Hayashi J, Werbin H, Shay J W
Cancer Res. 1986 Aug;46(8):4001-6.
We isolated hybrids and cybrids using HeLaTG cells and human normal primary fibroblasts to examine the functional differences between the mitochondrial genomes of tumor and normal cells with respect to their possible involvement in the regulation of tumorigenicity. Hybrids contained mitochondrial DNA (mtDNA) predominantly from the fibroblast parent and their tumorigenicity was suppressed completely. Then, cytoplasmic transmission of primary fibroblast mtDNA to HeLaTG cells was carried out using toxin-antitoxin selection. Two cybrid clones containing a HeLaTG nucleus only and more than 60% of transmitted fibroblast mtDNA were isolated and injected into nude mice to test their tumorigenicity. They formed tumors when 2 X 10(6) cells were injected, whereas no tumors were formed after injection of 5 X 10(5) cells (a concentration at which HeLaTG subclones formed tumors). These cybrids were cultivated in normal medium for two additional months and the content of fibroblast mtDNA increased gradually, resulting in HeLaTG mtDNA eventually being lost from both cybrid clones. We again examined their tumorigenicity and found that they recovered tumorigenicity completely. These results indicate that tumorigenicity of HeLaTG cells could not be suppressed by replacing their mitochondrial genomes with those of normal primary fibroblasts. Further, the partial suppression of tumorigenicity observed in the cybrid clones was temporary and may be due to cytoplasmic factors other than the mitochondrial genomes. Although we can find no difference between the mitochondrial genomes of normal and tumor cells regarding the regulation of tumorigenicity, the segregation pattern of the mtDNA in the cybrids was of interest: in the absence of any mitochondrial selection, HeLaTG mtDNA was lost while fibroblast mtDNA was retained, even though the nuclear component of these cybrids was from the HeLaTG cells. Thus, there should be some functional differences between the mitochondrial genomes of HeLaTG cells and primary fibroblasts that are responsible for the preferential segregation of HeLaTG mtDNA from the cybrids.
我们使用HeLaTG细胞和人正常原代成纤维细胞分离杂种细胞和胞质杂种细胞，以研究肿瘤细胞和正常细胞线粒体基因组在可能参与肿瘤发生调控方面的功能差异。杂种细胞主要含有来自成纤维细胞亲本的线粒体DNA（mtDNA），其致瘤性被完全抑制。然后，利用毒素-抗毒素选择法将原代成纤维细胞mtDNA细胞质传递至HeLaTG细胞。分离出两个仅含有HeLaTG细胞核且超过60%传递的成纤维细胞mtDNA的胞质杂种细胞克隆，并将其注射到裸鼠体内以测试其致瘤性。当注射2×10⁶个细胞时它们形成了肿瘤，而注射5×10⁵个细胞（HeLaTG亚克隆形成肿瘤的浓度）后未形成肿瘤。这些胞质杂种细胞在正常培养基中再培养两个月，成纤维细胞mtDNA的含量逐渐增加，最终两个胞质杂种细胞克隆中的HeLaTG mtDNA都丢失了。我们再次检测它们的致瘤性，发现它们完全恢复了致瘤性。这些结果表明，用正常原代成纤维细胞的线粒体基因组替换HeLaTG细胞的线粒体基因组并不能抑制其致瘤性。此外，在胞质杂种细胞克隆中观察到的致瘤性部分抑制是暂时的，可能是由于线粒体基因组以外的细胞质因子。虽然我们未发现正常细胞和肿瘤细胞的线粒体基因组在肿瘤发生调控方面存在差异，但胞质杂种细胞中mtDNA的分离模式令人感兴趣：在没有任何线粒体选择的情况下，HeLaTG mtDNA丢失而成纤维细胞mtDNA保留，尽管这些胞质杂种细胞的核成分来自HeLaTG细胞。因此，HeLaTG细胞和成纤维细胞的线粒体基因组之间应该存在一些功能差异，这些差异导致了HeLaTG mtDNA在胞质杂种细胞中的优先分离。
