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More active human L1 retrotransposons produce longer insertions.

ABSTRACT: The vast majority of L1 insertions are 5' truncated and thus inactive. Yet, the mechanism of 5' truncation is unknown. To examine whether the frequency of L1 retrotransposition is directly correlated with the length of genomic L1 insertions, we used a cell culture assay to measure retrotransposition frequency and a PCR-based assay to measure L1 insertion length. We tested five full-length human L1 elements that retrotranspose at different frequencies: LRE3, L1(RP), L1.3, L1.2A and L1.2B. Our data suggest that L1 insertion length correlates with L1 retrotransposition frequency for insertions >1 kb in length. For two elements, L1(RP) and L1.2A, we found that swapping the reverse transcriptase domains had little effect. Instead, we found that genomic insertion length and retrotransposition frequency are substantially affected by amino acid substitutions at positions 363, 1220 and 1259 in ORF2. We suggest that the region containing residues 1220 and 1259 may be important in the binding of ORF2p to L1 RNA to facilitate reverse transcription.

SUBMITTER: Farley AH

PROVIDER: S-EPMC373329 | biostudies-literature | 2004

REPOSITORIES: biostudies-literature

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More active human L1 retrotransposons produce longer insertions.

Farley Alexander H AH Luning Prak Eline T ET Kazazian Haig H HH

Nucleic acids research 20040123 2

The vast majority of L1 insertions are 5' truncated and thus inactive. Yet, the mechanism of 5' truncation is unknown. To examine whether the frequency of L1 retrotransposition is directly correlated with the length of genomic L1 insertions, we used a cell culture assay to measure retrotransposition frequency and a PCR-based assay to measure L1 insertion length. We tested five full-length human L1 elements that retrotranspose at different frequencies: LRE3, L1(RP), L1.3, L1.2A and L1.2B. Our dat ...[more]

PMID: 14742665

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Project description:BackgroundPatients with systemic lupus erythematosus (SLE) have autoantibodies against the L1-encoded open-reading frame 1 protein (ORF1p). Here, we report (i) which immune cells ORF1p emanates from, (ii) which L1 loci are transcriptionally active, (iii) whether the cells express L1-dependent interferon and interferon-stimulated genes, and (iv) the effect of inhibition of L1 ORF2p by reverse transcriptase inhibitors.ResultsL1 ORF1p was detected by flow cytometry primarily in SLE CD66b+CD15+ regular and low-density granulocytes, but much less in other immune cell lineages. The amount of ORF1p was higher in neutrophils from patients with SLE disease activity index (SLEDAI) > 6 (p = 0.011) compared to patients with inactive disease, SLEDAI < 4. Patient neutrophils transcribed seven to twelve human-specific L1 loci (L1Hs), but only 3 that are full-length and with an intact ORF1. Besides serving as a source of detectable ORF1p, the most abundant transcript encoded a truncated ORF2p reverse transcriptase predicted to remain cytosolic, while the two other encoded an intact full-length ORF2p. A number of genes encoding proteins that influence L1 transcription positively or negatively were altered in patients, particularly those with active disease, compared to healthy controls. Components of nucleic acid sensing and interferon induction were also altered. SLE neutrophils also expressed type I interferon-inducible genes and interferon β, which were substantially reduced after treatment of the cells with drugs known to inhibit ORF2p reverse transcriptase activity.ConclusionsWe identified L1Hs loci that are transcriptionally active in SLE neutrophils, and a reduction in the epigenetic silencing mechanisms that normally counteract L1 transcription. SLE neutrophils contained L1-encoded ORF1p protein, as well as activation of the type I interferon system, which was inhibited by treatment with reverse transcriptase inhibitors. Our findings will enable a deeper analysis of L1 dysregulation and its potential role in SLE pathogenesis.

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More active human L1 retrotransposons produce longer insertions.

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More active human L1 retrotransposons produce longer insertions.

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