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E Cukerman

Publications and source records attributed to E Cukerman.

At least 19 recordsLinked to original sources
PubMed1998

Identification of sequence-tagged transcripts differentially expressed within the human hematopoietic hierarchy.

Hematopoiesis is regulated by a complex gene expression program. To gain further insight into the molecular mechanisms underlying this process in humans, we sampled the transcriptional activity of the CD34+ hematopoietic progenitor line KG1a by single-pass sequencing the 5' ends of 1018 clones from a unidirectional cDNA library. Searches of public databases with the resulting expressed sequence tags (ESTs) identified 101 clones that showed no sequence similarity to any of the existing entries and that were therefore considered to derive from previously undescribed genes. Of the remaining 917 ESTs, 553 (a total of 485 distinct transcripts) corresponded to known genes. A further 279 KG1a ESTs matched or exhibited sequence similarity to ESTs or genomic sequences from humans and other species. Among the latter were putative human orthologs of developmental and cell cycle control genes from Caenorhabditis elegans, Drosophila, and yeast, as well as genes whose predicted amino acid sequences showed similarity to mammalian transcription factors. Hybridization of selected novel KG1a ESTs to globally amplified cDNAs prepared from single primary human hematopoietic precursors and homogeneous populations of terminally maturing hematopoietic cells revealed transcripts that are expressed preferentially at a specific stage or in a particular lineage within the hematopoietic hierarchy. Thus, included in the KG1a EST dataset are candidates for new human genes that may play roles in hematopoietic differentiative progression and lineage commitment.

Animals↗
PubMed1997

A genome-based resource for molecular cardiovascular medicine: toward a compendium of cardiovascular genes.

BACKGROUND: Large-scale partial sequencing of cDNA libraries to generate expressed sequence tags (ESTs) is an effective means of discovering novel genes and characterizing transcription patterns in different tissues. To catalogue the identities and expression levels of genes in the cardiovascular system, we initiated large-scale sequencing and analysis of human cardiac cDNA libraries. METHODS AND RESULTS: Using automated DNA sequencing, we generated 43,285 ESTs from human heart cDNA libraries. An additional 41,619 ESTs were retrieved from public databases, for a total of 84,904 ESTs representing more than 26 million nucleotides of raw cDNA sequence data from 13 independent cardiovascular system-based cDNA libraries. Of these, 55% matched to known genes in the Genbank/EMBL/DDBJ databases, 33% matched only to other ESTs, and 12% did not match to any known sequences (designated cardiovascular system-based ESTs, or CVbESTs). ESTs that matched to known genes were classified according to function, allowing for detection of differences in general transcription patterns between various tissues and developmental stages of the cardiovascular system. In silico Northern analysis of known gene matches identified widely expressed cardiovascular genes as well as genes putatively exhibiting greater tissue specificity or developmental stage specificity. More detailed analysis identified 48 genes potentially overexpressed in cardiac hypertrophy, at least 10 of which were previously documented as differentially expressed. Computer-based chromosomal localizations of 1048 cardiac ESTs were performed to further assist in the search for disease-related genes. CONCLUSIONS: These data represent the most extensive compilation of cardiovascular gene expression information to date. They further demonstrate the untapped potential of genome research for investigating questions related to cardiovascular biology and represent a first-generation genome-based resource for molecular cardiovascular medicine.

Blotting, Northern↗
PubMed1997

Identification of genes encoding zinc finger motifs in the cardiovascular system.

The Zn2+-finger DNA-binding domain has been identified in several developmental control proteins, transcription factors and gene products associated with diseases, as well as in several RNA-binding proteins. We applied library screening, expressed sequence tagging (EST sequencing), Zn2+-binding assays and Northern blot hybridization, in order to characterize novel cDNA clones of the human cardiovascular system which contain Zn2+-finger motifs. An embryonic (8-10 weeks gestation) heart lambda ZAP Express cDNA library was screened with an oligonucleotide probe deduced from a consensus amino acid sequence which is highly conserved for Zn2+-finger proteins, and approximately 350 positive clones were isolated from 1 x 10(4) plaque-forming units (pfu) initially plated. The isolated clones were classified as known and novel following single pass automated DNA sequencing. Analysis of Northern blot hybridization delineated the tissue specificity of these clones, as well as their association with cardiac growth and development. Existence of Zn2+-finger motifs in the novel clones was confirmed by Zn2+-binding assay. In this report, we present the characterization of eight novel clones, including the complete cDNA sequences of one of these clones (HHZ-123).

Amino Acid Sequence↗
PubMed1997

Construction of a human heart cDNA library and identification of cardiovascular based genes (CVBest).

The availability of high quality cDNA libraries is often crucial to the successful identification and characterization of genes. The concepts and potential pitfalls of constructing cDNA libraries are presented. Various applications requiring high quality cDNA libraries are outlined, including large-scale single pass sequencing of cDNA clones to generate expressed sequence tags (ESTs) and differential screening of cDNA libraries. The usefulness of combining such approaches for the discovery of novel disease-related and cardiovascular-based ESTs (CVBest) is discussed.

Base Sequence↗
PubMed1996

Localization of a novel zinc finger gene to the human chromosome 7p11.2-p12 by fluorescence in situ hybridization.

A novel zinc finger gene (ZNF182) was isolated from a human fetal cardiac cDNA library, using a consensus C2H2 zinc finger oligonucleotide probe. This gene was assigned to human chromosome 7p11.1-p12 by fluorescence in situ hybridization (FISH). Additional FISH signals were identified on both the long and the short arms of chromosome 19, suggesting the presence of homologous genes at these loci.

Chromosome Mapping↗
PubMed1996

Localization of three novel zinc finger genes to the centromeric region of human chromosome 10 by fluorescence in situ hybridization.

An oligonucleotide probe for the consensus sequence of the linker region of zinc finger proteins was used to isolate cDNA clones from a human fetal heart cDNA library. Following DNA sequencing analysis and comparison, genes for the novel clones were mapped by fluorescence in situ hybridization. We report the chromosomal localization of three zinc finger-coding genes to the region of centromere on human chromosome 10p11.1q-11.2, indicating involvement in gene duplication and chromosome rearrangement during primate evolution.

Amino Acid Sequence↗
PubMed1996

Identification of a locus of zinc finger genes in human chromosome 19q13.1-q13.3 region by fluorescence in situ hybridization.

A group of zinc-binding cDNA clones from a human fetal heart library was isolated using an oligonucleotide probe to the consensus sequence of the linker region of zinc finger proteins. Genes for novel clones were mapped by fluorescence in situ hybridization. In the process, we identified a previously unrecognized locus for two zinc finger-coding genes in human chromosome 19q13.1-q13.3 (ZNF180,ZNF181), where genomic rearrangements were shown to be accompanied by various developmental abnormalities, DNA repair deficiencies, and cellular malignancies.

Amino Acid Sequence↗
PubMed1995

Identification of a GATA motif in the cardiac alpha-myosin heavy-chain-encoding gene and isolation of a human GATA-4 cDNA.

In an attempt to identify the cardiac-specific genes regulated by the transcription factor GATA-4, we have identified a putative GATA-binding site located within the 5' flanking sequence of the human cardiac alpha-myosin heavy-chain-encoding gene. The 23-bp sequence surrounding the core GATA-binding site is conserved across species. The core motif and flanking sequences of this GATA-binding site are almost identical to that of a well-established GATA-binding site located within the 3' enhancer of the human beta-globin gene. Using electrophoretic mobility shift analysis, two distinct nuclear factors were found to bind specifically to this element. We have isolated a full-length cDNA clone for human GATA-4 (hGATA-4) by screening a human heart cDNA library. The hGATA-4 cDNA sequence shows 85% identity with murine GATA-4 in the protein coding region. The deduced amino-acid sequence within the two zinc-finger DNA-binding domains of human GATA-4 is 100% identical with murine GATA-4. Northern blot analysis reveals that this 4.4-kb transcript has higher expression in adult heart than in fetal heart. Our results suggest that GATA-4 may regulate a set of cardiac-specific genes and play a crucial role in cardiogenesis.

Amino Acid Sequence↗
PubMed1994

A catalogue of genes in the cardiovascular system as identified by expressed sequence tags.

The heart, which is composed of all the cellular components of the circulatory system, is a representative organ for obtaining genes expressed in the cardiovascular system in normal and disease states. We used partial sequences of cDNA clones, or expressed sequence tags, to identify and tag genes expressed in this organ. More than 3500 partial sequences representing > 3000 cDNA clones have been obtained from either the 5' or 3' end of inserts derived from human heart cDNA libraries. Of 3132 cDNA clones analyzed by sequence similarity searching against the GenBank/EMBL data bases, 1485 (47.4%) were found to represent additional, previously undiscovered genes, whereas 267 clones were matched to human brain expressed sequence tags. Clones matching to known genes were catalogued according to their putative structural and cellular functions. cDNA probes from reverse-transcribed mRNAs of fetal and adult hearts were used to study differential expression of selected clones in cardiac development. Cataloguing genes expressed in the heart may provide insight into the genes involved in health and cardiovascular disease.

Adult↗
PubMed1994

Characterization and nucleotide sequence of the cardiac alpha-myosin heavy chain gene from Syrian hamster.

In small mammals, the cardiac alpha-myosin heavy chain (MyHC) gene is predominantly expressed in the adult ventricle, while the beta-MyHC gene is predominantly expressed in the fetal ventricle. In order to perform comparative studies of these genes, we sequenced the Syrian hamster alpha-MyHC gene by determining a continuous 32,415 base pair (bp) sequence. It contained 39 exons encoding a predicted polypeptide of 1939 amino acid residues (aa) with a molecular weight of 223,644 Da. Sequence comparison revealed that the coding sequence is highly identical to that of the human alpha-MyHC gene. A marked feature of the Syrian hamster alpha-MyHC gene was the abundance of reiterated sequences within introns. Eleven B1 and four B2 retroposons were identified, a variety of di-, tri-, and tetra-nucleotide repeats were present, and three longer direct repeats were found. Some of them appear to be associated with genetic recombination events.

Amino Acid Sequence↗
PubMed1993

Characterization of Zn(2+)-binding nuclear proteins present in the myocardium.

Nonhistone nuclear proteins were isolated from 3-5 day old neonatal as well as 3 month-old adult myocardium. The nuclear proteins were separated and analyzed by two-dimensional polyacrylamide gel electrophoresis. Using a blot transfer technique equilibrated with 65Zn2+, at least four polypeptides exhibited Zn(2+)-binding activity over the spectrum of nonhistone nuclear proteins. A protein with a molecular weight of 68kDa pI7.8, which has been characterized for its involvement in nucleosome structure, consistently binds Zn2+ in both the neonatal and adult myocardium. This nuclear protein has now been further characterized by partial amino acid microsequencing. It was found that this novel polypeptide is distinct from the pore-complex lamina proteins. Three other polypeptides with M tau 90kDa, pI7.8, M tau 68kDa, pI6.5 and M tau 35kDa, pI7.5 exhibited increased Zn(2+)-binding activity in neonatal myocardium as compared to adult myocardium. Together with results from our previous studies, this study provides the first evidence implicating Zn(++)-binding nuclear proteins in the processes of growth and differentiation of myocardial development.

Amino Acid Sequence↗
PubMed1987

Hypophysectomy does not alter the effects of somatostatin or growth hormone on canine splanchnic biogenic amine release.

We have demonstrated previously that growth hormone (GH) and somatostatin (somatotropin release inhibitory factor, SRIF) exert comparable effects on the release of splanchnic biogenic amines. The purpose of the present investigation was to study further the response of the two hormones and see whether the similarity persists in dogs completely deprived of endogenous GH. Experiments were conducted in seven hypophysectomized dogs fitted with an indwelling portal catheter. Two to 4 weeks after surgery the responsiveness of their catecholaminergic neurons was tested by an injection of human beta-endorphin (20 micrograms/kg); it caused a rise in portal catecholamine levels equivalent to that seen in intact dogs. Then the effect of a spike concentration of SRIF or GH on hepatic portal and peripheral levels of free serotonin and catecholamines was studied, all by radioenzymatic methods. The intravenous injection of ovine GH (100 micrograms/kg) or equimolar amounts of SRIF (7.5 micrograms/kg) produced in the hepatic portal circulation a transient but statistically significant rise of serotonin and a concomitant reduction in the concentration of dopamine, norepinephrine, and epinephrine. No changes were found in the peripheral circulation. The response patterns to SRIF or GH were virtually identical, which is in keeping with our other data, suggesting that the effect of GH on splanchnic biogenic amine secretion is SRIF-dependent and mediated by SRIF-containing neurons.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1987

Secretory pattern of canine growth hormone.

Our aim was to define the secretory pattern of growth hormone (GH) under basal conditions in fasted, conscious, male dogs accustomed to handling. Blood samples were withdrawn from a cephalic vein at 15-min intervals. In this way, any ultradian rhythms, if present, could be detected within the frequency range of 0.042-2 cycles/h. In addition, samples were drawn at either 1- or 2.5-min intervals for 2.5 or 5 h to determine whether frequency components greater than 2 cycles/h were present. GH was measured by radioimmunoassay and the raw data were submitted to time series analysis employing power spectral estimation by means of fast Fourier transformation techniques. Peak plasma levels were up to 12 times higher than the baseline concentration of approximately 1 ng/ml. Spectral analysis revealed an endogenous frequency of 0.22 cycles/h, i.e., a periodicity of 4.5 h/cycle. The results indicate that under basal conditions the secretory bursts of canine GH are limited to one peak every 4.5 h.

Animals↗
PubMed1986

Insulin dependence of the actions of growth hormone and somatostatin on splanchnic biogenic amines of the dog.

It is known from studies previously conducted in this laboratory that an iv injection of ovine growth hormone (GH, 100 micrograms/kg BW) or an equimolar amount of somatostatin (SRIF, 7.5 micrograms/kg BW), given to normal conscious dogs into a saphenous vein, leads to a significant increase in hepatic portal plasma serotonin and a simultaneous decrease in the concentrations of dopamine, norepinephrine and epinephrine. The changes take place within 12 minutes after the injection and are observed only in the portal circulation. The purpose of the present experiment was to investigate whether or not similar results could be obtained in diabetic animals. Mongrel dogs were rendered diabetic by surgical pancreatectomy and fitted with an indwelling hepatic portal catheter. Radioenzymatic methods were employed for quantitative measurements of plasma free serotonin and catecholamines. No response was noted when the same type of experiments as those conducted in normal dogs were now carried out in trained, fully conscious totally pancreatectomized dogs deprived of exogenous insulin supply. When the same animals were given an injection into a peripheral vein of 50 mU/kg BW regular crystalline insulin (a small dose that affected neither plasma glucose nor biogenic amine levels) 10 minutes prior to the administration of the other hormones, the usual response to both GH and SRIF was restored, i.e. the data were comparable to those of normal dogs. It is concluded that the GH/SRIF effect on gut biogenic amines is insulin dependent.

Animals↗
PubMed1986

The effect of beta-endorphin on biogenic amines, insulin, and glucagon levels in the hepatic portal circulation of normal and pancreatectomized dogs.

The effect of peak concentrations of beta-endorphin on hepatic portal and peripheral levels of plasma catecholamines, free serotonin, glucose, insulin, and glucagon was studied in trained, conscious, normal adult dogs fitted with an indwelling portal catheter. An injection of synthetic human beta-endorphin (20 micrograms/kg BW) into a cephalic vein produced a significant rise in the portal concentration of dopamine, norepinephrine, and epinephrine. The rise was accompanied by a reduction of portal free serotonin levels. The changes were not seen in the peripheral circulation. No appreciable changes in plasma insulin, glucagon, and glucose concentrations were noticed either in the hepatic portal or in the peripheral circulation. The response of the biogenic amines to beta-endorphin was abolished by pretreatment with Naltrexone (1 mg/kg BW). A dose of somatostatin antiserum given before beta-endorphin did not alter the biogenic amine response to the opioid peptide. When beta-endorphin was administered to pancreatectomized dogs devoid of exogenous and endogenous insulin supply, the biogenic amine response remained virtually the same as in normal intact dogs. It is concluded that in the dog a pulse of beta-endorphin causes profound alterations of splanchnic biogenic amine concentrations that are independent of the ambient levels of insulin, somatostatin, and pancreatic glucagon.

Amines↗
PubMed1985

The effect of growth hormone on biogenic amines in the hepatic portal circulation of the dog.

The effects of a spike concentration of growth hormone (GH) on hepatic portal and peripheral levels of free serotonin and catecholamines were studied by improved radioenzymatic methods in trained, conscious, normal, adult dogs fitted with an indwelling portal catheter. An injection of ovine GH (6 or 100 micrograms/kg) into a cephalic vein produced in the hepatic portal circulation a transient, statistically significant rise of serotonin and a concomitant significant reduction in the concentration of dopamine, norepinephrine, and epinephrine. No change was found in the peripheral circulation, partly because the amines were conjugated to sulfates and glucuronides and these derivatives are not detectable by our assays. Thus, a pulse of GH not only stimulates the release of pancreatic hormones and glucose turnover, but also affects the portal profile of glucoregulatory bioamines. The present investigation lends further support to our view that the splanchnic area represents an endocrine system whose preferential target is the liver.

Animals↗
PubMed1985

The effect of tryptophan on biogenic amines in the hepatic portal circulation of the dog.

Mongrel dogs were fitted with indwelling hepatic portal catheters. After recovery from surgery, experiments were conducted in fasting, unrestrained, fully conscious, normal dogs which were accustomed to handling and withdrawal of blood samples. L-Tryptophan, a specific serotonin precursor, was injected into a saphenous vein, 10 microM/kg body weight, dissolved in saline. Plasma serotonin, dopamine, norepinephrine, and epinephrine were determined by radioenzymatic assays in blood samples withdrawn at frequent intervals for 2 h, simultaneously from the indwelling catheter and from a catheter temporarily inserted into a saphenous vein other than the one used for the injection of tryptophan. The injection of the amino acid caused a significant elevation of the concentration of platelet-free serotonin within 60 min and this was accompanied by a reduction in the concentration of the catecholamines, dopamine, norepinephrine, and epinephrine. The changes occurred only in the portal circulation and were not detected in peripheral blood samples. The results of these experiments indicate the existence of a cause and effect related interdependence between the splanchnic serotonergic and adrenergic systems in that the tryptophan-stimulated increase in serotonergic activity resulted in a concomitant reduction in gut adrenergic activity.

Animals↗