Repeat-induced gene silencing of L1 transgenes is correlated with differential promoter methylation

• Published in: Gene Vol. 456; no. 1; pp. 15 - 23
• Main Authors: Rosser, James M., An, Wenfeng
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 15.05.2010
• Subjects: Animals; Copy number; CpG Islands; DNA Methylation; Gene Dosage; Gene Silencing; Humans; LINE-1; Long Interspersed Nucleotide Elements; Mice; Mice, Inbred C57BL; Promoter Regions, Genetic; Retroelements; Retrotransposon; Transgenes; Transgenic mouse; LINE-1; Retrotransposon; LINE-1 or L1; ORF; LSL; RIGS; Copy number; CAG; Transgenic mouse; non-LTR; UTR; B6; Kb; DNA methylation; qPCR; Zp3; wt; gDNA
• ISSN: 0378-1119; 1879-0038; 1879-0038
• DOI: 10.1016/j.gene.2010.02.005

Recent transgenic studies on L1 retrotransposons have afforded exciting insights into L1 biology, and a unique opportunity to model their function and regulation in vivo. Thus far, the majority of the transgenic L1 mouse lines are constructed via pronuclear microinjection, a procedure that typically results in the integration of tandem arrayed transgenes. Transgene arrays are susceptible to repeat-induced gene silencing (RIGS) in both plants and animals. In order to examine the potential impact of RIGS on L1 retrotransposition, we derived a cohort of animals carrying reduced copies of ORFeus transgene at the same genomic locus by Cre-mediated recombination. The copy number reduction of ORFeus transgenes did not decrease the overall retrotransposition activity. Using a sensitive and reproducible quantitative PCR assay, an average frequency of 0.45 insertions per cell was observed for animals carrying the donor transgene at a single copy, representing a 9-fold increase of retrotransposition frequency on a per-copy basis. DNA methylation analyses revealed that the observed retrotransposition activity was correlated with differential CpG methylation at the heterologous promoter: the promoter region was largely methylated in animals with the high-copy array but significantly hypomethylated in animals with the single-copy counterpart. In contrast, the ORF2 region, which represents the body of the ORFeus transgene, and the 3′ end of the transgene showed high level of methylation in both high-copy and single-copy samples. The observed methylation patterns were metastable across generations. In summary, our data suggest that tandem arrayed L1 transgenes are subject to RIGS, and transgenes present at a single copy in the genome are thus recommended for modeling L1 in animals.