Key role of the achievement of an appropriate ribosomal RNA complement for G1-S phase transition in H4-II-E-C3 rat hepatoma cells
Key role of the achievement of an appropriate ribosomal RNA complement for G1-S phase transition in H4-II-E-C3 rat hepatoma cells
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DOI：
10.1002/jcp.20144
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发表时间：
2005-02-01
影响因子：
5.6
通讯作者：
Treré, D
中科院分区：
文献类型：
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作者：
Derenzini, M;Montanaro, L;Treré, D
细胞生长与细胞增殖密切相关，充足的核糖体生物合成似乎是细胞复制所必需的。在本研究中，我们研究了rRNA合成和细胞周期进程之间的关系。为此，在第一组实验中，我们评估了rRNA合成变异对异步生长的H4-II-E-C3大鼠肝癌细胞周期持续时间的影响。用胰岛素或胰岛素加放线菌素D（AMD）处理细胞。该激素刺激核糖体生物合成，随后增加DNA合成和缩短细胞倍增时间（DT）。双变量流式细胞术表明，细胞周期的长度减少主要是由于较短的G1期。浓度为0.04 μ g/ml的AMD阻碍核糖体生物合成，而不影响异质RNA的产生。AMD使核糖体生物合成水平降低12小时，导致DNA合成降低，细胞DT延长，G1期延长。在第二组实验中，我们分析了H4-II-E-C3细胞G1期28 S和18 S rRNA转录本的细胞含量变化，通过血清剥夺同步化，然后通过血清、血清加胰岛素以及血清加胰岛素和AMD刺激。在对照细胞中，rRNA含量逐渐增加，直到血清刺激后21 h达到rRNA含量的最高值。在胰岛素处理的细胞中，rRNA值在12 h达到最高，而在AMD处理的细胞中，rRNA量持续较低，直到18 h，然后在21 h急剧增加。在三种实验条件下，rRNA含量的最高值均出现在G1期末期，且三者之间的差异很小。我们还评估，通过实时RT-PCR，细胞周期蛋白E mRNA的表达，这似乎急剧增加，在这些时间中，观察到的rRNA含量的最大增加。我们的结果表明，获得适量的rRNA允许G1/S期转变，可能是通过调节细胞周期蛋白E mRNA的表达。(C)2004 Wiley-Liss，Inc.
Cell growth is closely related to cell proliferation and an, adequate ribosome biogenesis appears to be necessary for cell duplication. In the present study, we have investigated the relationship between rRNA synthesis and cell cycle progression. For this purpose, in a first set of experiments, we evaluated the effect of rRNA synthesis variation on cycle duration in asynchronously growing H4-II-E-C3 rat hepatoma cells. Cells were either treated with insulin or insulin plus actinomycin D (AMD). The hormone stimulated ribosome biogenesis, which was later followed by an increased synthesis of DNA and a shortening of cell doubling time (DT). Bivariate flow cytometry indicated that the reduced length of the cell cycle was mainly due to the shorter G1-phase. AMD, at the concentration of 0.04 mug/ml, hindered ribosome biogenesis without affecting heterogeneous RNA production. A 12-h reduction in ribosome biogenesis level by AMD caused a lowering of DNA synthesis and a lengthening of cell DT with a longer G1-phase. In a second set of experiments, we analyzed the cell content variations of 28S and 18S rRNA transcripts during G1 phase in H4-II-E-C3 cells, synchronized by serum deprivation, and then stimulated by serum, serum plus insulin, and serum plus insulin and AMD. In control cells, a progressive increase in rRNA content occurred until the highest value of rRNA content was reached 21 h after serum stimulation. In insulin-treated cells, the highest rRNA value was reached at 12 h whereas in AMD-treated cells, the rRNA quantity was constantly low until 18 h and then sharply increased at 21 h. In the three experimental conditions, the highest values of rRNA amount were reached at the end of G1 phase and were quite similar to one another. We also evaluated, by real-time RT-PCR, cyclin E mRNA expression, which appeared to sharply increase at those times in which the maximum increase in the rRNA content was observed. Our results indicated that the achievement of an appropriate amount of rRNA allows G1/S phase transition, probably by modulating the expression of cyclin E mRNA. (C) 2004 Wiley-Liss, Inc.