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J C Courvalin

Publications and source records attributed to J C Courvalin.

51 records · Page 3Linked to original sources
PubMed1988

Expression of nuclear lamins during mouse preimplantation development.

The expression of nuclear lamins during mouse preimplantation development was studied by immunofluorescence, immunoblotting and immunoprecipitation. Two sera were used, specific either for lamin B or lamins A and C. Both sera gave a positive staining of the nuclear periphery throughout preimplantation development (fertilized eggs to late blastocysts). Immunoblots revealed that the three lamins were present in eggs and blastocysts. However, lamin A from eggs was found to have a higher apparent Mr than lamin A from blastocysts and other mouse cells. Using immunoprecipitation, synthesis of lamin A was detected in eggs while synthesis of lamin B was detected in 8-cell embryos and blastocysts, indicating that at least some of the lamins used during early development do not come from a store in the egg. These results are discussed in relation to the possible role of lamins during cell differentiation.

Animals↗
PubMed1988

Antinuclear autoantibodies specific for lamins. Characterization and clinical significance.

In 11 patients, sera displaying a ringlike nuclear immunofluorescent staining on sections of rat liver tissue were shown by Western blotting to contain antibodies to lamins. Sera from 8 patients contained autoantibodies reacting with lamin B, whereas sera from the other 3 patients reacted with lamins A and C. All patients (9 women and 2 men) had a chronic autoimmune disorder, which rarely fulfilled the usual criteria for a diagnosis of systemic lupus erythematosus. The disorder was characterized by acute or chronic (active or granulomatous) hepatitis; steroid-responsive blood cytopenia, often associated with a circulating anticoagulant, or anticardiolipin antibodies, or both; and cutaneous leukocytoclastic angiitis or probable brain vasculitis. Eight patients had at least two of these three conditions. Antilamin autoantibodies may thus be a marker for an unusual subset of autoimmune diseases.

Adult↗
PubMed1988

Autoantibodies to 200 kD polypeptide(s) of the nuclear envelope: a new serologic marker of primary biliary cirrhosis.

Fifty sera which gave ring-like nuclear staining in immunofluorescence on rat liver tissue sections were characterized. Using immunoprecipitation and Western blotting we showed that 10 of the sera contained antibodies to 200 kD polypeptide(s) of nuclear envelope. Clinical and biological data were available for nine of the patients. Strikingly, all of these patients suffered from primary biliary cirrhosis with eight of them having anti-mitochondrial antibodies. As no control serum displayed such a reactivity, anti-200 kD polypeptide(s) antibodies can be considered as a new marker specific of a subset of primary biliary cirrhosis, being present even when anti-mitochondrial antibodies are absent. The exact identity of the target remains to be established, since several polypeptides of similar molecular weight have been reported to belong to the nuclear envelope.

Adult↗
PubMed1987

A human T lymphoblastic cell line lacks lamins A and C.

Lamins A, B and C, the three major proteins of nuclear envelope, constitute a class of intermediate filament polypeptides. We have compared the amount of these polypeptides in two human cell lines, epithelial HeLa cells and T lymphoblasts KE 37. It was found that the three lamins were present in roughly equimolar stoichiometry in HeLa cells, while lamin B was the unique lamin component in T lymphoblasts. Moreover, 3-kb mRNA of lamin A and 2.1-kb mRNA of lamin C were detected with a human cDNA probe in HeLa cells but not in T lymphoblasts. These results suggest that (i) lamin B can build up the lamina structure in actively dividing somatic cells by itself, and (ii) lamin expression in lymphoid cells may be subject to important quantitative variations. Comparison of the lamin composition of human cloned T lymphocytes and Epstein-Barr virus-transformed human B lymphocytes confirmed this statement. The lamin B level was nearly equivalent in both cells but the content of lamins A and C varied to a large extent, being low in T cells and high in B cells.

Cell Line↗
PubMed1987

Centrosomal proteins and lactate dehydrogenase possess a common epitope in human cell lines.

A spontaneously arising rabbit anti-centrosome serum with strong human specificity, used to identify specific antigens in isolated centrosomes, was shown to react with several noncentrosomal proteins including a 36-kDa protein that appeared to be the major cellular antigen. To explore the immunological relationship between noncentrosomal and centrosomal antigens, immunoglobulins were affinity purified using the individual noncentrosomal antigens (from lymphoblastoma KE37 cells) and were tested for their capacity to bind to human centrosomes in situ and to proteins from isolated centrosomes. In this way, the 36-kDa antigen, an abundant cytosolic protein, was shown to share at least one antigenic determinant with high molecular weight centrosomal proteins. This antigen was further identified by mild proteolysis as the glycolytic enzyme lactate dehydrogenase. In all the analyzed human cell lines, the centrosomal staining in situ was correlated with a strong labeling of purified lactate dehydrogenase in immunoblots. Conversely, the absence of centrosomal staining in rodent cells was always correlated with the absence of lactate dehydrogenase labeling. These data suggest an evolutionary relationship between centrosomal proteins and this "housekeeping" enzyme.

Animals↗
PubMed1987

Autoantibodies to nuclear lamin B in a patient with thrombopenia.

We report the characterization of novel nucleus specific autoantibodies in the serum of a patient with systemic lupus erythematosus. Immunofluorescent staining of cycling cells and absorption experiments localized the antigen to the nuclear envelope. Two-dimensional gel electrophoretic analysis of immunoprecipitated nuclear proteins show the antigen to be an acidic polypeptide (IP approximately 5.4) of 68 kDa molecular mass. It has been identified as lamin B, one of the three major nuclear envelope polypeptides of mammalian cells. Antibodies shown to be polyclonal immunoglobulin Gs, were directed against determinant(s) of the protein that have apparently been conserved during evolution. They do not appear to be related to other autoantibodies present in the serum (anti-DNA and anti-platelet). The nuclear specificity shown by these antibodies further demonstrates the antigenicity of proteins related to intermediate filament proteins in patients with autoimmune disorders.

Adult↗
PubMed1986

A protein of Mr 80,000 is associated with the nucleolus organizer of human cell lines.

A rabbit serum which had previously been reported to have an immunological affinity for centrosomes of human cell lines was shown also to be specific for the nucleus. Optical and ultrastructural immunolocalization in HeLa cells showed that this specificity is restricted to the fibrillar centre of nucleoli either in untreated or actinomycin D treated interphase cells. In mitotic cells discrete labelling was observed on chromosomes and shown to correspond, on spread metaphase plates, to the short arms of acrocentric chromosomes, i.e. to the nucleolar organizer regions (NORs). Using independent cell fractionation procedures in the human T-lymphoblastic KE 37 cell line and purification of immunoglobulins by affinity to antigens detected by electrophoresis and blotting, a strict correlation between immunoreactive proteins and cytological staining was established. The nucleolar specificity was shown to correspond to a protein with an Mr of 80,000 while the centrosomal specificity corresponded principally to a protein doublet of 60,000-65,000. These antigens share common epitopes as shown by the staining of both NOR and centrosome by immunoglobulins purified by affinity to either type of protein.

Animals↗
PubMed1982

Solubilization of nuclear structures by the polyanion heparin.

Rat liver nuclei were treated with different concentrations of the polyanion heparin, at low ionic strength and in the absence of divalent cations. Heparin at the optimal concentration (Heparin:DNA greater than or equal to 1) completely solubilized the intranuclear structures, allowing the preparation of pure nuclear membranes. The majority of the nuclear proteins, including all of H1 and 70% of the core histone oligomers, were solubilized. By contrast, suboptimal concentrations of heparin (heparin:DNA less than 0.3) resulted in the selective solubilization of non-histone proteins and the partial solubilization of chromatin without dissociation of histones from DNA. Among the RNA- and DNA-associated non-histone proteins revealed by the heparin fractionation procedure, some are noteworthy. A DNA-associated protein of 40,000 daltons was found to be associated with core histone oligomer. Three proteins of Mr = 62,000, 70,000, and 74,000, identical with that of the lamina proteins, were distinguished by their presence in all the fractions obtained, their predominance in the less soluble fractions, and their tight association with DNA and RNA. More than the solubilization of the histones, their extraction seems to be a prerequisite for chromatin solubilization by heparin.

Animals↗
PubMed1978

Isolation of nuclear envelopes with polyanions.

Optimal conditions for the isolation of nuclear envelopes by the action of heparin on nuclei are established and a morphological and biochemical study of such isolated envelopes is presented. An almost 100% yield of pure nuclear envelopes can be obtained by a single sedimentation step after incubation of nuclei with heparin for 40 min at 4 degrees C. The nuclear membrane pellet obtained in this way contains whole envelopes with a preserved perinuclear space and with ribosomes present on the outher leaflet. A single band with an apparent buoyant density of 1.18 is obtained by sucrose density gradient analysis. The chemical composition of the pellet is similar to that of the purified membranes and corresponds to 62% proteins, 34% phospholipids, 3% RNA, and 0.5% DNA. The presence of low concentrations of sodium phosphate (2-10 mM) is critical for a complete solubilization of the chromatin. A less rapid and complete solubilization is obtained with the potassium salt. Low concentrations of Mg++ (1-3 mM) counteract chromatin solubilization by heparin mainly at the level of chromatin-nuclear membrane association. The presence of EDTA in the medium leads to isolated nuclear envelopes on which neither ribosomes nor nuclear pores are visible, indicating the pore structure is dependent on the presence of Ca++ or Mg++. A comparison with other polyanions indicates a decisive advantage of heparin. However, pure nuclear envelopes can also be obtained by the action of dextran sulfate (mol wt 500,000) on nuclei incubated for 5 min at 37 degrees C, in the presence of phosphate ions.

Animals↗
PubMed1976

Effect of estradiol on rat uterus DNA-dependent RNA polymerases. Studies on solubilized enzymes.

DNA-dependent RNA polymerases of immature and castrated rat uteri were studied after estradiol administration. The enzymes were solubilized from either whole uterus homogenate or nuclei and their activities were measured on an exogenous DNA template. alpha-Amanitin was used to distinguish alpha-amanitin-resistant from alpha-amanitin-senitive forms of the enzyme. The number of alpha-amanitin-sensitive RNA polymerase molecules was measured by a binding assay using labeled amanitin. In the first series of experiments RNA polymerases were solubilized from whole uterus homogenate. alpha-Amanitin-sensitive and -resistant activities were constant during the first 6 hours after estradiol treatment, followed by a late and moderate increase in their activities (50% at 12 hours for the resistant form and 40% at 24 hours for the sensitive form). The number of sensitive polymerase molecules evolved in an identical manner to its activity (+40% at 24 hours), suggesting that the increase in activity is due to the synthesis of new enzyme molecules. For both forms, no diffusible stimulatory factor was detected in the uterus of hormone-treated animals. In the second series of experiments, disrupted nuclei were washed with 0.15 M (NH4)2SO4 in order to release only enzyme molecules which were not firmly bound to DNA in a transcription complex. The amount of the sensitive form of polymerase which remains firmly bound to chromatin, was constant for 6 hours after estradiol administration and was doubled by 24 hours. The firmly bound alpha-amanitin-resistant activity was solubilized and was measured in the presence of an exogenous template. There was a progressive increase in activity first detectable in 1 to 2 hours, amounting to 50% at 6 hours and 100% at 24 hours. The reported results show that during the first 6 hours of hormone treatment: (a) the total content of RNA polymerases remains unchanged in the uterus; (b) the number of alpha-amanitin resistant molecules tightly bound to DNA increases progressively while the alpha-amanitin sensitive remains constant. At a later time (24 hours), an increase is observed both for the total amount of enzymes and for their fraction engaged in a transcription complex.

Amanitins↗