A novel active L1 retrotransposon subfamily in the mouse

Abstract

Unlike human L1 retrotransposons, the 5' UTR of mouse L1 elements contains tandem repeats of approximately 200 bp in length called monomers. Multiple L1 subfamilies exist in the mouse which are distinguished by their monomer sequences. We previously described a young subfamily, called the T(F) subfamily, which contains approximately 1800 active elements among its 3000 full-length members. Here we characterize a novel subfamily of mouse L1 elements, G(F), which has unique monomer sequence and unusual patterns of monomer organization. A majority of these G(F) elements also have a unique length polymorphism in ORF1. Polymorphism analysis of G(F) elements in various mouse subspecies and laboratory strains revealed that, like T(F), the G(F) subfamily is young and expanding. About 1500 full-length G(F) elements exist in the diploid mouse genome and, based on the results of a cell culture assay, approximately 400 G(F) elements are potentially capable of retrotransposition. We also tested 14 A-type subfamily elements in the assay and estimate that about 900 active A elements may be present in the mouse genome. Thus, it is now known that there are three large active subfamilies of mouse L1s; T(F), A, and G(F), and that in total approximately 3000 full-length elements are potentially capable of active retrotransposition. This number is in great excess to the number of L1 elements thought to be active in the human genome.

Figure 1

(A) Alignment of the 206-bp G_F monomer consensus sequence, the 206-bp reconstructed F-consensus monomer (Adey et al. 1994b), and the 212-bp T_F monomer consensus (DeBerardinis and Kazazian 1999). The location of the consensus YY1-binding sequence is underlined. (B) G_F element variants (labeled I– IV) with different patterns of 5′ UTR monomer organization. The fragment shown with horizontal stripes in each element is in the nonmonomeric 5′ UTR and is 82% identical to sequence in the F-consensus monomer. G_F monomers are either full-length (204–207 bp) or truncated (∼64 bp). The monomeric regions of some clones (indicated by “?”) are too short to be certain of their variant group.

Figure 1

(A) Alignment of the 206-bp G_F monomer consensus sequence, the 206-bp reconstructed F-consensus monomer (Adey et al. 1994b), and the 212-bp T_F monomer consensus (DeBerardinis and Kazazian 1999). The location of the consensus YY1-binding sequence is underlined. (B) G_F element variants (labeled I– IV) with different patterns of 5′ UTR monomer organization. The fragment shown with horizontal stripes in each element is in the nonmonomeric 5′ UTR and is 82% identical to sequence in the F-consensus monomer. G_F monomers are either full-length (204–207 bp) or truncated (∼64 bp). The monomeric regions of some clones (indicated by “?”) are too short to be certain of their variant group.

Figure 2

(A) Classification scheme for mouse L1 ORF1 length polymorphism region (LPR) groups, as defined by Mears and Hutchison (2001). LPRs consists of tandem arrays of 66-bp and/or 42-bp blocks, and each begins with a 66-bp block. The 42-bp block is homologous with the 3′ portion of the 66-bp repeat. The novel G_F variant has been designated LPR Group V. (B) Alignment of the ORF1 LPR repeats from the G_F consensus sequence. As shown in the first line of the alignment, the 5′-most 24 bp of the 66-bp repeat has partial homology with the 42-bp repeat.

Figure 2

(A) Classification scheme for mouse L1 ORF1 length polymorphism region (LPR) groups, as defined by Mears and Hutchison (2001). LPRs consists of tandem arrays of 66-bp and/or 42-bp blocks, and each begins with a 66-bp block. The 42-bp block is homologous with the 3′ portion of the 66-bp repeat. The novel G_F variant has been designated LPR Group V. (B) Alignment of the ORF1 LPR repeats from the G_F consensus sequence. As shown in the first line of the alignment, the 5′-most 24 bp of the 66-bp repeat has partial homology with the 42-bp repeat.

Figure 3

(A) Phylogenetic analysis of that portion of the 5′ UTR of mouse L1s defined as region α by Mears and Hutchison (2001) and corresponding to nucleotides 1539–1740 of the A-type element L1Md-A2. The unrooted tree was constructed by the nearest neighbor-joining method (Saitou and Nei 1987) and includes the 17 members of the G_F dataset and members of the dataset used by Mears and Hutchison (2001). G_F clades identified by the analysis are named G-I, G-II, and G-III, and their corresponding LPR groups are indicated. Significant bootstrap values (of 1000 replicates) are shown at the nodes. (B) Phylogenetic analysis of 3′ UTR region corresponding to nts 6701–7333 of L1MdA2. Also included in this analysis are L1s classified by Hardies et al. (2000) as members of the “Z subfamily”: MMconreg2, L1_bg (Perou et al. 1997), and L1MdZ (Kraft et al. 1992). These group with T_F-type elements.

Figure 3

(A) Phylogenetic analysis of that portion of the 5′ UTR of mouse L1s defined as region α by Mears and Hutchison (2001) and corresponding to nucleotides 1539–1740 of the A-type element L1Md-A2. The unrooted tree was constructed by the nearest neighbor-joining method (Saitou and Nei 1987) and includes the 17 members of the G_F dataset and members of the dataset used by Mears and Hutchison (2001). G_F clades identified by the analysis are named G-I, G-II, and G-III, and their corresponding LPR groups are indicated. Significant bootstrap values (of 1000 replicates) are shown at the nodes. (B) Phylogenetic analysis of 3′ UTR region corresponding to nts 6701–7333 of L1MdA2. Also included in this analysis are L1s classified by Hardies et al. (2000) as members of the “Z subfamily”: MMconreg2, L1_bg (Perou et al. 1997), and L1MdZ (Kraft et al. 1992). These group with T_F-type elements.

Figure 4

Two G_F elements, G_F27 and G_F71, have transduced DNA flanking their 3′ ends to new genomic locations. Consecutive retrotransposition events have created the composite transduction of G_F27. TSDs are shown in bold lowercase letters, and putative hexanucleotide poly(A) signals are in bold uppercase letters.

Figure 5

Some G_F elements are recent insertions. Shown are PCR reaction products which verify the presence (7.5-kb product) or absence (450-bp product) in the genomes of individual mice of different subspecies and laboratory strains. A faint, ∼7.5-kb product is present in lane 5 (M. m. domesticus) of G_F21 which we have not been able to confirm as L1-related by sequencing. In the case of G_F46, no PCR product is seen in lane 9 (strain SJL/J), although in other reactions we have been able to amplify an obvious 450-bp product. The 7.5-kb product labeled 129/Sv BAC (lane 13 of G_F46) was amplified from bacmid DNA of CITB/Research Genetics clone #437P9.