Unequal recombination and evolution of the mating-type (MAT) loci in the pathogenic fungus Grosmannia clavigera and relatives

Abstract

Sexual reproduction in fungi is regulated by the mating-type (MAT) locus where recombination is suppressed. We investigated the evolution of MAT loci in eight fungal species belonging to Grosmannia and Ophiostoma (Sordariomycetes, Ascomycota) that include conifer pathogens and beetle symbionts. The MAT1-2 idiomorph/allele was identified from the assembled and annotated Grosmannia clavigera genome, and the MAT locus is flanked by genes coding for cytoskeleton protein (SLA) and DNA lyase. The synteny of these genes is conserved and consistent with other members in Ascomycota. Using sequences from SLA and flanking regions, we characterized the MAT1-1 idiomorph from other isolates of G. clavigera and performed dotplot analysis between the two idiomorphs. Unexpectedly, the MAT1-2 idiomorph contains a truncated MAT1-1-1 gene upstream of the MAT1-2-1 gene that bears the high-mobility-group domain. The nucleotide and amino acid sequence of the truncated MAT1-1-1 gene is similar to its homologous copy in the MAT1-1 idiomorph in the opposite mating-type isolate, except that positive selection is acting on the truncated gene and the alpha(α)-box that encodes the transcription factor has been deleted. The MAT idiomorphs sharing identical gene organization were present in seven additional species in the Ophiostomatales, suggesting that the presence of truncated MAT1-1-1 gene is a general pattern in this order. We propose that an ancient unequal recombination event resulted in the ancestral MAT1-1-1 gene integrated into the MAT1-2 idiomorph and surviving as the truncated MAT1-1-1 genes. The α-box domain of MAT1-1-1 gene, located at the same MAT locus adjacent to the MAT1-2-1 gene, could have been removed by deletion after recombination due to mating signal interference. Our data confirmed a 1:1 MAT/sex ratio in two pathogen populations, and showed that all members of the Ophiostomatales studied here including those that were previously deemed asexual have the potential to reproduce sexually. This ability can potentially increase genetic variability and can enhance fitness in new, ecological niches.

Keywords: heterothallism; homothallism; mating system evolution; outcrossing; selfing.

Figure 1

(A) Sequence arrangement and annotation of the MAT idiomorph and adjacent genes from annotated genome of Grosmannia clavigera SL-KW1407. (B) The MAT genes of opposite mating-type isolates as determined from long-range PCR and primer walking using primers ER and UP2-2F. Genes are indicated by different colors, and the arrows indicate the predicted directions of gene translation. (C) Amplicons of MAT fragments after long-range PCR were run in an agarose gel to indicate the size variation between MAT1-2 and MAT1-1 isolates.

Figure 2

Comparison of MAT loci in Grosmannia clavigera. (A) Dotplot comparison/pairwise alignment of DNA sequence data for MAT1-1 and MAT1-2 idiomorphs of G. clavigera. Sequence lengths are given along the axes. (B) The amino acid alignment of MAT1-1-1 in MAT1-2 idiomorph to the truncated MAT1-1-1 in MAT1-2 idiomorph of G. clavigera by Clustal W. The comparison indicates the deletion of α-box domain (in square) in truncated MAT1-1-1.

Figure 3

Homology among the MAT locus. (A) MAT1-1 idiomorph; (B) MAT1-2 idiomorph, of G. huntii, G. clavigera, and L. longiclavatum. The diagram was prepared from the output of Artemis Comparison Tool. Regions of strong homology are shaded and connected by lines. The intensity of shading indicates the strength of homology. Genes are represented by box arrows. The MAT1-2 sequence of G. clavigera SL-KW1407 was obtained from the genome sequence, while the MAT1-1 sequence of G. clavigera and opposite MAT isolates of other fungi were obtained in this investigation.

Figure 4

Comparison of MAT loci in Ophiostoma montium. (A) Dotplot comparison/pairwise alignment of DNA sequence data for MAT1-1 and MAT1-2 idiomorphs of O. montium. Sequence lengths are given along the axes. (B) The amino acid alignment of MAT1-1-1 in MAT1-2 idiomorph to the truncated MAT1-1-1 in MAT1-2 idiomorph of O. montium by Clustal W. The comparison indicates the truncated MAT1-1-1 was highly eroded and the absence of α-box domain (in square in full length MAT1-1-1).

Figure 5

NJ tree generated from MEGA showing the phylogenetic relationships among ascomycetes inferred from the HMG domain (129 amino acid characters) of the MAT1-2-1. Number on branches indicated bootstrap support (1000 pseudoreplicates) more than 60% from NJ and PhyML (from left to right). The taxa shaded in gray indicate the presence of truncated MAT genes in opposite MAT isolates.

Figure 6

Gene genealogies showing the phylogenetic relationships between G. clavigera and relatives. (A) Intergenic sequences after 3′ end of SLA to the 5′ end of the truncated MAT1-1-1 and MAT1-1-1 genes (1349 characters), and (B) the full-length and truncated MAT1-1-1 genes. MAT1-2 isolates are indicated in green while MAT1-1 isolates are highlighted in gray (1968 characters). Number on branches indicated bootstrap support (500 pseudoreplicates) greater than 60%.

Figure 7

Proposed model for the evolution of MAT locus in the common ancestor of Grosmannia and Ophiostoma (Ophiostomatales).