Biosynthesis of nuclear proteins after stimulation of quiescent Swiss mouse 3T3 cells (original) (raw)
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Two independent techniques have been used to study the effects of nucleolar damage on the entry of cells into the DNA synthetic phase. Experiments have been performed with HEp/Z cells, mouse L cells, freshly cultured embryonic mouse fibroblast cells &d freshly cultured monkey kidney cells. All the results confirm previous reports that when cells have proceeded more than 1-2 h into the G 1 phase the onset of the next DNA synthetic phase is not delayed by nucleolar damage. However, nucleolar damage at about the time of cell division appears to delay entry into S phase for certain cell types.
The role of nuclear proteins in RNA synthesis
Experimental Cell Research, 1972
When epithelial cells from baby mice are cultured on a solid substrate in vitro, DNA synthesis and cell division take place in a majority of the cells 24 to 48 h after inoculation. This cell proliferation is regularly preceded by a sequence of nuclear changes likely to be involved in gene activation during the process of growth-transformation. These nuclear changes, which in the present investigation were studied by quantitative cytochemical methods, are characterized by: (1) increased binding of acridine orange (AO) very soon after cell attachment, reflecting initial changes of the deoxvribonucleoprotein (DNP) complex: (2) accumulation of uroteins in the cell nucl&s; (3) dispersion of the nuclear cdromatin; (4) increased rate of 14C-uridine incorporation. The initial DNP change. reflected by the increased AO-binding, was per se not a sufficient factor for stimulation of-14C-uridine incorporation. An increasedgH-uridine incorporation was only observed when protein began to accumulate in the nucleus and when the chromatin changed from a condensed to a more dispersed state. The rate of 14C-uridine incorporation was, during the whole process of growth transformation, found to be directly proportional to the amount of protein that had accumulated in the nucleus. However, the initial DNP change was a necessary prerequisite for the subsequent accumulation of protein in the nucleus. Attachment of the cells to the solid substrate was found to be necessary in expressing this sequence of nuclear events. Furthermore, in areas on the glass slide where many cells had attached, these nuclear changes occurred more rapidly.
Phosphorylation of nuclear and DNA-binding proteins in proliferating and quiescent mammalian cells
Biochemical Journal, 1981
The dependence of cell proliferation on nuclear protein phosphorylation was studied with exponential-phase and stationary-phase cultures of Chinese-hamster ovary cells. Nuclear proteins were fractionated, according to their DNA-binding affinities, by using sequential extractions of isolated nuclei with increasing concentrations of NaCl. When viable whole cells were labelled with H332PO4, phosphorylation of nuclear proteins was found to be lower in quiescent cells than in proliferating cells. Phosphorylation of nuclear proteins soluble in 0.30M-NaCl (less than 50% of these proteins bind to DNA) was greater than for those proteins soluble in higher salt concentrations (80-100% of these proteins bind to DNA). Cyclic AMP enhanced the phosphorylation of nuclear proteins soluble in 0.3 m-NaCl by 40-50%, and this stimulation was independent of cell growth. Cyclic AMP also increased the phosphorylation of nuclear proteins soluble in 0.6M-NaCl and 2.0M-NaCl by 40-50% in exponential-phase cul...
Molecular and Cellular Biochemistry, 1992
The nuclear fraction isolated from Krebs II ascites cells following cell disruption by nitrogen cavitation was separated into four fractions by salt/detergent extraction: NP-40 soluble fraction, 130mM KC1 extract, DOC/Triton x 100 soluble fraction and salt/detergent treated nuclei. The protein composition of the individual fractions was studied by SDS-PAGE and the relative amounts of actin and a 35 kDa protein (p35) were measured from gel scans. There was a time-dependent shift of actin from the 130 mM KCI extract to the NP-40 soluble fraction upon storage of the nuclear fraction on ice, indicating a progressive depolymerization of microfilaments. Compared with actin there was a slower release of p35 into the NP-40 soluble fraction. The results suggest that p35 is not integrated in the microfilament network. Phalloidin, which stabilizes the microfilaments, enriched the amount of both proteins in the 130 mM KC1 extracts, together with a series of other proteins in the range 50-205 kDa. The presence of phalloidin also resulted in a large increase in the actin content in both the DOC/Triton x 100 extract and the fraction containing salt/detergent treated nuclei. Incubation of cells with insulin and/or cycloheximide enriched the amount of actin in the 130 mM KC1 fraction. The results show that short term incubation of cells with phalloidin, insulin or cycloheximide increases the actin content of the nuclear fraction and also affects the presence of several other proteins. (Mol Cell Biochem 115: 187-194, 1992)