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N C Popescu

Publications and source records attributed to N C Popescu.

At least 73 records · Page 4Linked to original sources
PubMed1997

TrnR2, a novel receptor that mediates neurturin and GDNF signaling through Ret.

Glial cell line-derived neurotrophic factor (GDNF) and neurturin (NTN) comprise a family of TGF-beta-related neurotrophic factors (TRNs), which have trophic influences on a variety of neuronal populations. A receptor complex comprised of TrnR1 (GDNFR alpha) and Ret was recently identified and found to be capable of mediating both GDNF and NTN signaling. We have identified a novel receptor based on homology to TrnR1, called TrnR2, that is 48% identical to TrnR1, and is located on the short arm of chromosome 8. TrnR2 is attached to the cell surface via a GPI-linkage, and can mediate both NTN and GDNF signaling through Ret in vitro. Fibroblasts expressing TrnR2 and Ret are approximately 30-fold more sensitive to NTN than to GDNF treatment, whereas those expressing TrnR1 and Ret respond equivalently to both factors, suggesting the TrnR2-Ret complex acts preferentially as a receptor for NTN. TrnR2 and Ret are expressed in neurons of the superior cervical and dorsal root ganglia, and in the adult brain. Comparative analysis of TrnR1, TrnR2, and Ret expression indicates that multiple receptor complexes, capable of mediating GDNF and NTN signaling, exist in vivo.

3T3 Cells↗
PubMed1997

Human uteroglobin gene: structure, subchromosomal localization, and polymorphism.

Human uteroglobin (hUG) or Clara cell 10-kD protein (cc10 kDa) is a steroid-dependent, immunomodulatory, cytokine-like protein. It is secreted by mucosal epithelial cells of all vertebrates studied. The cDNA encoding hUG and the 5' promoter region of the gene have been characterized previously. Here, we report that the structure of the entire hUG gene is virtually identical to those of rabbit, rat, and mouse. It is localized on human chromosome 11q12.3-13.1, a region in which several important candidate disease genes have been mapped by linkage analyses. Our data indicate that candidate genes for atopic (allergic) asthma and Best's vitelliform macular dystrophy are in closest proximity to the hUG gene. To determine whether hUG gene mutation may be involved in the pathogenesis of these diseases, we studied two isolated groups of patients, each afflicted with either atopy or Best's disease, respectively. We detected a single base-pair change in the hUG gene in Best's disease patients and normal controls but no such change was detected in atopy patients. This alteration in hUG gene-sequence in Best disease family appears to be a polymorphism. Although the results of our investigation did not uncover mutations in hUG gene that could be causally related to the pathogenesis of either of these diseases, its conservation throughout vertebrate phyla implies that this gene is of physiological importance. Moreover, the close proximity of this gene to several candidate disease genes makes it an important chromosomal marker in cloning and characterization of those genes.

Animals↗
PubMed1996

Tissue-specific expression, evolutionary conservation and localization of the cph proto-oncogene on Syrian hamster chromosome X.

Treatment of Syrian hamster embryo fibroblasts with a single dose of 3-methylcholanthrene caused the activation of the transforming potential of cellular sequences (Notario et al, Oncogene 5: 1425-1430, 1990), which were subsequently isolated by cosmid rescue techniques, and further identified as a novel oncogene, termed cph because of its involvement in the carcinogenic progression of hamster embryo cells (Velasco et al, Oncogene 9: 2065-2069, 1994). We have analysed the expression of the cph proto-oncogene in adult Syrian hamster tissues by northern hybridization using cph-specific genomic probes. The three cph transcripts expressed in normal and neoplastic Syrian hamster embryo cells in culture (5.0, 3.5 and 2.0 kb) were also present in most adult tissues, although different mRNA species, most likely resulting from alternative splicing events, were expressed in testes. The highest steady-state level of cph mRNA was found in kidney, whereas cph expression was nearly undetectable in skin and skeletal muscle. Southern blot analyses of DNAs from other eucaryotic organisms were performed under moderate stringency conditions with a Syrian hamster-specific cph probe. Discrete cph-hybridizing sequences were present in genomes from yeast to mammalian species, including humans, thus demonstrating that cph is a highly conserved gene in eucaryotic evolution. Using fluorescence in situ hybridization (FISH), we have determined also the chromosomal localization of the cph proto-oncogene in the hamster genome. FISH experiments demonstrated that cph is a single copy gene, localized on the euchromatic short arm of the X chromosome, at region Xpa7. Because chromosome X is frequently involved in structural alterations in neoplastic Syrian hamster cells transformed by chemical carcinogens and oncogenic viruses, the localization of the cph locus on this chromosome supports the notion that the cph oncogene plays a role in the malignant conversion of chemically transformed hamster fibroblasts. The wide range of tissue-specific expression and species-specific distribution of cph strongly suggest that the normal function of the cph protein product(s) may be essential for metabolic processes involved in the regulation of cell proliferation and survival.

Animals↗
PubMed1996

Deletion and translocation involving chromosome 3 (p14) in two tumorigenic Kaposi's sarcoma cell lines.

BACKGROUND: Two neoplastic Kaposi's sarcoma (KS) cell lines, KS Y-1 (derived from a patient with KS associated with acquired immunodeficiency syndrome) and KS SLK (derived from an immunosuppressed patient with a renal transplant and KS or iatrogenic KS), have been shown to have abnormal chromosome constitution and to require no exogenous growth factors. They produce malignant tumors in immunodeficient mice. In contrast, all other cell cultures prepared in the past from KS specimens have shown to have normal diploid characteristics are hyperplastic, and depends on cytokines for growth, but they do not produce malignant tumors in immunodeficient mice. PURPOSE: We investigated whether the chromosomal changes that occurred in these KS cell lines were random contribute to the pathogenesis of KS. METHODS: We used the conventional G-banding technique and fluorescence in sti hybridization to identify structural and numerical chromosomal changes in the KS cell lines. RESULTS: We demonstrated that both cell lines are aneuploid and have some additional features in common, i.e., loss of copies of chromosomes 14 and 21 and nonrandom translocations and deletions in the short arm of chromosome 3 at region 3p14. These KS cell lines also exhibits loss of heterozygosity of loci at region 3p14-ter. CONCLUSION: This is the first time nonrandom chromosomal alterations have been described in KS neoplastic cells. On the basis of information available on other available on other cancers, the chromosome 3 alterations observed here can be expected to contribute to the neoplastic process in KS. IMPLICATIONS: Future research should focus on the identification cytogenetic markers, thus facilitating generation of specific molecular probes for detecting neoplastic cells early in the disease process.

Acquired Immunodeficiency Syndrome↗
PubMed1996

Expression of alternative forms of Ras exchange factors GRF and SOS1 in different human tissues and cell lines.

DNA probes and antibodies specific for different coding regions of human SOS1 and GRF genes were used to screen expression of these genes in a variety of adult and fetal human tissues and cell lines. Despite previous reports of the exclusive expression of hGRF RNA in brain, we also observed expression of this gene in various other tissues including lung and pancreas, as well as several tumor cell lines. At least three different hGRF mRNA transcripts were observed depending on the probe used, with the larger transcripts being detected by probes corresponding to the 5' end of the gene while smaller transcripts were detected by probes corresponding to the 3' end. Expression of hSOS1-related transcripts was more ubiquitous and homogeneous than with hGRF, with similar levels of specific transcripts being detected in most tissues and cell fines tested. Three to five different transcripts were detected in human tissues when using probes for the 5' end and middle regions of this gene, whereas only two were detected with probes corresponding to the 3' end. Screening of multiple human tumor cell lines showed ubiquitous expression of three specific transcripts, although the level and ratio of each of these transcripts varied widely among individual cell lines. Consistent with the variety of transcripts detected, several protein forms were also identified in Western immunoblots with antisera raised against specific domains of hSOS1 and human Ras-GRF gene products. Fluorescence in situ chromosomal hybridization suggested that, in both cases, the multiple forms arise from single chromosomal loci. The heterogeneity of hGRF and hSOS1 gene products detected (which appear to retain in most cases a functional catalytic domain), suggests that differentially expressed, alternatively spliced hSOS1 and hGRF forms may contribute to fine regulation of Ras activation in different tissues or at different stages of development.

3T3 Cells↗
PubMed1996

Papillomaviruses and potential copathogens.

An in vitro multistage genital epithelial cell model for cervical cancer that parallels the in vivo neoplastic process has been developed using recombinant human papillomavirus (HPV) DNA and genital cells. HPV-16-immortalized genital cells are responsive to the genotoxic action of known chemical carcinogens (polycyclic hydrocarbons, alkylating agents or cigarette smoke condensate), but are not converted to malignancy. Ras oncogene and human herpes virus-2 did convert HPV immortalized cells to malignancy, whereas human herpes virus-6 infection only increased HPV expression. Human immunodeficiency virus did not infect genital cells.

Carcinogens↗
PubMed1996

NAB2, a corepressor of NGFI-A (Egr-1) and Krox20, is induced by proliferative and differentiative stimuli.

Previous work had identified a corepressor, NAB1, which represses transcriptional activation mediated by NGFI-A (also known as Egr-1, zif268, and Krox24) and Krox20. These zinc finger transcription factors are encoded by immediate-early genes and have been implicated in a wide variety of proliferative and differentiative processes. We have isolated and characterized another corepressor, NAB2, which is highly related to NAB1 within two discrete domains. The first conserved domain of NAB2 mediates an interaction with the R1 domain of NGFI-A. NAB2 represses the activity of both NGFI-A and Krox20, and its expression is regulated by some of the same stimuli that induce NGFI-A expression, including serum stimulation of fibroblasts and nerve growth factor stimulation of PC12 cells. The human NAB2 gene has been localized to chromosome 12ql3.3-14.1, a region that is rearranged in several solid tumors, lipomas, uterine leiomyomata, and liposarcomas. Sequencing of the Caenorhabditis elegans genome has identified a gene that bears high homology to both NAB1 and NAB2, suggesting that NAB molecules fulfill an evolutionarily conserved role.

Amino Acid Sequence↗
PubMed1996

Localization of the human vascular endothelial growth factor gene, VEGF, at chromosome 6p12.

Using overlapping cosmids representing the vascular endothelial growth factor (VEGF) locus, the VEGF gene was mapped by fluorescence in situ hybridization to chromosome 6p12. This localization permits linkage analysis and the identification of gene interaction in the region, as well as alterations of the VEGF structure or expression in cancer cells with chromosome abnormalities.

Base Sequence↗
PubMed1996

Silencing and selective methylation of the normal topoisomerase I gene in camptothecin-resistant CEM/C2 human leukemia cells.

Camptothecin resistance of the human leukemia CEM/C2 cells is associated with a topoisomerase I (top1) mutation: Asn722Ser (Fujimori, A. et al. Cancer Res. 55:1339-1346; 1995). The corresponding DNA point mutation generates a novel site for the restriction endonuclease DdeI. We found that only the mutated top1 transcript was detectable in CEM/C2 by reverse transcriptase-polymerase chain reaction. Genomic DNA analysis by Southern blotting with DdeI showed that both the mutated and normal top1 genes were present in CEM/C2 cells. The mechanism of normal top1 allele silencing was further investigated. Cytogenetic analysis with a human chromosome 20 specific probe and restriction mapping by Southern blotting showed that both cell lines had a similar copy number of chromosome 20, with the predominant population containing 5-6 copies, and no detectable top1 gene rearrangement. Southern blotting using methylcytosine-sensitive restriction endonuclease (HpaII) indicated differential top1 methylation in CEM/C2 cells. Global cytosine methylation, however, appeared similar in CEM/C2 and wild-type CEM cells. These results indicate that gene-specific DNA methylation can play a role in downregulating top1 gene(s) and in the cellular resistance to camptothecins.

Antineoplastic Agents, Phytogenic↗
PubMed1995

CDKN2 in HPV-positive and HPV-negative cervical-carcinoma cell lines.

Human cervical cancers frequently contain retinoblastoma protein (Rb) that is inactivated by binding with human papilloma virus (HPV) E7 protein or through mutation. The CDKN2 gene encodes p16INK4 which inhibits cdk4-cyclin D phosphorylation of Rb, preventing the G1-S transition. To determine whether abnormalities of CDKN2 occur in cervical-cancer cells, II cervical cell lines, including 8 HPV-positive cell lines, 2 HPV-negative cell lines containing mutant Rb, and one tumorigenic cell line derived from normal cervical cells following transfection with HPV-16 and v-H-ras (CX16-2HR), were analyzed. No cell line had a homozygous deletion of exon 1 or 2 of CDKN2, and only one cell line, CX16-2HR, had an altered DNA sequence, which represents a common polymorphism at codon 148. To exclude the possibility of other subtle inactivating mutations, immunoblot analysis of protein lysates was performed using a polyclonal anti-p16INK4 rabbit anti-serum. Abundant levels of normal-sized p16INK4 were observed in all cell samples. Thus, no alterations of CDKN2 were detected in these cervical cell lines. These results confirm that mutational inactivation of p16INK4 is a rare event in tumor samples with compromised Rb activity.

Base Sequence↗
PubMed1995

Molecular cytogenetics of human papillomavirus-negative cervical carcinoma cell lines.

Cervical cancer is a malignancy closely associated with human papillomavirus (HPV). However, some cervical carcinomas occur in the absence of the virus. Two HPV-negative cervical carcinoma cell lines, C-33A and HT-3, were examined by G-banding and fluorescence in situ hybridization (FISH) using several whole chromosome and satellite probes. Combined conventional banding and FISH analysis significantly enhanced identification of complex and cryptic chromosome rearrangements. Common to both cell lines were translocations of chromosome 1, involving a specific site on the short arm and partial or complete loss of the short arm of chromosome 9, as well as loss of chromosome 13. The C-33A line had a relatively simple chromosomal constitution, with chromosome 1 being involved in translocations with chromosomes 9, 18, and 21. Karyotype reconstruction of G-banded and painted chromosomes indicate a net loss of DNA as a result of unbalanced translocations, which occurred only at site 9p24 and loss of one copy of chromosomes 13 and 14. In contrast, HT-3 cells had complex rearrangements and deletions of chromosomes 1p, 3p, 9p, 10q/p, 11p/q, and 17p, all regions with known tumor suppressor genes. The deletions observed in these HPV-negative cervical carcinomas will be important in delineating regions of tumor suppressor genes.

Carcinoma↗
PubMed1995

Localization of the human HER4/erbB-4 gene to chromosome 2.

The HER4/erbB-4 gene has been isolated as the fourth member of the human EGFR subfamily of tyrosine kinases and has been reported to encode a receptor for NDF/heregulin. In the present study we determined the chromosomal location of the HER4/erbB-4 gene within the human genome. Using human cDNA probes in fluorescence in situ hybridization (FISH), we mapped the HER4/erbB-4 gene to human chromosome 2q33.3-34. This finding established that also the HER4/erbB-4 gene is located in close vicinity of homeobox and collagen gene loci, as is the case for the related EGFR, erbB-2/neu and erbB-3. Aberrations of this chromosomal region associated with T cell leukemias and lymphomas as well as alveolar rhabdomyosarcomas raise the possibility that HER4/erbB-4 might be activated in these tumour types.

Chromosome Mapping↗
PubMed1995

Structural organization of the human sorbitol dehydrogenase gene (SORD).

The primary structure of human sorbitol dehydrogenase (SORD) was determined by cDNA and genomic cloning. The nucleotide sequence of the mRNA covers 2471 bp including an open reading frame that yields a protein of 356 amino acid residues. The gene structure of SORD spans approximately 30 kb divided into 9 exons and 8 introns. The gene was localized to chromosome 15q21.1 by in situ hybridization. Two transcription initiation sites were detected. Three Sp1 sites and a repetitive sequence (CAAA)5 were observed in the 5' noncoding region; no classical TATAA or CCAAT elements were found. The related alcohol dehydrogenases and zeta-crystallin have the same gene organization split by 8 introns, but no splice points coincide between SORD and these gene types. The deduced amino acid sequence of the SORD structure differs at a few positions from the directly determined protein sequence, suggesting allelic forms of the enzyme. High levels of SORD transcripts were observed in lens and kidney, as judged from Northern blot analysis.

Amino Acid Sequence↗
PubMed1995

Acquisition of multiple copies of a mutant topoisomerase IIalpha allele by chromosome 17 aneuploidy is associated with etoposide resistance in human melanoma cell lines.

Mutants of the human melanoma cell line, FEM-X, selected in multiple steps with VP- 16 (etoposide), are cross resistant to the epipodophyllotoxins and doxorubicin. Complementary DNA's for topoisomerase IIalpha were cloned from both FEM-X and FVP3, the most resistant mutant. Deletion of nucleotides 1320-1322 (or Ala429 from the resulting topoisomerase IIalpha protein) was unique to the cDNA from the drug resistant cell line. Expression of the mutant mRNA increases in parallel with VP-16 resistance in this series of cell lines. Restriction analysis and Southern analysis with allele-specific oligonucleotide probes were used to quantify the ratio of wild-type to mutant topoisomerase IIalpha alleles present in DNA amplified by PCR from both FEM-X and the drug resistant sublines. This analysis shows that in cell lines of increasing drug resistance, the number of mutant topoisomerase IIalpha alleles increases incrementally along with a concomitant decrease in the number of wild-type alleles. By quantitative Southern analysis of genomic DNA the total number of topoisomerase IIalpha alleles in FVP3 is approximately 2-fold that in the parental cells. Fluorescence in situ hybridization with a chromosome l7 paint reveals that amplification of the topoisomerase IIalpha locus in FVP3 correlates with an increase in the number of chromosome 17's, specifically the long arm. Cytogenetic analysis demonstrates that FEM-X contains three copies of chromosome 17, two of which are morphologically normal. During drug selection, FVP3 has gained 2-3 additional copies of the long arm of chromosome 17, the chromosomal location of the topoisomerase IIalpha locus. In this subline it is likely that three copies of the topoisomerase IIalpha gene are found on normal chromosome 17's and two on an isochromosome of the long arm of 17. By pulsed field gel electrophoresis, we were able to detect changes in the restriction pattern of the region of the long arm of chromosome 17 around the topoisomerase IIalpha locus that correlate with observed cytogenetic changes in FVP3. These results suggest that the acquisition of the mutant allele of topoisomerase IIalpha confers a selective advantage to cells in the presence of VP-16. As the drug concentration increased during the selection process, surviving sublines show preferential expression of the mutant topoisomerase IIalpha mRNA over that of the wild-type which is associated with a concomitant increase in the number of mutant topoisomerase IIalpha alleles.

Alleles↗
PubMed1995

Refined localization of the erbB-3 proto-oncogene by direct visualization of FISH signals on LUT-inverted and contrast-enhanced digital images of DAPI-banded chromosomes.

Contrast-enhanced, look-up-table (LUT)-inverted digital images of DAPI-banded chromosomes after fluorescence in situ hybridization (FISH) permit direct regional chromosomal localization of the fluorescent signals of single-copy gene probes. Improved quality and resolution of chromosome banding allowed a refined localization of the erbB-3 protooncogene from chromosome band 12q13 to sub-bands 12q13.2-13.3. This procedure can be used for direct and precise mapping of single-copy genes on both normal and cancer-cell-rearranged chromosomes.

Chromosome Banding↗