Tseng B Y, Prussak C E, Almazan M T
Eukaryotic Regulatory Biology Program, School of Medicine, University of California, San Diego, La Jolla 92093.
Mol Cell Biol. 1989 May;9(5):1940-5. doi: 10.1128/mcb.9.5.1940-1945.1989.
Expression of the small-subunit p49 mRNA of primase, the enzyme that synthesizes oligoribonucleotides for initiation of DNA replication, was examined in mouse cells stimulated to proliferate by serum and in growing cells. The level of p49 mRNA increased approximately 10-fold after serum stimulation and preceded synthesis of DNA and histone H3 mRNA by several hours. Expression of p49 mRNA was not sensitive to inhibition by low concentrations of cycloheximide, which suggested that the increase in mRNA occurred before the restriction point control for cell cycle progression described for mammalian cells and was not under its control. p49 mRNA levels were not coupled to DNA synthesis, as observed for the replication-dependent histone genes, since hydroxyurea or aphidicolin had no effect on p49 mRNA levels when added before or during S phase. These inhibitors did have an effect, however, on the stability of p49 mRNA and increased the half-life from 3.5 h to about 20 h, which suggested an interdependence of p49 mRNA degradation and DNA synthesis. When growing cells were examined after separation by centrifugal elutriation, little difference was detected for p49 mRNA levels in different phases of the cell cycle. This was also observed when elutriated G1 cells were allowed to continue growth and then were blocked in M phase with colcemid. Only a small decrease in p49 mRNA occurred, whereas H3 mRNA rapidly decreased, when cells entered G2/M. These results indicate that the level of primase p49 mRNA is not cell cycle regulated but is present constitutively in proliferating cells.
引发酶的小亚基p49 mRNA的表达情况在受血清刺激而增殖的小鼠细胞以及生长中的细胞中进行了检测。引发酶是一种合成寡核糖核苷酸以启动DNA复制的酶。血清刺激后，p49 mRNA的水平增加了约10倍，并且在DNA和组蛋白H3 mRNA合成之前数小时就已升高。p49 mRNA的表达对低浓度环己酰亚胺的抑制不敏感，这表明mRNA的增加发生在哺乳动物细胞所描述的细胞周期进程的限制点控制之前，且不受其控制。与复制依赖性组蛋白基因的情况不同，p49 mRNA水平与DNA合成不相关，因为在S期之前或期间添加羟基脲或阿非迪霉素对p49 mRNA水平没有影响。然而，这些抑制剂确实对p49 mRNA的稳定性有影响，并将其半衰期从3.5小时延长至约20小时，这表明p49 mRNA降解与DNA合成之间存在相互依存关系。当通过离心淘析分离后对生长中的细胞进行检测时，在细胞周期的不同阶段未检测到p49 mRNA水平有明显差异。当让淘析得到的G1期细胞继续生长，然后用秋水仙酰胺将其阻断在M期时，也观察到了同样的情况。当细胞进入G2/M期时，p49 mRNA仅出现小幅下降，而H3 mRNA迅速下降。这些结果表明，引发酶p49 mRNA的水平不受细胞周期调控，而是在增殖细胞中持续存在。
