摘要:

Summary—The entry into, and exit from, mitosis are controlled by a universal M‐phase promoting factor (MPF) composed of at least p34cdc2and a cyclin. Embryonic systems are convenient for studying the association and dissociation of the active MPF complex because oocytes and eggs are naturally arrested at a specific point of the cell cycle until progression to the next point is triggered by a hormonal signal or sperm. In amphibians, eggs prior to fertilization are arrested at metaphase 2 of meiosis due to the presence of a stabilized MPF complex. Fertilization (egg activation) produces a transient increase in intracellular free Ca2+, a propagating Ca2+wave, that specifically triggers the destruction of cyclin, leading to MPF inactivation and entry into the first embryonic interphase. We have recently shown that intracellular pH (pHi) variations in amphibian eggs, a large increase at fertilization and small oscillations during the embryonic cell cycle, were temporally and functionally related to the corresponding changes in MPF activity. In addition, the recent finding that the pHiincrease at fertilization inXenopuseggs is a propagating, Ca2+‐dependent pH wave which closely follows the Ca2+wave, together with the absence in the egg plasma membrane of pHi‐regulating systems responsible for that pHiincrease, suggest the existence of cortical or subcortical vesicles acidifying in the wake of the Ca2+wave, thus producing the pH wave. Given the known functions of intracellular acidic compartments in various systems and the existence of acidic vesicles in marine invertebrate eggs, we propose that these hypothetic acidic vesicles inXenopuseggs might serve as a storage for the p34cdc2released in the cytoplasm following egg activation, both the pHiincrease and MPF inactivation being triggered and synchronized by the same signal, the C

ISSN:0248-4900    

DOI:10.1016/0248-4900(92)90137-P

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

Summary—Xenopusoocytes are blocked in prophase of the first meiotic division. During the G2/M transition drastic changes occur both in the cytoskeletal organization and in the capacity of tubulin to polymerize. Posttranslational modification of tubulin isoforms might be one of the factors that control the dynamic properties of microtubules. We have therefore analysed, by two‐dimensional polyacrylamide gel electrophoresis, the isotubulins purified fromXenopusoocytes, and we show that tubulin is resolved into at least four α‐isoforms and four β‐isoforms. We have identified a basic α (αb)‐tubulin isoform which is specific to prophase arrested oocyte and that progressively disappears during meiotic maturation; its decrease is initiated when the nuclear envelope breaks down and is controlled by the nucleus. Using 35S methionine labelled oocytes we demonstrate that the disappearance of the αbisotubulin results from both an arrest of its biosynthesis after maturation, and from posttranslational modification which induces a shift of this α‐isoform to a more acidic pI. Moreover,in vitroexperiments using 35S prelabelled tubulin purified from prophase oocytes show that metaphase extracts containing MPF activity are able to induce the acidification of the αb‐isoform, suggesting that the observed post‐translational modification might be regulated by p34cdc2. However, the nature of this modification re

ISSN:0248-4900    

DOI:10.1016/0248-4900(92)90138-Q

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

Summary—Anti‐Sm antibodies recognize the major small nuclear RNA‐protein particles (snRNPs) involved in pre‐mRNA processing. The spatial distribution of the snRNPs has been investigated inDrosophilaembryos up to the cellularization stage (cycle 14), using the Y12 anti‐Sm antibody. Our results show that: 1) all or most of the Sm antigen is localized in the cytoplasm of the syncitial blastoderm until the 12th cycle of division, in both the nuclear and cytoplasmic compartments at cycle 13, and then in the nuclei at cycle 14 and later. This relocalization takes place when zygotic transcriptional activation occurs; 2) at the subcellular level, the Sm antigen localizes in a speckled pattern and in foci‐like structures within the nucleus ofDrosophilablastoderm embryos; 3) strikingly, some nuclei of embryos at the 14th cycle appear to contain more snRNPs than others. The position of these nuclei differs from one embryo to another, and their distribution does not resemble any known developmental pattern ofDrosophilaembryogenesis. We propose that random differences in snRNP concentration may serve as an epigenetic signal for stochastic events occurring during

ISSN:0248-4900    

DOI:10.1016/0248-4900(92)90139-R

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

Summary—Immunofluorescent techniques have been used in the analysis of DNA‐RNA hybrids occurrence and its relationship to transcriptional events on polytene chromosomes ofDrosophila subobscura. We have studied the distribution of these hybrids on uninduced/induced chromosomes. Two different indirect immunofluorescence methods for the detection of DNA‐RNA hybrids were used. Our data confirm the positive correlation between localization of DNA‐RNA hybrids and transcriptional activity by following the Büsenet alprocedure 1982. Using the other protocol, which allows chromosomal DNA‐RNA to denature and renature, makes DNA‐RNA hybrids detectable not exclusively in active chromosomal regions. Taking Büsen as method of choice, this technique allowed to localize the exact transcriptional active sites on puffs: hybrid fluorescence was restricted to marginal or central puff areas. Moreover, no correlation between fluorescence and puffs size was found. However, our studies on induced chromosomes indicate that: 1) the 15DE puff, previously described as t‐puff, was not really a heat shock puff, since no transcriptional activity was detected; 2) hybrid fluorescence at 2C and 31CD regions was observed. No labelling was found in these loci in the autoradiography data, reported b

ISSN:0248-4900    

DOI:10.1016/0248-4900(92)90140-V

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

Summary—Madin‐Darby canine kidney (MDCK) cells have been extensively used as a model for the study of epithelial polarization. The contacts between the cell and extra‐cellular matrix (ECM) provide a signal for the polarization of apical membrane markers. In order to study the molecular basis of these contacts, MDCK cells extracts in Triton X‐100 were affinity‐purified on laminin, yielding polypeptides of 100–110 and 36 kDa, but only the second one could be enzymatically iodinated from the cell surface. This protein was also recognized by an antibody against the 37/67‐kDa laminin/elastin family of proteins. Different polypeptides were purified by the same method on type I collagen. An antibody developed against the polypeptides purified on laminin recognized also a 67‐kDa protein, blocked 125I‐laminin binding to a population of high affinity (1.5 nMKD) binding sites and caused a significant decrease in cell attachment and spreading to laminin or endogenous ECM. This antibody did not interfere with MDCK cell attachment to fibronectin or collagen matrices, but still impaired cell spreading. An apical MDCK plasma membrane protein (184 kDa), fully polarized in untreated cells, was partially mispolarized after treatment with anti‐36 kDa antibody. These results are consistent with a model of various ECM receptors operating together in these cells, and show an important role of a non‐integrin 36‐kDa laminin binding protein related to the 67‐kDa laminin receptor family in cell attachment, s

ISSN:0248-4900    

DOI:10.1016/0248-4900(92)90141-M

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

Summary—Functional relations between exocytotic vesicle membranes, plasmalemma and milk fat globule membranes (MFGM) were studied during the final stages of mouse mammary gland differentiation, in the gland during full lactation and in the postpartum gland in which the synthesis of secretory products was partly inhibited by application of 2‐Br‐α‐ergokryptin. Analysis of ultrathin sections, freeze‐fracture replicas, scanning electron microscopy and application of a cytochemical marker filipin showed that the apocrine secretion of lipid globules was closely related to the exocytosis of milk proteins. During the last days of gestation the secretion of lipid globules resulted from many exocytotic events of the secretory vesicles that accumulated and fused around the cytoplasmic lipid droplets. Seldom the lipid droplet protruded partly into the gland lumen and a part of its surface became covered with the apical plasmalemma. Although apical plasmalemma became more important in the formation of MFGM in the postpartum period, we could still confirm a direct contribution of secretory vesicle membranes to the final detachment of the lipid globule. The application of 2‐Br‐α‐ergokryptin hindered the apocrine secretion of the lipid globules and a situation similar to the situation in the prepartum g

ISSN:0248-4900    

DOI:10.1016/0248-4900(92)90142-N

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

Summary—Biosynthetic labelling experiments performed onP primaureliastrain 156, expressing the temperature‐specific G surface antigen, 156G SAg, demonstrated that the purified 156G SAg contained the components characteristic of a GPI‐anchor. [3h]ethanolamine, [3h]myo‐inositol, [32p]phosphoric acid and [3h]myristic acid could all be incorporated into the surface antigen. Myristic acid labelling was lost after treatmentin vitrowithBacillus thuringiensisphosphatidylinositol‐specific phospholipase C (PI‐PLC). After complete digestion by pronase, a fragment containing the intact GPI‐anchor of 156G surface antigen was isolated. This fragment was shown to be hydrophobic and glycosylated and to possess an epitope found specifically in the GPI component of GPI‐anchored proteins. The role of the GPI‐tail in anchoring the 156G surface antigen into the membrane was assessed by determining that purified 156G molecules with the GPI‐anchor could be incorporated into lipid vesicles and red cell ghosts whereas the 156G molecules lacking the GPI‐anchor, as result of treatment withB thuringiensisPI‐PLC, could not. It has also been shown that the membrane‐bound form and the soluble form, obtained after cleavage of the 156G SAg lipid moiety either by an endogenous PI‐PLC or by a bacterial PI‐PLC, displayed identi

ISSN:0248-4900    

DOI:10.1016/0248-4900(92)90143-O

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

Summary—The three‐dimensional architecture of the nucleolonema ofVicia fabahas been studied by applying a silver impregnation technique to serial ultrathin sections. This technique disclosed lateral and transverse segments of the nucleolonema which were heavily impregnated with silver. The lateral profiles of the nucleolonema segments were classified into three main categories; a segment made up of one to several rod‐like filaments (type I); a ladder‐like segment consisting of two parallel and of transverse filaments (type II); and a last type constructed from two parallel filaments (type III). Tracing of the lateral segments through serial sections has indicated that type I first appears, then either type II or III and finally type I reappears at the corresponding sites on sections. Types II and III remained constant in width, about 1.0 μm, along their longitudinal axes whereas the width of type I was significantly smaller than that of the two former. The lateral filaments of both types II and III showed heterogeneity in width on account of the presence of knobs intermittently distributed along them. The thickness of these knobs was about 0.35 μm. Combining the observations on serial ultrathin sections and the morphometrical data it is very probable that the elementary structure of the nucleolonema is a 0.35‐μm thick filament that tightly coils up into a solenoid structure with a thickness of approximately 1.0 μm. This model can explain the appearance of open‐ and closed‐argyrophilic rings in serial sections since transverse segments of the solenoid are expected to show the argyrophilic rings. The elementary filament of the nucleolonema solenoid was sometimes loosened. Judging from our cytochemical data at the electron microscope level, some argyrophilic proteins appear to reside in the axial space of the solenoid but both DNA and RNA were not detecta

ISSN:0248-4900    

DOI:10.1016/0248-4900(92)90144-P

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

Summary—Positional and structural modifications were demonstrated in nuclei of leek cells, after establishment of a symbiosis with two vesicular‐arbuscular fungi,Glomus versiformeandGlomus E3. By combining light, immuno‐electron microscopy and morphometry, the fungi were shown to have a direct effect on the host nuclear morphology: the effect was confined to a specific plant tissue (the cortical parenchyma) and to a moment of the fungal morphogenesis (the arbuscule). When they branch to form the complex structures called arbuscules in the inner parenchyma cells, the host nucleus migrates from the periphery of these cells towards their centre. In addition, it becomes larger and lobed, with a decondensed chromatin. A monoclonal antibody that mostly binds to the condensed chromatin revealed a significant decrease in gold labelling intensity over the nuclei of the colonized cells. These modifications suggest that the nuclear migration and the changes in chromatin organization are related to the modifications in gene expression observed during the establishment of mycorrhizal symb

ISSN:0248-4900    

DOI:10.1016/0248-4900(92)90145-Q

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

Summary—Transformations of the actin cytoskeleton of the pollen ofHyacinthus orientalisduring cycles of vapour‐phase hydration and dehydration have been examined using a non‐fixation, DMSO permeabilisation method for TRITC‐phalloidin staining, coupled with microwave stabilisation. In freshly shed pollen actin appears: a) at the plasmalemma in the form of extended, thin fibrils co‐oriented with the cellulosic microfibrils of the contiguous intine; b) as a sheath investing the generative cell; c) as spicules around the vegetative nucleus; and d) in scattered spicules in the cytoplasm. During hydration in 85–95% relative humidity (RH), actin from all of these sites is progressively translated into a system of extended fibrils in the vegetative cell, concurrently with the onset of movement in the cytoplasm. Dehydration at 5–7% RH reverses this process, actin accumulating in rodlets, spicules or larger fusiform bodies in close association with the generative cell and vegetative nucleus, and also in the cytoplasm. The fibril system initially present at the plasmalemma is not restored. After ten cycles of hydration and dehydration 3.6% of the grains remained germinable. The ecological significance of the findings is noted, and the possibility that the observed transformations result from variation in the Ca2+concentration in the cytosol as the water content of the cytosol changes

ISSN:0248-4900    

DOI:10.1016/0248-4900(92)90146-R

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY