low-molecular weight, alkaline xylanase (XYN II) of Trichoderma reesei RUT C-30 was investigated during growth on xylan, using immunoelectron microscopy. A monoclonal antibody, produced against XYN II, was used for this purpose. The enzyme was found at the endoplasmic reticulum and in electron dense 0.2 to 0.8 m vesicles, as well as in the vacuole, at the plasma membrane and in the fungal cell-wall. No staining occured in the cytoplasm, the mitochondria and the nucleus. No Golgi-like structures could be seen. Addition of the carboxylic ionophore monensin blocked xylanase as well as total protein secretion. The results are discussed with respect to XYN II being secreted by T. reesei via a pathway involving the endoplasmic reticulum and secretory vesicles and/or the vacuole.     

Characterization of the antigen recognized by a monoclonal antibody MN9: unique transport pathway to the equatorial segment of sperm head during spermiogenesis

Summary MN9, a monoclonal antibody raised against mouse spermatozoa, specifically recognizes the equatorial segment of sperm head in several mammalian species, including humans. Colloidal gold-immuno-electron microscopy of mouse spermatozoa has shown that the antigen is localized in the space between the outer and inner acrosome membranes and on the acrosome membranes at the equatorial segment. Immunoblotting after electrophoresis of spermatozoa from the cauda epididymidis has identified two immunoreactive bands: 38 kDa and 48 kDa in mouse, and 48 kDa in rat. During spermiogenesis in rat, this antigen is transported to the equatorial segment via a unique pathway, first appearing in some cisternae of the endoplasmic reticulum and in the Golgi apparatus of spermatids at around step 3. The antigen can further be found on the vesicles at thetrans-side of the Golgi apparatus, in the matrix of the head cap, and on the head cap membrane in step-4 to step-7 spermatids. The antigen appears to be concentrated at the equatorial segment during late spermiogenesis. Neither the (pro-)acrosomic granule nor the surrounding membrane are required in this pathway. This pathway can be termed the Golgi-head cap tract.     

β-1-4-and β-1-3-glucan synthases are associated with the plasma membrane of the fungus Saprolegnia

Cytoplasmic membranes from mycelium or protoplasts of Saprolegnia monoica (a cellulosic cell-wall fungus) were separated by continuous sucrose-density-gradient centrifugation. Glucan synthases assayed at low (micromolar uridine 5-diphosphate (UDP) glucose for -1-4-glucan synthase) and high (millimolar UDP glucose for -1-3-glucan synthase) substrate concentrations were associated with membranes exhibiting vanadate-sensitive, oligomycin-insensitive ATPase and equilibrating at density 1.16 g cm^-3. Synthase activities were also bound to membranes of lower density (1.10 and 1.145 g cm^-3). Plasma membranes were stabilized by coating protoplasts with concanavalin A. After lysis of the protoplasts, plasma membranes recovered by low centrifugal forces were isolated in continuous isopycinic gradients. Both synthase activities peaked with [³H]concanavalin A and Na-vanadate ATPase indicating that the synthetases are located at the plasma membrane. Treatments of intact protoplasts with cold glutaraldehyde or proteases before disruption lead to a diminution of glucan-synthase activities indicating that at least part of the enzymes of plasma membrane face the outside of the cell.Abbreviations ConA concanavalin A - ER endoplasmic reticulum - GSI -1,4-glucan synthase - GSH -1,3-glucan synthase - UDP uridine 5-diphosphate     

In vitro binding of riboflavin to subcellular particles from maize coleoptiles and Cucurbita hypocotyls

Saturable and reversible in vitro binding of [¹⁴C]riboflavin was found to occur on subcellular, sedimentable particles from maize coleoptiles and Cucurbita hypocotyls. The K_D was ca. 6 M, the pH optimum was near 6.0, and the number of binding sites amounted to 0.1–0.5 M on a fresh-weight basis. When the reducing agent dithionite was present, riboflavin binding increased-the K_D was 2.5 M, and the pH optimum above 8.0. The binding was specific: flavin mononucleotide (FMN) and flavin adenosine-dinucleotide (FAD) bound less tightly to these sites than riboflavin and another major soluble flavin, the previously described riboflavin-analog FX, occurring in grass coleoptiles. These flavin-binding sites were localized on vesicles derived from plasmalemma and endoplasmic reticulum by analyzing sucrose and metrizamide density gradients and marker enzymes.Abbreviations CCO cytochrome-c oxidase - CCR NADH-cytochrome-c oxidoreductase - ER endoplasmic reticulum - FAD flavin-adenosinedinucleotide - FMN flavin mononucleotide - MOPS N-morpholino-3-propansulfonic acid - NADH reduced -nicotinamide dinucleotide - nKP n thousand times g pellet - NPA l-naphthylphthalamic acid - PM plasma membrane, plasmalemma - RBF riboflavin - IAA indoleacetic acid - BA benzoic acid     

Germination and early tube development in vitro of Lycopersicum peruvianum pollen: Ultrastructural features

Morphologic changes occurring during pollen grain activation and ultrastructural features of Lycopersicum peruvianum Mill. pollen tube during the first stages of growth in vitro have been studied. The more evident morphologic changes during activation, in comparison to those already described for mature inactive pollen, concern dictyosomes, rough endoplasmic reticulum (RER), and ribosomes. The dictyosomes are very abundant and produce large and small vesicles. Near the germinative pores both types of vesicles are present, while all along the remaining cell wall only the large type is observed. These latter react weakly to Thiéry's test and probably contain a callose precursor necessary for the deposition of a callosic layer lining at first only the inner side of the functioning pore and occasionally the other two pores, and subsequently the entire inner surface of the cell wall. The small vesicles, highly positive to Thiéry's test, are present only near the pores and could be involved in the formation of the pectocellulosic layer of the tube wall. The setting free of RER cisterns, which in the mature inactive pollen were aggregated in stacks, coinciding with polysome formation and resumption of protein synthesis, is in accord with the hypothesized role of RER cistern stacks as a reserve of synthesizing machinery. The pollen tube reaches a definitive spatial arrangement soon after the generative cell and vegetative nucleus have moved into it. At this stage four different zones that reflect a functional specialization are present. In the apical and subapical zone two types of dictysosome-originated vesicles, similar to those found in the activated pollen grain, are present. Their role in the formation of the callosic and pectocellulosic wall layers seems to be the same as in the activated pollen grain.Abbreviations ER endoplasmic reticulum - RER rough endoplasmic reticulum Research performed under CNR program Biology of Reproduction     

Ultrastructure of freeze-substituted pollen tubes ofLilium longiflorum

Summary In view of the importance of the lily pollen tube as an experimental model and the improvements in ultrastructural detail that can now be attained by the use of rapid freeze fixation and freeze substitution (RF-FS), we have reexamined the ultrastructure of these cells in material prepared by RF-FS. Several previously unreported details have been revealed: (1) the cytoplasm is organized into axial slow and fast lanes, each with a distinct structure; (2) long, straight microtubule (MT) and microfilament (MF) bundles occur in the cytoplasm of the fast lanes and are coaligned with every organelle present; (3) the cortical cytoplasm contains complexes of coaligned MTs, MFs, and endoplasmic reticulum (ER); (4) the cortical ER is arranged in a tight hexagonal pattern and individual elements are closely appressed to the plasma membrane with no space between; (5) mitochondria and ER extend into the extreme apex along the flanks of the pollen tube, and vesicles and ER are packed into an inverted cone-shaped area at the center of the apex; (6) MF bundles in the tip region are fewer, finer, and in random orientation in comparison to those of the fast lanes; (7) the generative cell (GC) cell wall complex contains patches of plasmodesmata; (8) The GC cytoplasm contains groups of spiny vesicles that are closely associated with and seem to be fusing with or pinching off from mitochondria, and (9) the vegetative nucleus (VN) contains internal MT-like structures as well as numerous cytoplasmic MTs associated with its membrane and also located between the VN and GC.Abbrevations CF chemical fixation - ER endoplasmic reticulum - GC generative cell - MF microfilament - MT microtubule - PD plasmodesmata - PM plasma membrane - RF-FS rapid freeze fixation-freeze substitution - VN vegetative nucleus     

Mechanism of transport of vegetative storage proteins to the vacuole of the paraveinal mesophyll of soybean leaf

Summary Microscopy techniques were used to identify the pathway of transport of soybean leaf vegetative storage proteins (VSP/ and VSP94) to the vacuoles of a specialized cell type, the paraveinal mesophyll (PVM), where they accumulate. PVM cells are enriched in endoplasmic reticulum and Golgi bodies relative to surrounding mesophyll cells. The margins of medial and trans Golgi cisternae had attached or closely associated noncoated vesicles with densely staining membranes and lumenal contents of the same appearance as material that accumulated in the vacuole. These vesicles appeared to be transported preferentially to the tonoplast, where fusion with the membrane released the granular contents into the vacuole. Cytochemical staining with phosphotungstic acid and silver methenamine supported this interpretation as both the Golgi vesicles and the tonoplast stained intensely with these reagents, unlike the tonoplast of mesophyll cells which do not accumulate VSP. Immunocytochemical localization for VSP/ labeled the Golgi bodies and associated vesicles, and vacuolar material in PVM cells, but not in mesophyll. Similar labeling was seen in PVM of another legume species previously found to accumulate antigenically similar VSPs. Immunolocalization for VSP94, a lipoxygenase, labeled the PVM cytosol and material in the PVM vacuole, but not the Golgi or vesicles. The results of this study demonstrate that the Golgi pathway is utilized for transport of VSP/ in the PVM, which follows the mechanism of deposition demonstrated for certain seed storage proteins. VSP94 appeared to follow a separate path for accumulation in PVM vacuoles.Abbreviations LOX lipoxygenase - PVM paraveinal mesophyll - RER rough endoplasmic reticulum - TEM transmission electron     

In-vivo observations of a spherical aggregate of endoplasmic reticulum and of Golgi vesicles in the tip of fast-growing Chara rhizoids

In-vivo differential interference contrast microscopy was used to detect individual Golgi vesicles and a new structure in the tip of fast-growing rhizoids of Chara fragilis Desvaux. This structure is a spherical clear zone which is free of Golgi vesicles, has a diameter of 5 m and is positioned in the center of the apical Golgi-vesicle accumulation (Spitzenkörper). After glutaraldehyde fixation and osmium tetroxide-potassium ferricyanide staining of the rhizoid, followed by serial sectioning and three-dimensional reconstruction, the spherical zone shows a tight accumulation of anastomosing endoplasmic reticulum (ER) membranes. The ER membranes radiate from this aggregate towards the apical plasmalemma and to the membranes of the statolith compartments. Upon gravistimulation the ER aggregate changes its position according to the new growth direction, indicating its participation in growth determination. After treatment of the rhizoid with cytochalasin B or phalloidin the ER aggregate disappears and the statoliths sediment. It is concluded that the integrity of the ER aggregate is actin-dependent and that it is related to the polar organisation of the gravitropically growing cell tip.Abbreviations CB cytochalasin B - DIC differential interference contrast microscopy - DMSO dimethyl sulfoxide - ER endoplasmic reticulum     

Transglycosylation reactions by exoglycosidases from the termite Macrotermes subhyalinus

The ability of four exoglycosidases (-galactosidase, -glucosidase, -glucosidase and invertase) from the termite Macrotermes subhyalinus to catalyse tranglycosylation reactions was tested using lactose, cellobiose, maltose and sucrose as glycosyl donors and 2-phenylethanol as glycosyl acceptor. The experimental conditions were optimized in relation to the time course of the reaction, pH and concentrations of glycosyl donor and acceptor. Whereas the hydrolytic activity was largely predominant over the transferase activity with -galactosidase and -glucosidase, the transglycosylation activity represented 68% with -glucosidase. In addition, as demonstrated by the transglycosylation product formed, the hydrolysis of sucrose was catalysed by -glucosidase and not by invertase. On the basis of this work, -glucosidase from M. subhyalinus appears to be a valuable tool for the preparation of neoglycoconjugates.     

Organisation scalariforme du réticulum endoplasmique dans certains processus nerveux de l'hypothalamus deGasterosteus aculeatus L.

Résumé Dans l'hypothalamus de l'Epinoche, certains prolongements nerveux présentent une organisation particulière du réticulum endoplasmique. Des séries de vésicules interconnectées sont disposées en stries transverses espacées de 0,3 à 1,3 . Cette organisation particulière du réticulum endoplasmique existe dans les prolongements nerveux de fort diamètre (1 ), même dans les fibres neurosécrétrices.

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Special organization of the endoplasmic reticulum in some hypothalamic nerve processes ofGasterosteus aculeatus

Summary In some nerve processes of the hypothalamus ofGasterosteus aculeatus, the endoplasmic reticulum displays a special organization. Small interconnected smooth vesicles constitute transverse lines spaced between 0.3 and 1.3 . This special organization of the endoplasmic reticulum has been found in nerve processes with a diameter larger than 1 even in neurosecretory ones.

     

Ultrastructural investigations on Lycopersicum peruvianum pollen activation and pollen tube organization after self-and cross-pollination

No differences have been observed in vivo between Lycopersicum peruvianum compatible and incompatible pollen during activation and pollen tube emission and organization, that is until 4 h and 30 min after pollination. During pollen activation the main events are the setting free of rough endoplasmic reticulum (RER) cisterns which were stacked in the mature pollen, the increase in the number of polysomes, and a great activity of the dictyosomes. Immediately after germination of the vegetative nucleus and the generative cell move into the tube, the generative cell diviting to form the male gametes; the tube then becomes organized in four zones. This series of changes is similar to what has already been observed in vitro except that in vitro the generative cell remains undivided and the whole process from seeding to tube organization takes 3 h instead of 4 h and 30 min after pollination, as it does in vivo. Our findings are compatible with the main models of the tube inhibition mechanism proposed till now.Abbreviations RER rough endoplasmic reticulum - GC generative cell - VN vegetative nucleus - GP germinative pore Research performed under C.N.R. (Italian National Research Council) program Biology of Reproduction     

Die Spermiohistogenese des Süßwasser-Ostracoden Notodromas monacha O. F. Müller

Zusammenfassung Bei Notodromas monacha (Ostracoda) wird die Entwicklung der Spermatiden von der Spindelbis zur Schlauchform mit Phasenkontrast und elektronenmikroskopisch untersucht. Am Kern der spindelförmigen Spermatiden sind an zwei gegenüberliegenden Seiten jeweils zwei Ausstülpungen der Membran zu beobachten. Aus ihnen werden offenbar unter Beteiligung von Vesikeln des endoplasmatischen Reticulum zwei Flügelstrukturen aufgebaut. Jede Flügelstruktur besteht aus zwei Lamellen, die mit tubulären Elementen besetzt sind. An den schlauchförmigen Spermatiden läßt sich ein dicker Körper und ein dünner Faden unterscheiden. Kern, Nebenkern-Derivate und Flügelstrukturen durchziehen den Körper der Länge nach. Der Faden enthält nur noch den Kern und die Ausläufer der Flügelstrukturen. Im Bereich des freien Körperendes liegt im Kern ein als Chromatin gedeuteter Kolben. Diesem folgt in Richtung zum Fadenende ein dichter Achsenstab, der von einer Doppelmembran umhüllt ist. Im dorso-lateralen Kernraum finden sich elektronendichte Brocken unbekannter Bedeutung. Nach Beendigung des Aufbaues der Flügelstrukturen wird das endoplasmatische Reticulum samt Mikrotubuli weitgehend abgebaut. Gleichzeitig verschwindet die den Achsenstab umhüllende Doppelmembran. Die Flügelstrukturen werden als das für die Motilität der Spermien verantwortliche Organell gedeutet.

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The spermiogenesis of the fresh-water ostracod Notodromas monacha O. F. Müller

Summary The transformation of spindle-like spermatids into tubular forms in Notodromas monacha (Ostracoda) has been studied by means of phase contrast and electron microscopy. At each of two opposite sides of the nucleus of spindle-like spermatids, two extensions of the nuclear membrane arise. These extensions form two Flügelstrukturen (wing-like structures) obviously by participation of vesicles of the endoplasmic reticulum. Each Flügelstruktur consists of two lamellae, each bearing numerous tubules. The tube-like spermatids consist of a large body and a small thread. The nucleus, the Nebenkern-derivatives and the Flügelstrukturen extend through the whole body. Within the thread only the nucleus and the outermost parts of the Flügelstrukturen are found. A columnar body, supposed to be chromatin, is located within that part of the nucleus which is situated in the free end of the body. A dense axial rod extends between the columnar body and the free end of the thread. The dorso-lateral sphere of the nucleus contains electron dense particles of unknown function. As soon as the construction of the Flügelstrukturen is finished, the endoplasmic reticulum and the microtubules are reduced. Simultaneously the double-membrane surrounding the axial rod disappears. The Flügelstrukturen are interpreted as organelles responsible for the motility of the sperm.

     

Covisualization in living onion cells of putative integrin, putative spectrin, actin, putative intermediate filaments, and other proteins at the cell membrane and in an endomembrane sheath

Summary Covisualizations with wide-field computational opticalsectioning microscopy of living epidermal cells of the onion bulb scale have evidenced two major new cellular features. First, a sheath of cytoskeletal elements clads the endomembrane system. Similar elements clad the inner faces of punctate plasmalemmal sites interpreted as plasmalemmal control centers. One component of the endomembrane sheath and plasmalemmal control center cladding is antigenicity-recognized by two injected antibodies against animal spectrin. Immunoblots of separated epidermal protein also showed bands recognized by these antibodies. Injected phalloidin identified F-actin with the same cellular distribution pattern, as did antibodies against intermediate-filament protein and other cytoskeletal elements known from animal cells. Injection of general protein stains demonstrated the abundance of endomembrane sheath protein. Second, the endomembrane system, like the plasmalemmal puncta, contains antigen recognized by an anti-₁ integrin injected into the cytoplasm. Previously, immunoblots of separated epidermal protein were shown to have a major band recognized both by this antibody prepared against a peptide representing the cytosolic region of ₁ integrin and an antibody against the matrix region of ₁ integrin. The latter antibody also identified puncta at the external face of protoplasts. It is proposed that integrin and associated transmembrane proteins secure the endomembrane sheath and transmit signals between it and the lumen or matrix of the endoplasmic reticulum and organellar matrices. This function is comparable to that proposed for such transmembrane linkers in the plasmalemmal control centers, which also appear to bind cytoskeleton and a host of related molecules and transmit signals between them and the wall matrix. It is at the plasmalemmal control centers that the endoplasmic reticulum, a major component of the endomembrane system, attaches to the plasma membrane.Abbreviations DiOC₆ 3,3-dihexyloxacarbocyanine iodide - GF Gaussian filtering - ML maximum likelihood (algorithm or method) - PBS phosphate-buffered saline     

Comparison of [35S]GTPγS binding to plasma membranes and endomembranes prepared from soybean and rat

Summary Plasma membranes were prepared from soybean hypocotyls and roots by aqueous two-phase partitioning and subsequent free-flow electrophoresis. The highly purified plasma membranes bound [³⁵S]GTPS with a relatively high affinity (K_d10nM). The binding was saturable and specific as it was indicated by the displacement of bound [³⁵S]GTPS by unlabeled GTPS and GTP, but not by ATPS, ATP, UTP or CTP. ITP was intermediate in its ability to displace [³⁵S]GTPS. When soybean plasma membrane proteins were separated by SDS-PAGE and displayed by autoradiography, two major [³⁵S]GTPS binding proteins were revealed with apparent molecular weights of 24 and 28 kDa. Results with plasma membranes from soybean hypocotyls and roots were similar but differed from those with plasma membranes prepared from rat liver and adipocytes where only a single major [³⁵S]GTPS binding activity with a molecular weight of 28 kDa was observed.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - G protein hetero-trimeric GTP binding protein with , , subunits - G_n protein GTP binding protein detected on nitrocellulose blots - GTPS guanosine 5-[-thio]triphosphate - IAA 3-indoleacetic acid - SDS-PAGE sodium dodecylsulfate-polyacrylamide gel electrophoresis     

On the fine structure of the aglomerular renal tubule in Lophius piscatorius

Summary The fine structure of the secretory tubules in the kidney of the aglomerular goose-fish (Lophius piscatorius) is described. The cells have a pyramidal shape, are joined together by multiple desmosomes, and share as main characteristics: abundant and deep inflections of the basal and lateral cell membranes; coated luminal plasma membranes forming multiple microvilli or a genuine brush border; moderate numbers of comparatively small mitochondria, usually unassociated with the basal and lateral plasma membrane specializations; numerous multivesicular bodies occuring in the apical cytoplasm; abundant large lysosome-like bodies in the intermediate regions of the cytoplasm; and comparatively poor development of endoplasmic reticulum and Golgi apparatus.The observations suggest that the cells perform both absorptive and secretory functions and are metabolically unusually active in autolytic and heterolytic work. Comparisons with other aglomerular species indicate that the ability for active secretory function is not necessarily dependent on a close association between plasma membrane and mitochondria; however, this ability does appear to require a markedly increased basal and/or lateral cell surface created by multiple invaginations of the plasma membrane. The abundance of desmosomes and associated structures appears to represent a unique structural specialization of the goosefish tubule, and indicates that the cells must be firmly anchored to one another to supply a rigid and mechanically continuous lining of the tubule. The multivesicular bodies probably represent endocytic vacuoles which fuse with apical vesicles and invaginate their outer membrane to form the internal vesicles; they appear to transform to ambilysosomes via a function as heterophagosomes and — later — combined hetero- and autophagosomes.Supported by grants from Karolinska Institutet, Fonden til Videnskabens Fremme and Konsul Johannes Fogh-Nielsen og fru Ella Fogh-Nielsens Légat. Part of the study was performed at the Zoological Station at Naples, Italy. The assistance of Mrs. Britt-Marie Karlsson is gratefully acknowledged.