Retrohoming of a Mobile Group II Intron in Human Cells Suggests How Eukaryotes Limit Group II Intron Proliferation

Mobile bacterial group II introns are evolutionary ancestors of spliceosomal introns and retroelements in eukaryotes. They consist of an autocatalytic intron RNA (a "ribozyme") and an intron-encoded reverse transcriptase, which function together to promote intron integration into new DNA s...

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Bibliographic Details
Published in:PLoS genetics 2015-08, Vol.11 (8), p.e1005422-e1005422
Main Authors: Truong, David M, Hewitt, F Curtis, Hanson, Joseph H, Cui, Xiaoxia, Lambowitz, Alan M
Format: Article
Language:English
Subjects:
Academic libraries
Active Transport, Cell Nucleus
Bacteria
Bacterial Proteins - genetics
Base Sequence
Cell proliferation
Cell Survival
Colleges & universities
Deoxyribonucleic acid
Directed Molecular Evolution
DNA
Gene expression
Genomes
Health aspects
HEK293 Cells
Humans
Introns
Inverted Repeat Sequences
Methods
Molecular Sequence Data
Mutation
Nonsense Mediated mRNA Decay
Observations
Plasmids - genetics
Retroelements
RNA polymerase
RNA sequencing
RNA-Directed DNA Polymerase - genetics
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Summary:Mobile bacterial group II introns are evolutionary ancestors of spliceosomal introns and retroelements in eukaryotes. They consist of an autocatalytic intron RNA (a "ribozyme") and an intron-encoded reverse transcriptase, which function together to promote intron integration into new DNA sites by a mechanism termed "retrohoming". Although mobile group II introns splice and retrohome efficiently in bacteria, all examined thus far function inefficiently in eukaryotes, where their ribozyme activity is limited by low Mg2+ concentrations, and intron-containing transcripts are subject to nonsense-mediated decay (NMD) and translational repression. Here, by using RNA polymerase II to express a humanized group II intron reverse transcriptase and T7 RNA polymerase to express intron transcripts resistant to NMD, we find that simply supplementing culture medium with Mg2+ induces the Lactococcus lactis Ll.LtrB intron to retrohome into plasmid and chromosomal sites, the latter at frequencies up to ~0.1%, in viable HEK-293 cells. Surprisingly, under these conditions, the Ll.LtrB intron reverse transcriptase is required for retrohoming but not for RNA splicing as in bacteria. By using a genetic assay for in vivo selections combined with deep sequencing, we identified intron RNA mutations that enhance retrohoming in human cells, but
ISSN:1553-7404
1553-7390
1553-7404
DOI:10.1371/journal.pgen.1005422