Academic journal article Genetics

Mobile Introns Shape the Genetic Diversity of Their Host Genes

Academic journal article Genetics

Mobile Introns Shape the Genetic Diversity of Their Host Genes

Article excerpt

SELF-splicing introns are selfish elements with a broad but patchy phylogenetic distribution. Found in transfer RNA (tRNA), ribosomal RNA (rRNA) and (occasionally) proteincoding genes in bacteria and archaea, they are particularly numerous in mitochondrial genomes of fungi and plants where they have invaded genes encoding components of the electron transport chain (ETC) (Lambowitz and Belfort 1993). In many instances, fungal self-splicing introns have remained mobile, as demonstrated by experiments that track invasion capacity by crossing intron-containing with intronfree yeast strains (Jacquier and Dujon 1985; Wenzlau et al. 1989; Lazowska et al. 1994; Paschke et al. 1994) and intron presence/absence polymorphisms across natural populations of Saccharomyces cerevisiae (Wolters et al. 2015), Schizosaccharomyces pombe (Zimmer et al. 1987), and Lachancea kluyveri (Jung et al. 2012). For the majority of self-splicing introns in S. cerevisiae, spreading to an intron-free location is initiated by a homing endonuclease that is encoded in the intron itself and binds a large (~20-30 bp), often singular target motif with high affinity (Jacquier and Dujon 1985; Moran et al. 1992). Following cleavage of the intron-free homing site, the intron-containing copy of the mitochondrial genome is used as a template for homologous recombination (HR), resulting in the conversion of an intron-free to an introncontaining locus. Once gained, introns can be lost again either through fortuitous deletion or through a gene conversion event that involves an intronless complementary DNA (cDNA) produced by reverse transcriptase (RT) activity (Levra-Juillet et al. 1989), as has been proposed for spliceosomal introns (Cohen et al. 2012). In the absence of selection for intron retention, cycles ofintron gain and loss ensue (Goddard and Burt 1999), accompanied by recurrent endonuclease activity that predictably targets the very same recognition site.

Here, prompted by reports of possible mutational hotspots in the vicinity of self-splicing intron (Hensgens et al. 1983; Zimmer et al. 1987; Foury et al. 1998), we consider what impact these invasion-loss cycles have on the genetic diversity of the host gene. In particular, we consider the possibility that endonuclease-mediated cleavage and subsequent repair might be mutagenic. Although HR is generally considered to be high fidelity, it can carry nonnegligible mutagenic risks, depending on the precise nature of the repair process and whether error-prone polymerases are involved in DNA resynthesis (Rodgers and McVey 2016). Pertinently, Hicks et al. (2010) observed increased mutation rates during doublestrand break (DSB) repair at the mating type (MAT) locus of S. cerevisiae, which is cleaved by the endonuclease HO and subsequently repaired via HR (Hicks et al. 2010).

Mutagenic side effects associated with endonuclease activity have also come into sharp focus recently with the widespread adoption of targetable endonucleases for genome engineering. The principal concern here has been to identify and reduce off-target activity (Cho et al. 2014; Kleinstiver et al. 2016). However, endonuclease activity can also have undesired on-target effects. Notably, nonhomologous end joining (NHEJ) downstream of Cas9-mediated cleavage is associated with an increased risk of indel formation (van Overbeek et al. 2016). This has prompted the development of Cas9 derivatives that nick rather than cleave DNA (Cong et al. 2013; Mali et al. 2013), shifting repair pathway choice away from NHEJ and toward HR.

We reasoned that one way to develop a greater understanding of such on-target mutagenicity would be to study endonucleases in their native genomic context. If endonuclease activity is indeed mutagenic, cleavage and repair might have left a detectable imprint on population-wide genetic variation around the cleavage site. In search of such an imprint, we survey recent high-quality population genomic data from S. cerevisiae, S. …

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