Mo Y Y, Beck W T
Division of Developmental Therapeutics, University of Illinois at Chicago 60607, USA.
Oncol Res. 1997;9(4):193-204.
DNA topoisomerase II alpha is a nuclear enzyme essential for DNA metabolism and cell cycle progression. Previous studies have shown that human tumor cell lines can express more than one topoisomerase II alpha isoform through alternative splicing. A 160-kDa isoform of topoisomerase II alpha has been described in several cell lines selected for resistance to inhibitors of DNA topoisomerase, but its physiological function has not been defined. In the present study, we have identified two major (160 and 140 kDa) and two minor (150 and 145 kDa) isoforms of topoisomerase II alpha in drug-sensitive human leukemic CEM cells, all of which have lost C-terminal regions that produce epitopes recognized by specific antibodies. Reverse transcription-polymerase chain reaction and molecular cloning identified four alternatively spliced transcripts of topoisomerase II alpha from CEM cells. Furthermore, nucleotide sequencing indicated that the 160-kDa isoform is encoded by two transcripts derived from alternative splicing at a different C-terminal site and that the other two transcripts likely code for the 150-kDa isoform. Although the full-length topoisomerase II alpha resided in the cell nucleus, all altered isoforms, except the 160 kDa that was located in both cytoplasmic and nuclear extracts in about equal amount, were shown to be present predominantly in the cytosol. In contrast to the observations of other groups, we have not found an association of the topoisomerase II alpha isoforms with drug resistance. Rather, our results suggest that expression of topoisomerase II alpha isoforms is cell type specific or might be associated with the neoplastic phenotype of the cells. Thus, although T-lineage tumor cell lines examined (CEM, Jurkat, and H9) displayed altered topoisomerase II alpha isoforms, normal T cells expressed only a full-length copy of the gene. Together, these results suggest that expression of altered topoisomerase II alpha isoforms is not limited to drug resistance, but might be a feature of neoplastic cells.
DNA拓扑异构酶IIα是一种对DNA代谢和细胞周期进程至关重要的核酶。先前的研究表明，人类肿瘤细胞系可通过可变剪接表达不止一种拓扑异构酶IIα同工型。在几种对DNA拓扑异构酶抑制剂具有抗性的细胞系中已描述了一种160 kDa的拓扑异构酶IIα同工型，但其生理功能尚未明确。在本研究中，我们在对药物敏感的人类白血病CEM细胞中鉴定出了拓扑异构酶IIα的两种主要同工型（160 kDa和140 kDa）以及两种次要同工型（150 kDa和145 kDa），所有这些同工型均缺失了产生特定抗体可识别表位的C末端区域。逆转录-聚合酶链反应和分子克隆鉴定出了来自CEM细胞的拓扑异构酶IIα的四种可变剪接转录本。此外，核苷酸测序表明，160 kDa同工型由两个在不同C末端位点通过可变剪接产生的转录本编码，另外两个转录本可能编码150 kDa同工型。尽管全长拓扑异构酶IIα定位于细胞核，但所有改变的同工型，除了160 kDa在细胞质和细胞核提取物中含量大致相等外，主要存在于细胞质中。与其他研究小组的观察结果相反，我们未发现拓扑异构酶IIα同工型与耐药性之间存在关联。相反，我们的结果表明拓扑异构酶IIα同工型的表达具有细胞类型特异性，或者可能与细胞的肿瘤表型相关。因此，尽管所检测的T细胞系肿瘤细胞系（CEM、Jurkat和H9）显示出拓扑异构酶IIα同工型发生改变，但正常T细胞仅表达该基因的全长拷贝。总之，这些结果表明，改变的拓扑异构酶IIα同工型的表达不仅限于耐药性，可能是肿瘤细胞的一个特征。
