Genetic mapping of thymine DNA glycosylase (Tdg) gene and of one pseudogene in the mouse.
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Animals↗
Biomedical subjects
M A Pierotti
Publications and source records attributed to M A Pierotti.
Genetic mapping of a pulmonary adenoma resistance (Par1) in mouse.
Lung cancer, a major cause of death in the Western world, has a poor prognosis. So far, therapeutic strategies have had only a limited effect. Lung cancer risk is strongly associated with cigarette smoking and lung cancer pedigrees are rare. However, a possible polygenic nature of inherited predisposition to this cancer has been envisaged. Mouse inbred strains with inherited predisposition and resistance to lung cancer provide an important tool for the dissection of the genetics of this complex disease. The A/J strain carries the pulmonary adenoma susceptibility 1 (Pas1) locus and develops many lung tumours. We have mapped the M. spretus-derived locus that strongly resists the lung tumorigenesis in Pas1/+ mice. This locus, pulmonary adenoma resistance 1 (Par1) maps to mouse chromosome 11, near the Rara locus, with a lod score of 5.3. In Pas1/+ mice Par1 accounts for 23% of the phenotypic variance and 10 fold reduction in total tumour volume. These results provide evidence for a major resistance locus affecting the expression of an inherited predisposition to lung cancer.
Adenoma↗
p53 activity and chemotherapy.
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Animals↗
A polygenic model of inherited predisposition to cancer.
Polygenic inheritance of predisposition to cancer is demonstrated in experimental animals for different tumor types. Genetic susceptibility to hepatocarcinogenesis, lung tumorigenesis, skin and intestine carcinogenesis, and plasmacytomagenesis is determined by inheritance of multiple cancer predisposition and resistance alleles, whose chromosomal locations have been found by genetic linkage analysis. In some of these experimental models, genetic heterogeneity has also been reported. In humans, increased risk of lung cancer associated with multiple genes coding for drug metabolizing enzymes, increased risk of cancer in relatives of cancer patients, and genetic heterogeneity are compatible with polygenic inheritance of cancer predisposition. Polygenic inheritance based on the combination of multiple alleles that give predisposition and resistance to cancer would predict a very high risk of cancer in carrier individuals and a marginal increase in the relative risk of cancer in the progeny of the cancer patients. Therefore, predisposition to cancer may be genetically determined even in the absence of familial clustering of cases.
Animals↗
The full oncogenic activity of Ret/ptc2 depends on tyrosine 539, a docking site for phospholipase Cgamma.
RET/PTC oncogenes, generated by chromosomal rearrangements in papillary thyroid carcinomas, are constitutively activated versions of proto-RET, a gene coding for a receptor-type tyrosine kinase (TK) whose ligand is still unknown. RET/PTCs encode fusion proteins in which proto-RET TK and C-terminal domains are fused to different donor genes. The respective Ret/ptc oncoproteins display constitutive TK activity and tyrosine phosphorylation. We found that Ret/ptcs associate with and phosphorylate the SH2-containing transducer phospholipase Cgamma (PLCgamma). Two putative PLCgamma docking sites, Tyr-505 and Tyr-539, have been identified on Ret/ptc2 by competition experiments using phosphorylated peptides modelled on Ret sequence. Transfection experiments and biochemical analysis using Tyr-->Phe mutants of Ret/ptc2 allowed us to rule out Tyr-505 and to identify Tyr-539 as a functional PLCgamma docking site in vivo. Moreover, kinetic measurements showed that Tyr-539 is able to mediate high-affinity interaction with PLCgamma. Mutation of Tyr-539 resulted in a drastically reduced oncogenic activity of Ret/ptc2 on NIH 3T3 cells (75 to 90% reduction) both in vitro and in vivo, which correlates with impaired ability of Ret/ptc2 to activate PLCgamma. In conclusion, this paper demonstrates that Tyr-539 of Ret/ptc2 (Tyr-761 on the proto-RET product) is an essential docking site for the full transforming potential of the oncogene. In addition, the present data identify PLCgamma as a downstream effector of Ret/ptcs and suggest that this transducing molecule could play a crucial role in neoplastic signalling triggered by Ret/ptc oncoproteins.
3T3 Cells↗
Rapid differential diagnosis of myxoid liposarcoma by fluorescence in situ hybridisation on cytological preparations.
In two cases of suspected myxoid liposarcoma, where chromosomal metaphase preparations were not available, fluorescence in situ hybridisation was performed on interphase nuclei of cytological preparations for the detection of the specific translocation, t(12;16), characteristic of this tumour and of trisomy 8, which is the most frequent secondary chromosome aberration. Probes directed against chromosomes 12 and 16 and against the centromeres of chromosomes 12 and 8 were hybridised on cell brushings and cytocentrifuge preparations. The finding of three painting domains of both chromosomes 12 and 16 and of only two signals with the centromeric probe directed against chromosome 12, suggested the presence of t(12;16) in both cases. In one case trisomy 8 was inferred from the occurrence of three centromere 8 signals. This approach can be used to detect specific chromosomal abnormalities when an urgent differential diagnosis is requested or when chromosome preparations are not available, or both.
Journal Article↗
Age-related activation of the tyrosine kinase receptor protooncogenes RET and NTRK1 in papillary thyroid carcinoma.
Oncogenic rearrangements of RET and NTRK1 proto-oncogenes are frequently detected in papillary thyroid carcinomas. Several studies have shown an association between ionizing radiation and development of this tumor type. In addition in vitro irradiation of tumor cell lines induced rearrangements of RET similar to those observed in human papillary thyroid carcinomas. These two observations could be related to the reported increased incidence of papillary thyroid carcinomas in children living in contaminated areas around Chernobyl, given that it has been demonstrated that about 60% of them presents a RET oncogenic activation. However, this high frequency of RET positivity in radiation exposed children does not rule out the possibility that age could also play a role in the development of RET positive tumors. To assess this possibility we looked for a relationship between the presence of RET and NTRK1 oncogenic rearrangements and age at surgery in a sample of 92 consecutive patients. Our results show that, in papillary thyroid carcinoma, the frequency of RET and NTRK1 activation is significantly higher in the group of patients aged 4-30 years, thas supporting the concept that age could be contributing to this thyroid specific carcinogenic process.
3T3 Cells↗
Polygenic inheritance of predisposition to lung cancer.
Inherited predisposition to lung tumor development appears to be a complex genetic trait, in humans as well as in experimental animals. In humans, tobacco smoking represents the main risk factor for lung tumors. Familial clusterings of lung cancer cases are rare, although affected sib pairs are reported. However, several studies have reported an increased risk of lung cancer in first degree relatives of lung cancer patients, as compared with appropriate controls. Inheritence of particular alleles of genes or phenotypes for drug metabolizing enzymes are also associated with an increased lung cancer risk. In experimental models, murine inbred strains with high genetic predisposition to lung cancer are available, as well as resistant strains. A major locus affecting inherited predisposition to lung cancer in mice has been mapped on chromosome 6, near the Kras2 gene. The locus, pulmonary adenoma susceptibility 1 (Pas1) derives from the A/J strain. Additional "minor" loci have also been mapped. Dominant lung tumor resistance loci have also demonstrated in the mouse genome. Therefore, evidence for polygenic inheritance of predisposition to lung cancer is demonstrated in the mouse model, and it is strongly suggested for human lung cancer.
Adenocarcinoma↗
Tyrosines outside the kinase core and dimerization are required for the mitogenic activity of RET/ptc2.
Defects in the c-ret proto-oncogene, a member of the protein tyrosine kinase receptor family, have recently been linked to two types of genetic syndromes, Hirschsprung's disease and the multiple endocrine neoplasia family of inherited cancers. RET/ptc2 is the product of a papillary thyroid carcinoma translocation event between the genes coding for c-ret and the type I alpha regulatory subunit of protein kinase A (RI alpha) (Lanzi, C., Borrello, M., Bongarzone, I., Migliazza, A., Fusco, A., Grieco, M., Santoro, M., Gambetta, R., Zunino, F., Della Porta, G., and Pierotti, M. (1992) Oncogene 7, 2189-2194). The resulting 596-residue protein contains the first two-thirds of RI alpha and the entire tyrosine kinase domain of c-ret (RETtk). An in vivo assay of growth stimulatory effects was developed, which consisted of microinjecting a RET/ptc2 expression plasmid into the nuclei of 10T1/2 mouse fibroblasts and observing the incorporation of 5-bromodeoxyuridine. This assay was used to determine that only the dimerization domain of RI alpha fused to RETtk is required for RET/ptc2's mitogenic activity. In addition, all of the reported Hirschsprung's disease point mutations in the RETtk (S289P, R421Q, and R496G) inactivate RET/ptc2 in our assay, confirming that these are loss of function mutations. Two tyrosines outside the conserved kinase core were also identified that are essential for full mitogenic activity of RET/ptc2. These two tyrosines, Tyr-350 and Tyr-586, are potential sites for Src homology 2 and phosphotyrosine binding domain interactions.
Animals↗
Different susceptibility to lung tumorigenesis in mice with an identical Kras2 intron 2.
The A/J mouse strain is genetically susceptible to pulmonary tumorigenesis. We have performed a genetic linkage analysis to map pulmonary adenoma susceptibility (Pas) loci in an urethane-treated (A/J x Mus spretus) x C57BL/6J (ASB) interspecific testcross. In this interspecific cross we have confirmed our previous results in AC3F2 mice on the mapping of the Pas1 locus to the distal region of chromosome 6, near Kras2 (Nature Genetics 3: 132-136, 1993). The A/J and M. spretus strains differed at the Pas1 locus, with the M. spretus providing the resistant allele. In the latter strain, we studied the nucleotide sequence of a portion of the second intron of Kras2 that contains polymorphisms associated with lung tumor susceptibility in several inbred strains. The lung tumor-resistant M. spretus strain had the same specific nucleotide sequence of susceptible strains. Mutations in codon 61 of Kras2 in urethane-induced lung tumors from ASF1 hybrids involved the A/J allele in all cases, while the M. spretus allele was never affected. Our results indicate that the M. spretus and A/J mice have an identical structure of the second intron of the Kras2 gene, but they differ in genetic susceptibility to pulmonary tumorigenesis and in mutability of their Kras2 allele.
Analysis of Variance↗
A sequence analysis of the genomic regions involved in the rearrangements between TPM3 and NTRK1 genes producing TRK oncogenes in papillary thyroid carcinomas.
Papillary thyroid carcinomas have frequently been found to display oncogenic rearrangements of the NTRK1 gene, which encodes the high-affinity nerve growth factor receptor. Replacement of its extracellular domain by sequences coding for the 221 amino-terminal residues of the TPM3 gene was responsible for the oncogenic NTRK1 activation in three of eight of these tumors. In all of them, the illegitimate recombination involved the 611-bp NTRK1 intron placed upstream of the transmembrane domain and the TPM3 intron located between exons 7 and 8. Therefore, due to the splicing mechanism, all of the TPM3/NTRK1 gene fusions encoded an invariable transcript and the same chimeric protein of 70 kDa, which was constitutively phosphorylated on tyrosine. In two of the three tumors the simultaneous presence of the reciprocal products of the TPM3/NTRK1 recombination, 5'TPM3-3'NTRK1 and 5'NTRK1-3'TPM3 sequences, respectively, and the previously demonstrated localization of both genes on the long arm of chromosome 1 lead us to suggest that an intrachromosomal inversion could be responsible for their recombination. In an attempt to understand the molecular basis that predisposes NTRK1 and TPM3 genes to be a recurrent target of illegitimate recombination, we have determined the nucleotide sequence around the breakpoints of the recombination products in all three patients as well as those of the corresponding regions from the normal TPM3 and NTRK1 genes. In these regions, a search for common features usually involved in illegitimate recombination in mammalian cells revealed the presence of some recombinogenic elements as well as pal-indromes, direct and inverted repeats, and Alu family sequences.
3T3 Cells↗
Allelotyping in Wilms tumors identifies a putative third tumor suppressor gene on chromosome 11.
An analysis of loss of heterozygosity for markers on both the short and the long arm of chromosome 11 was performed in 24 sporadic Wilms tumors. Six cases (25%) showed allelic losses involving the entire chromosome. In one case (4%) the loss was restricted solely to the WT1 gene on band p13. Two cases (8%) displayed allelic losses for WT1 and for markers on band p15.5, where the putative tumor suppressor gene WT2 has been mapped, but retained heterozygosity for markers on the long arm. In three tumors (13%) the loss of heterozygosity involved markers mapped to chromosomal regions p15.5 and q23.3-qter, but did not affect WT1 and markers on q12-q13. Altogether, the proportion of cases showing allelic losses at the distal region of 11q (37%) was comparable to that of cases with LOH affecting the WT1 (37%) or the WT2 (46%) loci, thus suggesting the existence of a third chromosome 11 tumor suppressor gene involved in the pathogenesis of Wilms tumors.
Alleles↗
Ring 22 chromosomes in dermatofibrosarcoma protuberans are low-level amplifiers of chromosome 17 and 22 sequences.
Ring chromosomes have been found with some regularity as solid tumors have come increasingly under cytogenetic study. The full genetic content and significance of these rings remain unclear. Dermatofibrosarcoma protuberans, a tumor of the deep dermis, consistently has supernumerary ring chromosomes, sometimes as the sole detectable cytogenetic change. Using a modified method for comparative genomic hybridization and fluorescent in situ hybridization with a panel of various probes, we found that these ring chromosomes consistently contain the chromosome 22 centromere along with interstitial sequences from chromosomes 17 and 22, specifically from regions 17q23-24 and 22q11-12. The ring chromosomes in dermatofibrosarcoma protuberans are vehicles for a particular pattern of relatively low-level genomic amplification of selected sequences.
Adult↗
The human archain gene, ARCN1, has highly conserved homologs in rice and Drosophila.
A novel human gene, ARCN1, has been identified in chromosome band 11q23.3. It maps approximately 50 kb telomeric to MLL, a gene that is disrupted in a number of leukemia-associated translocation chromosomes. cDNA clones representing ARCN1 hybridize to 4-kb mRNA species present in all tissues tested. Sequencing of cDNAs suggests that at least two forms of mRNA with alternative 5' ends are present within the cell. The mRNA with the longest open reading frame gives rise to a protein of 57 kDa. Although the sequence reported is novel, remarkable similarity is observed with two predicted protein sequences from partial DNA sequences generated by rice (Oryza sativa) and fruit fly (Drosophila melanogaster) genome projects. The degree of sequence conservation is comparable to that observed for highly conserved structural proteins, such as heat shock protein HSP70, and is greater than that of gamma-tubulin and heat shock protein HSP60. A more distant relationship to the group of clathrin-associated proteins suggests a possible role in vesicle structure or trafficking. In view of its ancient pedigree and a potential involvement in cellular architecture, we propose that the ARCN1 protein be named archain.
Amino Acid Sequence↗
Regulation of apoptosis induced by the retinoid N-(4-hydroxyphenyl) retinamide and effect of deregulated bcl-2.
The cancer chemopreventive retinoid N-(4-hydroxyphenyl)-all-trans retinamide (HPR) was recently shown by us to have antiproliferative and apoptotic effects on human leukemic cell lines, including those unresponsive to all-trans retinoic acid (ATRA). We have now characterized further the process of HPR-induced cell death. We report that inhibitors of RNA transcription and of protein synthesis, activators of protein kinase C (PKC), inhibitors of tyrosine kinases, Zn++, and the antioxidants acetylcysteine, ascorbic acid, alpha-tocopherol, and deferoxamine suppressed HPR-induced apoptosis. HL60 cells induced toward monocytic differentiation by 1,25 dihydroxyvitamin-D3 [1,25(OH)2D3], but not those induced toward the granulocytic differentiation by ATRA, showed reduced responses to HPR. The transport of HPR by cells with different sensitivity to the retinoid, however, was similar, even after treatment with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA), which induces unresponsiveness to HPR. The expression of the apoptosis-related genes bcl-2, p53, and c-myc was examined to determine their role in HPR-triggered cell death. The levels of bcl-2 mRNA were markedly diminished by 24 hours of HPR treatment in all cell lines except in the relatively HPR-insensitive line K422. However, probably because of its long half-life, bcl-2 protein levels were either unchanged or only slightly decreased. Downregulation of p53 mRNA was also observed within 24 hours of HPR exposure in NB4 but not K422 cells, but no changes in the amount of p53 protein were found. Suppression of c-myc transcription was observed in all cells except K422. The protective role of bcl-2 on cell death by HPR was investigated in HL60 as well as 697 pre-B leukemia and Jurkat T-acute lymphocytic leukemia (T-ALL) cells constitutively expressing high levels of bcl-2 proteins due to gene transfer manipulation. Compared with control cells, the onset of apoptosis in these cells with deregulated bcl-2 production was delayed by at least 24 hours. These findings establish that cell death by HPR requires RNA transcription and protein synthesis and is regulated by the activation of PKC. Although changes in bcl-2, p53, and c-myc expression are found in cells treated with HPR, the time-course of these events suggests that HPR-triggered apoptosis is not directly controlled by these genes. Finally, while ectopic overexpression of bcl-2 does not protect cells from death by HPR, it markedly delays its onset.(ABSTRACT TRUNCATED AT 400 WORDS)
Anticarcinogenic Agents↗
Involvement of chromosomes 17 and 22 in dermatofibrosarcoma protuberans.
Literature on the cytogenetics of dermatofibrosarcoma protuberans (DFSP) is limited; only 10 cases with chromosome aberrations have been reported. They are karyotypically characterized by the presence of supernumerary ring(s), either as the sole cytogenetic abnormality or together with a few additional structural or numerical changes. We report the cytogenetic and fluorescence in situ hybridization (FISH) analysis of three new DFSP, one primary and two recurrent tumors. In two cases we found a supernumerary ring as the sole change, whereas the third had two copies of a marker chromosome and monosomy of chromosome 22. Sequences of chromosomes 17 and 22 were identified by FISH in the supernumerary rings and in the markers. The fluorescence pattern suggested that additional sequences were present in the two rings, but showed that the marker chromosomes were entirely painted by chromosome 17 and 22 probes. The findings indicate that juxtaposition and/or amplification of chromosome 17 and 22 sequences could be crucial in the pathogenesis of DFSP.
Adult↗
Analysis of loss of heterozygosity in murine hepatocellular tumors.
Because allelotype analysis of tumors has been important in the identification of new tumor suppressor genes, here we studied loss of heterozygosity (LOH) in a well-defined animal experimental system. We analyzed spontaneous liver tumors from C3HHc x C57BL/6J (B6C3F1) mice and urethane-induced hepatocellular tumors from (C3H/He x Mus spretus) x C57BL/6JBy (HSB) interspecific mice. A total of 95 different genetic markers were tested: 13 in 24 B6C3F1 tumors, 76 in 58 HSB tumors, and six in both groups. Minisatellite finger-printing analysis detected one case of LOH and less than 1% genomic rearrangements in polymorphic and nonpolymorphic bands, respectively. There were no changes at hepatocellular susceptibility loci or at markers homologous to loci frequently lost in human hepatocellular carcinomas. Therefore, our results suggest that LOH and genomic rearrangements are uncommon in mouse hepatocellular tumors.
Alleles↗
Genetic mapping and expression analysis of the murine DNA ligase I gene.
We mapped the murine DNA ligase I gene (Lig1) in the mouse genome by using a mapping panel from an interspecific cross. Lig1 mapped to a centromeric part of chromosome 7, a region homologous to human chromosome 19q, where the human homologue LIG1 was localized. In addition, Lig1 expression was analyzed during the course of mouse liver-cell regeneration induced by partial hepatectomy, necrogenic doses of carbon tetrachloride, or the mitogen 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene. The results demonstrate that Lig1 is expressed in the liver during active cell proliferation.
Animals↗