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C M Wolff

Publications and source records attributed to C M Wolff.

13 recordsLinked to original sources
PubMed2000

Heterotopic expression of the Xl-Fli transcription factor during Xenopus embryogenesis: modification of cell adhesion and engagement in the apoptotic pathway.

In the Xenopus laevis embryo, the overexpression of the Xl-FLI protein, a transcription factor of the ETS family, provokes severe developmental anomalies, which affect anteroposterior and dorsoventral polarities, optic cup formation, head cartilage morphogenesis, and erythrocyte differentiation. It has been proposed that these effects could be correlated to modifications of cell adhesion properties and/or to an increased engagement of cells in the apoptotic pathway during early development (Remy et al., Int. J. Dev. Biol. 40, 577-589, 1996). To address these questions, we have first analyzed the behavior of cells overexpressing the protein in both aggregation and adhesion assays. We observe perturbations of cell-cell interactions as well as perturbations of cell adhesion and spreading on fibronectin and extracellular matrix (ECM). Second, we have analyzed apoptosis of cells overexpressing the Xl-FLI protein, by testing DNA fragmentation, caspase-3 activity and by performing TUNEL assay. We show that Xl-Fli overexpression results in the appearance of hallmarks of apoptosis, including exclusion of cells from the interior of the embryo, internucleosomal fragmentation of DNA and dose-dependent induction of caspase-3, resulting in the hydrolysis of poly(ADP-ribose) polymerase. In addition, a dominant-negative mutation of BMPs receptors decreases the effects of Xl-Fli overexpression, suggesting that a modification of the BMP signalling could be responsible for increased apoptosis. The latter appears to affect predominantly ventral and ventrolateral regions of the embryo.

Animals↗
PubMed1999

Protection of immunoreactivity of dry immobilized proteins on microtitration plates in ELISA: application for detection of autoantibodies in myasthenia gravis.

We show the ability of the BSA-trehalose film to convert normally fragile proteins such as mouse monoclonal antibody to the Alzheimer precursor protein A4 (APP695) and cell line TE671 acetylcholine receptor (AChRTE671) into a stable reagent, after its immobilization on microtitration plates. The remarkable property of the dry immobilized proteins are their stability under prolonged exposure to temperatures as high as 50 degrees C. Using the AChRTE671, the proposed method was applied for the measurement of anti-AChR autoantibodies in Myasthenia gravis by means of an enzyme-linked immunosorbent assay (ELISA). The test was shown to be specific and able to detect anti-AChR autoantibodies at concentrations as low as 3 nM. Using the same AchRTE671 as antigen, the results of examination of 34 serum samples for detection of anti-AChR autoantibodies by ELISA were compared with those of the conventional radioimmunoprecipitation assay (RIA). It was concluded that ELISA is another useful method for the diagnosis of M. gravis. The ELISA method offers a rapid, simple, safe and inexpensive means for mass screening of M. gravis.

Autoantibodies↗
PubMed1998

The avian fli gene is specifically expressed during embryogenesis in a subset of neural crest cells giving rise to mesenchyme.

The ets-family of transcription factors is involved in the development of endothelial and hematopoietic cells. Among these genes, fliwas shown to be responsible for erythroblastomas and Ewing's sarcomas. Its involvement in Ewing's sarcoma, a putative neurectodermal tumor, as well as the in situ hybridization studies performed in mice and Xenopus suggested a role in neural crest development. We cloned quail fli cDNA in order to analyze in more detail its expression in neural crest cells, which have been extensively studied in avian species. Fli gene maps on chicken chromosome 1 to band q31->q33. Two RNAs are transcribed, most likely arising from two different promoters. The analysis of its expression in neural crest cells reveals that it is expressed rather late, when the neural crest cells reach their target. Among the various lineages derived from the crest, it is restricted to the mesenchymal one. It is maintained at later stages in the cartilage of neural crest but also of mesodermal origin. In addition, fli is expressed in several mesoderm-derived cells: endothelial cells as well as intermediate and splanchnopleural mesoderm.

Amino Acid Sequence↗
PubMed1993

The c-ets-1 proto-oncogenes in Xenopus laevis: expression during oogenesis and embryogenesis.

We previously reported the cloning and sequencing of two cDNAs derived from the Xenopus laevis ets-1 gene (Stiegler et al., 1990). The Xl-ets-1a cDNA encodes a polypeptide highly homologous to known ets-1 proteins. The 3'-UTR contains two AATAAA polyadenylation signals together with three copies of the TTTTTAT sequence thought to confer a maturation-specific polyadenylation and implicated in the deadenylation of dormant mRNAs. Several transcripts with maternal characteristics were detected in oogenesis and early embryogenesis. A marked augmentation of the major transcript in the poly(A)+ fraction was detected at fertilization. Ets-1 transcripts were observed at constant levels during the cleavage stages but decreased abruptly at gastrulation, to reappear from neurulation to late embryogenesis. The possible contribution of 3'-UTR sequence elements to this behavior is discussed.

Adenosine↗
PubMed1993

Xl-fli, the Xenopus homologue of the fli-1 gene, is expressed during embryogenesis in a restricted pattern evocative of neural crest cell distribution.

The Xenopus laevis fli cDNA, belonging to the ets family of transcription factors, was isolated from a library prepared from unfertilized eggs. It encodes a polypeptide with extensive homology to murine and human Fli proteins. The long 3'-untranslated region contains five nuclear polyadenylation signals and three cytoplasmic polyadenylation elements, as well as many A/T rich elements. Two polyadenylated transcripts appear at the early neurula and accumulate up to the tadpole stage. In situ hybridization reveals an expression in territories invaded by neural crest cells. In the head region, fli is expressed in the peri-ocular zone, in the branchial buds and at the level of the brain floor. In the trunk, a metamerized expression is detected in the dorsum. At a lower level, the tailbud and the peri-cardiac region also appear positive.

Animals↗
PubMed1991

Immobilized enzyme electrode for creatinine determination in serum.

An immobilized enzyme electrode for continuous creatinine determination in blood serum is described. The enzymes creatinine amidohydrolase, creatine amidinohydrolase, and sarcosine oxidase are coimmobilized to the surface of the polypropylene membrane of a Clark-type electrode responsive to oxygen. The immobilized enzymes catalyze the decomposition of creatinine with the consumption of oxygen and thus permit the creatinine measurement. The whole assay takes less than 1 min. Effects of pH and temperature on electrode response are also described. The proposed technique offers a rapid, simple, and inexpensive means to determine creatinine in blood serum within the normal and abnormal ranges. The repeatability of the creatine determination in serum is 2.5% (relative standard deviation), and the detection limit is 3 x 10(-6) mol L-1. The results obtained by this method were compared to those obtained with the Technicon AutoAnalyzer SMAC system based on the Jaffé reaction; the correlation factor between the two methods was found to be r = 0.9997.

Amidohydrolases↗
PubMed1991

Cloning, sequencing, and expression of two Xenopus laevis c-ets-2 protooncogenes.

In a general approach to identify genes important in the control of genetic expression during development of Xenopus laevis, two complementary DNAs corresponding to two different c-ets-2 genes were cloned and sequenced. One of these complementary DNAs appears to be almost full length. The two variant genes differ in their overlapping sequences by 87 nucleotide substitutions, leading to 17 amino acid modifications in the proteins, 8 of them being conservative. All but one of these changes map outside of the 142 COOH-terminal residues, a region critical for nuclear localization and DNA binding in the ets proteins. Features potentially important for the biological activity of the gene products are conserved. Two transcripts (3.2 and 1.7 kilobases) with maternal characteristics are detected at a constant level from stages II/III of oogenesis to stage 10 of embryogenesis. They later decline to hardly detectable levels at stages 30-40. Variable amounts of the same transcripts are observed in many adult tissues. All of these characteristics support the idea that the ets-2 gene products play an important role during embryogenesis, as well as in adult life. Indeed, they act as ubiquitous transcriptional activators, as recently demonstrated by several investigators (C. V. Gunther, J. A. Nye, R. S. Bryner, and B. J. Graves, Genes & Dev., 4: 667-679, 1990; R. Bosselut, J.F. Duvall, A. Gegonne, M. Bailly, A. Hemar, J. Brady, and J. Ghysdael, EMBO J., 9: 3137-3144, 1990; R. Wasylyk, C. Wasylyk, P. Florès, A. Bègue, D. Leprince, and D. Stéhelin, Nature (Lond.), 346: 191-193, 1990).

Amino Acid Sequence↗
PubMed1990

Repair of acetyl-aminofluorene modified pBR322 DNA in Xenopus laevis oocytes and eggs; effect of diadenosine tetraphosphate.

Using Xenopus laevis oocytes and unfertilized eggs, we have developed a system which allows the study of DNA repair upon microinjection of pBR 322 DNA which has been previously modified in vitro by N-acetyl-aminofluorene, under controlled conditions. In unfertilized eggs, an efficient repair of pBR-18AAF DNA takes place, leading to a restoration of the transforming activity of the plasmid DNA towards Escherichia coli. The repaired DNA is even efficiently replicated, the egg being "activated" by the microinjection. In the oocyte, a partial repair is observed as shown by the incorporation of labelled dCTP in the modified plasmid DNA, even in the presence of aphidicolin, an inhibitor of DNA polymerase alpha. However, the repair appears to be very limited, since it does not restore the transforming activity of the modified plasmid DNA. This inefficient repair in the oocyte may be due to the rapid packaging of foreign DNA into a minichromosome and/or to a very low level of DNA polymerase beta. This system was used to study the effect of diadenosine tetraphosphate (Ap4A) on DNA repair. Ap4A seems not to interfere with repair processes in the oocyte, but significantly inhibits the replication following the repair of AAF-modified plasmid DNA in unfertilized eggs. These results suggest that Ap4A could be involved in switching off the replication machinery when DNA is badly damaged, thus helping to avoid the perpetuation of DNA modifications in the daughter cells. This hypothesis is consistent with many previous reports on the accumulation of dinucleoside polyphosphates under stress conditions, which are known to result in modification of DNA.

2-Acetylaminofluorene↗
PubMed1990

Significance of dinucleoside tetraphosphate production by cultured tumor cells exposed to the presence of ethanol.

The use of 30 to 50% ethanol solutions to extract the nucleotides from HTC and A-459 cells results in dinucleoside tetraphosphate (Ap4X) levels 3-30-fold as high as those obtained by 5% classical trichloracetic acid extraction, while ATP levels are identical in both cases. The amplification factor varies with the percentage of ethanol and duration of contact between the cells and the extraction mixture. It remains constant for the HTC cells during cell growth, but exhibits a maximum for the A-459 cells towards the end of the exponential growth period. The incorporation of radioactivity in Ap4X when [alpha-32P]ATP is added to the extraction mixture suggests an Ap4X neosynthesis in the presence of ethanol. The results carried out in the presence of pyrophosphate, EDTA and zinc acetate strongly suggest that aminoacyl-tRNA synthetases could be responsible for the increase in Ap4A content with ethanol treatment. Nevertheless, the effect of ethanol is probably not the result of an activation of these enzymes, but rather, as already suggested by earlier results in our laboratory, the result of a fast inactivation of the degradation enzymes.

Adenosine Triphosphate↗
PubMed1986

Factor V inhibitor associated with immune complex formation.

A 72-year-old man was noted, shortly after surgery, to have a bleeding diathesis secondary to the development of a high-titer anti-factor V inhibitor. We documented the presence of factor V:anti-factor V IgG immune complexes and their disappearance following a short course of steroid therapy.

Abdomen↗
PubMed1984

Kinetic approach for the enzymic determination of creatinine.

By exploiting the kinetics of NADH consumption in the enzyme system creatinine amidohydrolase + creatine kinase + pyruvate kinase + lactate dehydrogenase, one can determine creatinine in serum in the range of 1 to 90 mg/L. By using the conditions defined here, this determination can be made with a single measurement of the rate of disappearance of NADH. The analytical rate measurement is made at a fixed time (2 min) after the sample is introduced into the enzyme solution. For this fixed time, the interferences of creatine and pyruvate are eliminated. Results so obtained for creatinine in human blood serum samples correlated well (r = 0.98) with those obtained by the classical Jaffé-reaction method. Run-to-run reproducibility (CV) was 3%, and the limit of detection was 1 mg/L.

Amidohydrolases↗