Leduc Y, Lawrence J J, De Murcia G, Poirier G G
Centre de recherche sur les mécanismes de sécrétion, Faculté des Sciences, Université de Sherbrooke, Québec, Canada.
Biochim Biophys Acta. 1988 Mar 11;968(3):275-82. doi: 10.1016/0167-4889(88)90018-3.
The properties of poly(ADP-ribose) synthetase were studied throughout the cell cycle progression of non-synchronized rat FR3T3 fibroblasts using an immunological and biochemical approach. Cells in the various phases of the cell cycle were sorted from an asynchronously growing population by using flow cytofluorometry. G1, S and G2 + M fractions were used for enzymatic assays in the presence of saturating concentrations of DNAase I for the analysis of total poly(ADP-ribose) synthetase; maximal enzyme activity was found in the G2 + M phase. Purified IgG, specific for the FR3T3 poly(ADP-ribose) synthetase were used for the labelling of endogenous synthetase in order to quantify the enzyme immunologically. Localization of nuclear immunofluorescence was observed and analysis of poly(ADP-ribose) synthetase content throughout the cell cycle were carried out using double fluorescent staining and cytofluorometry. Poly(ADP-ribose) synthetase content as measured immunologically was found to increase from G1 to S and G2 + M phases. Quiescent cells showed a lower content as measured immunologically of poly(ADP-ribose) synthetase than cells in the G1 phase. In exponentially growing cells, the ratio between enzyme activity of poly(ADP-ribose) synthetase over the amount of enzyme measured immunologically was found to be higher in the G2 + M phase. These results show that a cell-cycle specific event activates poly(ADP-ribose) synthetase in the G2 + M phase.
采用免疫学和生物化学方法，研究了非同步化大鼠FR3T3成纤维细胞在整个细胞周期进程中聚（ADP - 核糖）合成酶的特性。通过流式细胞荧光术从异步生长的细胞群体中分选处于细胞周期各个阶段的细胞。利用饱和浓度的DNA酶I，将G1、S和G2 + M组分用于酶活性测定，以分析总聚（ADP - 核糖）合成酶；发现最大酶活性出现在G2 + M期。用对FR3T3聚（ADP - 核糖）合成酶特异的纯化IgG对内源性合成酶进行标记，以便从免疫学角度对该酶进行定量。观察核免疫荧光的定位，并使用双重荧光染色和细胞荧光术对整个细胞周期中的聚（ADP - 核糖）合成酶含量进行分析。从免疫学角度测定，聚（ADP - 核糖）合成酶的含量从G1期到S期和G2 + M期增加。静止细胞的聚（ADP - 核糖）合成酶免疫学测定含量低于G1期细胞。在指数生长的细胞中，发现G2 + M期聚（ADP - 核糖）合成酶的酶活性与免疫学测定的酶量之比更高。这些结果表明，细胞周期特异性事件在G2 + M期激活聚（ADP - 核糖）合成酶。
