RESUMO
Alu elements are non-autonomous Short INterspersed Elements (SINEs) derived from the 7SL RNA gene that are present at over one million copies in human genomic DNA. Alu mobilizes by a mechanism known as retrotransposition, which requires the Long INterspersed Element-1 (LINE-1) ORF2-encoded protein (ORF2p). Here, we demonstrate that HeLa strains differ in their capacity to support Alu retrotransposition. Human Alu elements 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 other 7SL RNA-derived SINEs and tRNA-derived SINEs. In contrast, mammalian LINE-1s, a zebrafish LINE, a human SINE-VNTR-Alu (SVA) element, and an L1 ORF1-containing mRNA can retrotranspose in all four HeLa strains. Using an in vitro reverse transcriptase-based assay, we show that Alu RNAs associate with ORF2p and are converted into cDNAs in both Alu-permissive and Alu-nonpermissive HeLa strains, suggesting that 7SL- and tRNA-derived SINEs use strategies to 'hijack' L1 ORF2p that are distinct from those used by SVA elements and ORF1-containing mRNAs. These data further suggest ORF2p associates with the Alu RNA poly(A) tract in both Alu-permissive and Alu-nonpermissive HeLa strains, but that Alu retrotransposition is blocked after this critical step in Alu-nonpermissive HeLa strains.
Assuntos
Elementos Alu , Elementos Nucleotídeos Longos e Dispersos , Humanos , Células HeLa , Elementos Alu/genética , Elementos Nucleotídeos Longos e Dispersos/genética , Elementos Nucleotídeos Curtos e Dispersos/genética , Animais , Retroelementos/genética , RNA/genética , RNA/metabolismo , RNA de Transferência/genética , RNA de Transferência/metabolismo , Peixe-Zebra/genéticaRESUMO
Alu elements are non-autonomous Short INterspersed Elements (SINEs) derived from the 7SL RNA gene that are present at over one million copies in human genomic DNA. Alu mobilizes 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 support Alu retrotransposition. Human Alu elements 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 other 7SL RNA -derived SINEs and tRNA -derived SINEs. In contrast, mammalian LINE-1s, a zebrafish LINE, a human SINE-VNTR - Alu ( SVA ) element, and an L1 ORF1 -containing messenger RNA can retrotranspose in all four HeLa strains. Using an in vitro reverse transcriptase-based assay, we show that Alu RNAs associate with ORF2p and are converted into cDNAs in both Alu -permissive and Alu -nonpermissive HeLa strains, suggesting that 7SL - and tRNA -derived SINE RNAs use strategies to 'hijack' L1 ORF2p that are distinct from those used by SVA elements and ORF1 -containing mRNAs. These data further suggest ORF2p associates with the Alu RNA poly(A) tract in both Alu -permissive and Alu -nonpermissive HeLa strains, but that Alu retrotransposition is blocked after this critical step in Alu -nonpermissive HeLa strains.