Lost in Translation: population genomics and long-read sequencing reveals relaxation of concerted evolution of the ribosomal DNA cistron

Abstract

<jats:title>Abstract</jats:title><jats:p>Concerted evolution of the ribosomal DNA array has been studied in numerous eukaryotic taxa, yet is still poorly understood. rDNA genes are repeated dozens to hundreds of times in the eukaryotic genome [1] and it is believed that these arrays are homogenized through concerted evolution [2, 3] preventing the accumulation of intragenomic, and intraspecific, variation. However, numerous studies have reported rampant intragenomic and intraspecific variation in the rDNA array [4–10], contradicting our current understanding of concerted evolution. The internal transcribed spacers (ITS) of the rDNA cistron are the most commonly used DNA barcoding region in <jats:italic>Fungi</jats:italic> [11], and rely on concerted evolution to homogenize the rDNA array leading to a “barcode gap” [12]. Here we show that in <jats:italic>Boletus edulis</jats:italic> Bull., ITS intragenomic variation persists at low allele frequencies throughout the rDNA array, this variation does not correlate with genomic relatedness between populations, and rDNA genes may not evolve in a strictly concerted fashion despite the presence of unequal recombination and gene conversion. Under normal assumptions, heterozygous positions found in ITS sequences represent hybridization between populations, yet through allelic mapping of the rDNA array we found numerous heterozygous alleles to be stochastically introgressed throughout, presenting a dishonest signal of gene flow. Moreover, despite the signal of gene flow in ITS, our organisms were highly inbred, indicating a disconnect between true gene flow and barcoding signals. In addition, we show that while the mechanisms of concerted evolution are ongoing in pseudo-heterozygous individuals, they are not fully homogenizing the ITS array. Concerted evolution of the rDNA array may insufficiently homogenize the ITS gene, allowing for misleading signals of gene flow to persist, vastly complicating the use of the ITS locus for DNA barcoding in <jats:italic>Fungi</jats:italic>.</jats:p>

Contrasting continental patterns of adaptive population divergence in the holarctic ectomycorrhizal fungus Boletus edulis

“…Previous attempts to characterize population genetic structure within Boletus edulis Bull. have been inconclusive (Tremble et al ., 2020). Therefore, we first used a coalescent‐based phylogenomic approach to segregate B. edulis specimens into lineages.…”

“…To characterize patterns of admixture and haplotype diversity across the landscape, we used the highly contiguous B. edulis assembly (Tremble et al ., 2020). For improved gene prediction, total RNA was extracted from frozen hymenophore tissue of sample BD747 (Tremble et al ., 2020) using TRI Reagent ® (Sigma‐Aldrich) following the manufacturer's instructions. Short‐read mRNA enrichment, library prep, and paired‐end sequencing were performed by Novogene USA (Sacramento, CA, USA).…”

“…1), the taxonomy of B. edulis is a matter of some controversy with many alternative classifications (Arora, 2008;Dentinger et al, 2010;Arora & Frank, 2014). Current molecular evidence is equally ambiguous, indicating that this complex group may be in the early stages of speciation (Coyne et al, 2004;Beugelsdijk et al, 2008;Dentinger et al, 2010;Tremble et al, 2020). Taken together, the combination of morphological variation, enormous ecological amplitude, holarctic distribution, and small genome (c. 50 Mbp) makes B. edulis an ideal natural system to investigate the roles of demography, introgression, adaptation, and isolation in the speciation process at a global scale.…”

ITS-based assessment of Madagascar’s fungal diversity and arrival of ectomycorrhizal fungi to the island

“…However, an equal sampling across regions and lineages would be necessary for this approach, which will require major investment since documentation of these diverse families remains highly incomplete. Moreover, using a multi-gene approach would allow for a deeper resolution of the phylogeny (Frøslev et al 2005, Tremble et al 2019), allowing for fossil-calibrated dating of the trees that may result in more accurate dates (Zhao et al 2017, Varga et al 2019).…”

Delimiting species in Basidiomycota: a review

“…Cryptic speciation is present in many genera, including Astraeus (Phosri et al 2014), Paxillus (Hedh et al 2008;Jargeat et al 2014Jargeat et al , 2016, Pisolithus (Martin et al 2002), Rhizopogon (Dowie et al 2017, Serpula (Carlsen et al 2011;Balasundaram et al 2015), and Strobilomyces (Sato et al 2007). Balasundaram et al (2015) and Tremble et al (2020) questioned the employment of the ITS locus alone in separating cryptic species in Boletales. In particular, Balasundaram et al (2015) suggested using ITS only in combination with other loci, after pointing out that btub, hsp, rpb2, and tef1 loci are more informative than ITS at the species level (even in single-locus trees).…”