Department of Evolutionary Experimental Biology, University of Bologna, Bologna, Italy.
Exp Cell Res. 2010 Dec 10;316(20):3358-67. doi: 10.1016/j.yexcr.2010.06.024. Epub 2010 Jul 8.
Long Interspersed Nuclear Elements (L1) are retroelements generally repressed in most differentiated somatic cells. Their activity has been observed in some undifferentiated and tumour cells and could be involved in tumour onset and progression. Growing evidences show that the L1 activation can occur in neuronal precursor cells during differentiation process. Neuroblastoma is a tumour originating from neuronal precursor cells, and, although the molecular basis of its progression is still poorly understood, the implication of L1 activation has not yet been investigated. In this study L1 mobility in neuroblastoma BE(2)C cells was assessed using the in vitro retrotransposition assay consisting in an episomal EGFP-tagged L1(RP) element, whose mobility can be evaluated by cytofluorimetric analysis (FACS) of EGFP expression. FACS results have shown a low retrotransposition activity. To detect L1(RP) integrated in transcriptionally repressed genomic sites, both a cell treatment with a stimulator of reporter gene promoter, and a quantitative Real-Time PCR analysis were performed. A retrotransposition activity ten and one thousand times that of FACS was found, respectively. These results point out that the real rate of L1 retrotransposition events in tumour cells might be considerably higher than that reported so far by evaluating only the reporter gene expression.
长散布核元件（L1）是一种通常在大多数分化体细胞中受到抑制的反转录元件。它们的活性已在一些未分化和肿瘤细胞中观察到，可能参与肿瘤的发生和进展。越来越多的证据表明，L1 的激活可能发生在神经元前体细胞的分化过程中。神经母细胞瘤是一种起源于神经元前体细胞的肿瘤，尽管其进展的分子基础仍不清楚，但 L1 的激活尚未得到研究。在这项研究中，使用体外反转录转座测定法评估了神经母细胞瘤 BE(2)C 细胞中的 L1 迁移，该测定法包括一个带有 EGFP 标签的 L1（RP）顺式元件，其迁移可以通过 EGFP 表达的细胞荧光分析（FACS）来评估。FACS 结果显示反转录转座活性较低。为了检测 L1（RP）整合到转录受抑制的基因组位点，进行了细胞处理和报告基因启动子刺激剂的处理，以及定量实时 PCR 分析。分别发现反转录转座活性是 FACS 的十倍和一千倍。这些结果表明，在肿瘤细胞中，L1 反转录转座事件的真实率可能远远高于迄今为止仅通过评估报告基因表达所报告的水平。
