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E Jost

Publications and source records attributed to E Jost.

At least 37 records · Page 2Linked to original sources
PubMed1991

Identification of phosphorylation sites on murine nuclear lamin C by RP-HPLC and microsequencing.

Isolated interphase lamin C, obtained from Ehrlich ascites tumor cells, was digested by Lys-C endoproteinase, the resulting peptides separated by reversed-phase HPLC and subjected to microsequencing in order to identify phosphorylation sites in interphase and following phosphorylation in vitro by cdc2-kinase, protein kinase C (PKC) and protein kinase A (PKA), respectively. Nuclear lamin C showed partial phosphorylation of Ser392 and Ser409, and possibly Ser407 in interphase. Phosphorylation was increased in response to cdc2-kinase at Ser390 and Ser392 and to PKC at Ser572. The N-terminal peptide (aa 1-32) containing consensus sequences for the 3 kinases was phosphorylated by cdc2-kinase, PKC and PKA. The sequence data suggests that multiple molecular switches via lamina modification control the dynamic behaviour of the nucleoskeleton during the cell cycle.

Amino Acid Sequence↗
PubMed1989

Alteration of nuclear lamina protein in human fibroblasts infected with cytomegalovirus (HCMV).

Immunoblotting with monoclonal as well as polyclonal lamin antibodies revealed that nuclear lamina proteins of human foreskin fibroblasts (HFF) were differentially affected after infection with human cytomegalovirus (HCMV). Lamin A immunoreactivity was progressively lost during the course of the infectious cycle whereas that of lamin C was comparatively stable. This process was not observed in herpes simplex virus-infected HFF. On the other hand, noninfected arrested HFF stimulated by serum to enter S-phase also exhibited loss of lamin A immunoreactivity. Selective in vivo proteolysis of lamin A is suggested as the possible underlying mechanism.

Cells, Cultured↗
PubMed1988

Lamin disassembly kinetics: a cell-free system with extracts from mitotic HeLa cells.

We describe a cell-free system in which a postribosomal supernatant from metaphase HeLa cells induces prophase-like changes in permeabilized HeLa cell populations as evidenced by the nuclear lamin disassembly and chromatin condensation. We have attempted to characterize the cell-free system with permeabilized HeLa cells. First, by extracting lamins with agents known to disrupt the noncovalent interactions in the supramolecular lamin aggregate in interphase using polyclonal and a newly established monoclonal anti-lamin Ab 2E3, uniform extraction of lamins was achieved with urea and deoxycholate whereas the cation Mg2+ and 2-mercaptoethanol had little effect on the disassembly of interphase lamins. Second, cytoplasmic extract from mitotic HeLa cells, synchronized by a nitrous oxide metaphase arrest, was tested. It had a differential effect on interphase lamin depolymerization. Nuclei in G1 phase of the cell cycle were more resistant against the mitotic extracts than cells in S and G2 phase. The results are discussed in terms of a possible inactivation of mitotic extracts by factors present in nuclei in early interphase.

Adenosine Triphosphate↗
PubMed1986

Redistribution of nuclear lamins in mitotic cells.

The nuclear lamins are directed from the cytoplasm to chromosomes as part of the maturation pathway of the interphase nucleoskeleton. In mitosis, the three polypeptides lamin A, B and C were found in the cytoplasm from prophase until anaphase and shifted to chromosomal surfaces at telophase (Ely, D'Arcy and Jost, 1978; Gerace, Blum and Blobel, 1978). We show here that early events in nucleoskeleton formation could be regulated by extracellular pH. When exponentially growing tissue culture cells and cells arrested in mitosis were exposed to different extracellular pH values, three patterns of distribution of lamins were observed in mitotic cells: exclusively cytoplasmic distribution of mitotic lamins at low pH (6.8 to 7.3); a premature association of a lamin subfraction with metaphase chromosomes at intermediate pH 7.5; a more prominent relocation of lamins onto chromosomes in metaphase and in disorganized metaphase at pH 8.0. Reassembly of lamins occurred at telomeric ends of mitotic chromosomes followed by a lateral fusion to form a nuclear cage. Using immunogold localization, we show that pH-induced, premature, partial deposition of lamins onto condensed chromosomes may occur prior to the formation of the bilamellar nuclear envelope. These results suggest that the pH-induced redistribution of lamins acts to trigger early events of mitosis to interphase transition.

Cell Cycle↗
PubMed1986

Nuclear lamins during prophasing and telophasing in heterophasic HeLa and Chinese hamster homokaryons.

We have perturbed the dynamics of the nuclear lamins by means of cell fusion between mitotic and interphase cells and have studied redistribution of lamins in fused cells as a function of extracellular pH levels. We show here that in heterophasic M-1 HeLa homokaryons disassembly of interphase lamins predominates at low pH levels between 7.0 to 7.3, whereas deposition of cytoplasmic lamins around condensed metaphase chromosomes was observed at pH 8.0. In HeLa homokaryons lamina disassembly and lamina deposition around chromosomes are mutually exclusive. Using heterophasic M-1 homokaryons of the Chinese hamster cell line DON we observed that disassembly of interphase lamins and deposition of lamins around condensed chromosomes coexisted in the same homokaryon kept at pH 7.0. Disassembly of lamins developed synchronously with premature chromosome condensation (PCC) whereas lamina deposition around the condensed M-chromosomes was followed by telophasing. In fusions kept at pH 8.0 cytoplasmic lamins were exclusively deposited around mitotic chromosomes. The results are interpreted as showing that pH regulates the lamina dynamics in homokaryons of mitotic and interphase cells.

Animals↗
PubMed1984

Patterns of nuclear lamins in diverse animal and plant cells and in germ cells as revealed by immunofluorescence microscopy with polyclonal and monoclonal antibodies.

Polyclonal and monoclonal antibodies against rat liver nuclear lamins have been used to evaluate the immunological cross-reactivity of lamins with a given antibody in a variety of animal and plant cells. The results indicated that lamins of all vertebrate cells but not invertebrate cells share at least one antigenic determinant, resulting in immunological cross-reaction with polyclonal antisera to lamina from rat liver. The range of cross-reaction with monoclonal antibody to rat lamins includes all mammalian cells tested but we observed no reaction with other vertebrate and invertebrate cells. Thus, by means of immunological cross-reaction a less stringently conserved pattern is observed for lamins than, for example, cytoskeletal proteins. We have also investigated the fate of the nuclear lamins during meiosis in testes and ovaries of the mouse. Lamins are absent from male meiotic cells and during oogenesis in meiotic prophases.

Animals↗
PubMed1983

New patterns of nuclear lamina induced by cell fusion.

During the eukaryote cell cycle the nuclear envelope displays a series of major morphogenetic changes, the most significant of which include its breakdown and reconstitution as cells move up to, pass through and emerge from division. The three polypeptides, lamins A, B and C, are major components of the nuclear pore complex-lamina fraction of the nuclear envelope and their association with the nuclear membrane or their dispersal in the cytoplasm reflects the existing balance between polymerization and depolymerization in the envelope. We have perturbed the lamina polymerization cycle by means of cell fusion between mitotic and interphase cells, following the redistribution of nuclear lamina protein by means of immunofluorescence techniques. In these heterophasic heterokaryons changes in the distribution of lamina occur as a function of (1) the time elapsed after fusion; (2) the ratio of mitotic to interphase elements in the cell, and (3) the stage in the cell cycle occupied by the interphase partner at the time of fusion. Depolymerization of nuclear lamina occurs most rapidly in cells with high ratios of mitotic to interphase elements, and especially in G1 rather than S-phase nuclei. While lamina depolymerization predominates early after fusion, at later times lamina is deposited around both the original metaphase and interphase nuclear masses and this is associated with the resumption of interphase activity in the form of limited DNA synthesis. These observations lead us to conclude that lamina depolymerization is under positive control mediated by diffusible factors in the cytoplasm of the metaphase partner. Repolymerization is likely to be associated with the inactivation of these factors as the heterokaryons age and, as a result, pass into an interphase-like state.

Basement Membrane↗
PubMed1979

Transfer of mouse nuclear envelope specific proteins to nuclei of chick erythrocytes during reactivation in heterokaryons with mouse A9 cells.

When chick erythrocyte nuclei are introduced into the cytoplasm of mouse A9 cells by cell fusion, proteins present in a fraction of the mouse nuclear envelope begin to appear in the envelope of the chick erythrocyte. The protein uptake was examined using antisera raised in chickens against the 3 major polypeptides of the nuclear pore complex-fibrous lamina fraction from rat liver nuclei. In indirect immunofluorescence studies these antisera give a strong envelope-specific staining with various mammalian but not chicken cells. Eighteen hours after cell fusion the first murine antigens can be observed in the erythrocyte nucleus. Two days after cell fusion the vast majority of the erythrocyte nuclei in cell hybrids contain some antigen and by 3 days the fluorescence of the reactivated erythrocyte nuclei reaches a level comparable to that of the mouse A9 nuclei. The rate of appearance of fluorescence in the chick nuclei depends upon the ratio of A9 cytoplasm to chick nuclei. Antigen uptake by the erythrocyte envelope is inhibited when protein synthesis is blocked suggesting that synthesis of mouse antigen, rather than a redistribution, determines the velocity or erythrocyte envelope reactivation. The early uptake of nucleospecific protein into the reactivating chick erythrocyte may not require any alteration in the nuclear envelope.

Animals↗
PubMed1978

The dissociation of nuclear proteins from superhelical DNA.

Structures retaining many of the morphological features of nuclei may be released by gently lysing human cells in solutions containing non-ionic detergents and high concentrations of salt. These nucleoids contain superhelical DNA. Using a double-labelling procedure we have compared, at different salt concentrations, the amounts and types of protein associated with human nucleoides containings superhelical or relaxed DNA. We find that the slightly lysine-rich histones (H2A and H2B) but not the arginine-rich histones (H3 and H4) dissociate more slowly from nucleoids containing superhelical DNA than from those containing relaxed DNA. A protein of apparent molecular weight of 22000 also binds more tightly to superhelical DNA. We conclude that this protein and the slightly lysine-rich histones transmute free energy of supercoiling into binding energy when they bind to superhelical DNA.

Arginine↗
PubMed1978

Localization of a nuclear envelope-associated protein by indirect immunofluorescence microscopy using antibodies against a major polypeptide from rat liver fractions enriched in nuclear envelope-associated material.

The location of a specific major polypeptide present in nuclear pore complex-enriched fractions from rat liver was examined by indirect immunofluorescence microscopy using chicken antibodies against this polypeptide. In both whole cell preparations of cultured cells grown on cover slips (mouse 3 T 3, rat kangaroo PtK2) and in frozen sections through liver and mammary gland tissue a strongly preferential, if not exclusive, binding to the nuclear periphery of interphase cells was observed. The specificity of this localization was demonstrated in these cells by the decoration of chromatin with antibodies against histones and of elements of the endoplasmic reticulum--outer mitochondrial membrane--system with antibodies to cytochrome b5. In addition, the localization was examined by electron microscopy using frozen sections and "immunoperoxidase" techniques. The results suggest that this polypeptide is contained in a protein specific for the nuclear periphery, probably closely associated with the peripheral chromatin.

Animals↗
PubMed1976

Characterization of nuclear structures containing superhelical DNA.

Structures resembling nuclei but depleted of protein may be released by gently lysing cells in solutions containing non-ionic detergents and high concentrations of salt. These nucleoids sediment in gradients containing intercalating agents in a manner characteristic of DNA that is intact, supercoiled and circular. The concentration of salt present during isolation of human nucleoids affects their protein content. When made in I-95 M NaCl they lack histones and most of the proteins characteristic of chromatin; in 1-0 M NaCl they contain variable amounts of histones. The effects of various treatments on nucleoid integrity were investigated.

Acridines↗
PubMed1975

Affinity chromatography of DNA-binding proteins from human, murine and man-mouse hybrid cell lines.

About 60 proteins from human and murine cell lines were isolated by their ability to bind to different preparations of DNA. In the intact cell, the majority of these proteins are to be found in the cell nucleus. The electrophoretic mobilities of the DNA-binding proteins from human, murine and man-mouse hybrid cell lines were compared in two-dimensional acrylamide gels. Few, if any, species-specific differences were found. These observations suggest that the structures of the vast majority of the proteins that interact with DNA are conserved through evolution. A molecular basis is thus provided for the intracellular of hybrid cells derived from different animal species.

Animals↗