Structural organization of ovarian follicle cells in the cotton bug (Dysdercus intermedius) and the honeybee (Apis mellifera)

Summary The somatic epithelia of Dysdercus and Apis follicles were analyzed by electron microscopy, and the patterns of F-actin and microtubules were studied by fluorescence microscopy. The epithelia in both species differ considerably in shape and in the organization of the cytoskeleton. During previtellogenic stages, the epithelium consists of columnar-shaped cells with small (Dysdercus) or no (Apis) lateral intercellular spaces. During vitellogenesis, the follicle cells round up; the intercellular spaces increase in size in Dysdercus follicles, whereas in Apis follicles they remain small. Along the basal surface of the follicle cells, there are conspicuous parallel bundles of microfilaments perpendicular to the anteroposterior axis of the follicles. In the honeybee, these microfilament bundles are present in long filopodia, most of which are embedded in thickenings of the basement membrane and extend over the surfaces of neighbouring cells. In the cotton bug, the basal surface of the follicle cells is thrown into parallel folds. The microfilament bundles are located just underneath the cell membrane where the folds contact the basement membrane. In the polar regions of the Dysdercus follicle, the epithelial cells become flat and adhere to each other without forming intercellular spaces. The basement membrane is particularly thick in the polar areas; this has also been observed in Apis follicles around the intercellular bridge connecting oocyte and nurse cells.     

Ultrastructure of the gametes ofCoelomomyces dodgei Couch (Blastocladiales,Chytridiomycetes)

Summary As part of an investigation on the developmental biology ofCoelomomyces dodgei Couch (Blastocladiales, Chytridiomycetes), the ultrastructure of the male and female gametes was studied. The nucleus is central and conical in shape except for a basal spur that curves back towards the large plate-like mitochondrion. A nuclear cap of ribosomes sits on the flat anterior end of the nucleus. Approximately seven lipid globules are partially embedded in the mitochondrion and are interconnected by membrane cisternae. The lipid globules are covered by a single fenestrated microbody and a backing membrane lies between the microbody and the gamete plasma membrane. The kinetosome is at the base of the nucleus and is connected to a single, posterior, whiplash flagellum. A nonkinetosomal centriole is absent. In the peripheral cytoplasm of both mating types there is a paracrystalline body of unknown composition and function. No significant ultrastructural differences were found between the male and female gametes.     

Membranous appendices of spherosomes (oleosomes)

Spherosomes (oleosomes) of cotyledons of rape (Brassica napus L.), sunflower (Helianthus annuus L.), and watermelon (Citrullus vulgaris, Schrad.) seedlings are delimited by a half unit membrane that appears to be continuous with each of the osmiophilic layers of a tripartite unit membrane forming a handlelike appendix of the spherosomes. Prior to any noticeable utilization of the spherosomal storage fat, ribosomes were found to be attached to these handles. At later stages appendices of the spherosomes are smooth, showing a diameter of about 22 nm that greatly exceeds the thickness of any other unit membrane profiles identical in structure and diameter osomes appears to be continuous with the thick lipid layer of the handles. In intermediate stages of fat depletion the spherosomal bodies become invaginated with cytoplasmic material. Finally vesicles with cytoplasmic contents surrounded by a membrane with a typically thick lipid layer are left in the cells. Membrane profiles indentical in structure and diameter to the spherosomal appendices were also present in electron micrographs of the lipolytic membrane fraction recovered from sucrose density gradients after centrifugation of a microsomal cell fraction. The ultrastructural observations are taken for evidence that the spherosomal appendices represent the lipase-carrying membranes isolated previously (Theimer and Rosnitschek, 1978). A novel hypothesis for development and utilization of fat-storing spherosomes is also proposed.     

The tapetum inSchizaea pectinata (Schizaeaceae) and a comparison with the tapetum inPsilotum nudum (Psilotaceae)

A combination tapetum consisting of a cellular, parietal component and a plasmodial component occurs inSchizaea pectinata. A single, tapetal initial layer divides to form an outer parietal layer which maintains its cellular integrity until late in spore wall development. The inner tapetal layer differentiates into a plasmodium which disappears after the outer exospore has developed. In the final stages of spore wall development, granular material occurs in large masses and is dispersed as small granules throughout the sporangial loculus. No tapetal membrane develops. Comparisons are drawn with the combination tapetum found inPsilotum nudum.     

Two new species of the genusAmblyeleotris (Pisces: Gobiidae) from the Ryukyu Islands,Japan

Two new shrimp-associated gobies,Amblyeleotris yanoi sp. nov. andA. masuii sp. nov. are described on the basis of specimens from Iriomote-jima Island and Okinawa-jima Islands, Okinawa Prefecture, Japan.A. yanoi is distinguished from other members of the genus by the combination of the following characters: 14 anal fin soft rays, 19 pectoral fin rays, 97–103 longitudinal scales, a candle flame-shaped marking on the caudal fin, a very low membrane connecting the pelvic fins and absence of a ventral frenum.A. masuii differs from all other congeners by having 92–97 longitudinal scales, the length of the interpelvic connecting membrane relative to the longest pelvic fin ray (0.43–0.66), black blotches on the sides of the chin, and blue spots on the opercle and preopercle.     

Ferrioxamine transport mutants and the identification of the ferrioxamine receptor protein (FoxA) inErwinia herbicola (Enterobacter agglomerans)

Summary Iron deprivation ofErwinia herbicola (Enterobacter agglomerans) induces the biosynthesis of six high-M _r outer-membrane proteins and large amounts of ferrioxamine E. Mutagenesis withN-methyl-N-nitro-N-nitrosoguanidine and selection with ferrimycin A yielded mutants ofE. herbicola K4 (wild type), defective in the expression of a 76-kDa outer-membrane protein, as determined by SDS/polyacrylamide gel electrophoresis. While in bioassays wild-type cells showed growth promotion in the presence of ferrioxamines (B, D₁, D₂, E, G), enterobactin, citrate, ferrichrome and coprogen, these mutants failed to respond to ferrioxamines. Moreover, experiments with⁵⁵Fe-labelled siderophores confirmed that iron transport mediated by ferrioxamine E and B in the mutants was completely inhibited, whereas iron transport by other hydroxamate siderophores, such as ferrichrome and coprogen was unaffected. The results are evidence that the 76-kDa protein in the outer membrane represents the receptor protein (FoxA) for ferrioxamines inE. herbicola.     

Haustoria inUrocystis (Tilletiales)

Haustoria of severalUrocystis spp. have been investigated by transmission electron microscopy. The haustoria are botryose and have an extrahaustorial matrix with vesiclelike bodies. The extrahaustorial membrane shows high ATPase activity in contrast to the haustorial plasmalemma. In walled off haustoria the haustorial plasmalemma stains more intensely than the extrahaustorial membrane. The vesicle-like bodies are ATPase negative. The role of the vesicle-like bodies is discussed.Dedicated to Prof. DrLothar Geitler on the occasion of the 90th anniversary of his birthday. Part 55 of a series Studies inHeterobasidiomycetes.     

The fine structure of the carapace integument of Daphnia magna straus (Crustacea branchiopoda)

Summary The structure of the two integumental layers comprising the carapace of female D. magna was examined at several points through the molt cycle. The epicuticle and procuticle are simple in organisation; pore canals are absent but intracuticular fibres are present, forming complexes with invaginations of the epidermal plasma membrane similar to such complexes described in the literature for other arthropods. The epidermis consists almost entirely of cuticle-secreting cells. Secretion of the new cuticle begins when 50–67% of the instar has elapsed by which time the epidermal cells have increased in height and their nuclei have become more rounded. However, other presumed secretory phenomena observed viz. the formation of dense core vesicles by Golgi bodies, and the occurrence of these and coated vesicles near the apical plasma membrane are not restricted to any particular period during the molt cycle. This suggests that the mechanisms of cuticle secretion do not undergo marked changes in activity as they do in decapods; presumably this relative continuity is related to the much shorter molt cycle of cladocerans.The technical assistance of G.A. Bance, and the financial support provided by the National Research Council of Canada are gratefully acknowledged     

Iron(III) hydroxamate transport across the cytoplasmic membrane ofEscherichia coli

Summary Transport of iron(III) hydroxamates across the inner membrane into the cytoplasm ofEscherichia coli is mediated by the FhuC, FhuD and FhuB proteins and displays characteristics typical of a periplasmic-binding-protein-dependent transport mechanism. In contrast to the highly specific receptor proteins in the outer membrane, at least six different siderophores of the hydroxamate type and the antibiotic albomycin are accepted as substrates. AfhuB mutant (deficient in transport of substrates across the inner membrane) which overproduced the periplasmic FhuD 30-kDa protein, bound [⁵⁵Fe] iron(III) ferrichrome. Resistance of FhuD to proteinase K in the presence of ferrichrome, aerobactin, and coprogen indicated binding of these substrates to FhuD. FhuD displays significant similarity to the periplasmic FecB, FepB, and BtuE proteins. The extremely hydrophobic FhuB 70-kDa protein is located in the cytoplasmic membrane and consists of two apparently duplicated halves. The N-and C-terminal halves [FhuB(N) and FhuB(C)] were expressed separately infhuB mutants. Only combinations of FhuB(N) and FhuB(C) polypeptides restored sensitivity to albomycin and growth on iron hydroxamate as a sole iron source, indicating that both halves of FhuB were essential for substrate translocation and that they combined to form an active permease. In addition, a FhuB derivative with a large internal duplication of 271 amino acids was found to be transport-active, indicating that the extra portion did not disturb proper insertion of the active FhuB segments into the cytoplasmic membrane. A region of considerable similarity, present twice in FhuB, was identified near the C-terminus of 20 analyzed hydrophobic proteins of periplasmic-binding-protein-dependent systems. The FhuC 30 kDa protein, most likely involved in ATP binding, contains two domains representing consensus sequences among all peripheral cytoplasmic membrane proteins of these systems. Amino acid replacements in domain I (LysGlu and Gln) and domain II (AspAsn and Glu) resulted in a transport-deficient phenotype.     

The presence of vasoactive intestinal polypeptide (VIP)-like-immunoreactive nerve fibres and VIP-receptors in the pineal gland of the Mongolian gerbil (Meriones unguiculatus)

Summary The photocytes and other endodermal cells composing the wall of the meridional canals of the comb-jelly, Mnemiopsis leidyi, were investigated by transmission electron microscopy. Although many of these cells possess distinctive features such as a ciliary apparatus, lysosome-like bodies or vacuoles, they share with photocytes the presence of a network of rough endoplasmic reticulum (RER) whose cisternae enwrap large mitochondria and are aligned along the subsurface of the plasma membrane. A stereological analysis of organelle content in photocytes confirms the prominence of the RER in these cells and a shift of RER from mitochondria to plasma membrane subsurface in photocytes induced to luminesce by the mitochondrial inhibitor dinitrophenol. Photocytes and other endodermal cells of the meridional canals are interconnected by numerous gap junctions which, among photocytes, often form symmetrical triads with cortical cisternae and mitochondria. The gap junctions and RER/mitochondria assemblages are interpreted as possible substrates for, respectively, conduction of luminescence excitation along the canals and for excitation-luminescence coupling. Neuntes occasionally make synapses with photocytes and other endodermal cells lying adjacent to the mesoglea.     

Ornamentation of the testa inSedum (Crassulaceae)

A SEM investigation of the seeds of 40 European and 5 Asiatic species ofSedum revealed three fundamentally different types of ornamentation of the testa. The most prominent difference in the ontogeny of the seeds differing in ornamentation is the ovular envelope, which is a mucous membrane enclosing the ovules and young seeds and occurring only in plants whose seeds have a simple type of ornamentation. The ornamentation of the testa was found to be a very constant character. Because the ornamentation is highly correlated with the hybridization pattern of the species it is considered to be an important character in the infrageneric classification ofSedum.     

Comparison of auditory sense organs in parasitoid Tachinidae (Diptera) hosted by Tettigoniidae (Orthoptera) and homologous structures in a non-hearing Phoridae (Diptera)

The dipteran parasitoids Therobia leonidei and Homotrixa alleni (Tachinidae) use acoustic cues to locate their calling tettigoniid (Ensifera, Orthoptera) hosts. The sexually dimorphic tympanal organs of both fly species are located at the prosternum. For comparison a homologous chordotonal organ in the non-hearing fly Phormia regina, Meigen (Phoridae) is also described. The scolopidial sense organs of the ears have approximately 180 sensory cells in Th. leonidei and 250 cells in H. alleni. Interspecific analysis indicates that the cell number and arrangement might be genus specific in Tachinidae. The mononematic scolopidia, each with one sensory cell, are of different sizes and insert at the tympanal membrane. Large scolopidial units (diameter of sensory cells up to 50 μm) extend longitudinally from the centre of the sensory organ towards the ligament, whereas small units (sensory cell diameter up to 10 μm) are arranged sequentially within the sensory organ. This arrangement is discussed to be a possible basis for frequency discrimination. The ultrastructure of the scolopidia is similar in the hearing and non-hearing flies. In both groups, the majority of scolopales has a diameter from 2 to 2.9 μm, although hearing species have additionally wider scolopales. The homologous chordotonal organ of Ph. regina consists of approximately 55 sensory cells of uniform direction. The data are discussed in comparison to the ears of other Diptera.     

Eukaryotic integral membrane protein expression utilizing the <Emphasis Type="Italic">Escherichia coli</Emphasis> glycerol-conducting channel protein (GlpF)

A fusion protein expression system is described that allows for production of eukaryotic integral membrane proteins in Escherichia coli (E. coli). The eukaryotic membrane protein targets are fused to the C terminus of the highly expressed E. coli inner membrane protein, GlpF (the glycerol-conducting channel protein). The generic utility of this system for heterologous membrane-protein expression is demonstrated by the expression and insertion into the E. coli cell membrane of the human membrane proteins: occludin, claudin 4, duodenal ferric reductase and a J-type inwardly rectifying potassium channel. The proteins are produced with C-terminal hexahistidine tags (to permit purification of the expressed fusion proteins using immobilized metal affinity chromatography) and a peptidase cleavage site (to allow recovery of the unfused eukaryotic protein).     

Membrane maintenance and electrical properties of photoreceptors of wild-type andrpa (receptor potential absent) mutant blowflies (Calliphora erythrocephala)

Summary The maintenance of photoreceptor cell membranes in the blowfly was investigated in relation to the diurnal cycle, age, and therpa (receptor potential absent) phototransduction mutation. The effect of disturbed membrane assembly on the electrical membrane properties was examined using single-electrode discontinuous current-clamp techniques. In wild-type flies the cross-sectional dimensions of the rhabdomeres were markedly reduced with age, and the quantity of synthetic organelles decreased concurrently, whereas no correlation was found between the diurnal cycle and membrane turnover. Therpa mutation is thought to block the visual transduction cascade in photoreceptor cells and to lead to degeneration of the photoreceptor cell bodies. The volume of rhabdomeres decreased markedly inrpa mutants and the quantity of synthetic organelles was reduced significantly, indicating an imbalance between photoreceptive membrane renewal and degradation. Also, the plasma membrane underwent degenerative changes. The passive electrical properties of photoreceptor cells — resting membrane voltages and input resistances — were only slightly changed from those of wild-type flies, although the photoreceptive membrane did not depolarize in response to light. This indicates no apparent disturbance in the function of the ionic channels in these membranes. Taken together, these results suggest that the photoreceptor cells need a functional phototransduction cascade with its feedback controls to maintain continuous renewal of rhabdomeres, but that the plasma membrane maintains its normal electrochemical properties despite extreme morphological degeneration of photoreceptor cell.     

The eyespot of the flagellateTetraselmis cordiformis stein (Chlorophyceae): Structural spezialization of the outer chloroplast membrane and its possible significance in phototaxis of green algae

Summary The eyespot region of the flagellateTetraselmis cordiformis Stein (Chlorophyceae) was investigated with the freeze-fracture technique. The only fracture faces observed in this region were the two complementary fracture faces (PF and EF) of the outer chloroplast envelope membrane. Intramembranous particle numbers on both fracture faces of this membrane were significantly higher in the eyespot region as compared to regions outside the eye-spot. Higher numbers of particles on the PF face in the eyespot region were mainly caused by an increase in particle numbers of the size class 6–8 mm, while on the EF face particle size distribution was not significantly different between eyespot and other regions. Functional implications are discussed and evidence is presented that the outer chloroplast envelope membrane may be the site of photoreceptor location in green algal phototaxis.     

Alkaline phosphatase which lacks its own signal sequence becomes enzymatically active when fused to N-terminal sequences of Escherichia coli haemolysin (HlyA)

Summary Fusion of the alkaline phosphatase gene (phoA) which lacks its own signal peptide sequence to the N-terminal region of hlyA, the structural gene for Escherichia coli haemolysin, leads to active alkaline phosphatase (AP). AP activity depends on the length of the N-terminal region of hlyA. An optimum is reached when 100–200 amino acids of HlyA are fused to PhoA but fusion of as little as 13 amino acids of HlyA to PhoA is sufficient to yield appreciable AP activity. When cells are treated with lysozyme most of the AP activity is found associated with the membrane fraction but a substantial amount is also found in the soluble fraction, most of which may represent, a periplasmic pool of AP. The soluble portion of AP activity is significantly increased when the cells are disrupted by ultrasonication, which indicates that the fusion proteins are only loosely associated with the membrane and that large parts are already located on the outside of the cytoplasmic membrane. The expected fusion proteins were identified in the soluble and the membrane fractions and their amounts in these fractions correlated well with AP activity.     

珍稀濒危植物堇叶紫金牛对持续干旱的生理响应

采用盆栽控水法,研究了珍稀濒危植物堇叶紫金牛(Ardisia violacea)在持续干旱条件下的生理响应。随着持续干旱时间的延长,堇叶紫金牛应对持续干旱的阶段可分为适应期、轻度干旱期、中度干旱期和重度干旱期。在适应期和轻度干旱期,堇叶紫金牛叶片游离脯氨酸和可溶性糖含量稳定在一个较低水平,可溶性蛋白质含量先下降后快速上升,细胞膜系统和抗氧化酶系统能主动进行生理调节;中度干旱期,丙二醛(MDA)含量和质膜相对透性迅速升高,细胞膜系统受损加剧,游离脯氨酸、可溶性糖含量均急剧增加,对抵御干旱起到重要的渗透调节作用。在轻度干旱期和中度干旱期,光合色素中叶绿素a和叶绿素b含量显著提高,以抵抗干旱胁迫。重度干旱期,细胞膜系统、抗氧化酶SOD、游离脯氨酸和可溶性糖含量上升,但MDA略微下降,这时可能达到植物耐受干旱的极限,不再发生膜脂过氧化作用。综上表明,堇叶紫金牛具有较强的耐旱性,RWC为49.94%是细胞膜系统、抗氧化酶系统和渗透调节物质含量变化的拐点,渗透调节和抗氧化酶系统的主动适应是其耐旱的主要机制。     

Freeze-fracture of the generative cell of Phoenix dactylifera (Arecaceae)

Summary Mature pollen of Phoenix dactylifera was freeze-fractured in germination medium. The surface of the generative cell was highly convoluted. Microtubules were not in close contact with the indentations. The vegetative cell membrane was appressed to the generative cell. Ordered ridges appeared in both fracture faces of the vegetative cell inner plasma membrane at the indentations. No ordered ridges were observed in the fracture faces of the generative cell. The nuclear envelopes of the vegetative and generative cells differed, with the generative cell having fewer and larger nuclear pores irregularly dispersed among porefree areas. These differences in plasma membrane and in nuclear envelope correlate with the subsequent differentiation of the two cells.Acknowledgements. We thank J.I. Stillman for technical assistance with the thin section preparations. This research was supported in part by NSF grant DCB-8607765 to W.W.T.     

Single-molecule investigation of the interactions between reconstituted planar lipid membranes and an analogue of the HP(2-20) antimicrobial peptide

In this study, we employed electrophysiology experiments carried out at the single-molecule level to study the mechanism of action of the HPA3 peptide, an analogue of the linear antimicrobial peptide, HP(2–20), isolated from the N-terminal region of the Helicobacter pylori ribosomal protein. Amplitude analysis of currents fluctuations induced by HPA3 peptide at various potentials in zwitterionic lipid membranes reveal the existence of reproducible conductive states in the stochastic behavior of such events, which directly supports the existence of transmembrane pores induced the peptide. From our data recorded both at the single-pore and macroscopic levels, we propose that the HPA3 pore formation is electrophoretically facilitated by trans-negative transmembrane potentials, and HPA3 peptides translocate into the trans monolayers after forming the pores. We present evidence according to which the decrease in the membrane dipole potential of a reconstituted lipid membranes leads to an augmentation of the membrane activity of HPA3 peptides, and propose that a lower electric dipole field of the interfacial region of the membrane caused by phloretin facilitates the surface-bound HPA3 peptides to break free from one leaflet of the membrane, insert into the membrane and contribute to pore formation spanning the entire thickness of the membrane.     

A study on pollen grains ofCrocus cartwrightianus (Iridaceae)

Crocus cartwrightianus andC. cartwrightianus cv.albus pollen have been studied from structural and ultrastructural points of view and the germination assayed in vivo and in vitro.C. cartwrightianus pollen is regularly shaped and sized, has a low percentage of anomalous grains and has a high germination rate in vitro, whileC. cartwrightianus cv.albus is less regularly shaped with some variation in size and has a high percentage of anomalous grains and a low germination percentage in vitro. Ultrastructural observations have revealed, in the pollen of both the taxa, the presence of a thin elongated vegetative nucleus and a generative cell surrounded by a thin membrane. However,C. cartwrightianus pollen shows a thicker intine, andC. cartwrightianus cv.albus shows numerous pollen germination anomalies which are in common withC. sativus.