Characterization of a synthetic human LINE-1 retrotransposon ORFeus-Hs

The human genome is littered with transposable element sequences; some are mere fossil records of ancient insertion events, whereas others remain active. Of these active elements, the long interspersed elements, type 1 (LINE-1 or L1) remain among the most active, and are capable of autonomous retrotransposition [1] and of providing enzymatic activities for the non-autonomous retrotransposition of short interspersed nucleotide elements (SINE) such as Alu elements [2]. Full-length versions of L1 elements are approximately 6 kb long, and consist of a 5' (untranslated region) UTR containing an internal promoter sequence, two open reading frames (ORFs), ORF1 and ORF2, and a 3'UTR followed by a poly(A) tail encoded in the DNA [3-8]. The L1 ORF1 protein (ORF1p) is a non-specific nucleic acid binding protein with nucleic acid chaperone activity [9-12]. The ORF2 protein (ORF2p) is responsible for the catalytic activity necessary for retrotransposition, and contains both endonuclease and reverse transcriptase activities [13,14].L1s make up approximately 17% of the human genome. However, despite their abundance, the replication and control mechanisms of these elements are poorly understood, partly because of their low expression levels of messenger (m)RNA and protein [15]. We have previously linked inefficient L1 expression to a transcription elongation defect potentially caused by high adenosine content in the ORFs. We subsequently constructed a synthetic L1, termed ORFeus, in which the codons of both ORFs were synonymously optimized, based on a mouse L1 protein sequence [16,17]. This element was at least 200-fold more active for retrotransposition than the native mouse element L1spa [18].In this paper, we describe our use of similar techniques to construct a synthetic human L1 (ORFeus-Hs) element and several synthetic/native chimeric L1 elements. Although we observed increased levels of L1 mRNA and ORF1p, the levels of L1 retrotransposition, as measured by two different retr