Wallace Nicholas, Wagstaff Bradley J, Deininger Prescott L, Roy-Engel Astrid M
Tulane Cancer Center SL-66, Deparment of Epidemiology, Tulane University Health Sciences Center, 1430 Tulane Ave., New Orleans, LA 70112, USA.
Gene. 2008 Aug 1;419(1-2):1-6. doi: 10.1016/j.gene.2008.04.007. Epub 2008 Apr 24.
Retroelements have contributed over one third of the human genome mass. The currently active LINE-1 (L1) codes for two proteins (ORF1p and ORF2p), both strictly required for retrotransposition. In contrast, the non-coding parasitic SINE (Alu) only appears to need the L1 ORF2p for its own amplification. This requirement was previously determined using a tissue culture assay system in human cells (HeLa). Because HeLa are likely to express functional L1 proteins, it is possible that low levels of endogenous ORF1p are necessary for the observed tagged Alu mobilization. By individually expressing ORF1 and ORF2 proteins from both human (L1RP and LRE3) and rodent (L1A102 and L1spa) L1 sources, we demonstrate that increasing amounts of ORF1 expressing vector enhances tagged Alu mobilization in HeLa cells. In addition, using chicken fibroblast cells as an alternate cell culture source, we confirmed that ORF1p is not strictly required for Alu mobilization in our assay. Supporting our observations in HeLa cells, we find that tagged Alu retrotransposition is improved by supplementation of ORF1p in the cultured chicken cells. We postulate that L1 ORF1p plays either a direct or indirect role in enhancing the interaction between the Alu RNA and the required factors needed for its retrotransposition.
逆转录元件占人类基因组质量的三分之一以上。目前活跃的LINE-1（L1）编码两种蛋白质（ORF1p和ORF2p），这两种蛋白质都是逆转录转座所严格必需的。相比之下，非编码寄生性SINE（Alu）似乎仅需L1 ORF2p来实现自身扩增。这一需求先前是通过在人类细胞（HeLa）中进行的组织培养检测系统确定的。由于HeLa细胞可能表达功能性L1蛋白，因此观察到的标记Alu转座可能需要低水平的内源性ORF1p。通过分别从人类（L1RP和LRE3）和啮齿动物（L1A102和L1spa）的L1来源表达ORF1和ORF2蛋白，我们证明，表达ORF1的载体数量增加会增强HeLa细胞中标记Alu的转座。此外，使用鸡成纤维细胞作为另一种细胞培养来源，我们证实，在我们的检测中，Alu转座并不严格需要ORF1p。与我们在HeLa细胞中的观察结果一致，我们发现，在培养的鸡细胞中补充ORF1p可改善标记Alu的逆转录转座。我们推测，L1 ORF1p在增强Alu RNA与其逆转录转座所需因子之间的相互作用中发挥直接或间接作用。
