Electron-microscope observations on the organization of the nucleus in chicken erythrocytes and a superunit thread hypothesis for chromosome structure.
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Haplopappus gracilis (n = 2), Haplopappus ravenii (n = 4), and Haplopappus wigginsii (n = 4) are isolated by F1 hybrid sterility due mainly to translocation heterozygosity. There is no evidence that this can be overcome at the diploid level so that introgression can occur among them. They are also separated geographically, but occasional populations of H. gracilis and H. ravenii may be brought together along roadways to form sterile hybrids. There were no statistically significant differences in nuclear DNA content among the same or structurally different aneuploid n = 2 and n = 3 chromosome races or ecotypes of H. gracilis. Some of the H. gracilis races were not significantly different from one race of the ancestral H. ravenii, and these samples of both species were from plants growing on poor soils in contrast to accessions from normal habitats. How much and which classes of DNA in these species are subject to changes induced by environmental effects is not known. There were no correlations between DNA amounts and altitude, latitude, and longitude. H. wigginsii had a greater amount of DNA per nucleus than either H. ravenii or H. gracilis, and its increased DNA content may reflect a more rapid accumulation of noncoding sequences due to facultative self-compatibility not found in the other two species.
As visualized by in situ hybridization with fluorescence detection, newly amplified CAD genes in 10(5) cell colonies are contained in multiple copies of very large regions of DNA, each tens of megabases long. The extra DNA is usually linked to the short arm of chromosome B9, which retains CAD at its normal site. The widely spaced genes are often interspersed with new G-negative regions. Individual cells within a clone have highly variable numbers of CAD genes (range 2-15). When resistant clones are examined later, at the 10(15) cell stage, the amplified genes are usually found in much more condensed structures. We propose that, in the initial event of CAD gene amplification, much of the short arm is transferred from one B9 chromosome to another. In subsequent cell cycles this initial duplication expands rapidly through unequal but homologous sister chromatid exchanges. Relatively rare secondary events lead to more condensed structures.
Gastric cancer is of major importance world-wide representing the second most common cause of cancer related death in the world. Data concerning the chromosomal changes in gastric cancer are limited and no specific change has been identified to date. We have studied cytogenetically 15 cases of primary gastric cancer by direct culture of tumors cells and G-banding technique. We focused on structural aberrations observed in order to identify non-random aberrations presenting common chromosomal breakpoints. Chromosomes most commonly involved were according to frequency 1,11,14,7,17,6,8 and 13. Chromosome 11 was involved as add(11)(p15), while the pericentromeric area of chromosome 14 was constantly participated in aberrations. Isochromosomes i(1q), i(8q), i(13q), i(14q) and i(17q) were constantly found. Furthermore translocations t(1;7), t(7;14), t(6;17) and t(5;14) were identified. Conventional cytogenetics continues to be valuable in cancer study detecting genomic areas potentially candidate for the isolation of genes related to carcinogenesis.
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We examined the genetic, morphological, and molecular effects of position effect variegation in Drosophila, and the effects of mutations that either suppress [Su(var)] or enhance [E(var)] this phenomenon. All eight Su(var) mutations examined strongly suppress the inactivation of variegating alleles of the genes white [In(l) wm4], brown [In(2R) bwVDe2] and Stubble [T(2; 3) SbV]. The E(var) mutation enhances variegation of these loci. The chromosomal region 3C-E (26 bands) which includes the white locus is usually packaged as heterochromatin in salivary glands of the variegating strain wm4. Addition of any of the Su(var) mutations restores a more euchromatic morphology to this region. In situ hybridization to polytene chromosomes and DNA blot analyses of gene copy number demonstrate that the DNA of the w+ gene is less accessible to its probe in the variegating wm4 strain than it is in the wild-type or variegation-suppressed strains. Blot analysis of larval salivary gland DNA indicates that the white gene copy number does not vary among the strains. Hence, the differences in binding of the w+ gene probe in the variegating and variegation-suppressed strains reflect differences in chromosomal packaging rather than alterations in gene number. The effects of variegation and the Su(var) mutations on chromatin structure were analyzed further by DNAse I digestion and DNA blot hybridization. In contrast to their dramatic effects on chromosomal morphology and gene expression, the Su(var) mutations had negligible effects on nuclease sensitivity of the white gene chromatin. We suggest that the changes in gene expression resulting from position effect variegation and the action of the Su(var) mutations involve alterations in chromosomal packaging.
The effect of caffeine (2mM) on the frequency of structural mutations induced by UV light (lambda = 265 nm at an incident dose of 40 erg/mm2) in the primary culture of mouse embryonic fibroblasts is studied. A half-hour treatment with caffeine of cells at the time of the first mitosis metaphase decreased approximately by 2 times the frequency of chromosome aberrations induced by UV light at the S stage and observed at the metaphase of this or the next C-mitosis. The frequency of both breaks and exchanges decreased as the result of caffeine treatment. The persistence of the protective effect of caffeine at the time of the second C-mitosis suggests that the observed decrease of the aberration rate is accompanied by the true reparation of pre-mutational lesions in chromosomes; the nature of the reparative process and the time when it takes place is as yet not clear. Caffeine did not decrease the frequency of spontaneous structural mutations.
cDNA of mitochondrial glycerophosphate dehydrogenase (mGPDH), a defect of which is a possible cause of non-insulin dependent diabetes mellitus, was cloned from a human insulinoma cDNA library. The deduced amino acid sequence showed 91% and 92% homology with those of rat and mouse mGPDH, respectively. The mGPDH gene was mapped to chromosome 2q23 by FISH analysis. Genomic clones for mGPDH were then isolated using mouse mGPDH cDNA and PCR products of human mGPDH cDNA as probes. Genomic structure was studied by sequencing the exon-intron boundaries and by PCR amplification of intronic regions using genomic clones as templates. The human mGPDH gene was shown to be composed of 15 coding exons, containing a (CA)n repeat region inside the gene, which was not polymorphic in the Japanese population. Genomic cloning also identified a pseudogene located on chromosome 19q13.4. These results provide information useful for analyzing the mGPDH gene in patients with non-insulin dependent diabetes mellitus.
HES-1 is a mammalian helix-loop-helix factor structurally related to the Drosophila hairy and Enhancer of split proteins. It binds more preferentially to the N box (CACNAG) than to the E box (CANNTG) and acts as a negative regulator. In this study, we have isolated and characterized the mouse HES-1 gene. This gene consists of four exons, and the positions of introns are well conserved when compared with those of the Drosophila hairy gene, except for the third intron. Southern blot and interspecies backcross analyses suggest that the mouse HES-1 gene is a single-copy gene and is located around position 26 on chromosome 16. The transcription initiation site, determined by the S1 nuclease and primer extension experiments, is located 31 nucleotides downstream of a TATA box. In the 5'-regulatory region, there are four N box sequences, and the DNase I foot-printing and gel mobility shift analyses show that HES-1 binds to these sequences. Transient transfection assays using C3H10T1/2 cells suggest that there are several positive regulatory regions in the HES-1 gene. However, cotransfection of the HES-1 expression vector leads to approximately 40-fold repression in promoter activity. Furthermore, when the N box sequences are disrupted, this negative regulation is severely impaired. These results raise the possibility that HES-1 gene expression may be negatively autoregulated through the N box sequences.
Results of fluorescence in situ hybridization (FISH) of Bos taurus and B. indicus Y chromosomes using the bovine locus-specific Y probes BC1.2 and lambda ES6.0 and region-specific probes of B. indicus and B. taurus Y chromosomes, which were generated by microdissection and DOP-PCR, indicate that the Y chromosomes of B. indicus (BIN Y) and B. taurus (BTA Y) differ by a pericentric inversion. Parts of the short and long arms of the Y chromosome in B. taurus and the distal half of the Y chromosome in B. indicus were microdissected, amplified by DOP-PCR, biotinylated, and rehybridized in situ to the corresponding metaphase chromosomes to test the chromosome fragment specificity of the DNA probes. The region-specific painting probes were used for hybridization to metaphase chromosomes of the other species. The DNA painting probes BTA Yp12 and BTA Yq12.1-ter derived from BTA Y hybridized to the distal and proximal halves of BIN Y, respectively. Complex hybridization signals on BTA Yq12.1-->qter were generated with the DNA probe BIN Yqcen-centr (centromere-central) after FISH. The results demonstrate that BTA Yp is homologous to the distal half of BIN Y and that BTA Yq corresponds to the proximal part of BIN Yq. Hybridization of the Y chromosome-specific DNA probes lambda ES6.0 to BTA Yp12-->p11 and near to the telomere of BIN Y and BC1.2 to BTA Yq12-->q13 and to the telomere of BIN Y indicate an opposite orientation of the homologous chromosome fragments BTA Yp and of the distal half of BIN Yq.