The nature and mechanisms of chromosome banding.
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Chromosome Banding↗
Biomedical subjects
A T Sumner
Publications and source records attributed to A T Sumner.
The role of proteins in the production of different types of chromosome bands.
Experiments have been carried out to try and answer two questions on the role of proteins in chromosome banding: firstly, what degree of protein extraction is required before banding can be produced; and secondly, to what extent are redistribution and reorganization of chromosomal components required for the production of banding. Partial extraction of all histones, and of a group of non-histones with molecular weights mainly between 50,000 and 70,000 appears to be necessary before G-, C- or R-banding can be produced. More extensive 'dehistonization' to produce chromosome scaffolds inhibits the production of all types of bands. Protein-protein and protein-DNA cross-linking inhibits all types of banding tested, the degree of inhibition being roughly related to the degree of cross-linking, but not apparently to the type of cross-linking. The results of both sets of experiments indicate that chromosome banding of all types is dependent on the prior loss from chromosomes of a specific set of proteins, and on some alteration of the arrangement of remaining chromosomal components during the banding procedure.
Chromosomal Proteins, Non-Histone↗
Application of X-ray microanalysis to the study of histochemical staining reactions.
The application of X-ray microanalysis to the study of histochemical staining reactions is reviewed. Advantages of the technique are the ability to examine colourless intermediate reaction products, and to study the efficiency of their conversion to the final reaction product. The number of X-rays emitted by an element is, under appropriate conditions, proportional to the amount of that element. Errors such as glare and deviations from Beer's Law in microdensitometry and quenching of fluorescence in microfluorimetry, are avoided. The possibility of analysing several elements simultaneously is a further advantage, as the amount of reaction product can be related to the quantity of some other cellular constituent. The application of X-ray microanalysis to the study of the binding of the fluorochrome quinacrine to nuclei is described. The extinction of this fluorochrome is too low for it to be measured by microdensitometry. X-ray microanalysis shows that its fluorescence is not proportional to the amount of quinacrine bound, and thus fluorescence cannot be used for this study. Quinacrine was shown to be strongly bound to nuclei, and the strength of its affinity is consistent with intercalative binding.
Adenosine Triphosphatases↗
The distribution of quinacrine on chromosomes as determined by X-ray microanalysis. I. Q-bands on CHO chromosomes.
The distribution of quinacrine in relation to Q-banding on CHO chromosomes has been investigated using X-ray microanalysis. Technical problems involved in this type of experiment were studied in detail. It was necessary to use a solution of quinacrine acetate in acetic acid to ensure that the only chlorine detectable in quinacrine-stained chromosomes was in the quinacrine molecule. Electron irradiation during analysis rapidly destroys quinacrine fluorescence, but the chlorine is not lost from the chromosomes, and there are several reasons for supporting that a reliable distribution of quinacrine on the chromosome can be obtained by the method. - Small variations along the chromosome in the amounts of chlorine (representing quinacrine) and of phosphorus (mainly DNA) occur. The distribution patterns for chlorine and phosphorous show a good resemblance to each other for each homologous chromosome; quinacrine fluorescence patterns (Q-bands) do not resemble chlorine distribution patterns, however. The results of this study therefore support the view that Q-bands result from the differential quenching of fluorescence along chromosomes to which the quinacrine is essentially uniformly bound, and do not reflect differential binding of quinacrine along the chromosome.
Animals↗
Protein-depleted chromosomes. I. Structure of isolated protein-depleted chromosomes.
Protein-depleted isolated Chinese hamster chromosomes have been obtained by different protein extraction procedures and examined by electron microscopy and SDS-polyacrylamide gel electrophoresis. Salt-resistant centromeric and telomeric structures are visible in protein-depleted chromosomes and the protein-depleted chromosomes appear to have a regular, longitudinal pattern in critical point dried preparations. The scaffold-like structure of protein-depleted chromosomes is highly affected by the ionic strength and composition of the extraction medium and by the spreading conditions. Nucleosomal histones of isolated chromosomes proved to be more sensitive to the sodium chloride treatment than histones of isolated chromatin. A small, but constant quantity of core histones was detected in 2 M salt extracted chromosomes and H3 and H4 histones of isolated chromosomes appeared to be resistant to the sodium deoxycholate treatment.
Animals↗
Protein-depleted chromosomes. II. Experiments concerning the reality of chromosome scaffolds.
Chromosome scaffolds are chromosome-shaped bodies, composed of non-histone proteins, which remain when the histones are extracted from chromosomes. Because of the well-known tendency of chromosomal proteins to aggregate, we have tested the possibility that chromosome scaffolds might be produced by aggregation of proteins during the preparation of scaffolds. Extraction of histones in the presence of sucrose, which inhibits aggregation, results in a much looser structure lacking the characteristic appearance of a scaffold, although sucrose does not extract any extra proteins. Extraction of histones from chromosomes in situ on EM grids produced only a network of fine fibres without a scaffold-like structure, while digestion of DNA from typical chromosome scaffolds in situ fell only discrete particles of protein and not a continuous structure. We conclude, therefore, that the typical appearance of chromosome scaffolds produced by histone extraction may well represent an artefact resulting from protein aggregation. Our experiments suggest further that DNA, as well as protein, is a structural component of whatever type of core structure is responsible for maintaining the form of chromosome.
Animals↗
Preparation and assessment of frozen-hydrated sections of mammalian tissue for electron microscopy and X-ray microprobe analysis.
A system is described for preparing and examining frozen-hydrated sections of mammalian tissue, using commercially available equipment. An essential part of the system is a liquid nitrogen cooled vacuum transfer system. Minor modifications were required to interface the various manufacturers' equipment, and also to facilitate handling of specimens in the cryoultramicrotome. A variety of criteria indicates that our sections are truly frozen-hydrated, but negligible image contrast in the sections is a serious problem both for morphological work and X-ray microanalysis.
Animals↗
The nature of chromosome bands and their significance for cancer research.
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Chromosome Banding↗
Dye binding mechanisms in G-banding of chromosomes.
The mechanisms whereby Giemsa is bound to chromosomes to produce G-banding patterns have been studied. The magenta colour produced in chromosomes by Giemsa staining appears to be due to the same 2:1 thiazine-eosin compound that precipitates from Giemsa solutions. This precipitate is formed in chromosomes in regions in which DNA phosphates occur at the correct distance apart to bind two thiazine molecules, which are subsequently able to bind to the same eosin molecule. Banding appears to be produced as a result of this precipitation occurring preferentially in hydrophobic regions of chromosomes.
Animals↗
Predicting treatment costs and life expectancy for end-stage renal disease.
To estimate the cumulative 10-year direct medical costs and life expectancy associated with different methods of treatment for end-stage renal disease, we assessed predictively three treatment transition options. It is predicted that if 1000 patients shift from facility to home dialysis for each of 10 years, life expectancy of the cohort will not be reduced, but there will be a reduction of $241 million in total costs. The same number shifting from facility dialysis to cadaveric transplantation are predicted to have a $279 to $330 million reduction in total costs but a reduction of 7 to 17 per cent in life expectancy. Shifting from home dialysis to transplantation is predicted to reduce total costs by +103 to $142 million, and life expectancy by 10 to 20 per cent. As new program policies for treatment of end-stage renal disease are developed, their effect on both costs and life expectancy needs to be considered.
Cadaver↗
Changes in elemental composition of human chromosomes during a G-banding (ASG) and a C-banding (BSG) procedure.
Human chromosomes fixed in methanol-acetic acid have been examined by X-ray microanalysis, before, during and after a G-banding and a C-banding procedure. Phosphorus (representing mainly DNA), sulphur and calcium are the most prominent elements in untreated chromosomes. In the G-banding procedure, the calcium is lost during 2 x SSC treatment. In the C-banding procedure, calcium is lost in the preliminary HCl treatment. During the following barium hydroxide treatment a large amount of barium becomes attached to the chromosomes, but is lost again during the subsequent 2 x SSC treatment. In both banding techniques Giemsa staining produces large peaks for sulphur (thiazine dyes) and bromine (eosin), showing that both types of dyes are involved in the staining. Reduction in the phosphorus peak during these procedures may be partly due to extraction of DNA and other chromosomal components, but could also be due to absorption of phosphorus X-rays by heavy elements (barium and bromine).
Chromosomes, Human↗
Quantitation in biological X-ray microanalysis, with particular reference to histochemistry.
Thin specimens consisting of various light and heavy elements in gelatine have been subjected to X-ray microanalysis to determine the relationship between the number of X-ray counts for a specific element expressed as a percentage of the continuum (the percentage counts) and the concentration of that element. For light elements, the relationship between the percentage counts and concentration is strictly linear. For heavier elements, the relationship is not linear, because of the increase of the continuum counts with (formula: see text). If a correction is made for the effect of (formula:see text), heavy elements also show a linear relationship between percentage counts and concentration. Within the limits of atomic number (Z = 56) and concentration (approximately 10%) studied here, it is shown that when X-ray microanalysis is carried out on bulk specimens consisting of various elements in gelatine, the relationship between X-ray counts and concentration for a particular element is linear. The problems in quantitation of the results of X-ray microanalysis caused by exogenous continuum and mass loss induced by irradiation are discussed. It is pointed out that when X-ray microanalysis is used to study histochemical and other staining procedures, allowance must also be made for the reduction in concentration of other elements in the specimen as a result of the addition of the stain to the specimen.
Chromosomes, Human↗
Suppression of quinacrine banding of human chromosomes by mounting in organic media.
When A-banded human chromosomes mounted in water are transferred to an organic mounting medium, the chromosomes show uniform bright quinacrine fluorescence. This change is reversible. It is inferred that quinacrine is bound uniformly along the chromosomes, and that Q-banding is a consequence of a non-uniform distribution along the chromosomes of chemical groups, probably proteinaceous, which affect the fluorescence efficiency of the bound quinacrine.
Chemical Phenomena↗
Estimation of the sizes of polymorphic C-bands in man by measurement of DNA content of whole chromosomes.
A method is proposed for estimating the sizes of polymorphic C-bands of human chromosomes by microdensitometric measurement of the DNA content of whole chromosomes. The method requires measurements of: a chromosome known not to by polymorphic, as a standard against which to normalize all measurements; the polymorphic chromosomes of interest; and a homologous polymorphic chromosome lacking the C-band. In an example studies here, the C-band of chromosome 9 was estimated to contain about 0.0296 pg of DNA.
Chromosomes, Human↗
Relative DNA contents of somatic nuclei of ox, sheep and goat.
Diploid ox nuclei contain about 14% more DNA than nuclei from sheep of the same sex. Goat nuclei have a similar DNA content to those of sheep. In view of the similar chromosome banding patterns in these species, it appears that chromosome evolution must have involved numerous minute interstitial deletions of additions of DNA. Although chromosomes which have similar banding patterns in these three species may be regarded as homologous in this respect, and can be regarded as having a common evolutionary origin, they are not homologous for the quantity of their DNA.
Animals↗
A difference in dry mass between the heads of X- and Y-bearing human spermatozoa.
An integrating microinterferometer was used to measure the dry mass of sperm heads. The dry mass was found to be proportional to DNA content, and thus provides a useful method of estimating sperm DNA content. Using this technique we have confirmed that human spermatozoa which show none and one quinacrine-fluorescent spot are X- and Y-bearing respectively. However, the measurements suggest that many of the spermatozoa with two quinacrine-fluorescent spots are not YY-bearing, as previously thought, but might be incompletely condensed Y-bearing spermatozoa.
DNA↗
Involvement of protein disulphides and sulphydryls in chromosome banding.
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Chlorides↗
Chromosome homology and heterochromatin in goat, sheep and ox studied by banding techniques.
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Animals↗