Gong J, Traganos F, Darzynkiewicz Z
Cancer Research Institute, New York Medical College, Valhalla 10523, USA.
Exp Cell Res. 1995 Sep;220(1):226-31. doi: 10.1006/excr.1995.1310.
Cyclins, the regulatory subunits of their respective cyclin-dependent kinases, are the key components of the cell-cycle progression machinery. Some cyclins are expressed discontinuously during the cell cycle, their synthesis and degradation being strictly scheduled. The presence of these cyclins in the cell, therefore, provides landmarks of the cell cycle, in addition to DNA replication and mitosis. Cyclin A is expressed in late S and G2 phase and degraded during mitosis just prior to metaphase. Degradation of another "mitotic" cyclin, cyclin B1, occurs later, at the transition from metaphase to anaphase. Based on the difference in time of degradation of cyclin A versus cyclin B1 it was possible, in the present study, to discriminate between G2 and mitotic (postprophase) MOLT-4 leukemic cells, by multiparameter (cellular DNA content versus cyclin expression) flow cytometry. The cells arrested in metaphase by Vinblastine were cyclin A negative and had an elevated level of cyclin B1. The cells arrested in G2 by the DNA topoisomerase II inhibitor m-AMSA had a very high level of cyclin B1 expression and unchanged expression of cyclin A. During stathmokinesis induced by Vinblastine the percentage of mitotic cells estimated by analysis of cellular DNA content and cyclin A expression was identical to that estimated by the alternative method based on in situ DNA denaturation followed by staining with acridine orange. Thus, differences in expression of cyclins A and B1 make it possible to discriminate cells that have the same DNA content but reside in different phases of the cycle, such as DNA diploid cells in G2 versus tetraploid G1 cells or mitotic versus G2 cells.
细胞周期蛋白作为各自依赖细胞周期蛋白激酶的调节亚基，是细胞周期进程机制的关键组成部分。一些细胞周期蛋白在细胞周期中呈间断性表达，其合成和降解严格按计划进行。因此，除了DNA复制和有丝分裂外，这些细胞周期蛋白在细胞内的存在为细胞周期提供了标志。细胞周期蛋白A在S期晚期和G2期表达，并在有丝分裂中期之前的有丝分裂过程中降解。另一种“有丝分裂”细胞周期蛋白，即细胞周期蛋白B1，在后期降解，发生在中期向后期的转变过程中。基于细胞周期蛋白A与细胞周期蛋白B1降解时间的差异，在本研究中，通过多参数（细胞DNA含量与细胞周期蛋白表达）流式细胞术，可以区分G2期和有丝分裂期（前期后）的MOLT-4白血病细胞。长春碱使细胞停滞在中期，这些细胞细胞周期蛋白A呈阴性，细胞周期蛋白B1水平升高。DNA拓扑异构酶II抑制剂m-AMSA使细胞停滞在G2期，这些细胞细胞周期蛋白B1表达水平非常高，细胞周期蛋白A表达未改变。在长春碱诱导的静止期，通过分析细胞DNA含量和细胞周期蛋白A表达估计的有丝分裂细胞百分比，与基于原位DNA变性后用吖啶橙染色的替代方法估计的百分比相同。因此，细胞周期蛋白A和B1表达的差异使得区分具有相同DNA含量但处于细胞周期不同阶段的细胞成为可能，例如G2期的DNA二倍体细胞与四倍体G1期细胞，或有丝分裂期细胞与G2期细胞。
