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Arbuscular mycorrhizal fungi (AMF) are an ancient and ubiquitous group of obligate plant symbionts that are thought to have assisted the colonization of land by plants c. 800 million yr ago through the establishment of the mycorrhizal symbiosis; a close association of AMF with the roots of over 80% of land plants, and many important crops (Smith et al., 1997; Sanders, 2002, 2003; Corradi & Charest, 2011). The hallmark of this symbiosis is the transfer of photosynthetically produced carbohydrates from the plant to the fungus in exchange for increased supplies of water and mineralized nutrients, and for this reason AMF are considered key mutualists in terrestrial ecosystems (van der Heijden et al., 1998; Smith et al., 2000; Munkvold et al., 2004; Wagg et al., 2011).
Besides their ecological relevance, AMF are also intriguing from a cellular point of view, as they harbour coenocytic hyphae (i.e. aseptate) that are perpetually multinucleate (Rosendahl, 2008). These multinucleated hyphae have been proposed to be able to fuse through a process called anastomosis, which could allow nuclear exchange between genetically dissimilar AMF (Croll et al., 2009). AMF are also characterized by an unusually elevated molecular diversity within single spores, whose origin is currently debated (Lloyd-Macgilp et al., 1996; Lanfranco et al., 1999; Kuhn et al., 2001; Pawlowska & Taylor, 2004; Rodriguez et al., 2004; Hijri & Sanders, 2005; Pawlowska, 2005; Stukenbrock & Rosendahl, 2005; Heitman et al., 2007).
AMF are also peculiar from an evolutionary perspective, as they are currently thought to have propagated for the past 500 million yr in the absence of sexual reproduction. This remarkable long-term clonal evolution has resulted in these curious fungi being referred to as ‘ancient asexuals’ – that is, evolutionary ‘anomalies’ that should have long gone extinct as a consequence of their intrinsic incapacity to offset the accumulation of deleterious mutations over time (e.g. through meiotic recombination; Smith, 1986; Judson & Normark, 1996; Butlin et al., 1998; Welch & Meselson, 2000; Normark et al., 2003). Although sexual reproduction has yet to be formally observed in AMF, alternative scenarios for their long-term evolutionary and ecological success in the sexual stages have started to emerge. In particular, AMF have been recently proposed to be capable of undergoing a cryptic sexual cycle following the identification of events of recombination in some natural populations (Vandenkoornhuyse et al., 2001; Croll & Sanders, 2009; den Bakker et al., 2010), and the detection of many homologues of genes essential for meiosis (Halary et al., 2011; Corradi & Lildhar, 2012) and those normally found in the mating type (MAT) loci of early diverging fungi (Tisserant et al., 2012).
Among these potential signatures of sex, the identification of Rhizophagus homologues of SexM and SexP is particularly intriguing, as it suggested an intrinsic potential for these fungi to harbour a bona fide MAT locus. Briefly, a fungal MAT locus is a genomic region found in most fungi that serves the common purpose of determining sexual compatibility between two individuals of outcrossing species, or a region required for sexual differentiation in self-fertile species. Sexual identity is determined by allelic variation at this locus, but the gene content and structure of the locus can differ quite drastically across members of the fungal kingdom (Lee et al., 2010). Alleles of the MAT locus are often called idiomorphs to denote sequences that occupy the same locus on a chromosome, but do not necessarily share conservation in sequence, gene order, or a common descent (Metzenberg & Glass, 1990). Overall, the MAT loci of diverse fungi can contain different combinations of genes, including homeodomain proteins, high-mobility group (HMG) transcription factors, and an alpha-box (which has recently been reclassified as an HMG domain; (T. Martin et al., 2010); but the putative ancestral version of them all is currently found in zygomycetes, where the two idiomorphs are represented by MATA-HMG genes known as SexM or SexP (Idnurm et al., 2008).
The ancestral role of MATA-HMGs in fungal mating, combined with the fact that homologues of these genes are often found in most known fungal MAT loci, makes them ideal candidates for new signatures of sexual reproduction that might be found in surveys of AMF genomes. Indeed, homologues of SexM and SexP have been identified in Rhizophagus irregularis, but their diversity and genomic context in AMF are completely unknown. Here, we surveyed the transcriptomes of two isolates of R. irregularis (SwiC2, this study; and DAOM 197198, Tisserant et al. (2012)) and one of the closely related species Rhizophagus diaphanus (MUCL 43196, this study) for the presence of homologues of MATA-HMGs, and our explorations revealed the presence of a surprisingly elevated number of gene homologues in all strains investigated.