Georg Mohr1,2,3, Eman Ghanem1,2,3, Alan M. Lambowitz1,2,3*
1 Institute for Cellular and Molecular Biology, University of Texas at Austin, Austin, Texas, United States of America, 2 Department of Chemistry and Biochemistry, University of Texas at Austin, Austin, Texas, United States of America, 3 Section of Molecular Genetics and Microbiology, School of Biological Sciences, University of Texas at Austin, Austin, Texas, United States of America
T. elongatus./Georg Mohr
Abstract
Mobile group II introns, which are found in bacterial and organellar genomes, are site-specific retroelments hypothesized to be evolutionary ancestors of spliceosomal introns and retrotransposons in higher organisms. Most bacteria, however, contain no more than one or a few group II introns, making it unclear how introns could have proliferated to higher copy numbers in eukaryotic genomes. An exception is the thermophilic cyanobacterium Thermosynechococcus elongatus, which contains 28 closely related copies of a group II intron, constituting ~1.3% of the genome. Here, by using a combination of bioinformatics and mobility assays at different temperatures, we identified mechanisms that contribute to the proliferation of T. elongatus group II introns. These mechanisms include divergence of DNA target specificity to avoid target site saturation; adaptation of some intron-encoded reverse transcriptases to splice and mobilize multiple degenerate introns that do not encode reverse transcriptases, leading to a common splicing apparatus; and preferential insertion within other mobile introns or insertion elements, which provide new unoccupied sites in expanding non-essential DNA regions. Additionally, unlike mesophilic group II introns, the thermophilic T. elongatus introns rely on elevated temperatures to help promote DNA strand separation, enabling access to a larger number of DNA target sites by base pairing of the intron RNA, with minimal constraint from the reverse transcriptase. Our results provide insight into group II intron proliferation mechanisms and show that higher temperatures, which are thought to have prevailed on Earth during the emergence of eukaryotes, favor intron proliferation by increasing the accessibility of DNA target sites. We also identify actively mobile thermophilic introns, which may be useful for structural studies, gene targeting in thermophiles, and as a source of thermostable reverse transcriptases.
Author Summary
Group II introns are bacterial mobile elements thought to be ancestors of introns and retroelements in higher organisms. They comprise a catalytically active intron RNA and an intron-encoded reverse transcriptase, which promotes splicing of the intron from precursor RNA and integration of the excised intron into new genomic sites. While most bacteria have small numbers of group II introns, in the thermophilic cyanobacterium Thermosynechococcus elongatus, a single intron has proliferated and constitutes 1.3% of the genome. Here, we investigated how the T. elongatus introns proliferated to such high copy numbers. We found divergence of DNA target specificity, evolution of reverse transcriptases that splice and mobilize multiple degenerate introns, and preferential insertion into other mobile introns or insertion elements, which provide new integration sites in non-essential regions of the genome. Further, unlike mesophilic group II introns, the thermophilic T. elongatus introns rely on higher temperatures to help promote DNA strand separation, facilitating access to DNA target sites. We speculate how these mechanisms, including elevated temperature, might have contributed to intron proliferation in early eukaryotes. We also identify actively mobile thermophilic introns, which may be useful for structural studies and biotechnological applications.
Citation: Mohr G, Ghanem E, Lambowitz AM (2010) Mechanisms Used for Genomic Proliferation by Thermophilic Group II Introns. PLoS Biol 8(6): e1000391. doi:10.1371/journal.pbio.1000391
Academic Editor: Eugene V. Koonin, National Center for Biotechnology Information, United States of America
Received: January 25, 2010; Accepted: April 28, 2010; Published: June 8, 2010
Copyright: © 2010 Mohr et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Funding: This work was supported by National Institutes of Health grants GM037949 and GM037951 and Welch Foundation grant F-1607. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Competing interests: The authors have declared that no competing interests exist.
Abbreviations: AmpR, ampicillin resistant; CapR, chloramphenicol resistant; CL, chloroplast-like; cp, chloroplast; E1, 5′ exon; E2, 3′ exon; EBS, exon-binding site; IBS, intron-binding site; IEP, intron-encoded protein; IPTG, isopropyl β-D-1-thiogalactopyranoside; LB medium, Luria-Bertani medium; ML, mitochondrial-like; mt, mitochondrial; ORF, open reading frame; RNP, ribonucleoprotein particle; RT, reverse transcriptase; TetR, tetracycline resistant
* E-mail: lambowitz@mail.utexas.edu
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