Chromosome 17p13.2 transfer reverts transformation phenotypes and fas‐mediated apoptosis in breast epithelial cells
Mohamed H. LareefQuivo TahinJoon Ho SongIrma H. RussoDana MihailaCarolyn SlaterBinaifer BalsaraJoseph R. TestaDominique BroccoliJennifer V. GrobelnyGil MorAndrew CuthbertJosé Russo
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Abstract Transformation of the human breast epithelial cells (HBEC) MCF‐10F with the carcinogen benz(a)pyrene (BP) into BP1‐E cells resulted in the loss of the chromosome 17 p13.2 locus (D17S796 marker) and formation of colonies in agar‐methocel (colony efficiency (CE)), loss of ductulogenic capacity in collagen matrix, and resistance to anti‐Fas monoclonal antibody (Mab)‐induced apoptosis. For testing the role of that specific region of chromosome 17 in the expression of transformation phenotypes, we transferred chromosome 17 from mouse fibroblast donors to BP1‐E cells. Chromosome 11 was used as negative control. After G418 selection, nine clones each were randomly selected from BP1‐E‐11 neo and BP1‐E‐17 neo hybrids, respectively, and tested for the presence of the donor chromosomes by fluorescent in situ hybridization and polymerase chain reaction‐based restriction fragment length polymorphism (PCR‐RFLP) analyses. Sensitivity to Fas Mab–induced apoptosis and evaluation of transformation phenotype expression were tested in MCF‐10F, BP1‐E, and nine BP1‐E‐11 neo and BP1‐E‐17 neo clones each. Six BP1‐E‐17 neo clones exhibited a reversion of transformation phenotypes and a dose dependent sensitivity to Fas Mab‐induced apoptosis, behaving similarly to MCF‐10F cells. All BP1‐E‐11 neo , and three BP1‐E‐17 neo cell clones, like BP1‐E cells, retained a high CE, loss of ductulogenic capacity, and were resistant to all Fas Mab doses tested. Genomic analysis revealed that those six BP1‐E‐17 neo clones that were Fas‐sensitive and reverted their transformed phenotypes had retained the 17p13.2 (D17S796 marker) region, whereas it was absent in all resistant clones, indicating that the expression of transformation phenotypes and the sensitivity of the cells to Fas‐mediated apoptosis were under the control of genes located in this region. © 2004 Wiley‐Liss, Inc.Cite
Craniosynostosis, the abnormal development of the calvarial sutures, occurs as an autosomal dominant trait in many clinically distinct syndromes. We performed linkage analysis of a provisionally novel type of autosomal dominant craniosynostosis in a large three generational family. Linkage was established between the craniosynostotic locus and D5S211, a locus defined by the short tandem repeat polymorphism (STRP) marker Mfd 154 in distal 5q. The maximum LOD score, Zmax, was 4.8 at a recombination fraction of zero. No significant linkage was found with markers located 30 cM and more proximal to D5S211. The findings assign the craniosynostotic locus in this family to a telomeric region in the long arm of chromosome 5. Linkage analysis with Mfd 154 in other autosomal dominant craniosynostotic syndromes should reveal whether these disorders are caused by mutations of genes at the same or at different loci.
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Recombination Fraction
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ABSTRACT We isolated, in E. coli K12, new alleles of the ilvB locus, the structural gene for acetolactate synthase isoenzyme I, and showed them to map at or near the ilvB619 site. The map position of the ilvB locus was redetermined because plasmids containing the ilvC-cya portion of the chromosome did not complement mutations at the ilvB locus. Furthermore, diploids for the ilvEDAC genes formed with these plasmids in an ilvHI background facilitated the isolation of the new ilvB alleles. The ilvB locus was remapped and found to be located at 81.5 minutes, between the uhp and dnaA loci. This location was determined by two- and three-point transductional crosses, deletion mapping and complementation with newly isolated plasmids. One of the new alleles of the ilvB gene is a mu-1 insertion. When present in the donor strain, this allele interferes with the linkage of genes flanking the mu-1 insertion, as well as the linkage of genes to either side of the mu-1 insertion.
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Abstract In Prunus, the self-incompatibility (S-) locus region is <70 kb. Two genes—the S-RNase, which encodes the functional female recognition component, and the SFB gene, which encodes the pollen recognition component—must co-evolve as a genetic unit to maintain functional incompatibility. Therefore, recombination must be severely repressed at the S-locus. Levels of recombination at genes in the vicinity of the S-locus have not yet been rigorously tested; thus it is unknown whether recombination is also severely repressed at these loci. In this work, we looked at variability levels and patterns at the Prunus spinosa SLFL1 gene, which is physically close to the S-RNase gene. Our results suggest that the recombination level increases near the SLFL1 coding region. These findings are discussed in the context of theoretical models predicting an effect of linked weakly deleterious mutations on the relatedness of S-locus specificities. Moreover, we show that SLFL1 belongs to a gene family of at least five functional genes and that SLFL1 pseudogenes are frequently found in the S-locus region.
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Abstract The A mating type locus of Coprinus cinereus determines mating compatibility by regulating essential steps in sexual development. Each A locus contains several genes separated into two functionally independent complexes termed Aα and Aβ, and the multiple alleles of these genes generate an estimated 160 A mating specificities. The genes encode two classes of homeodomain-containing proteins designated HD1 and HD2. In this report we describe two newly cloned loci, A2 and A5, and compare them with A42, A43 and A6 that we have described previously. An Aβ-null locus, retaining just a single active HD1 gene from the α-complex, was generated by mutation. Using this as a transformation host, gene combinations that promote A-regulated development were identified. We demonstrate that each A locus contains members of three paralogous pairs of HD1 and HD2 genes. Different allelic versions of gene pairs are compatible but paralogous genes are incompatible. The genes present in four uncloned A loci were deduced using Southern analyses and transformations with available cloned genes. The combined analysis of nine A factors identifies sufficient A gene alleles to generate at least 72 A mating specificities.
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The mating-type information residing at the HML and HMR loci in Saccharomyces cerevisiae is kept unexpressed by the action of at least four MAR (or SIR) loci. To determine possible interactions between the MAR/SIR gene products and to find new regulatory loci, we sought extragenic suppressors of the mar1-1 mutation. A strain with the genotype HMLa MAT alpha HMRa mar1-1 is unable to mate because of the simultaneous expression of a and alpha information. A mutant of this strain was isolated that exhibits an alpha phenotype and, therefore, presumably fails to express the HML and HMR loci. We designate the new locus SUM1 (suppressor of mar). The mutation is recessive, centromere unlinked and does not correspond to the MAT, HML, HMR, SIR1, MAR1, MAR2 (SIR3) or SIR4 loci. The sum1 mutation affects expression of both a and alpha information at the HM loci. Suppression by sum1-1 is neither allele specific nor locus specific as it suppresses a deletion mutation of the MAR1 locus and mutations in SIR3 and SIR4. The sum1-1 mutation has no discernible phenotype in a Mar+ strain. We propose that the MAR/SIR gene products negatively regulate the SUM1 locus, the gene product of which is necessary for expression of the HM loci.
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ABSTRACT Fifty seven mutations at the ade3 locus have been crossed to ochre, amber and ochre-amber suppressors. 70% (39/56) of the mutations at this locus are nonsense mutations; 61% (34/56) are ochre mutations and 9% (5/56) are amber mutations. The frequency of nonsense mutations among ade3 alleles recovered is very high and raises the interesting possibility that only polar mutations at this locus are recovered. An hypothesis to explain these genetical findings as well as physiological properties of these mutations is proposed.
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Cosmid
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The mouse Ulnaless locus is a semidominant mutation which displays defects in patterning along the proximal-distal and anterior-posterior axes of all four limbs. The first Ulnaless homozygotes have been generated, and they display a similar, though slightly more severe, limb phenotype than the heterozygotes. To create a refined genetic map of the Ulnaless region using molecular markers, four backcrosses segregating Ulnaless were established. A 0.4-cM interval containing the Ulnaless locus has been defined on mouse chromosome 2, which has identified Ulnaless as a possible allele of a Hoxd cluster gene(s). With this genetic map as a framework, a physical map of the Ulnaless region has been completed. Yeast artificial chromosomes covering this region have been isolated and ordered into a 2 Mb contig. Therefore, the region that must contain the Ulnaless locus has been defined and cloned, which will be invaluable for the identification of the molecular nature of the Ulnaless mutation.
Yeast artificial chromosome
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