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M Veron

Publications and source records attributed to M Veron.

At least 19 recordsLinked to original sources
PubMed2002

X-ray diffraction from rectangular slits.

It is shown that for micrometre-sized beams the X-ray diffraction from slits is a source of strong parasitic background, even for slits of high quality. In order to illustrate this effect, the coherent diffraction from rectangular slits has been studied in detail. A large number of interference fringes with strong visibility have been observed using a single set of slits made of polished cylinders. For very small apertures, asymmetrical slits generate asymmetrical patterns. This pattern is calculated from the theory of electromagnetic field propagation and compared with experiment in the far-field regime. The use of guard slits to remove Fraunhofer diffraction from the beam-defining slits is treated theoretically. Numerical simulations yield the optimum aperture of the guard slits with respect to the distance to the primary slits. Diffraction theory is shown to be essential to understand how to reduce the background-to-signal ratio in high-resolution experiments.

Journal Article↗
PubMed2001

NDP kinase reactivity towards 3TC nucleotides.

Nucleoside diphosphate (NDP) kinase is usually considered as the enzyme responsible for the last step of the cellular phosphorylation pathway leading to the synthesis of biologically active triphospho-derivatives of nucleoside analogs used in antiviral therapies and in particular in the treatment of AIDS. NDP kinase lacks specificity for the nucleobase and can use as substrate both ribo- or 2'-deoxyribonucleotides. However, only nucleoside analogs with a sugar moiety in the D-configuration (e.g. 3'-deoxy-3'-azidothymidine (AZT), 2',3'-didehydro-2',3'-dideoxythymidine (d4T)) have so far been analyzed as substrates of NDP kinase. In contrast, beta-L-2',3'-dideoxy-3'-thiacytidine (3TC), also called lamivudine, is a nucleoside analog that is now widely used in AIDS therapy and has a sugar moiety in the L-configuration. Using protein fluorescence to monitor the phosphotransfer between the enzyme and the nucleotide derivative at the presteady state, we have studied the reactivity of 3TC triphosphate and of other L-dideoxynucleotides with NDP kinase. We found that L-dideoxynucleoside triphosphates have a poor affinity for NDP kinase and that the catalytic efficiency of the phosphorylation of L-dideoxyderivatives is very low as compared with their D-enantiomers. We discuss these results using a computer model of 3TC diphosphate bound to the NDP kinase active site. NDP kinase may not seem to be the major enzyme phosphorylating 3TC-DP, in contrast to current opinion.

Animals↗
PubMed2001

Addition and correction: the NF-kappa B-like DNA binding activity observed in Dictyostelium nuclear extracts is due to the GBF transcription factor.

We have previously reported that a NF-kappa B transduction pathway was likely to be present in the cellular slime mold Dictyostelium discoideum. This conclusion was based on several observations, including the detection of developmentally regulated DNA binding proteins in Dictyostelium nuclear extracts that bound to bona fide kappa B sequences. We have now performed additional experiments which demonstrate that the protein responsible for this NF-kappa B-like DNA binding activity is the Dictyostelium GBF (G box regulatory element binding factor) transcription factor. This result, along with the fact that no sequence with significant similarity to components of the mammalian NF-kappa B pathway can be found in Dictyostelium genome, now almost entirely sequenced, led us to reconsider our previous conclusion on the occurrence of a NF-kappa B signal transduction pathway in Dictyostelium.

Published Erratum↗
PubMed2000

The mechanism of phosphorylation of anti-HIV D4T by nucleoside diphosphate kinase.

The last step in the intracellular activation of antiviral nucleoside analogs is the addition of the third phosphate by nucleoside diphosphate (NDP) kinase resulting in the synthesis of the viral reverse transcriptase substrates. We have previously shown that dideoxynucleotide analogs and 3'-deoxy-3'-azidothymidine (AZT) as di- or triphosphate are poor substrates for NDP kinase. By use of protein fluorescence, we monitor the phosphotransfer between the enzyme and the nucleotide analog. Here, we have studied the reactivity of D4T (2',3'-dideoxy-2',3'-didehydrothymidine; stavudine) as di- (DP) or triphosphate (TP) at the pre-steady state. The catalytic efficiency of D4T-DP or -TP is increased by a factor of 10 compared with AZT-DP or -TP, respectively. We use an inactive mutant of NDP kinase to monitor the binding of a TP derivative, and show that the affinity for D4T-TP is in the same range as for the natural substrate deoxythymidine triphosphate, but is 30 times higher than for AZT-TP. Our results indicate that D4T should be efficiently phosphorylated after intracellular maturation of a prodrug into D4T-monophosphate.

Adenosine Triphosphate↗
PubMed1999

Evidence for the presence of an NF-kappaB signal transduction system in Dictyostelium discoideum.

The Rel/NF-kappaB family of transcription factors and regulators has so far only been described in vertebrates and arthropods, where they mediate responses to many extracellular signals. No counterparts of genes coding for such proteins have been identified in the Caenorhabditis elegans genome and no NF-kappaB activity was found in Saccharomyces cerevisiae. We describe here the presence of an NF-kappaB transduction pathway in the lower eukaryote Dictyostelium discoideum. Using antibodies raised against components of the mammalian NF-kappaB pathway, we demonstrate in Dictyostelium cells extracts the presence of proteins homologous to Rel/NF-kappaB, IkappaB and IKK components. Using gel-shift experiments in nuclear extracts of developing Dictyostelium cells, we demonstrate the presence of proteins binding to kappaB consensus oligonucleotides and to a GC-rich kappaB-like sequence, lying in the promoter of cbpA, a developmentally regulated Dictyostelium gene encoding the Ca(2+)-binding protein CBP1. Using immunofluorescence, we show specific nuclear translocation of the p65 and p50 homologues of the NF-kappaB transcription factors as vegetatively growing cells develop to the slug stage. Taken together, our results strongly indicate the presence of a complete NF-kappaB signal transduction system in Dictyostelium discoideum that could be involved in the developmental process.

Animals↗
PubMed1998

Pre-steady state of reaction of nucleoside diphosphate kinase with anti-HIV nucleotides.

The pre-steady-state reaction of Dictyostelium nucleoside diphosphate (NDP) kinase with dideoxynucleotide triphosphates (ddNTP) and AZT triphosphate was studied by quenching of protein fluorescence after manual mixing or by stopped flow. The fluorescence signal, which is correlated with the phosphorylation state of the catalytic histidine in the enzyme active site, decreases upon ddNTP addition according to a monoexponential time course. The pseudo-first order rate constant was determined for different concentrations of the various ddNTPs and was found to be saturable. The data are compatible with a two-step reaction scheme, where fast association of the enzyme with the dideoxynucleotide is followed by a rate-limiting phosphorylation step. The rate constants and dissociation equilibrium constants determined for each dideoxynucleotide were correlated with the steady-state kinetic parameters measured in the enzymatic assay in the presence of the two substrates. It is shown that ddNTPs and AZT triphosphate are poor substrates for NDP kinase with a rate of phosphate transfer of 0.02 to 3.5 s-1 and a KS of 1-5 mM. The equilibrium dissociation constants for ADP, GDP, ddADP, and ddGDP were also determined by fluorescence titration of a mutant F64W NDP kinase, where the introduction of a tryptophan at the nucleotide binding site provides a direct spectroscopic probe. The lack of the 3'-OH in ddNTP causes a 10-fold increase in KD. Contrary to "natural" NTPs, NDP kinase discriminates between various ddNTPs, with ddGTP the more efficient and ddCTP the least efficient substrate within a range of 100 in kcat values.

Animals↗
PubMed1998

Tyrosine hydroxylase in the european eel (Anguilla anguilla): cDNA cloning, brain distribution, and phylogenetic analysis.

We report the isolation of a full-length eel tyrosine hydroxylase (TH) cDNA that is characterized by a long 3' untranslated region and by a diversity restricted to the 3' end owing to the differential use of three polyadenylation signals. The longest eel TH mRNA was distinctive in the presence of four pentameric elements (AUUUA) in the AU-rich 3' noncoding region. Such a diversity could provide the basis of posttranscriptional or translational regulation of eel TH gene expression. Comparison of the eel TH sequence with those of other aromatic amino acid hydroxylases (TH, tryptophan hydroxylase, and phenylalanine hydroxylase) and phylogenetic analysis confirmed that the N-terminal regulatory domain is highly divergent, contrasting with the conservation of the catalytic core of the enzyme. Molecular phylogenies including the available sequences of the three hydroxylase genes suggested that the duplication of their common ancestor occurred before the emergence of arthropods. The regional expression of the eel TH mRNA was studied by semiquantitative PCR, northern blots, and in situ hybridization and compared with the immunocytochemical localization of TH protein. The data showed that TH mRNA is mostly expressed in the olfactory and hypothalamic areas, whereas sparse TH-expressing cell bodies are present in the telencephalic region and brainstem. No labeling was detected in the mesencephalic area, in striking contrast with that found in amphibians and amniotes.

Amino Acid Sequence↗
PubMed1997

NMR studies on the flexibility of nucleoside diphosphate kinase.

Human NDP kinase B, product of the nm23-H2 gene, binds DNA. It has been suggested that a helix hairpin on the protein surface, part of the nucleotide substrate binding site, could accommodate DNA binding by swinging away. The presence of flexible regions was therefore investigated by 1H NMR dynamic filtering. Although TOCSY peaks could be assigned to five residues at the N terminus of Dictyostelium NDP kinase, no flexible region was detected in the human enzyme. These data favor the idea that the protein offers different binding sites to mono- and polynucleotides.

Amino Acid Sequence↗
PubMed1996

Projection neurons of the mormyrid electrosensory lateral line lobe: morphology, immunohistochemistry, and synaptology.

This paper describes the morphological, immunohistochemical, and synaptic properties of projection neurons in the highly laminated medial and dorsolateral zones of the mormyrid electrosensory lateral line lobe (ELL). These structures are involved in active electrolocation, i.e., the detection and localization of objects in the nearby environment of the fish on the basis of changes in the reafferent electrosensory signal generated by the animal's own electric organ discharge. Electrosensory, corollary electromotor command-associated signals (corollary discharges), and a variety of other inputs are integrated within the ELL microcircuit. The organization of ELL projection neurons is analyzed at the light and electron microscopic levels based on Golgi impregnations, intracellular labeling, neuroanatomical tracer techniques, and gamma-aminobutyric acid (GABA), gamma-aminobutyric acid decarboxylase (GAD), and glutamate immunohistochemistry. Two main types of ELL projection neurons have been distinguished in mormyrids: large ganglionic (LG) and large fusiform (LF) cells. LG cells have a multipolar cell body (average diameter 13 microns) in the ganglionic layer, whereas LF cells have a fusiform cell body (on average, about 10 x 20 microns) in the granular layer. Apart from the location and shape of their soma, the morphological properties of these cell types are largely similar. They are glutamaterigic and project to the midbrain torus semicircularis, where their axon terminals make axodendritic synaptic contacts in the lateral nucleus. They have 6-12 apical dendrites in the molecular layer, with about 10,000 spines contacted by GABA-negative terminals and about 3,000 GABA-positive contacts on the smooth dendritic surface between the spines. Their somata and short, smooth basal dendrites, which arborize in the plexiform layer (LG cells) or in the granular layer (LF cells), are densely covered with GABA-positive, inhibitory terminals. Correlation with physiological data suggests that LG cells are I units, which are inhibited by stimulation of the center of their receptive fields, and LF cells are E units, excited by electric stimulation of the receptive field center. Comparison with the projection neurons of the ELL of gymnotiform fish, which constitute another group of active electrolocating teleosts, shows some striking differences, emphasizing the independent development of the ELL in both groups of teleosts.

Animals↗
PubMed1996

Interneurons of the ganglionic layer in the mormyrid electrosensory lateral line lobe: morphology, immunohistochemistry, and synaptology.

This is the second paper in a series that describes the morphology, immunohistochemistry, and synaptology of the mormyrid electrosensory lateral line lobe (ELL). The ELL is a highly laminated cerebellum-like structure in the rhombencephalon that subserves an active electric sense: Objects in the nearby environment of the fish are detected on the basis of changes in the reafferent electrosensory signals that are generated by the animal's own electric organ discharge. The present paper describes interneurons in the superficial (molecular, ganglionic, and plexiform) layers of the ELL cortex that were analyzed in the light and electron microscopes after Golgi impregnation, intracellular labeling, neuroanatomical tracing, and gamma-aminobutyric acid (GABA) immunohistochemistry. The most numerous interneurons in the ganglionic layer are GABAergic medium-sized ganglionic (MG) cells and small ganglionic (SG) cells. MG cells have 10-20 spiny apical dendrites in the molecular layer, a cell body of 10-12 microns diameter in the ganglionic layer, a single basal dendrite that gives rise to fine, beaded, axon-like branches in either the plexiform layer (MG1 subtype) or the deeper granular layer (MG2 subtype), and an axon that terminates in the plexiform layer. Their apical dendritic tree has 12,000-22,000 spines that are contacted by GABA-negative terminals, and it receives, 1,250-2,500 GABA-positive contacts on the smooth dendritic surface between the spines. The average ratio of GABA-negative to GABA-positive contacts on the interneuron apical dendrites (14:1) is significantly higher than that for the efferent projection cells that have been described previously (Grant et al. [1996] J. Comp. Neurol., this issue). The somata and basal dendrites of MG cells receive a low to moderate density of GABAergic synaptic input, and their axons make GABAergic synaptic contacts with the somata and cell bodies of MG as well as with large ganglionic (LG) cells. SG cells probably represent immature, growing MG cells. Other interneurons in the superficial ELL layers include GABAergic stellate cells in the molecular layer, two types of non-GABAergic cells with smooth dendrites in the deep molecular layer that are named thick-smooth dendrite cells and deep molecular layer cells, and horizontal cells that are encountered particularly in the plexiform layer. Comparison with the ELL of waveform gymnotiform fish, which is another group of active electrolocating teleosts that has been investigated thoroughly, shows striking differences. In these fish, no GABAergic interneurons are found in the ganglionic (pyramidal) layer of the ELL, and GABA-negative interneurons with smooth dendrites in the molecular layer also seem to be lacking. At present, the phylogenetic origin of the described superficial interneurons in the mormyrid ELL is uncertain.

Animals↗
PubMed1996

The enzymatic activity of Drosophila AWD/NDP kinase is necessary but not sufficient for its biological function.

The Drosophila abnormal wing discs (awd) gene encodes the subunit of a protein that has nucleoside diphosphate kinase (NDP kinase) activity. Null mutations of the awd gene cause lethality after puparium formation. Larvae homozygous for such mutations have small imaginal discs, lymph glands, and brain lobes. Neither the imaginal discs nor the ovaries from such null mutant larvae are capable of further growth or normal differentiation when transplanted into suitable host larvae. This null mutant phenotype can be entirely rescued by one copy of a transgene that has 750 bp of awd upstream regulatory DNA fused to a full-length awd cDNA. Tissue-specific expression of AWD protein from this rescue transgene is identical to tissue-specific expression of beta-galactosidase from a reporter transgene that has the same regulatory region fused to the bacterial lac Z gene. However, this rescue transgene or reporter transgene expression pattern is only a subset of the endogenous pattern of expression detected by either in situ hybridization or immunohistochemistry. This suggests that awd is normally expressed in some tissues where it is not required. The null mutant phenotype cannot be rescued at all by a transgene that has 750 bp of awd upstream regulatory DNA fused to a full-length awd cDNA with a mutation that eliminates NDP kinase activity by replacement of the active site histidine with alanine. This suggests that the enzymatic activity of the AWD protein is necessary for its biological function. The human genes nm23-H1 and nm23-H2 encode NDP kinase A and B subunits, respectively. The protein subunit encoded by either human nm23 gene is 78% identical to that encoded by the Drosophila awd gene. Transgenes that have the 750-bp awd upstream regulatory DNA fused to human nm23-H2 cDNA but not to nm23-H1 cDNA can rescue the imaginal disc phenotype and the zygotic lethality caused by homozygosis for an awd null mutation as efficiently as an awd transgene. However, rescue of female sterility requires twice as much nm23-H2 expression as awd expression. This implies that the enzymatic activity of the AWD protein is not sufficient for its biological function. The biological function may require nonconserved residues of the AWD protein that allow it to interact with other proteins.

Animals↗
PubMed1996

The Enzymatic Activity of Drosophila AWD/NDP Kinase Is Necessary but Not Sufficient for Its Biological Function

The Drosophila abnormal wing discs (awd) gene encodes the subunit of a protein that has nucleoside diphosphate kinase (NDP kinase) activity. Null mutations of the awd gene cause lethality after puparium formation. Larvae homozygous for such mutations have small imaginal discs, lymph glands, and brain lobes. Neither the imaginal discs nor the ovaries from such null mutant larvae are capable of further growth or normal differentiation when transplanted into suitable host larvae. This null mutant phenotype can be entirely rescued by one copy of a transgene that has 750 bp of awd upstream regulatory DNA fused to a full-length awd cDNA. Tissue-specific expression of AWD protein from this rescue transgene is identical to tissue-specific expression of beta-galactosidase from a reporter transgene that has the same regulatory region fused to the bacterial lac Z gene. However, this rescue transgene or reporter transgene expression pattern is only a subset of the endogenous pattern of expression detected by either in situ hybridization or immunohistochemistry. This suggests that awd is normally expressed in some tissues where it is not required. The null mutant phenotype cannot be rescued at all by a transgene that has 750 bp of awd upstream regulatory DNA fused to a full-length awd cDNA with a mutation that eliminates NDP kinase activity by replacement of the active site histidine with alanine. This suggests that the enzymatic activity of the AWD protein is necessary for its biological function. The human genes nm23-H1 and nm23-H2 encode NDP kinase A and B subunits, respectively. The protein subunit encoded by either human nm23 gene is 78% identical to that encoded by the Drosophila awd gene. Transgenes that have the 750-bp awd upstream regulatory DNA fused to human nm23-H2 cDNA but not to nm23-H1 cDNA can rescue the imaginal disc phenotype and the zygotic lethality caused by homozygosis for an awd null mutation as efficiently as an awd transgene. However, rescue of female sterility requires twice as much nm23-H2 expression as awd expression. This implies that the enzymatic activity of the AWD protein is not sufficient for its biological function. The biological function may require nonconserved residues of the AWD protein that allow it to interact with other proteins.

Journal Article↗
PubMed1996

Control of 6-(D-threo-1',2'-dihydroxypropyl) pterin (dictyopterin) synthesis during aggregation of Dictyostelium discoideum. Involvement of the G-protein-linked signalling pathway in the regulation of GTP cyclohydrolase I activity.

6-(D-threo-1',2'-Dihydroxypropylpterin (dictyopterin) has been identified in extracts of growing Dictyostelium dicoideum cells [Klein, Thiery and Tatischeff (1990) Eur. J. Biochem. 187, 665-669]. We demonstrate that it originates from GTP by de novo biosynthesis and that the first committed step is catalysed by GTP cyclohydrolase I, yielding dihydroneopterin triphosphate [neopterin is 6-(D-erythro-1',2',3'-trihydroxypropyl) pterin]. The GTP cyclohydrolase I activity is found in the cytosolic fraction and in a membrane-associated form. The level of a 0.9 kb mRNA coding for GTP cyclohydrolase I decreases to about 10% of its initial value within 2 h after Dictyostelium cells start development induced by starvation. In the cytosolic fraction, the specific activities of GTP cyclohydrolase I, as well as the concentrations of (6R/S)-5,6,7,8-tetrahydrodictyopterin (H4dictyopterin), follow this decline of the mRNA level. In the particulate fraction, however, the specific activities of GTP cyclohydrolase I and, in consequence, H4dictyopterin synthesis, transiently increase and reach a maximum after 4-5 h of development. The time-course of H4dictyopterin concentrations in the starvation medium closely correlates with its production in the membrane fraction. The activity of membrane-associated GTP cyclohydrolase I can be increased by pre-incubation of the cell lysate with guanosine 5'-[gamma-thio]triphosphate and Mg2+. This GTP analogue does not serve as a substrate and has no direct effect on the enzyme activity, indicating that a G-protein-linked signalling pathway is involved in the regulation of GTP cyclohydrolase I activity and thus in H4dictyopterin production during early development of D. discoideum.

Alcohol Oxidoreductases↗
PubMed1996

Increased levels of nm23 H1/nucleoside diphosphate kinase A mRNA associated with adenocarcinoma of the prostate.

Overexpression of the nm23H1 gene has been associated with the suppression of metastasis in several solid tumors. However, in colorectal carcinoma and neuroblastoma, increased levels of nm23 H1 nucleoside diphosphate kinase A (NDPKA) mRNA are associated with tumorigenesis. To determine the role of nm23 H1/NDPKA in the prostate, normal and/or malignant tissue samples from 29 consecutive patients were studied. Levels of nm23 H1/NDPKA mRNA and nm23 H1/NDPKA mRNA protein were determined in tissue from 18 and 27 patients, respectively. In all, 16 of the 18 tumor samples expressed increased levels of nm23 H1/NDPKA mRNA as compared with those measured in normal tissue. The level of nm23 H1/NDPKA mRNA was > 10-fold higher in a metastatic lymph node than in normal prostate tissue. All cancer specimens and areas of prostatic intraepithelial neoplasia showed immunoreactivity with the nm23 H1/NDPKA antibody; however, normal prostatic tissue was unreactive. These findings suggest that overexpression of the nm23 H1/NDPKA gene occurs frequently in adeno-carcinomas of the prostate and may be an early event in prostate cancer tumorigenesis.

Adenocarcinoma↗
PubMed1996

Overexpression of nucleoside diphosphate/kinase A/nm23-H1 protein in human lung tumors: association with tumor progression in squamous carcinoma.

Levels of nm23-H1/nucleoside diphosphate/kinase A expression have been reported to correlate inversely with metastatic potential in some tumors but not in others. To clarify the role of nm23 in lung carcinoma, the genetic abnormalities of nucleoside diphosphate/kinase A/nm23-H1 were investigated at the DNA and protein levels. A series of 104 human lung tumors (42 neuroendocrine (NE) and 62 non-NE tumors) was analyzed for nm23-H1 protein expression by immunohistochemistry using one polyclonal and two monoclonal Ab and for genomic alterations using Southern blotting and single-strand conformation polymorphism. Overexpression of the nm23-H1 protein relative to the normal lung epithelia (pneumocyte and bronchial epithelial cells) was observed in 83% (35/42) of NE carcinomas and in 89% (55/62) of non-NE carcinomas. Eight of nine carcinoids exhibited an increased expression of nm23-H1 protein, suggesting that this overexpression of the nm23 protein is necessary for proliferation in any tumors. No significant correlation was found between nm23 staining and any clinicopathologic parameters in NE carcinoma or in adenocarcinoma. In squamous carcinoma, high levels of nm23-H1 protein expression were associated with tumor stage (p = 0.0036). Allelic deletion or genetic amplification was never found. No altered mobility was detected using single-strand conformation polymorphism analysis. These data show that nm23-H1 protein is overexpressed in a large number of lung tumors of all histologic types, in association with advanced tumor stage in squamous carcinoma. They also suggest that nm23-H1 might play a role in the progression of lung tumors rather than in antimetastatic function.

Base Sequence↗
PubMed1995

Candida albicans nucleoside-diphosphate kinase: purification and characterization.

Soluble nucleoside-diphosphate kinase (NDP kinase) from Candida albicans was purified to electrophoretic homogeneity and a partial sequence was determined. The enzyme was kinetically and physically characterized. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the purified enzyme revealed a single polypeptide of 17 kDa upon staining, by immunodetection with heterologous and homologous antibodies, and by autoradiography of the phosphorylated enzyme. Furthermore, isoelectric focusing of the purified native enzyme showed a single acidic band (pI 4.5). These results together with a native molecular mass of 98 kDa suggest a hexameric native enzyme composed of identical subunits. Like NDP kinases from other sources, the catalysis involves a phosphoenzyme intermediate that is rapidly formed upon incubation of the enzyme with ATP. The transfer of phosphate from phosphoprotein intermediate to nucleoside diphosphates is equally fast. Kinetic experiments indicated that GTP and ATP had the lowest Km compared to UTP, dTTP, and CTP. GDP acted as a preferred acceptor as assessed by kinetic measurements as well as by competition experiments. Experimental data are presented indicating the existence of a membrane-associated NDP kinase. Preliminary characterization of this enzyme suggests that cytosolic and membrane-associated NDP kinases are similar proteins.

Amino Acid Sequence↗
PubMed1995

High nm23-H1/NDPK-A expression in Ewing tumors: paradoxical immunohistochemical reactivity and lack of prognostic significance.

Expression of nm23-H1/NDPK-A has been reported to correlate inversely with metastasizing potential of rodent experimental cells and some human tumors. In the search for reliable molecular prognostic indicators for Ewing tumors (ET), a group of aggressive presumably neuroectodermal malignancies in children and adolescents, we studied nm23-H1/NDPK-A expression. Northern-blot and RT-PCR analyses were employed to semi-quantificatively measure nm23-H1 mRNA levels in ET cell lines and tissue extracts. A panel of monoclonal antibodies (MAbs) were used to evaluate protein abundance by Western blotting and immunohistochemistry. The nm23-H1/NDPK-A gene was also investigated on the DNA level to define possible genomic alterations. Our results revealed neither nm23-H1 allelic loss nor gene amplification and failed to show any significant variation in nm23-H1 mRNA or NDPK-A protein levels of primary or metastatic ET. NDPK-A protein levels were high and comparable to those of MCF-7 breast-cancer cells and to aggressive stage-IV neuroblastoma cell lines. nm23-H2/NDPK-B expression in ET was slightly more variable but generally lower than in MCF-7 cells. In the immunohistochemical analysis, however, discrepancies in the reactivity patterns with different antibodies were observed. Differential sensitivity to various fixation methods and heat treatment pointed to a structurally polymorphic NDPK-A protein. nm23-H1 expression studies using immunohistochemistry for prognostic counselling should thus be interpreted with caution.

Adolescent↗
PubMed1995

A protein kinase from Dictyostelium discoideum with an unusual acidic repeat domain.

DdKinX codes for 1093 amino acids which are organized in four regions: the N-terminal catalytic domain, a region containing 30% acidic amino acids, tandem repeats of the motif VKVEEPVEE and the C-terminus. Identity with other protein kinases is 25 to 30%. Descendent trees show that DdKinX does not belong to any of the known kinase branches.

Amino Acid Sequence↗