O.J. Miller

Suppression of human nucleolus organizer activity in mouse-human somatic hybrid cells☆

Abstract Cells from four different mouse-human somatic cell hybrids were stained with quinacrine to identify each metaphase chromosome and with ammoniacal silver by the Ag-AS method to locate nucleolus organizer regions. Each of the hybrids contained human acrocentric chromosomes. None of these human acrocentric chromosomes was stained with silver in any hybrid cell. Diploid cells were available from the human parent of one of the hybrids. In these cells both copies of nos. 13 and 15 stained with silver; the same chromosomes in the hybrid cell were not stained. These results support earlier reports that the expression of human ribosomal RNA (rRNA) genes is suppressed in mouse-human hybrid cells. Further, they suggest that silver staining by the Ag-AS method reflects activity of rRNA genes rather than just the presence of these genes.

Detection of nucleolus organizer regions in chromosomes of human, chimpanzee, gorilla, orangutan and gibbon

Nucleolus organizer regions were detected by the Ag-AS silver method in fixed metaphase chromosomes from human and primates. In the human, silver was deposited in the secondary constriction of a maximum of five pairs of acrocentric chromosomes: 13, 14, 15, 21 and 22. The chimpanzee also had five pairs of acrocentric chromosomes stained, corresponding to human numbers 13, 14, 18, 21 and 22. A gibbon had a single pair of chromosomes with a secondary constriction, which corresponded to the nucleolus organizer region. In each case the Ag-AS method detected the sites which have been shown by in situ hybridization to contain the ribosomal RNA genes. An orangutan had eight pairs of acrocentric chromosomes stained with Ag-AS, probably corresponding to human numbers 13, 14, 15, 18, 21 and 22, plus two others. Two gorillas had silver stain over two pairs of small acrocentric chromosomes and at the telomere of one chromosome 1. The larger gorilla acrocentric chromosomes had no silver stain although they all had secondary constrictions and entered into satellite associations.

Preferential derivation of abnormal human G-group-like chromosomes from chromosome 15

SummaryThe marked binding of antibodies specific for 5-methylcytidine to the short arm of chromosome 15 distinguishes this chromosome from the other human acrocentrics. This method has been used to study over 60 individuals including 12 who did not have Downs syndrome, but who did have an extra G-group sized acrocentric chromosome. In six cases the extra chromosome did not show intensive binding of anti-5-methylcytidine. In the other six cases, the extra chromosome contained a 5-methylcytidine rich band at each end indicating that both ends were derived from chromosome 15 and contained centromeric heterochromatin normally present on the short arm of chromosome 15. The duplication of short arm material in the abnormal chromosomes was confirmed in all cases by quinacrine staining, nucleolar organizer (Ag-AS) staining or C-banding. In three cases, the abnormal chromosome appeared to arise from two different chromosomes 15. Several possible mechanisms for the production of the abnormal chromosome are discussed. The individuals with this abnormal chromosome all showed some degree of mental retardation, but few common physical findings.

Human Thymidine Kinase Gene Locus: Assignment to Chromosome 17 in a Hybrid of Man and Mouse Cells

The human chromosome retained in a hybrid clone derived from human cells and a moluse line deficiemt in thymidine kinase has the quinacrinefluorescence pattern characteristic of chromosome 17.

Quinacrine fluorescent karyotypes of human diploid and heteroploid cell lines

Quinacrine-fluorescence karyotypes were prepared on a series of human cell lines. WI-38 karyotypes were indistinguishable from those obtained from cultured XX blood leukocytes. WI-L2 lymphoblastoid ce

The use of antinucleoside antibodies to probe the organization of chromosomes denatured by ultraviolet irradiation

Abstract Ultraviolet irradiation of methanol: acetic acid-fixed human and mouse metaphase chromosomes rendered them capable of binding antibodies specific for purine or pyrimidine bases. Since these antibodies react with single-stranded but not with native DNA, our results indicate that UV irradiation generated single-stranded regions in chromosomal DNA. Using an indirect immuno-fluorescence technique to detect antibody binding, highly characteristic, nonrandom patterns of antibody binding were observed. Antibodies to adenosine (anti-A) and thymidine (anti-T) produced identical patterns of binding which in most respects matched the chromosome banding patterns produced by quinacrine. However, additional foci of intense fluorescence were seen in the paracentromeric regions of constitutive heterochromatin on chromosomes 1, 9 and 16, regions which had been shown by in situ DNA-RNA hybridization to be the locations of AT-rich human satellite DNA. Antibodies to cytidine also bound to the same region of chromosome 9. In mouse chromosome preparations, both anti-A and anti-T produced bright fluorescence of the region containing centromeric heterochromatin, which had been shown to be the location of the AT-rich satellite DNA of this species.

Familial translocation involving chromosome 6, 14 and 20, identified by quinacrine fluorescence

SummaryA 4 year old girl with physical and mental retardation but few other abnormalities was found to have an unbalanced karyotype, 47,XX,6-,t(6q,20p?)+,t(14q,6q)+, resulting in partial trisomy-14. This arose by aberrant segregation of chromosomes during meiosis in her mother, who has a complex translocation involving chromosomes 6, 14 and 20.

5-Methylcytosine in heterochromatic regions of chromosomes: chimpanzee and gorilla compared to the human

Fixed metaphase chromosomes of gorilla and chimpanzee were UV-irradiated to produce regions of single-stranded DNA and then treated with antibodies specific for the minor DNA base 5-methylcytosme (5 MeC). An indirect immunofluorescence technique was used to visualize sites of antibody binding. In the gorilla six pairs of autosomes contained major fluorescent regions, indicating localized regions of highly methylated DNA. These corresponded, with the exception of chromosome 19, to the major regions of constitutive heterochromatin as seen by C-banding. The Y chromosome also contained a highly fluorescent region which was located just proximal to the intense Q-band region. In the chimpanzee no comparable concentrations of highly methylated DNA were seen. Smaller regions of intense 5 MeC binding were present on perhaps six chimpanzee chromosomes, including the Y. Five of these corresponded to chromosomes which were highly methylated in the gorilla. — There is diversity among the human, gorilla and chimpanzee in both the size and location of concentrations of 5 MeC, supporting the idea that satellite DNA evolves more rapidly than DNA in the remainder of the chromosome.

Human chromosome structure as revealed by an immunoperoxidase staining procedure

Abstract An indirect immunoperoxidase technique has been used to visualize specific chromosome banding patterns produced by antinucleoside antisera. The patterns were observed with ultraviolet-irradiated chromosomes which were subsequently exposed to antibodies specific for 5-methylcytosine. Binding occurred mainly in the regions of constitutive heterochromatin on chromosomes 1, 9, and 16, on the short arm of chromosome 15 and the midportion of the Y. The patterns were comparable to those observed by indirect immunofluorescence, but the immunoperoxidase technique appears to give greater resolution and sensitivity.

Chromosome markers in Mus musculus: Differences in C-banding between the subspecies M. m. musculus and M. m. molossinus

Quinacrine (Q-band) and centromeric heterochromatin (C-band) patterns of metaphase chromosomes of two subspecies of Mus musculus were compared. M. m. musculus (the laboratory mouse) and M. m. molossinus (a subspecies from Southeast Asia) had similar Q-band patterns along the length of the chromosomes, but differences were observed in the centromeric region of some chromosomes. The two subspecies had very different distributions of C-band material. Antibodies to 5-methylcytosine were bound to regions of the chromosome corresponding to the C-bands in each animal. These findings support the idea that satellite DNA, which is concentrated in the C-band region, changes more quickly than bulk DNA. The interfertility of these two subspecies permits the development of a musculus strain carrying normal marker chromosomes for genetic studies.