Arita I, Tachibana A, Takebe H, Tatsumi K
Department of Molecular Oncology, Faculty of Medicine, Kyoto University, Japan.
Carcinogenesis. 1989 Nov;10(11):2067-73. doi: 10.1093/carcin/10.11.2067.
Previous studies have demonstrated that approximately one-third of human lymphoblastoid cell lines (LCLs) are deficient in removing O6-methylguanine residues because of the lack of O6-alkylguanine-DNA alkyltransferase (O6-AGT) activity. Such LCLs have been designated Mex-, while the proficient LCLs are Mex+. Our determinations of O6-AGT activity as a function of cellular protein concentration on 37 previously-established LCLs disclosed that the expression of the enzyme was high in 14 (Mex+) and barely detectable in 16 (Mex-). The other seven LCLs showed intermediate activity of the enzyme. By contrast, all of the 28 LCLs that we newly established contained high enzyme activity, implying that they consisted of mainly Mex+ cells. Since the conventional O6-AGT assay on Mex+ cell populations was not capable of detecting the co-existence of Mex- cells as a minor component, we attempted to determine the proportion of Mex- phenotype in newly-immortalized lymphoblastoid cell clones which had been established directly on semisolid agar. All of the 15 independent clones derived from a single blood sample also showed high O6-AGT activity, rendering it unlikely that Epstein-Barr virus transformation per se was responsible for the generation of Mex- LCLs. These results collectively indicate that Mex+ cells predominate in LCLs shortly after establishment and also suggest that the possible growth advantage for Mex- cells should play an important role in the subsequent development of Mex- LCLs during the long-term culture in vitro.
先前的研究表明，由于缺乏O6-烷基鸟嘌呤-DNA烷基转移酶（O6-AGT）活性，大约三分之一的人类淋巴母细胞系（LCLs）在去除O6-甲基鸟嘌呤残基方面存在缺陷。这类LCLs被指定为Mex-，而具有活性的LCLs则为Mex+。我们根据细胞蛋白质浓度测定了37个先前建立的LCLs的O6-AGT活性，结果显示，14个LCLs（Mex+）中该酶的表达较高，而16个LCLs（Mex-）中几乎检测不到该酶的表达。另外7个LCLs显示出该酶的中等活性。相比之下，我们新建立的28个LCLs都具有较高的酶活性，这意味着它们主要由Mex+细胞组成。由于对Mex+细胞群体进行的传统O6-AGT检测无法检测到作为次要成分的Mex-细胞的共存情况，我们试图确定直接在半固体琼脂上建立的新永生化淋巴母细胞克隆中Mex-表型的比例。从单一血样中获得的所有15个独立克隆也都显示出较高的O6-AGT活性，这表明爱泼斯坦-巴尔病毒转化本身不太可能是产生Mex- LCLs的原因。这些结果共同表明，Mex+细胞在LCLs建立后不久占主导地位，也表明Mex-细胞可能的生长优势在体外长期培养过程中Mex- LCLs的后续发展中应发挥重要作用。
