Gene expression in mouse neuroblastoma cells: properties of the genome.

Abstract

Chromatin was prepared from isolated nuclei of proliferating and differentiated cultures of C1300 mouse neuroblastoma cells. Differentiation (2-8), biochemical (4-7, 9-14), and electrical properties characteristic of neurons (15,16).The complex changes that accompany transition from the proliferating to the differentiated state may reflect alterations in the expression of information contained within the genome. One might anticipate modifications, therefore, in the macromolecules that comprise the genome and interact with DNA to regulate its function. Several lines of evidence suggest that the proteins associated with the genome play such a regulatory role in eukaryotic cells (17,18). Histones have been shown to be involved in the repression of I;DNAdependent RNA synthesis (19,20), maintenance of chromosome structure (21), and cytodifferentiation (22). Nonhistone chromosomal proteins may recognize defined gene loci and thus mediate specificity of gene read-out (18,(23)(24)(25)(26)(27)(28)(29). The nonhistone proteins have been shown to direct the tissue specific transcription of globin genes (41-43) and the cell cycle stage-specific transcription of histone genes (44,45). In the present study we have compared the histones and nonhistone chromosomal proteins associated with the genome in proliferating and differentiated mouse neuroblastoma C1300 cells. The possibility of a functional relationship between chromosomal proteins and differences observed in the transcriptional activity of chromatin from proliferating and differentiated mouse neuroblastoma cells is discussed. MATERIALS AND METHODSThe neuro 2a line of mouse neuroblastoma C1300 cells was grown in monolayer culture in 1-liter glass Blake flasks in Eagle's minimum essential medium (MEM) supplemented with 10% fetal calf serum in a moist CO2 incubator. To induce differentiation, cells were plated in MEM with 20% fetal calf serum and incubated for 24 hr. The monolayers were then washed with MEM and subsequently maintained for 24 hr in either serum-free MEM or in MEM with 20% fetal calf serum and 1 mM dibutyryl cyclic AMP. Proliferating cultures were treated in a similar manner, but 20% fetal calf serum was present during both 24-hr incubation periods.Nuclei and chromatin were prepared from mechanically harvested cells at 40. Details of the procedures have been described (30,31). Isolated nuclei were free of cytoplasmic contamination when examined by phase contrast microscopy. Little difference was observed in the patterns of proteins fractionated electrophoretically from either nuclei or isolated chromatin when comparisons were made with and without the inclusion of 50 Mg/ml of L-1-tosylamide-2-phenyl ethyl chloromethyl-ketone (TPCK) in the isolation and washing media to inhibit proteolysis (32). The histone:nonhistone chromosomal protein ratios in chromatin from proliferating and differentiated cultures were 0.73 and 0.79, respectively.The ability of chromatin from proliferating and differentiated cells to support RNA synthesis in vitro was determ...

High-Content Screening as a Universal Tool for Fingerprinting of Cytotoxicity of Nanoparticles

“…While the mitochondrial membrane potential of undifferentiated cells follows an inverse decay, the same parameter appears to decay exponentially in differentiated cells and could possibly explain their vulnerability to QD-induced apoptosis. We suspect these differences in cellular response arise from changes in biological machinery as cells differentiate . Changes involving mitochondria, membrane structures, and protein modifications may account for the higher sensitivity of differentiated neurons to cytotoxic substances …”

“…We suspect these differences in cellular response arise from changes in biological machinery as cells differentiate. 45 Changes involving mitochondria, membrane structures, and protein modifications may ac- count for the higher sensitivity of differentiated neurons to cytotoxic substances. 46 Gold Nanoparticles.…”

<i>In vitro</i> ‘Ageing’ and Nuclear Template Function

“…In addi tion, it is difficult to assign functional signif icance to such chromatin alterations. In this respect, it needs to be remembered that com positional changes have also been shown to occur during development (Bandy and Roberts, 1969;Bojanovic et al, 1970) and transforma tion (Zornetzer and Stein, 1975). Histone acet ylation, however, does seem to be correlated with transcriptional ability (Pochran et al, 1978).…”

Effect of Ultraviolet Irradiation on Chromatin and Its Components from Yoshida Ascites Tumour Cells

“…1 per cent SDS and 0 . 1 per cent fl-mercaptoethanol (Zornetzer and Stein 1975) . SDS-polyacrylamide gel electrophoresis of total chromosomal proteins was carried out in 7 .…”