摘要:

Summary—Mediatophore is a nerve terminal membrane protein purified fromTorpedoelectric organ on its ability to translocate acetylcholine upon calcium action. An antiserum able to immunoprecipitate mediatophore activity was used to study the subcellular distribution of this protein. The presynaptic membrane exhibited a strong and discontinuous immunogold labelling, especially at the active zone where ACh is thought to be released. Two antigens were recognized on immunoblots of synaptosomal membranes: the 15‐kDa subunit of mediatophore and a 14‐kDa membrane protein that has a wide non‐neuronal distribution. Antibodies purified from the serum on native mediatophore and monospecific towards the 15‐kDa antigen still gave a high presynaptic membrane localized labelling. In addition, a few 14‐kDa protein sites were present at the active zone. The Schwann cell finger interposed between the presynaptic membrane and the postsynaptic arch also exhibited the 14‐kDa antigen raising the question of a possible interaction of mediatophore with the 14‐kDa protein originating from t

ISSN:0248-4900    

DOI:10.1016/0248-4900(93)90125-X

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

Summary—In this study we describe the biochemical features of theToxoplasma gondiitachyzoite surface glycoprotein, gp23, demonstrating that it is attached to the parasite membrane by a glycosyl‐phosphatidyl inositol anchor. Gp23 was metabolically labeled with tritiated palmitate, myristate, ethanolamine, inositol, glucosamine, mannose and galactose, as expected for a GPI‐anchor structure. Gp23 was released from the surface of living parasites after treatment with phosphatidyl inositol‐specific phospholipase C (PI‐PLC) and the resulting water‐soluble protein was immunoprecipitated with a monoclonal antibody specific for gp23. The GPIcore glycan was generated after aqueous‐HF dephosphorylation followed by nitrous acid deamination and its carbohydrate structure was analyzed using selective exo‐ and endoglycosidase treatments. Finally, the phosphatidylinositol moiety of gp23 was characterized using PI‐PLC and phospholipase A2(PLA2) digestions. Our cumulative data suggest that gp23 ofT gondiitachyzoites contains a modified GPI‐backbone similar to the mammalian Thy‐1 anchor, consisting of a conserved core structure (ethanolaminePO4‐6‐Manαl‐2‐Manαl‐6‐Manαl‐4‐GIcNαl‐6‐PI) bear

ISSN:0248-4900    

DOI:10.1016/0248-4900(93)90126-Y

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

Summary—We are investigating the roles of RNA synthesis, chromatin structure and nuclear matrix organization in establishing and maintaining transcription domains, using mitogen stimulated lymphocytes as a model system. In a continuing study, the effects of the RNA polymerase inhibitor DRB and of its removal on nuclear organization have been examined by EM cytochemistry and by immunofluorescence labelling of the nuclear matrix P11, Sm and nucleolar fibrillarin antigens. Chromatin, interchromatin granules and nucleoli were extensively restructured after DRB, as were matrix antigens. According to cytochemical staining properties, the conformation of DRB‐induced condensed chromatin resembled that in partially stimulated lymphocytes. The nucleoplasmic fibrogranular RNP network appeared little altered, but the fibrillar proteinaceous interchromatinic regions, interpreted as representing the nuclear matrixin situ, were more affected. After removal of DRB, nuclei recovered the organization and transcriptional activity of controls within 8 h. These results suggest that the matrix subtending transcription domains remains stable when transcription is arrested, even though the chromatin and individual RNP components of the domains are disorganized. The data further indicate that absence of transcription is not solely accountable for the highly aggregated state of the chromatin in resting lymphocy

ISSN:0248-4900    

DOI:10.1016/0248-4900(93)90127-Z

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

Summary—Nasal cartilage cells from 21‐day‐old rat fetuses were cultured at high density in the presence of ascorbic acid and β‐glycerophosphate over a 12‐day period. Immediately after plating, the cells exhibited a fibroblastic morphology, lost their chondrocyte phenotype and expressed type I collagen. On day 3, clusters of enlarged polygonal cells were found. These cell clusters synthetised type II collagen and formed an alcian‐blue‐positive matrix. The following days, a progressive increase in the number of cells positive for type 11 collagen was noted and, on day 8, typical cartilaginous nodules were formed. These nodules increased in size and number, spreading outward, laying down a dense matrix which mineralized. Light and electron microscopy observations of cross‐sections of nodules confirmed the cartilaginous nature of this tissue formedin vitrowith typical chondrocytes embedded in a hyaline matrix. Furthermore, at the electron microscopic level, matrix vesicles were seen in extracellular matrix associated with the initiation of mineralization. Typical rod‐like crystals were present in the intercellular spaces along the collagen fibers. These results indicated that in a specific environment, dedifferentiated chondrocytes were able to redifferentiate and to form nodular structures with morphological ultrastructure of calcified cartila

ISSN:0248-4900    

DOI:10.1016/0248-4900(93)90128-2

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

Summary—The influence of triiodothyronine (T3) on avian myoblast proliferation and differentiation was studied in secondary cultures using plating densities of 2500 and 7000 cells/cm2. Culture media were depleted of T3 (control myoblasts) and increasing amounts were then added to concentrations of 0.6, 3 and 15 nM T3 (treated myoblasts). Independent of the cell density, T3 induced a dose‐related decrease in myoblast proliferation measured by cell number, doubling time and3H‐thymidine incorporation. However, with the lower plating density, this influence was delayed, occurring only after the third day of culture for 0.6 nM T3‐treated myoblasts and simultaneous with the onset of myosin heavy chain accumulation. Moreover, when myoblasts were exposed to BrdU for 48 h, the T3 growth inhibitory effect disappeared, thus showing that this effect was clearly linked to differentiation. In addition, we have shown that T3 induced an early fusion of myoblasts: 65% of the maximal value of the fusion index was reached on day 3 in the T3‐treated cells in comparison to 25% in the control myoblasts. This hormone also enhanced accumulation of muscle‐specific proteins (connectin, acetylcholine receptors, myosin heavy chain), tested by cytoimmunofluorescence, ELISA, binding experiments and Western blot. All these results show that T3 increased myoblast differentiation through a pathway including myoblast withdrawal from the cell cycle. The influence of T3 could partly explain its previously reported positive effect on the number of mu

ISSN:0248-4900    

DOI:10.1016/0248-4900(93)90129-3

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

Summary—It is well known that numerous germ cells degenerate during the first meiotic division and spermatid elongation is due mostly to an adverse physiological microenvironment in relation to hormonal deficiency. The present study is aimed at investigating the endocytic activity of germ cells, during the first wave of mouse spermatogenesis, using transferrin coupled to gold particles, in order to study the efficiency of this possible pathway of communication between Sertoli cells and germ cells. Labelling experiments in control animals confirmed a receptor‐mediated pathway in all germ cells during puberty. Furthermore, our morphological and quantitative data revealed that during spermatid elongation, degenerating germ cells possessed a highly developed endocytic apparatus which contained twice as many transferrin gold particles than normal adult cells. The fact that endocytosis of transferrin was increased in degenerating germ cells indicates that, most probably, germ cell degeneration during the first wave of spermatogenesis did not result from a deficiency in iron transport. The higher endocytic activity of degenerating germ cells, compared to adult control cells, could not only be the result of a simple process of plasma membrane internalization but also a complex mechanism which could be involved in the degradation of the ce

ISSN:0248-4900    

DOI:10.1016/0248-4900(93)90130-7

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

Summary—Secretory vesicles of the ciliatePseudomicrothorax dubius, called trichocysts, are separated into>40 proteins by two‐dimensional gel electrophoresis. The trichocyst, composed of a shaft and four arms, is in a condensed state when docked in the cell cortex, and it elongates into an extended state during exocytosis. Monoclonal antibodies (mAbs) were raised against trichocyst proteins. Their reactivities were analysed: I) on Western blots of extended, isolated trichocysts by immunolabeling; 2) on entire cells and extended trichocysts by indirect immunofluorescent binding assay (IFA); 3) on semi‐thin sectioned cells by IFA; and 4) on ultra‐thin sections of cells by immunogold labeling. mAb IV 4E5 labels major trichocyst proteins at 15–19, 22 and 24 kDa, pI4.6−6.6. The epitope recognized by mAb IV 4E5 is common to as many as 30 proteins and suggests a family of proteins with possible sequence homology. By IFA, the shafts of extended trichocysts are labeled. The shafts of condensed trichocysts are labeled on both semi‐thin sections in Lowicryl and ultrathin sections. On semi‐thin Epon sections, the part of the trichocyst which is labeled is arm‐like. mAb VI 2D12 labels three major trichocyst proteins at 31 kDa, pI5.0−5.4. The arms of extended trichocysts are labeled by IFA, but are only weakly labeled on ultrathin sections. The shaft of extended trichocysts is labeled by IFA, and the shaft of condensed trichocysts is labeled on

ISSN:0248-4900    

DOI:10.1016/0248-4900(93)90131-W

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

Summary—Based on immunoblotting procedure, the isolated epithelium of amphibian skin was found to contain a 180 kDa protein which cross‐reacts with a polyclonal antiserum raised against human erythrocyte Band 3. Immunoperoxidase and immunofluorescence staining techniques indicated that the Band 3‐related protein was localized in the mitochondria‐rich cells (MRC) of this epithelium, with characteristic apical labelling pattern. Our findings show that the putative apical anion exchanger of the MRC is immunologically related to the band 3 multigenic family, which catalyzes Cl‐HCO3−transmembranous exchange. It thus suggests a molecular basis for the role played by these cells in the transepithelial Cl pathway and acid‐ba

ISSN:0248-4900    

DOI:10.1016/0248-4900(93)90132-X

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

Summary—Rhodamine phalloidin‐staining was used to study the organization of the cortical actin cytoskeleton of the earlyCeratitis capitataembryo. The dynamics of the actin aggregates and their changes in distribution during the formation of the syncytial blastoderm, were followed in detail. It was found that these aggregates formed a shell‐like cluster around the interphase nuclei, and concentrated toward the poles of the mitotic apparatus when the nuclei divided. Laser scanning confocal microscopy revealed that aggregates not clustered at the poles of the mitotic apparatus were closely associated with fine fibers of a dense cytoplasmic network of actin fila

ISSN:0248-4900    

DOI:10.1016/0248-4900(93)90133-Y

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

Summary—The role of microtubules in intracellular transport of African swine fever virus (ASFV) and virus‐induced inclusions was studied by immunofluorescence using anti‐ASFV and anti‐tubulin antibodies, by electron microscopy of infected Vero cells and byin vitrobinding of virions to purified microtubules. MTC, a reversible colchicine analogue, was used to depolymerize microtubules. In cells treated with MTC multiple large inclusions containing ASFV antigens and particles were observed in the cytoplasm. Removal of the drug lead to migration and fusion of the inclusions at a perinuclear location. To study the effect of microtubule repolymerization on virus particle distribution, the particles were counted in thin sections of MTC treated cells and at different times after removal of the drug. In cells treated with MTC 6.8% and 3.6% of the virus particles were found respectively in the cytoplasm and at the cell membrane while 38% of the particles were located around the virosome. With reversal of the drug effect the number of virus particles around the virosomes progressively decreased to 10% at 2 h while the number of particles in the cytoplasm and at the cell membrane increased. At 2 h after removal of the drug 33.5% of the particles were found budding from the cell membrane. Virus particles were found closely associated with microtubules in cytoskeletons obtained by Triton X‐100 extraction of taxol treated cells. The association of virus particles with microtubules was also observedin vitrousing purified microtubules and virus particles. The results show that microtubules are involved in the transport of African swine fever virus particles from the assembly site to the cell surface and in the movement and fusion of the virus i

ISSN:0248-4900    

DOI:10.1016/0248-4900(93)90134-Z

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY