Phosphorylation Of Nonhistone Chromosomal Proteins Research Articles

A proposed model of cancer as the inappropriate expression of non-body introns

An a posteriori proposal is made that cancer represents an alien non-body phenotype erupting from “silent” gene groups within actively coding regions of a normal cell's genome. Relevant empirical data is garnered from the literature and twelve premises are derived from the data. On the basis of these premises it is hypothesized that the malignant neoplastic phenotype results from interference with non-histone chromosomal proteins causing retention and expression in a body cell of non-body introns. These are asserted to be the same introns which, as exons in a trophoblast cell, direct the normal development of the fetal placenta. Development of a malignant tumor is presented as a pathological recapitulation of the development of a normal placenta. Unrestrained tumor growth is attributed to the inability of endogenous body chalones to suppress the non-body gene groups coding for neoplastic mitosis, while invasion and metastasis result from failure of the non-body genes which code for implantation and mitosis to switch off at the proper time. The hypothesis asserts that all carcinogenic agents alter phosphorylation of non-histone chromosomal proteins, that cancer and normal trophoblast cells are genetically programmed to travel through the body's vascular system to effect immunosuppression in lymphoid tissues, that malignant neoplastic tissue will regress when exposed to trophoblast-based chalones, and that mammals immunized against trophoblast-based antigen will be resistant to the development of malignant tumors.

Chromosomal proteins of conifers

AbstractDuring imbibition and germination of jack pine seeds, the composition of the total extractable chromatin varied. Relative to DNA, the histone levels decreased as the nonhistone chromosomal proteins (NHCP) increased. New chromosomal proteins were synthesized after 2 days of imbibition as judged by recovery of 14C‐amino acids from the major protein fractions. Phosphorylation of histones from 32P‐phosphoric acid was detected before the incorporation of 14C‐amino acids. In the seed the synthesis and relative changes of chromatin coincided with a fall in total soluble protein and free arginine N. By contrast, adenylate energy charge, free glutamine N and in vitro template activity of chromatin increased during chromatin protein synthesis.When seeds had germinated for 4 days after the start of imbibition more radioactivity, derived from free 14C‐amino acids, was recovered from the NHCP than from the histones. The percentage amino acid composition of most histone fractions remained stable, whereas the composition of NHCP changed more with time. The phosphorylation of NHCP was 8‐ to 41‐fold greater than that of the histones. Phosphorylation of histone H4 was not detected at any stage of germination. Correlations between recovery of radioactivity (32P and 14C) from chromosomal proteins and higher adenylate energy charge were positive.

Phosphorylation and acetylation of nonhistone chromosomal proteins of the brain of rats of various ages and their modulation by calcium and estradiol

In vitro phosphorylation and acetylation of nonhistone chromosomal (NHC) proteins and their modulation by Ca ++ and estradiol were studied by incubating slices of cerebral cortex of 2-, 15- and 84-week female rats with 32Pi and 14C-Na-acetate. Phosphorylation pattern of NHC proteins is unique for each age. Ca ++ and estradiol stimulate phosphorylation of different NHC proteins which is also age-specific. Acetylation of NHC proteins decreases precipitously with age. No unique NHC protein is acetylated preferentially at any age, nor does Ca ++ stimulate acetylation. Estradiol, however, stimulates acetylation of a few NHC proteins. It is suggested that phosphorylation of NHC proteins and its modulation by effectors may be more important for gene expression than their acetylation.

Chapter 8. Assessment Of Methylation In Nonhistone Chromosomal Proteins

Covalent modification of proteins has been implicated in some regulatory role(s) in terms of cellular activity. The covalent modification of proteins should be independent of polypeptide synthesis. Studies of phosphorylation of non-histone chromosomal proteins (NHCPs) have been extensively investigated in an attempt to explore the cause–effect relationship in gene regulation. One report indicates that methylation of NHCP might correlate with DNA replication in salivary gland by use of autoradiographic studies. Others demonstrate the incorporation of [methyl- 14 C]methionine into acid-insoluble nuclear proteins after 4 days of consecutive injection in rat. Although extensive studies of methylation in histones are well demonstrated, the role(s) of this postsynthetic modification of NHCP has yet to be elucidated. These results together with the established procedures developed for the investigation of chromosomal proteins may provide some information for further exploration of the structure and function of NHCP.

Chapter 11. Phosphorylation of Nonhistone Chromosomal Proteins by Nuclear Phosphokinases Covalently Linked to Agarose

Publisher Summary Phosphorylation of nuclear proteins has been implicated in the control of gene expression. With increasing interest in nuclear protein phosphorylation, the availability of agarose-coupled protein kinase activity has become desirable. The method of attaching phosphoprotein kinase activity to the commercially available activated agarose, Affi Gel 10, as described in this chapter is an extremely effective procedure. The lack of harsh chemicals or extreme pH and the quick coupling time are extremely important for retaining activity of the rather sensitive nuclear protein kinases. By using partially purified nuclear phosphoproteins instead of total nonhistone chromosomal proteins for the source of nuclear protein kinase activity, the resulting agarose–protein kinase column is relatively free of protease or phosphatase activity. Moreover, a small amount of protein leakage occurs. Hence, nuclear protein kinases attached to agarose offer valuable tools for the study of nuclear protein phosphorylation with numerous applications.

Stimulation of uterine nonhistone protein phosphorylation and nuclear protein kinase activity by estradiol-17β

Changes in the phosphorylation of nonhistone chromosomal proteins have been followed in rat uterus stimulated by 17β-estradiol. Isolated uteri were found to incorporate 32P i into nonhistone proteins via an endogenous nuclear protein kinase reaction. The rate of 32P labeling of nonhistone proteins and the activity of nuclear protein kinase(s) were found to be elevated over three- and two-fold respectively in uteri obtained from ovariectomized animals treated with estrogen. A dramatic change was observed in the radioactivity profile of 32P-labeled proteins fractionated via sodium dodecyl sulfate-polyacrylamide gel electrophoresis. These observations are compatable with the hypothesis that phosphorylation of nonhistone proteins plays a role in the regulation of gene activity in the uterus.

Phosphorylation of nonhistone chromosomal proteins early during the prereplicative phase of the cell cycle of WI-38 human diploid fibroblasts

Stimulation of confluent monolayers of nondividing WI-38 cells to proliferate is accompanied by a doubling in the rate of phosphorylation of nonhistone chromosomal proteins during the first hour. The use of cycloheximide to inhibit protein synthesis does not appear to affect phosphorylation of the nonhistones. Nondividing and serum-stimulated cells exhibit qualitative differences in the phosphorylation and dephosphorylation of individual polypeptide peaks resolved by polyacrylamide gel electrophoresis. These results are consistent with other evidence implicating nonhistone protein phosphorylation as an early event involved in the activation of transcription occurring during cell proliferation.

Gene activation during muscle differentiation and the role of nonhistone chromosomal protein phosphorylation

The phosphorylation of nuclear proteins, in a standard assay with nuclei isolated from chick muscle cell cultures, increases steadily with time in culture. This increase is shown to reflect increased nuclear protein kinase activity. Most of the phosphorylated protein is chromatin bound and so are the enzymes responsible for the phosphorylation. Analysis of chromosomal proteins on hydroxylapatite columns shows that 32P is incorporated almost entirely into nonhistone proteins, while the specific radioactivity of the histones is low and does not increase during cell culture. Experiments in which cells were grown at different initial densities and, consequently, underwent cell fusion at different times show that phosphorylation is dependent neither on cell density nor on the degree of cell fusion. Chick thigh fibroblasts, prepared by repeated subculture of fibroblasts from muscle cell preparations to eliminate myogenic cells, do not show the increase in phosphorylation with culture time, suggesting that the changes may be specific for primary myoblast cultures. These changes are not shown by thigh muscle nuclei isolated during development in the whole embryo. In fact, nuclear protein phosphorylation activity decreases rapidly in ovo with increasing embryonic age. Possible roles for phosphorylation of nonhistone chromosomal proteins in gene activation during myogenesis are discussed in the light of these observations.

Phosphorylation of nonhistone chromosomal proteins in dog kidney cells infected with infectious canine hepatitis virus

Infectious canine hepatitis virus infection of dog kidney cells altered the metabolism of the nonhistone chromosomal proteins. The rate of incorporation of [ 32P]phosphate into the nonhistone chromosomal proteins began to increase between 6 and 7 hr after infection, and the specific activity of the proteins labeled between 8 and 9 hr after infection was 195% of that observed in the mock-infected control. Analysis by sodium dodecyl sulfate polyacrylamide gel electrophoresis showed that phosphorylation of the nonhistone chromosomal proteins was increased after infection especially in some particular components of the proteins. Alteration of the metabolism of the nonhistone chromosomal proteins correlated with an increase of RNA synthesis and template activity of chromatin in infected cells.

Properties of the genome in normal and SV-40 transformed WI-38 human diploid fibroblasts. III. Turnover of nonhistone chromosomal proteins and their phosphate groups

The turnover of nonhistone chromosomal proteins and their phosphate groups was compared in normal and in SV-40 virus transformed WI-38 human diploid fibroblasts. Cells were pulse labelled with tryptophan- 3H and 32P for 30 minutes and the specific activities of tryptophan- 3H and 32P in the various molecular weight classes of nonhistone chromosomal proteins were determined during the first four hours following termination of labelling. While a rapid turnover of high molecular weight nonhistone polypeptides (142, 000 to 200, 000 Daltons) is evident after one hour in SV_40 transformed cells, the specific activities of these nonhistone chromosomal polypeptides are not significantly decreased in normal cells. In contrast, a rapid turnover of low molecular weight (30, 000 to 51, 000 Daltons) nonhistone chromosomal proteins occurs during the first hour in normal WI-38 cells with no corresponding decrease in the specific activities of these proteins in SV-40 transformed cells. There is no apparent net turnover of phosphate groups on nonhistone chromosomal proteins in either normal or SV-40 transformed cells four hours following pulse labelling. Rather, during the first four hours significant fluctuations are observed in the 32P specific activities of defined molecular weight fractions. Taken together with previous reports of differences in the composition, synthesis and phosphorylation of nonhistone chromosomal proteins in normal and SV-40 transformed human diploid cells the present results further indicate the complex nature of the alterations in these proteins which accompany viral transformation.

Changes in the phosphorylation of non-histone chromosomal proteins in relationship to DNA and RNA synthesis in BHK 21 C 13 cells

We have compared the in vivo phosphorylation of the non-histone chromosomal (NHC) proteins in relation to nucleic acid synthesis, in cells stimulated to proliferate and in resting cells. When BHK 21C 13 cells in saturation density are stimulated to proliferate by a change of medium, a synchronized DNA synthesis occurs with a maximum at about 16 h. Early after stimulation the NHC protein phosphorylation increases and reaches a maximum in late G 1 before the onset of DNA synthesis. It remains high during early S and decreases in late S. The phosphorylation of total cellular proteins, on the contrary, is increased only at the time of maximum DNA synthesis and always remains lower than that of NHC proteins. Early after stimulation and through the G 1 phase an increase of total RNA synthesis is also observed, however the time of maximum RNA synthesis does not coincide with the maximum of NHC protein phosphorylation. The involvement of NHC protein phosphorylation in the regulation of some messenger RNA (mRNA) synthesis is discussed.