A Microphotometric Study of the Syntheses of Desoxyribonucleic Acid and Nuclear Histone

The incorporation of thymidine-H 8 and lysine-H 3 into human leukocyte chromosomes was studied in order to determine the temporal relationships between the syntheses of chromosomal deoxyribonucleic acid and chromosomal protein. The labeled compounds were incorporated into nuclei of interphase cells. Label from both precursors became apparent over the chromosomes of dividing cells. Incorporation of thymidine-H 3 occurred during a restricted period of midinterphasc (S) which was preceded by a nonsynthetic period (G1) and followed by a nonsynthetic period (G2). Incorporation of lysine-H 3 into chromosomal protein occurred throughout interphase. Grain counts made over chromosomes of dividing cells revealed that the rate of incorporation of lysine-H~ into chromosomal protein differed during various periods of interphase. The rate of incorporation was diminished during GI. During early S period the rate of incorporation increased, reaching a peak in late S. The high rate continued into G2. Thymidine-H 3 incorporated into DNA was distributed to mitotic chromosomes of daughter cells in a manner which has been referred to as a "semiconservative segregation." No such semi-conservative mechanism was found to affect the distribution of lysine-H 3 to the mitotic chromosomes of daughter cells. Therefore, it is concluded that synthesis of chromosomal protein and its distribution to chromosomes of daughter cells are not directly influenced by synthesis and distribution of the chromosomal DNA with which the protein is associated.The relationships between duplication of deoxyribonucleic acid (DNA) and syntheses or aggregation of various nuclear protein fractions have been studied in different tissues. The synthesis of DNA occurs during a restricted period of midinterphase referred to as S. The S period is preceded by a postmitotic G1 (gap1) period and followed by a premitotic G~ (gap~) period during which synthesis of DNA does not occur (24).Synthesis of nuclear protein has been studied by means of microspectrophotometry (1, 6, 19, 32, 56), interferometry (30, 45), radioautography (15, 23, 51, 56), and isotope tracer techniques (12,22, 54). The results of such experiments indicate that syntheses of various nuclear protein fractions occur during different stages of the cell cycle.The proteins associated with the mitotic chromosomes have not been identified. It is clear, however, that chromosomes of dividing cells contain proteins (9). Although the protein of the chromosomes is not identifiable in chemical terms, it is demonstrable morphologically. Chromosomal protein may be visualized in chromosomes of mitotic cells by means of histochemical methods (28), or by radioautographic localization of la-

Section: Discussionmentioning

confidence: 99%

“…Synthesis of nuclear protein has been studied by means of microspectrophotometry (1,6,19,32,56), interferometry (30,45), radioautography (15,23,51,56), and isotope tracer techniques (12,22,54). The results of such experiments indicate that syntheses of various nuclear protein fractions occur during different stages of the cell cycle.…”

mentioning

confidence: 99%

Incorporation of Tritium-Labeled Thymidine and Lysine Into Chromosomes of Cultured Human Leukocytes

Cave

1966

“…Such analyses of DNA, using Feulgen and methyl green staining and ultraviolet absorption, and of histone, by employing fast green under alkaline conditions, have demonstrated a doubling in amounts of both substances during an interphase period prior to cell division (44,47,19,34,17,8,2). Furthermore the increase in these substances occurs simultaneously during this period (8,2). Relatively little is known, however, of possible qualitative changes in the chromosomal components and their interassociations during the different stages of the cell cycle.…”

Section: Introductionmentioning

confidence: 99%

“…A number of investigations of the chromosomal components, desoxyribonucleic acid (DNA) and histone, suggest that both substances occur in relatively constant amounts in non-proliferating ceils of diploid tissues (10,30,44,12,4,8,2). While in the case of DNA this constancy per genome appears to hold for all such ceils within a species, differences in amounts and composition of histone among various tissues of the same organism (43,4,13) suggest that a histone constancy may hold only for cells of the same type.…”

Section: Introductionmentioning

confidence: 99%

Evidence of Differences in the Desoxyribonucleoprotein Complex of Rapidly Proliferating and Non-Dividing Cells

Bloch

Godman

1955

Self Cite

1. Quantitative cytophotometric analysis of the interphase cells of a rapidly proliferating differentiated tissue such as liver of new born rat, indicates that these cells can be separated into two groups on the basis of their staining characteristics after methanol fixation. 2. These groups are thought to correspond to two stages of interphase. The first, called "autosynthetic interphase," comprises cells which are duplicating chromosomal material in preparation for mitosis, and shows parallel increases in the methyl green and Feulgen staining of DNA and the fast green staining of histone from the diploid (2 C) to double these values (4 C). 3. The second group is designated the "heterosynthetic interphase," during which the cell ceases proliferating and functions in a manner commensurate with its state of differentiation. In this stage Feulgen staining indicates the diploid chromosomal complement, but there is a decreased capacity of the DNA to bind methyl green and of the histone to bind fast green. 4. The difference between the methyl green binding of the heterosynthetic and autosynthetic 2 C cells is due to the presence of a protein in the former which presumably inhibits staining by competing with the dye for binding sites on the DNA. The effect of this inhibition can be removed by extracting the protein, or by blocking the protein basic groups. 5. The decreased fast green staining of histone in the heterosynthetic cells can be minimized by prolonged fixation with formaldehyde. It is thought to stem either from a similar type of inhibition, or from an increased susceptibility of the histone to loss from the cell during this stage. 6. While histone inhibits methyl green staining of DNA in all cells, the differences between the staining properties of the autosynthetic and heterosynthetic interphase cells are believed to be due to another protein, whose properties appear similar to those of the chromosomal "residual protein." It is concluded that a complex of DNA and residual protein existing during the heterosynthetic interphase is dissociated during the autosynthetic interphase.

“…Histochemical and biochemical studies of DNA and histone indicate that these two substances are bound together by ionic bonds (15) and are in constant proportion to each other (1,2,26). Bloch and Godman (2) have suggested that this proportionality is maintained by simultaneous proportional synthesis of histone and DNA.…”

Section: Introductionmentioning

confidence: 99%

SIMULTANEOUS SYNTHESIS OF HISTONE AND DNA IN SYNCHRONOUSLY DIVIDING TETRAHYMENA PYRIFORMIS

Hardin

Einem

Lindsay

1967

Histone and DNA syntheses have been studied in synchronously dividing Tetrahymena pyriformis GL. During the heat treatment necessary to synchronize cultures of this amicronucleate protozoan, the DNA content of the already polyploid macronucleus increases. When the cells begin synchronous division, their DNA content is reduced in a stepwise process which is closely paralleled by reduction of macronuclear histone content. During cell division, the contents of DNA and histone decrease by slightly more than twofold, and in the subsequent S phase, DNA and histone increase simultaneously to 85 % of the values expected if all chromosomes were to double. The first step in the process of reduction of DNA and histone contents is their decrease in excess of twofold, and this is accomplished by removal of extrusion bodies from the nuclei of dividing cells. The second step is a mechanism which allows, in effect, only 70 % of the chromatin in the average nucleus to duplicate. Such partial duplication suggests that both histone and DNA syntheses in synchronous Tetrahymena depend upon a regulatory mechanism, the mediating elements of which are localized in only certain chromosomes.