Variable patterns of retrotransposition in different HeLa strains provide mechanistic insights into SINE RNA mobilization processes

Abstract

ABSTRACTAluelements are non-autonomous Short INterspersed Elements (SINEs) derived from the7SL RNAgene that are present at over one million copies in human genomic DNA.Alumobilizes by a mechanism known as retrotransposition, which requires the Long INterspersed Element-1 (LINE-1 or L1)ORF2-encoded protein (ORF2p). Here, we demonstrate that HeLa strains differ in their capacity to supportAluretrotransposition. HumanAluelements retrotranspose efficiently in HeLa-HA and HeLa-CCL2 (Alu-permissive) strains, but not in HeLa-JVM or HeLa-H1 (Alu-nonpermissive) strains. A similar pattern of retrotransposition was observed for other7SL RNA-derived SINEs andtRNA-derived SINEs. In contrast, mammalian LINE-1s, a zebrafish LINE, a humanSINE-VNTR-Alu(SVA) element, and anL1 ORF1-containing messenger RNA can retrotranspose in all four HeLa strains. Using anin vitroreverse transcriptase-based assay, we show thatAluRNAs associate with ORF2p and are converted into cDNAs in bothAlu-permissive andAlu-nonpermissive HeLa strains, suggesting that7SL- andtRNA-derived SINE RNAs use strategies to ‘hijack′ L1 ORF2p that are distinct from those used bySVAelements andORF1-containing mRNAs. These data further suggest ORF2p associates with theAluRNA poly(A) tract in bothAlu-permissive andAlu-nonpermissive HeLa strains, but thatAluretrotransposition is blocked after this critical step inAlu-nonpermissive HeLa strains.