Enzo Bard - Academia.edu (original) (raw)

Papers by Enzo Bard

Annals of the New York Academy of Sciences, May 1, 1987

The genus Leishmania includes several species that cause disease in man. These

Biochemistry and Cell Biology, Sep 1, 1989

Leishmania is a trypanosomatid protozoa with a digenetic life cycle. Sandflies inject promastigot... more Leishmania is a trypanosomatid protozoa with a digenetic life cycle. Sandflies inject promastigotes, the free living form present in their salivary glands, into mammals where the parasite colonizes macrophages, transforming into intracellular amastigotes. The cycle is completed when during a blood meal the insect ingests infected macrophages, the amastigotes are released in the gut where they transform back into promastigotes. Leishmania has to adapt to the changing life conditions, from free-living forms in the poikilothermic insect vector to obligatory intracellular parasite in the homeothermic mammalian host. It also has to adapt to the acidic pH of the macrophage's phagolysosome where amastigotes multiply. The adaptative response of Leishmania includes morphological, physiological, and biochemical changes. Promastigotes can be grown in culture medium. Studies of changes taking place during adaptation have been facilitated by the establishment of in vitro conditions that allow the transformation of amastigotes into promastigotes and vice versa. The system is well suited for studying regulation of gene expression during adaptative differentiation. Some mechanisms of mRNA processing are unique to these protozoa: trans-splicing and RNA editing. Several genes that are differentially expressed in the two stages have been studied. No obvious cis regulatory motifs have been found in the DNA.Key words: Leishmania, genes, differentiation, regulation.

Elsevier eBooks, 1977

Publisher Summary This chapter discusses that several types of structural and kinetic evidence in... more Publisher Summary This chapter discusses that several types of structural and kinetic evidence indicate that hnRNA contains mRNA precursors in a variety of forms. In some cases the precursors consist of hnRNA niolecules that are considerably larger than the mature mRNA. In other cases the size of the pre-mRNA is similar to that of the mature mRNA, either because processing events have closely followed transcription, or because the transcriptional units are not significantly larger than the mature mRNA. The mRNA segments may be located at the 5' termini of initial transcripts as well as at internal regions that are exposed by appropriate processing cleavages. Nuclear processing of the mRNA precursors is completed by cap formation on the 5' terminus and, for most mRNA species, methylation of internal adenylate residues and polyadenylylation of the 3'-OH terminus. The cellular content of any particular mRNA species and its precursors is determined by the relative rates of transcription, processing and turnover.

Journal of Cell Biology, 1984

The intracellular pathway followed by the influenza virus hemagglutinin (HA) to the apical surfac... more The intracellular pathway followed by the influenza virus hemagglutinin (HA) to the apical surface of Madin-Darby canine kidney cells was studied by radioimmunoassay, immunofluorescence, and immunoelectron microscopy. To synchronize the migration, we used a temperature-sensitive mutant of influenza WSN, ts61, which, at the nonpermissive temperature, 39.5°C, exhibits a defect in the HA that prevents its exit from the endoplasmic reticulum. Upon transfer to permissive temperature, 32°C, the HA appeared in the Golgi apparatus after 10 min, and on the apical surface after 30-40 min. In the presence of cycloheximide, the expression was not inhibited, indicating that the is defect is reversible; a wave of HA migrated to the cell surface, where it accumulated with a half time of 60 min. After passage through the Golgi apparatus the HA was detected in a population of smooth vesicles, about twice the size of coated vesicles, located in the apical half of the cytoplasm. These HA-containing vesicles did not react with anti-clathrin antibodies. Monensin (10 ' UM) delayed the surface appearance of HA by 2 h, but not the transport to the Golgi apparatus. Incubation at 20°C retarded the migration to the Golgi apparatus by^-30 min and blocked the surface appearance by acting at a late stage in the intracellular pathway, presumably at the level of the post-Golgi vesicles. The initial appearance of HA on the apical surface was in the center ; no preference was observed for the tight-junctional regions .

Current topics in membranes and transport, 1985

Elsevier has created a COVID-19 resource centre with free information in English and Mandarin on ... more Elsevier has created a COVID-19 resource centre with free information in English and Mandarin on the novel coronavirus COVID-19. The COVID-19 resource centre is hosted on Elsevier Connect, the company's public news and information website. Elsevier hereby grants permission to make all its COVID-19-related research that is available on the COVID-19 resource centre-including this research content-immediately available in PubMed Central and other publicly funded repositories, such as the WHO COVID database with rights for unrestricted research re-use and analyses in any form or by any means with acknowledgement of the original source. These permissions are granted for free by Elsevier for as long as the COVID-19 resource centre remains active.

Springer eBooks, 1973

The mechanisms of initiation and termination of the synthesis of protein chains have been the sub... more The mechanisms of initiation and termination of the synthesis of protein chains have been the subject of many studies in recent years, both in mammalian and in bacterial systems (for review, see ref. 1). Those studies emphasize the utilization of ribosomal subunits in the process of chain initiation and polyribosome formation either as derived subunits in the presence of defined initiation factors (2–6) or as native ribosomal subparticles pre-existent as such in the cytoplasm (7–10).

Proceedings ... annual meeting, Electron Microscopy Society of America, Aug 13, 1982

The polarized distribution of surface components between apical and basolateral domains of the pl... more The polarized distribution of surface components between apical and basolateral domains of the plasma membrane constitutes the basis of epithelial function. We are currently studying the mechanisms employed by epithelial cells to segregate different sets of integral proteins in two opposite regions of the plasma membrane. For this purpose, we are utilizing a model system which involves the infection of polarized epithelial cell lines, such as the dog kidney cell line MDCK, with enveloped RNA viruses. Influenza virus and two paramyxoviruses bud from the apical surface regions of MDCK cells, vesicular stomatitis virus (VSV), a rhabdovirus, is assembled instead from the basolateral surface. A main determinant of polarized budding appears to be the addressing of viral envelope glycoproteins to the surface domain that the virus utilizes for budding. The mechanisms and intracellular pathways involved in this sorting are probably the same as those utilized by the cell for its own surface proteins.

Elsevier eBooks, 1979

Publisher Summary This chapter discusses the synthesis and processing of RNA in stimulated fibrob... more Publisher Summary This chapter discusses the synthesis and processing of RNA in stimulated fibroblasts and lymphocytes. Several reviews have discussed the regulation of gene expression at various levels and in a variety of cell types. The primary gene transcripts, in the case of all kinds of RNA, must be converted to mature, functional molecules by modification or processing. Processing includes cleavage of nonconserved sequences, methylation of bases and sugars, and terminal addition of nucleotides to the 3′ OH and the 5′ P ends of the molecules. The chapter presents a comparison of the mechanisms of regulation of RNA metabolism of activated fibroblasts with those of lymphocytes. Extensive gene derepression and synthesis of new transcripts is unlikely to be characteristic of the relatively early events in lymphocytes. A major factor in the train of events following early upon the triggering of lymphocytes is an acceleration in the processing of RNA and in its transport from nucleus to cytoplasm.

Journal of Biological Chemistry, 1981

Treatment of rats with phenobarbital (PB) leads to a substantial increase in the hepatic levels o... more Treatment of rats with phenobarbital (PB) leads to a substantial increase in the hepatic levels of translatable polysomal poly(A) + cytochrome P-450 mRNA. An enriched fraction of P-450 mRNA was obtained by agarose gel electrophoresis and used to prepare a cDNA probe by differential hybridization to total mRNA from control and PB-treated rats. The majority of the sequences within the probe hybridized to recombinant DNA plasmids which contained a bona fide P-450 cDNA insert identified by positive hybridization selection and in vitro translation. The cDNA probe was used to demonstrate that PB treatment leads to a 30-fold increase in polysomal P-450 mRNA, which is not due to more efficient utilization of previously existing mRNA but to the appearance of new messenger in the cytoplasm. The induction of cytoplasmic P-450 mRNA by PB was rapid, with increases detected within 3 h of PB injection and steady state levels reached in approximately 20 h. The data suggest that the increase in cytoplasmic P-450 protein levels observed after PB treatment may be totally accounted for by an enhanced rate of synthesis resulting from translation of higher cytoplasmic levels of its specific mRNA. The P-450 mRNA was almost exclusively segregated into the membrane-bound polysome compartment as expected for an mRNA coding for an integral membrane protein of the endoplasmic reticulum.

From Gene to Protein: Information Transfer in Normal and Abnormal Cells, 1979

Publisher Summary This chapter describes the processing and transport of RNA following mitogenic ... more Publisher Summary This chapter describes the processing and transport of RNA following mitogenic stimulation of human lymphocytes. In a study described in the chapter, the kinetics of appearance of labeled RNA in the cytoplasm of human peripheral blood lymphocytes with Concanavalin A (Con A) was studied using an improved method that afforded clean separation of nuclei from cytoplasm. There was no significant increase over resting cell levels in labeled total cell RNA before 10 h exposure to Con A. An increase was detected in the 3 H-uridine-labeled nonpolyadenylated RNA in the cytoplasm following 6 h exposures to Con A. In stimulated cells prelabeled with 3 H-uridine, labeled cytoplasmic nonpolyadenylated RNA level increased after a 3 h exposure to Con A. 3 H-uridine-labeled polyadenylated RNA required at least 13 h before it appeared in cytoplasmic extracts of stimulated cells, and it did not appear in those of resting cells at least for the 8–9 h duration of the experiments. Increased levels of 3 H-adenine-labeled polyadenylated RNA were detectable in cytoplasm after 5 h exposure to mitogen. The lag between addition of isotope and appearance of labeled RNA in cytoplasm was inversely related to the duration of incubation of cells with mitogen.

Proceedings, annual meeting, Electron Microscopy Society of America, 1982

The polarized distribution of surface components between apical and basolateral domains of the pl... more The polarized distribution of surface components between apical and basolateral domains of the plasma membrane constitutes the basis of epithelial function. We are currently studying the mechanisms employed by epithelial cells to segregate different sets of integral proteins in two opposite regions of the plasma membrane. For this purpose, we are utilizing a model system which involves the infection of polarized epithelial cell lines, such as the dog kidney cell line MDCK, with enveloped RNA viruses. Influenza virus and two paramyxoviruses bud from the apical surface regions of MDCK cells, vesicular stomatitis virus (VSV), a rhabdovirus, is assembled instead from the basolateral surface. A main determinant of polarized budding appears to be the addressing of viral envelope glycoproteins to the surface domain that the virus utilizes for budding. The mechanisms and intracellular pathways involved in this sorting are probably the same as those utilized by the cell for its own surface p...

Cell Biology and Immunology of Leukocyte Function, 1979

Publisher Summary This chapter discusses the synthesis and processing of RNA in stimulated fibrob... more Publisher Summary This chapter discusses the synthesis and processing of RNA in stimulated fibroblasts and lymphocytes. Several reviews have discussed the regulation of gene expression at various levels and in a variety of cell types. The primary gene transcripts, in the case of all kinds of RNA, must be converted to mature, functional molecules by modification or processing. Processing includes cleavage of nonconserved sequences, methylation of bases and sugars, and terminal addition of nucleotides to the 3′ OH and the 5′ P ends of the molecules. The chapter presents a comparison of the mechanisms of regulation of RNA metabolism of activated fibroblasts with those of lymphocytes. Extensive gene derepression and synthesis of new transcripts is unlikely to be characteristic of the relatively early events in lymphocytes. A major factor in the train of events following early upon the triggering of lymphocytes is an acceleration in the processing of RNA and in its transport from nucleus to cytoplasm.

Canadian Journal of Biochemistry, 1978

In order for significant DNA synthesis to be observed in the case of human lymphocytes stimulated... more In order for significant DNA synthesis to be observed in the case of human lymphocytes stimulated for 36 h in presence of phytohemagglutinin (PHA), Ca2+ must be present in the medium continuously for at least 20 h; access to Ca2+ for 10-h periods during the first 30 h was not sufficient to permit DNA synthesis to occur. Addition of the chelator EGTA from 0 to 60 h after stimulation caused severe inhibition of incorporation of labelled thymidine when this was measured after 36 to 144 h of culture. Equimolar calcium reversed the inhibition caused by EGTA. Incorporation of labelled uridine and leucine showed a temporal pattern of dependence on the presence of Ca2+ in the medium similar to that of thymidine. Ca2+ appears not to be required in the medium during the last half (i.e.,20–36 h) of the presynthetic G1 phase nor during S phase since removal of Ca2+ from the medium after 20 h did not prevent a subpopulation of lymphocytes from entering S phase 16 h later.

mRNA: The Relation of Structure to Function, 1977

Publisher Summary This chapter discusses that several types of structural and kinetic evidence in... more Publisher Summary This chapter discusses that several types of structural and kinetic evidence indicate that hnRNA contains mRNA precursors in a variety of forms. In some cases the precursors consist of hnRNA niolecules that are considerably larger than the mature mRNA. In other cases the size of the pre-mRNA is similar to that of the mature mRNA, either because processing events have closely followed transcription, or because the transcriptional units are not significantly larger than the mature mRNA. The mRNA segments may be located at the 5' termini of initial transcripts as well as at internal regions that are exposed by appropriate processing cleavages. Nuclear processing of the mRNA precursors is completed by cap formation on the 5' terminus and, for most mRNA species, methylation of internal adenylate residues and polyadenylylation of the 3'-OH terminus. The cellular content of any particular mRNA species and its precursors is determined by the relative rates of transcription, processing and turnover.

Canadian Journal of Biochemistry, 1978

An improved method affording clean separation of nuclei from cytoplasm was utilized to study the ... more An improved method affording clean separation of nuclei from cytoplasm was utilized to study the kinetics of appearance of [3H]uridine-labelled polyadenylated (poly(A)+) and nonpolyadenylated (poly(A)−) RNA in the cytoplasm of human lymphocytes stimulated with the mitogen concanavalin A (Con A). While our previous and present studies utilizing lymphocyte preparations purified on Ficoll-Hypaque gradients had shown that the earliest observable increase in labelled total cell RNA occurred after 12 h of exposure to Con A, we were here able to detect highly significant increases in levels of uridine-labelled poly(A)− RNA in the cytoplasm of lymphocytes which had been exposed to mitogen for 6 h; these cells had been pulsed with [3H]uridine for 2 h prior to harvest. This suggested that the early effects of mitogen may be to increase processing and transport of RNA rather than to increase the production of gene transcripts. At least 13 h of exposure to Con A was required before labelled pol...

Cell, 1984

To study the biogenetic pathway of influenza hemagglutinin (HA), a model apical glycoprotein, in ... more To study the biogenetic pathway of influenza hemagglutinin (HA), a model apical glycoprotein, in polarized epithelial MDCK cells, anti-HA antibodies were added to the basolateral surface during influenza infection. In monolayers grown on collagen gels influenza and VSV plaque development was blocked only when the antibodies were added to the respective budding surface. Addition of anti-HA antibodies to the basal medium of monolayers grown on nitrocellulose filter chambers neither resulted in HA-coupled transport of antibody nor inhibited HA migration to the apical surface. These results indicate that the bulk of HA is vectorially inserted into the apical surface of MDCK cells by polarized exocytosis. Other apical proteins in epithelia may use a similar mechanism during biogenesis .

In Situ PCR and Related Technology, 1995

Current Topics in Membranes and Transport, 1985

Elsevier has created a COVID-19 resource centre with free information in English and Mandarin on ... more Elsevier has created a COVID-19 resource centre with free information in English and Mandarin on the novel coronavirus COVID-19. The COVID-19 resource centre is hosted on Elsevier Connect, the company's public news and information website. Elsevier hereby grants permission to make all its COVID-19-related research that is available on the COVID-19 resource centre-including this research content-immediately available in PubMed Central and other publicly funded repositories, such as the WHO COVID database with rights for unrestricted research re-use and analyses in any form or by any means with acknowledgement of the original source. These permissions are granted for free by Elsevier for as long as the COVID-19 resource centre remains active.

Annals of the New York Academy of Sciences, May 1, 1987

The genus Leishmania includes several species that cause disease in man. These

Biochemistry and Cell Biology, Sep 1, 1989

Leishmania is a trypanosomatid protozoa with a digenetic life cycle. Sandflies inject promastigot... more Leishmania is a trypanosomatid protozoa with a digenetic life cycle. Sandflies inject promastigotes, the free living form present in their salivary glands, into mammals where the parasite colonizes macrophages, transforming into intracellular amastigotes. The cycle is completed when during a blood meal the insect ingests infected macrophages, the amastigotes are released in the gut where they transform back into promastigotes. Leishmania has to adapt to the changing life conditions, from free-living forms in the poikilothermic insect vector to obligatory intracellular parasite in the homeothermic mammalian host. It also has to adapt to the acidic pH of the macrophage's phagolysosome where amastigotes multiply. The adaptative response of Leishmania includes morphological, physiological, and biochemical changes. Promastigotes can be grown in culture medium. Studies of changes taking place during adaptation have been facilitated by the establishment of in vitro conditions that allow the transformation of amastigotes into promastigotes and vice versa. The system is well suited for studying regulation of gene expression during adaptative differentiation. Some mechanisms of mRNA processing are unique to these protozoa: trans-splicing and RNA editing. Several genes that are differentially expressed in the two stages have been studied. No obvious cis regulatory motifs have been found in the DNA.Key words: Leishmania, genes, differentiation, regulation.

Elsevier eBooks, 1977

Publisher Summary This chapter discusses that several types of structural and kinetic evidence in... more Publisher Summary This chapter discusses that several types of structural and kinetic evidence indicate that hnRNA contains mRNA precursors in a variety of forms. In some cases the precursors consist of hnRNA niolecules that are considerably larger than the mature mRNA. In other cases the size of the pre-mRNA is similar to that of the mature mRNA, either because processing events have closely followed transcription, or because the transcriptional units are not significantly larger than the mature mRNA. The mRNA segments may be located at the 5' termini of initial transcripts as well as at internal regions that are exposed by appropriate processing cleavages. Nuclear processing of the mRNA precursors is completed by cap formation on the 5' terminus and, for most mRNA species, methylation of internal adenylate residues and polyadenylylation of the 3'-OH terminus. The cellular content of any particular mRNA species and its precursors is determined by the relative rates of transcription, processing and turnover.

Journal of Cell Biology, 1984

The intracellular pathway followed by the influenza virus hemagglutinin (HA) to the apical surfac... more The intracellular pathway followed by the influenza virus hemagglutinin (HA) to the apical surface of Madin-Darby canine kidney cells was studied by radioimmunoassay, immunofluorescence, and immunoelectron microscopy. To synchronize the migration, we used a temperature-sensitive mutant of influenza WSN, ts61, which, at the nonpermissive temperature, 39.5°C, exhibits a defect in the HA that prevents its exit from the endoplasmic reticulum. Upon transfer to permissive temperature, 32°C, the HA appeared in the Golgi apparatus after 10 min, and on the apical surface after 30-40 min. In the presence of cycloheximide, the expression was not inhibited, indicating that the is defect is reversible; a wave of HA migrated to the cell surface, where it accumulated with a half time of 60 min. After passage through the Golgi apparatus the HA was detected in a population of smooth vesicles, about twice the size of coated vesicles, located in the apical half of the cytoplasm. These HA-containing vesicles did not react with anti-clathrin antibodies. Monensin (10 ' UM) delayed the surface appearance of HA by 2 h, but not the transport to the Golgi apparatus. Incubation at 20°C retarded the migration to the Golgi apparatus by^-30 min and blocked the surface appearance by acting at a late stage in the intracellular pathway, presumably at the level of the post-Golgi vesicles. The initial appearance of HA on the apical surface was in the center ; no preference was observed for the tight-junctional regions .

Current topics in membranes and transport, 1985

Elsevier has created a COVID-19 resource centre with free information in English and Mandarin on ... more Elsevier has created a COVID-19 resource centre with free information in English and Mandarin on the novel coronavirus COVID-19. The COVID-19 resource centre is hosted on Elsevier Connect, the company's public news and information website. Elsevier hereby grants permission to make all its COVID-19-related research that is available on the COVID-19 resource centre-including this research content-immediately available in PubMed Central and other publicly funded repositories, such as the WHO COVID database with rights for unrestricted research re-use and analyses in any form or by any means with acknowledgement of the original source. These permissions are granted for free by Elsevier for as long as the COVID-19 resource centre remains active.

Springer eBooks, 1973

The mechanisms of initiation and termination of the synthesis of protein chains have been the sub... more The mechanisms of initiation and termination of the synthesis of protein chains have been the subject of many studies in recent years, both in mammalian and in bacterial systems (for review, see ref. 1). Those studies emphasize the utilization of ribosomal subunits in the process of chain initiation and polyribosome formation either as derived subunits in the presence of defined initiation factors (2–6) or as native ribosomal subparticles pre-existent as such in the cytoplasm (7–10).

Proceedings ... annual meeting, Electron Microscopy Society of America, Aug 13, 1982

The polarized distribution of surface components between apical and basolateral domains of the pl... more The polarized distribution of surface components between apical and basolateral domains of the plasma membrane constitutes the basis of epithelial function. We are currently studying the mechanisms employed by epithelial cells to segregate different sets of integral proteins in two opposite regions of the plasma membrane. For this purpose, we are utilizing a model system which involves the infection of polarized epithelial cell lines, such as the dog kidney cell line MDCK, with enveloped RNA viruses. Influenza virus and two paramyxoviruses bud from the apical surface regions of MDCK cells, vesicular stomatitis virus (VSV), a rhabdovirus, is assembled instead from the basolateral surface. A main determinant of polarized budding appears to be the addressing of viral envelope glycoproteins to the surface domain that the virus utilizes for budding. The mechanisms and intracellular pathways involved in this sorting are probably the same as those utilized by the cell for its own surface proteins.

Elsevier eBooks, 1979

Publisher Summary This chapter discusses the synthesis and processing of RNA in stimulated fibrob... more Publisher Summary This chapter discusses the synthesis and processing of RNA in stimulated fibroblasts and lymphocytes. Several reviews have discussed the regulation of gene expression at various levels and in a variety of cell types. The primary gene transcripts, in the case of all kinds of RNA, must be converted to mature, functional molecules by modification or processing. Processing includes cleavage of nonconserved sequences, methylation of bases and sugars, and terminal addition of nucleotides to the 3′ OH and the 5′ P ends of the molecules. The chapter presents a comparison of the mechanisms of regulation of RNA metabolism of activated fibroblasts with those of lymphocytes. Extensive gene derepression and synthesis of new transcripts is unlikely to be characteristic of the relatively early events in lymphocytes. A major factor in the train of events following early upon the triggering of lymphocytes is an acceleration in the processing of RNA and in its transport from nucleus to cytoplasm.

Journal of Biological Chemistry, 1981

Treatment of rats with phenobarbital (PB) leads to a substantial increase in the hepatic levels o... more Treatment of rats with phenobarbital (PB) leads to a substantial increase in the hepatic levels of translatable polysomal poly(A) + cytochrome P-450 mRNA. An enriched fraction of P-450 mRNA was obtained by agarose gel electrophoresis and used to prepare a cDNA probe by differential hybridization to total mRNA from control and PB-treated rats. The majority of the sequences within the probe hybridized to recombinant DNA plasmids which contained a bona fide P-450 cDNA insert identified by positive hybridization selection and in vitro translation. The cDNA probe was used to demonstrate that PB treatment leads to a 30-fold increase in polysomal P-450 mRNA, which is not due to more efficient utilization of previously existing mRNA but to the appearance of new messenger in the cytoplasm. The induction of cytoplasmic P-450 mRNA by PB was rapid, with increases detected within 3 h of PB injection and steady state levels reached in approximately 20 h. The data suggest that the increase in cytoplasmic P-450 protein levels observed after PB treatment may be totally accounted for by an enhanced rate of synthesis resulting from translation of higher cytoplasmic levels of its specific mRNA. The P-450 mRNA was almost exclusively segregated into the membrane-bound polysome compartment as expected for an mRNA coding for an integral membrane protein of the endoplasmic reticulum.

From Gene to Protein: Information Transfer in Normal and Abnormal Cells, 1979

Publisher Summary This chapter describes the processing and transport of RNA following mitogenic ... more Publisher Summary This chapter describes the processing and transport of RNA following mitogenic stimulation of human lymphocytes. In a study described in the chapter, the kinetics of appearance of labeled RNA in the cytoplasm of human peripheral blood lymphocytes with Concanavalin A (Con A) was studied using an improved method that afforded clean separation of nuclei from cytoplasm. There was no significant increase over resting cell levels in labeled total cell RNA before 10 h exposure to Con A. An increase was detected in the 3 H-uridine-labeled nonpolyadenylated RNA in the cytoplasm following 6 h exposures to Con A. In stimulated cells prelabeled with 3 H-uridine, labeled cytoplasmic nonpolyadenylated RNA level increased after a 3 h exposure to Con A. 3 H-uridine-labeled polyadenylated RNA required at least 13 h before it appeared in cytoplasmic extracts of stimulated cells, and it did not appear in those of resting cells at least for the 8–9 h duration of the experiments. Increased levels of 3 H-adenine-labeled polyadenylated RNA were detectable in cytoplasm after 5 h exposure to mitogen. The lag between addition of isotope and appearance of labeled RNA in cytoplasm was inversely related to the duration of incubation of cells with mitogen.

Proceedings, annual meeting, Electron Microscopy Society of America, 1982

The polarized distribution of surface components between apical and basolateral domains of the pl... more The polarized distribution of surface components between apical and basolateral domains of the plasma membrane constitutes the basis of epithelial function. We are currently studying the mechanisms employed by epithelial cells to segregate different sets of integral proteins in two opposite regions of the plasma membrane. For this purpose, we are utilizing a model system which involves the infection of polarized epithelial cell lines, such as the dog kidney cell line MDCK, with enveloped RNA viruses. Influenza virus and two paramyxoviruses bud from the apical surface regions of MDCK cells, vesicular stomatitis virus (VSV), a rhabdovirus, is assembled instead from the basolateral surface. A main determinant of polarized budding appears to be the addressing of viral envelope glycoproteins to the surface domain that the virus utilizes for budding. The mechanisms and intracellular pathways involved in this sorting are probably the same as those utilized by the cell for its own surface p...

Cell Biology and Immunology of Leukocyte Function, 1979

Publisher Summary This chapter discusses the synthesis and processing of RNA in stimulated fibrob... more Publisher Summary This chapter discusses the synthesis and processing of RNA in stimulated fibroblasts and lymphocytes. Several reviews have discussed the regulation of gene expression at various levels and in a variety of cell types. The primary gene transcripts, in the case of all kinds of RNA, must be converted to mature, functional molecules by modification or processing. Processing includes cleavage of nonconserved sequences, methylation of bases and sugars, and terminal addition of nucleotides to the 3′ OH and the 5′ P ends of the molecules. The chapter presents a comparison of the mechanisms of regulation of RNA metabolism of activated fibroblasts with those of lymphocytes. Extensive gene derepression and synthesis of new transcripts is unlikely to be characteristic of the relatively early events in lymphocytes. A major factor in the train of events following early upon the triggering of lymphocytes is an acceleration in the processing of RNA and in its transport from nucleus to cytoplasm.

Canadian Journal of Biochemistry, 1978

In order for significant DNA synthesis to be observed in the case of human lymphocytes stimulated... more In order for significant DNA synthesis to be observed in the case of human lymphocytes stimulated for 36 h in presence of phytohemagglutinin (PHA), Ca2+ must be present in the medium continuously for at least 20 h; access to Ca2+ for 10-h periods during the first 30 h was not sufficient to permit DNA synthesis to occur. Addition of the chelator EGTA from 0 to 60 h after stimulation caused severe inhibition of incorporation of labelled thymidine when this was measured after 36 to 144 h of culture. Equimolar calcium reversed the inhibition caused by EGTA. Incorporation of labelled uridine and leucine showed a temporal pattern of dependence on the presence of Ca2+ in the medium similar to that of thymidine. Ca2+ appears not to be required in the medium during the last half (i.e.,20–36 h) of the presynthetic G1 phase nor during S phase since removal of Ca2+ from the medium after 20 h did not prevent a subpopulation of lymphocytes from entering S phase 16 h later.

mRNA: The Relation of Structure to Function, 1977

Publisher Summary This chapter discusses that several types of structural and kinetic evidence in... more Publisher Summary This chapter discusses that several types of structural and kinetic evidence indicate that hnRNA contains mRNA precursors in a variety of forms. In some cases the precursors consist of hnRNA niolecules that are considerably larger than the mature mRNA. In other cases the size of the pre-mRNA is similar to that of the mature mRNA, either because processing events have closely followed transcription, or because the transcriptional units are not significantly larger than the mature mRNA. The mRNA segments may be located at the 5' termini of initial transcripts as well as at internal regions that are exposed by appropriate processing cleavages. Nuclear processing of the mRNA precursors is completed by cap formation on the 5' terminus and, for most mRNA species, methylation of internal adenylate residues and polyadenylylation of the 3'-OH terminus. The cellular content of any particular mRNA species and its precursors is determined by the relative rates of transcription, processing and turnover.

Canadian Journal of Biochemistry, 1978

An improved method affording clean separation of nuclei from cytoplasm was utilized to study the ... more An improved method affording clean separation of nuclei from cytoplasm was utilized to study the kinetics of appearance of [3H]uridine-labelled polyadenylated (poly(A)+) and nonpolyadenylated (poly(A)−) RNA in the cytoplasm of human lymphocytes stimulated with the mitogen concanavalin A (Con A). While our previous and present studies utilizing lymphocyte preparations purified on Ficoll-Hypaque gradients had shown that the earliest observable increase in labelled total cell RNA occurred after 12 h of exposure to Con A, we were here able to detect highly significant increases in levels of uridine-labelled poly(A)− RNA in the cytoplasm of lymphocytes which had been exposed to mitogen for 6 h; these cells had been pulsed with [3H]uridine for 2 h prior to harvest. This suggested that the early effects of mitogen may be to increase processing and transport of RNA rather than to increase the production of gene transcripts. At least 13 h of exposure to Con A was required before labelled pol...

Cell, 1984

To study the biogenetic pathway of influenza hemagglutinin (HA), a model apical glycoprotein, in ... more To study the biogenetic pathway of influenza hemagglutinin (HA), a model apical glycoprotein, in polarized epithelial MDCK cells, anti-HA antibodies were added to the basolateral surface during influenza infection. In monolayers grown on collagen gels influenza and VSV plaque development was blocked only when the antibodies were added to the respective budding surface. Addition of anti-HA antibodies to the basal medium of monolayers grown on nitrocellulose filter chambers neither resulted in HA-coupled transport of antibody nor inhibited HA migration to the apical surface. These results indicate that the bulk of HA is vectorially inserted into the apical surface of MDCK cells by polarized exocytosis. Other apical proteins in epithelia may use a similar mechanism during biogenesis .

In Situ PCR and Related Technology, 1995

Current Topics in Membranes and Transport, 1985

Elsevier has created a COVID-19 resource centre with free information in English and Mandarin on ... more Elsevier has created a COVID-19 resource centre with free information in English and Mandarin on the novel coronavirus COVID-19. The COVID-19 resource centre is hosted on Elsevier Connect, the company's public news and information website. Elsevier hereby grants permission to make all its COVID-19-related research that is available on the COVID-19 resource centre-including this research content-immediately available in PubMed Central and other publicly funded repositories, such as the WHO COVID database with rights for unrestricted research re-use and analyses in any form or by any means with acknowledgement of the original source. These permissions are granted for free by Elsevier for as long as the COVID-19 resource centre remains active.