Targeting cytoplasmic cap methylation. Homo sapiens cultivar:U2OS

To identify targets of cytoplasmic capping, we developed an inducible U2OS cell line overexpressing a dominant negative cytoplasmic form of cap methyltransferase. Overexpressing this dominant negative protein results in a lack of methylation on cytoplasmically recapped RNAs and thus their degradation through the cells quality control mechanisms. Comparing RNA expression in induced and uninduced cell lines thus reveals targets of cytoplasmic recapping.

为鉴定细胞质加帽（cytoplasmic capping）的作用靶标，我们构建了一株诱导型U2OS细胞系，该细胞系过表达一种显性负效的帽甲基转移酶（cap methyltransferase）胞质突变体。过表达该显性负效蛋白后，经细胞质重加帽修饰的RNA将无法发生甲基化，进而通过细胞的质量控制机制被降解。因此，对比诱导组与未诱导组细胞系的RNA表达谱，即可揭示细胞质重加帽的靶标分子。