The coxal glands of geophilomorpha (chilopoda): Organs of osmoregulation

Summary The organs terminating at the coxal pores of the tug-legs of Geophilomorpha are not repugnatorial glands, but possess typical transport epithelia with deep apical and basal infoldings of the cell membranes, between which numerous large mitochondria are located. Many transport vesicles are found in the basal region but fewer in the apical cytoplasm. The apex is characterized by bundles of longitudinally oriented microtubules, sparse endoplasmic reticulum and free ribosomes. Single neurosecretory axons with synaptoid areas are scattered among the cells. It is suggested that the coxal organs have a diuretic function in moist habitats and an antidiuretic effect in arid environments. The switch-over is evidently controlled by a neuroendocrine mechanism.     

Dynamics of the cytoskeleton of epidermal cells in situ and in culture

Summary The cytoskeleton of primary tissue-culture cells from the epidermis of Xenopus laevis tadpoles was investigated by phase-contrast, immunofluorescence, and electron microscopy. The connection between the arrangement of different types of filaments and the mechanical properties of the epidermis is discussed. The bilayered epidermis attains stability from thick bundles of tonofilaments interconnecting the basal desmosomes. Twisting of tonofilaments around each other can explain the occurrence of elastic filamentous curls forming a meshwork braced between rows of small desmosomes in the apical region of the epidermis. Actin is arranged as a diffuse meshwork and sometimes forms bundles intermingling with tonofilament bundles. Surface membranes and rows of small desmosomes are delineated by actin and contain -actinin. Actin raises the tension for rounding and spreading of cells. Microtubules stabilize already well-developed lamellae.     

Molecular characterization of β-thalassemia in Hungary

We have identified seven different -thalassemia mutations and one -thalassemia determinant (the Sicilian type) in 32 members of 17 Hungarian families. The most common mutation is the IVS-I-1 (GA) change; its high frequency is comparable to that observed in neighboring Czechoslovakia. Additional mutations are of Mediterranean origin. One rare mutation (initiation codonATGGTG) was identified as an independent mutation because of the absence of known polymorphisms in the -globin gene. One new frameshift at codon 51 (-C) was observed in a single individual; hematological data were as expected for a °-thalassemia heterozygosity.     

Arabidopsis thaliana ‘extra-large GTP-binding protein’ (AtXLG1): a new class of G-protein

Heterotrimeric GTP-binding proteins, composed of , , and subunits, are involved in signal transduction pathways in animal and plant systems. In plants, physiological analyses implicate heterotrimeric G-proteins in ion channel regulation, light signaling, and hormone and pathogen responses. However, only one class of plant G genes has been identified to date. We have cloned a novel gene, Arabidopsis thaliana extra-large GTP-binding protein (AtXLG1). AtXLG1 appears to be a member of a small gene family and is transcribed in all tissues assayed: roots, leaves, stems, flowers, and fruits. The conceptually translated protein from AtXLG1 is 99 kDa, twice as large as typical G proteins. The carboxy-terminal half of the AtXLG1 protein has significant homology to animal and plant G proteins. This region includes a GTP-binding domain, a predicted helical domain, and an aspartate/glutamate-rich loop, which are characteristics of G's. Despite the absence of some of the amino acids implicated in GTP binding and hydrolysis by crystallographic and mutational analyses of mammalian G's, recombinant AtXLGl binds GTP with specificity. The amino-terminal region of AtXLGl contains domains homologous to the bacterial TonB-box, which is involved in energy transduction between the inner and outer bacterial membranes, and to zinc-finger proteins. Given the unique structure of AtXLG1, it will be of interest to uncover its physiological functions.     

Localization of gamma-tubulin to the basal foot associated with the basal body extending a cilium

The human oviduct epithelium primarily consists of ciliated cells and secretory cells. Solitary cilia usually extend from the apical surface of the secretory cells. We investigated the localization of -tubulin in the ciliary basal apparatus of both cell types by fluorescence immunohistochemistry and immunoelectron microscopy. In addition to basal bodies, -tubulin was identified in the lateral basal foot, especially the basal foot cap. This observation is consistent with previous observations that microtubules radiate from the basal foot and the basal foot serves as the microtubule organizing centre.     

Qualitative Analysis of the Carbohydrate Composition of Apolipoprotein H

The specific binding of digoxigenin-labeled lectins to carbohydrate moieties is used to characterize the carbohydrate chains bound to apolipoprotein H. Our results show that apolipoprotein H is rich in sialic acid linked (2–6) to galactose or N-acetylgalactosamine. Sialic acid is not (2–3)-linked to galactose. Galactose is (1–4)-linked to N-acetylglucosamine and (1–3)-linked to N-acetylgalactosamine. High-mannose N-glycan chains are barely detectable. After N-glycosidase F treatment the molecular weight is substantially reduced. The main band is 32,500 daltons. Carbohydrate O-linked chains, which are mainly represented by sialic acid, are (2–6)-linked to galactose or N-acetylgalactosamine. Galactose is also organized in O-linked chains and it is (1–4)-linked to N-acetylglucosamine and (1–3)-linked to acetylgalactosamine. Biochemical analysis of carbohydrate structures reveals that no specific carbohydrate complex is bound to a single isoform.     

Variable Deposition of Amyloid β-Protein (Aβ) with the Carboxy-Terminus that Ends at Residue Valine40 (Aβ40) in the Cerebral Cortex of Patients with Alzheimer's Disease: A Double-Labeling Immunohistochemical Study with Antibodies Specific for Aβ40 and the Aβ that Ends at Residues Alanine42/Threonine43 (Aβ42)

Amyloid -protein (A) deposits in the cerebral cortices of patients with Alzheimer's disease (AD) were investigated immunohistochemically to determine their carboxy terminal sequences. Antibodies specific for A terminating at residue valine₄₀ (A40) and at residues alanine₄₂/threonine₄₃ (A42) were used. Virtually all parenchymal A deposits were positive for A42. Many of these deposits were also partially or completely labeled for A40. The degree of A40 labeling varied from area to area within a given brain and from AD case to AD case. In contrast to parenchymal deposits, A40 labeled essentially all the vascular deposits which constitute amyloid angiopathy (AA), with A42 occurring variably in some of these deposits. Occasional AA was found, however, in which A42 predominated or was exclusively deposited. Such a diversity of A species, both in brain parenchyma and in AA, suggests that multiple C-terminal processing mechanisms occur in the cell types responsible for these deposits.     

A phycourobilin-containing phycoerythrin from the cyanobacterium Oscillatoria sp.

A filamentous cyanobacterium Oscillatoria sp. was isolated from a thermal spring of the Kamchatka peninsula. It contained a phycoerythrin unusual for cyanobacteria in that it had a phycourobilin prosthetic group. The absorption spectrum of the native purified phycoerythrin displayed maxima at 498 and 567 nm. The phycoerythrin comprised - and -subunits of molecular weights 18,700 and 19,800, respectively, in 1:1 stoichiometry. Polyacrylamide gel isoelectric focusing revealed one protein band at pI 4.6. The - and -subunits differed in their chromophore composition and content: -subunit carried two phycoerythrobilins while the -subunit had three phycoerythrobilins and one phycourobilin. The chromophore composition of all known phycoerythrins of cyanobacteria and red algae were compared, and on the basis of this comparative study designations C₁- to C₅-phycoerythrin were proposed for cyanobacterial red pigments.     

Morphological aspects of chromosome spindle fibres inMesostoma: “microtubular fir-tree” structures and microtubule association with kinetochores and chromatin

Summary Bipolarly oriented bivalents in spermatocytes of the turbellarianMesostoma ehrenbergii, displaying fast oscillatory movements in metaphase, were studied with the electron microscope. Kinetochores and chromosome fibres were reconstructed using serial sections cut perpendicular to the spindle axis. Only a small proportion of kinetochore microtubules (kMT) is continuous between the kinetochore and the centrosome. kMTs intermingle with non-kinetochore microtubules (non-kMTs), partly inclined with regard to the kMTs, thus forming a chromosome fibre MT lattice. This resembles the microtubular fir-tree structures (MTFT) described by Bajer and Molè-Bajer inHaemanthus endosperm mitosis. A minimal function of the MTFT may be the anchorage of kMTs in the polar region. Regarding the association of MTs with the chromosome, three types of attachment can be discriminated: (1) normal insertion of kMT plus ends in the kinetochore, (2) penetration of kinetochores and deep insertion in the chromatin, and (3) lateral attachment with kinetochore and chromatin. Lateral association of MTs seems to be mediated by filamentous crossbridges. The observations are discussed in connection with possible behaviour of kMTs during kinetochore movement.Abbreviations kMT kinetochore microtubules - MAP microtubule-associated protein - non-kMT non-kinetochore microtubules - MTFT microtubular fir-tree - PCM pericentriolar material     

Relationship of nuclear membranes with filaments and microtubules

Summary In certain HeLa cells characteristic aggregates of cytoplasmic filaments, microfilaments and microtubules were found which preferentially encompass the nucleus. These structures are intimately associated with the membranes of the nuclear envelope and the endoplasmic reticulum as well as with ribosomes. Starting from this observation, a review is presented which demonstrates the nuclear membranes generally as being a type of membrane with a close relationship to proteinaceous structures of the actin-like type such as microtubules and (micro)filaments. As an explanation for the phenomena observed, it is hypothesized that the formation of such tubules and filaments is nucleated at cytomembraneous surfaces and/or that membrane (and possibly also ribosomal) material can be transformed to such tubular or filamentous structures. The concept of cocrystallization between membranes and the diverse tubular and filamentous categories and of a transformation of membrane protein moieties into such structures is thought to provide a basis for explaining many structural phenomena involving membranes and structures of actin-like proteins.The author gratefully acknowledges the careful technical assistance of MissSigrid Krien and MissMarianne Winter and valuable discussions with his colleagues, Drs. H.Falk, H.Kleinig, U.Scheer, as well as with Drs. M.Moses (Duke University), D. J.Morré (Purdue University), and K.Wolff (University of Vienna). The work was supported by the Deutsche Forschungsgesellschaft.     

Ultrastructure and behaviour of the spindle pole body of the aphid-pathogenic fungusErynia neoaphidis

Summary A detailed account of the ultrastructure and behaviour of the spindle pole body (SPB) of the entomophthoraceous fungusErynia neoaphidis is presented for the first time.The SPB consists of extranuclear (ENC) and intranuclear (INC) components. The ENC is a saucepan-shaped structure which lies in a pocket of the nuclear envelope. It is composed of a forked, fibrillar handle and a shallow, cylindrical pan. The pan has a wall of two layers, both of which are thickened with a regular periodicity so that they appear to be beaded. It is postulated that the pan is formed from rough endoplasmic reticulum and that it synthesizes the amorphous, electron-dense material coating the ENC.The INC is a saucer-shaped, electron-dense plaque in which the ends of the spindle microtubules terminate. During metaphase, a clear zone separates the INC from the nuclear envelope and persists until telophase. The roles of the amorphous, electron-dense material and the clear zone as well as the method of SPB replication are discussed.     

Integral and peripheral protein composition of the apical and basolateral membrane domains in MDCK cells

Summary Selective biotinylation of the apical or basolateral domains of confluent MDCK monolayers grown on polycarbonate filters with a water soluble biotin analog, sulfo-NHS-biotin, was employed to reveal strikingly distinct patterns of endogenous peripheral and integral membrane proteins. Peripheral proteins were found to be approximately fivefold more abundant with this procedure than integral membrane proteins, both on the apical and on the basolateral surface. The distinct apical and basal patterns were shown to depend upon the integrity of the monolayer; when the tight junctions were disrupted by preincubation in calcium-depleted medium, the patterns appeared practically indistinguishable. Two-dimensional gel electrophoresis demonstrated that only a very small percentage of the biotinylated proteins were found in similar amounts on both apical and basolateral domains. These results indicate that the sorting mechanisms that segregate apical and basolateral epithelial proteins are very strict. The simple procedure described here has clear advantages over other methods available to label apical and basal epithelial surface domains, namely, higher accessibility of the biotin probe to the basolateral membrane, possibility of purifying biotinylated proteins via immobilized streptavidin and minimal exposure of the researcher to isotopes. It should be very useful in characterizing the apical and basolateral protein compositions of other epithelial cells and in studies on the development of epithelial cell polarity.     

Ecological aspects of swidden cultivation among the Andoke and Witoto Indians of the Colombian Amazon

The investigation of crop and soil-crop conditions among Andoke and Witoto cultivators in southeast Colombia is used as a basis for assessing Geertz' (1963) model of swidden cultivation. In this respect, the extent to which maniocdominated swiddens in the study area simulate the structure and composition of the forest climax community is questioned. As Geertz (1963) indicates, an initial nutrient boost for crop cultivation results from the preliminary burning of forest debris, but weed competition, rather than progressive loss of soil fertility, is reported to be the primary cause of abandoning manioc cultivation after 2–3 years. While the Andoke and Witoto crop system remains adaptive at the individual field level, particularly in its constituent species, its fundamental adaptation is considered to be its integration into the broader field and fallow system that juxtaposes crop production with extended periods of forest regeneration.     

Ultrastructural and biochemical study of extracellular matrix vesicles in normal alveolar bone of rats

Summary The immunocharacteristics of androgen target cells in the anterior pituitary of male rats are assessed by a combined thaw-mount autoradiography and immunoperoxidase staining method permitting the simultaneous identification of steroid-hormone target cells and protein or polypeptide hormone-producing cells in the same preparation. After injection of ³H-dihydrotestosterone, nuclear concentration of radioactivity is observed in cells of the anterior lobe, in ectopic cells in the intermediate and posterior lobes as well as in certain pituicytes. The radioactively labeled cells in the anterior lobe are identified immuno-histochemically as gonadotropes and thyrotropes with the use of antiserum to ovine LH or its subunit -LH and bovine -TSH. The results suggest genomic effects of androgen on gonadotropes and thyrotropes as well as pituicytes.Supported by U.S. PHS Grant NS09914     

Preliminary crystallographic analysis of the Cks protein p13suc1P90AP92A from Schizosacharromyces pombe

The p13^suc1 is the fission yeast member of the Cks (Cdc28-dependant kinase subunit) family of proteins. The Cks proteins bind to and are required for the function of cyclin-dependant kinase (Cdk) proteins during cell cycle progression in eukaryotic cells. Two conformations of Cks have been detected crystallographically; a compact monomer with the C-terminal fourth -strand inserted into the core of the molecule between strands 2 and 3, and a strand-exchanged dimer where the fourth -strand is inserted into the core of the dimer partner in an equivalent position. There is a highly conserved hinge region consisting of the motif PEP, N-terminal to the fourth -strand. In the monomer this motif constitutes a -turn, while in the dimeric structure it is extended, allowing strand exchange. The mutant protein p13^suc1P90AP92A, in which alanine residues replace both prolines of the turn, provides an opportunity to examine the role of the prolines in this hinge region and how they may allow for the formation of strand-exchanged dimers by Cks proteins. We have expressed and purified this mutant protein. Two millimolar p13^suc1P90AP92A crystallised in 50 mM tris(hydroxymethyl)aminomethane pH 7.5, 30% poly(ethylene glycol) 1500. Diffraction data were collected at room temperature on an MAR345 image plate using Cu K radiation from a Rigaku RU200 rotating-anode generator source to 2.70Å. The crystal has unit cell parameters a=b=75.1 Å, c=34.9 Å, ==90°, =120°. Diffraction data were indexed to the space group P6 and systematic absences 00l indicate a screw axis consistent with P6₃.     

Ultrastructure of the tubule of the aglomerular teleost Nerophis ophidion

Summary The tubules in the aglomerular kidney of Nerophis ophidion are composed of cells showing different types of specializations of their plasma membrane. All cells possess a luminal brush border composed of microvilli, and show presence of vesicles with 100 Å thick unit membranes — some containing electron dense material —, tubular elements, multivesicular bodies, and plasma membrane invaginations in their apical cytoplasm. These features suggest an absorptive function of the cells. The apical portions of the cells are supplied with typical cilia.Some cells have abundant basilar plasma membrane invaginations usually lacking cytoplasmic organelles. Other cells appear to form interdigitating basilar cytoplasmic processes containing mitochondria; still other cells have smooth basilar cell membranes. These findings are discussed with reference to the known secretory function of the tubules and are compared with tubular fine structure in other species. It was concluded that urine formation by tubular secretion may occur in cells with different types of basilar cell membrane specializations.The occurrence of coated vesicles associated with invaginated basilar plasma membranes indicates transport of proteins (from peritubular blood vessels ?) at these sites.The tubule cells have abundant smooth surfaced endoplasmic reticulum and large and numerous active Golgi zones.Supported by grants from Fonden til Videnskabens Fremme and Therese och Johan Anderssons Minne. Part of this study was done at the Zoologica Stazione, Naples.The assistance of Miss Britt-Marie Petterson and Mr. Magnus Norman is gratefully acknowledged.     

Oligomerization of deoxynucleoside-bisphosphate dimers: Template and linkage specificity

Evidence is presented that a poly(U) template selectively favors the oligomerization of the activated, 3–5 pyrophosphate-linked dimer pdAppdAp, in comparison with the 3–3 and 5–5 linked dimers. In the absence of poly(U), the 5–5linked dimer is the most reactive, and chains are formed which are more than 60 monomer units in length.Nucleic Acid-Like Structures V. For the previous paper in this series see Visscher and Schwartz (1988).     

Cytochemical studies on the localization of 5′-nucleotidase in the acinar cells of the rat salivary glands

Summary The cytochemical localization of 5-nucleotidase (5-AMPase), and its validity, were investigated in parotid and submandibular acinar cells of a rat. Biochemical determinations showed that adequate treatment with glutaraldehyde could minimize the loss of enzymatic activity, and that 5-AMPase and non-specific alkaline phosphatase (-GPase) possessed different pH optima.The cytochemical distribution of the reaction products from the 5-AMPase activity was distinct from those of -GPase. 5-AMPase activity was localized on the surface membranes of acinar, ductal and myoepithelial cells of both salivary glands. -GPase activity was evenly distributed on the entire plasma membranes of myoepithelial cells and on the basal plasmalemma of acinar cells. The reaction products, which appeared on the luminal and lateral plasma membranes of the acinar cells, were presumed to reflect the presence of 5-AMPase, while those on the myoepithelial surface and basal plasma membranes of the acinar cells demonstrated both 5-AMPase and -GPase.The results indicate that 5-AMPase activity can be utilized as a reliable marker enzyme of plasma membranes in the salivary acinar cells.     

Effects of Lead on Adenylate Cyclase Activity in Rat Cerebral Cortex

Lead decreased in a dose dependent manner the basal AC activity in membranes of rat cerebral cortex (IC₅₀ = 2.5 ± 0.1 M). In membranes preincubated under basal conditions, AC activity was stimulated by approximately two and fourfold by 10 M Gpp(NH)p or forskolin, respectively. Under basal conditions, lead (3 M) inhibited enzyme activity up to 50%, but was not able to inhibit the Gpp(NH)p- or the forskolin-stimulated AC activity. However, in membranes preincubated with Gpp(NH)p (10 M), lead (3 M) had no significant effect on enzyme activity, but it partly blocked the stimulation of AC activity elicited by forskolin (10 M). In membranes preincubated with 10 M lead, the addition of 10 M Gpp(NH)p or forskolin in the incubation medium did not stimulate AC activity. However, when added together in the incubation medium Gpp(NH)p + forskolin produced an increase in enzyme activity. In membranes preincubated with 10 M lead + 10 M Gpp(NH)p, Gpp(NH)p (10 M) or forskolin (10 M) added alone or in combination to the incubation medium did not stimulate AC activity. Moreover, under these latter conditions lead had no further effect on enzyme activity. These results indicate that lead may interact with G-proteins and with the catalytic subunit of cerebral cortical AC to produce inhibition of the enzyme activity.     

Etude ultrastructurale d'une formation de soutien épidermo-conjonctive inédite chez les Insectes

Résumé Une formation de soutien (formation hypopharyngienne transverse ou FHT) est tendue transversalement dans la cavité hypopharyngienne de Blabera craniifer BURM. (Dictyoptère). Elle est constituée par les épidémies droit et gauche hypertrophiés reliés l'un à l'autre par un matériel conjonctif.Le cytoplasme épidermique est envahi par des microtubules qui s'attachent sur les membranes plasmatiques apicale et basale; les zones d'attache de microtubules sont comparables aux hémidesmosomes connus chez les Vertébrés.La jonction épidermo-cuticulaire est assurée par l'association des zones d'attache apicales et de fibres à structure périodique traversant toute la cuticule. La jonction épidermo-conjonctive est réalisée par l'association des zones d'attache basales et d'une couche de matériel dense extra-cellulaire par l'intermédiaire de laquelle le conjonctif (et en particulier les fibres collagènes) s'attache sur la membrane plasmatique.Cette liaison conjonctivo-épidermo-cuticulaire est comparable à la liaison myo-épidermo cuticulaire des Arthropodes.De telles formations de soutien semblent communes et il est probable que le rôle squelettique du conjonctif a été jusqu'ici sous-estime chez les Insectes.

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Ultrastructure of a supporting structure in insects

Summary A supporting structure (FHT) is stretched transversely in the hypopharyngeal cavity of Blabera craniifer Burm (Dictyoptera); the right and left epidermal cell layers are hypertrophied and connected by connective tissue.The cytoplasm of the epidermal cells is filled with an unusually large number of microtubules inserted both on the apical and basal plasma membranes; the attachment zones of microtubules are similar to the hemidesmosomes of Vertebrates.The epidermo-cuticular junction is formed through the association of intracuticular banded fibers with the apical attachment zones and the epidermo-connective tissue junction through the insertion of connective tissue structures (collagen fibers and others) on an extra-cellular dense material layer apposed to the basal attachment zones.This connectivo-epidermo-cuticular connection can be compared to the myo-epidermocuticular connection of Arthropods.Supporting formations of this type are not infrequent, so it appears that the skeletal functions of the connective tissue of Insects have been underestimated previously.

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Nous tenons à remercier le Prof. Ch. Noirot et Madame Noirot-TimothÉe pour leur aide efficace et les suggestions qu'ils nous ont apportées au cours de la réalisation de ce travail.