title: Characterization of a synthetic human LINE-1 retrotransposon ORFeus-Hs.
abstract: Long interspersed elements, type 1(LINE-1, L1) are the most abundant and only active autonomous retrotransposons in the human genome. Native L1 elements are inefficiently expressed because of a transcription elongation defect thought to be caused by high adenosine content in L1 sequences. Previously, we constructed a highly active synthetic mouse L1 element (ORFeus-Mm), partially by reducing the nucleotide composition bias. As a result, the transcript abundance of ORFeus-Mm was greatly increased, and its retrotransposition frequency was > 200-fold higher than its native counterpart. In this paper, we report a synthetic human L1 element (ORFeus-Hs) synthesized using a similar strategy. The adenosine content of the L1 open reading frames (ORFs) was reduced from 40% to 27% by changing 25% of the bases in the ORFs, without altering the amino acid sequence. By studying a series of native/synthetic chimeric elements, we observed increased levels of full-length L1 RNA and ORF1 protein and retrotransposition frequency, mostly proportional to increased fraction of synthetic sequence. Overall, the fully synthetic ORFeus-Hs has > 40-fold more RNA but is at most only ~threefold more active than its native counterpart (L1RP); however, its absolute retrotransposition activity is similar to ORFeus-Mm. Owing to the elevated expression of the L1 RNA/protein and its high retrotransposition ability, ORFeus-Hs and its chimeric derivatives will be useful tools for mechanistic L1 studies and mammalian genome manipulation.
authors.affiliation: Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.|School of Molecular Biosciences, Washington State University, Pullman WA 99164, USA.
authors.forename: Wenfeng
authors.initials: W
authors.lastname: An
authors.affiliation: Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
authors.forename: Lixin
authors.initials: L
authors.lastname: Dai
authors.affiliation: Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
authors.forename: Anna Maria
authors.initials: AM
authors.lastname: Niewiadomska
authors.affiliation: Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.|Stanford University School of Medicine, Stanford, CA 94305, USA.
authors.forename: Alper
authors.initials: A
authors.lastname: Yetil
authors.affiliation: Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
authors.forename: Kathryn A
authors.initials: KA
authors.lastname: O'Donnell
authors.affiliation: Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.|Carnegie Institution of Washington, 3520 San Martin Drive, Baltimore, MD 21218, USA.
authors.forename: Jeffrey S
authors.initials: JS
authors.lastname: Han
authors.affiliation: Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
authors.forename: Jef D
authors.initials: JD
authors.lastname: Boeke