De Novo Genome Assembly of Toniniopsis dissimilis (Ramalinaceae, Lecanoromycetes) from Long Reads Shows a Comparatively High Composition of Biosynthetic Genes Putatively Involved in Melanin Synthesis

Abstract

Lichens have developed numerous adaptations to optimize their survival in various environmental conditions, largely by producing secondary compounds by the fungal partner. They often have antibiotic properties and are involved in protection against intensive UV radiation, pathogens, and herbivores. To contribute to the knowledge of the arsenal of secondary compounds in a crustose lichen species, we sequenced and assembled the genome of Toniniopsis dissimilis, an indicator of old-growth forests, using Oxford Nanopore Technologies (ONT, Oxford, UK) long reads. Our analyses focused on biosynthetic gene clusters (BGCs) and specifically on Type I Polyketide (T1PKS) genes involved in the biosynthesis of polyketides. We used the comparative genomic approach to compare the genome of T. dissimilis with six other members of the family Ramalinaceae and twenty additional lichen genomes from the database. With only six T1PKS genes, a comparatively low number of biosynthetic genes are present in the T. dissimilis genome; from those, two-thirds are putatively involved in melanin biosynthesis. The comparative analyses showed at least three potential pathways of melanin biosynthesis in T. dissimilis, namely via the formation of 1,3,6,8-tetrahydroxynaphthalene, naphthopyrone, or YWA1 putative precursors, which highlights its importance in T. dissimilis. In addition, we report the occurrence of genes encoding ribosomally synthesized and posttranslationally modified peptides (RiPPs) in lichens, with their highest number in T. dissimilis compared to other Ramalinaceae genomes. So far, no function has been assigned to RiPP-like proteins in lichens, which leaves potential for future research on this topic.

Keywords: ecology; fungal RiPP-like proteins; genomics; lichen; long-read sequencing; symbiosis; type I polyketide gene.

Figure 1

(A) Detail of T. dissimilis (M-0355157): thallus, consisting of scattered, rounded to slightly flattened or subsquamulose granules, and dark apothecia. Scale: 0.5 mm. (B) Habitat of T. dissimilis: A mixed forest in the Oytal Valley, Allgäu (Bavaria, Germany), 47°23′08″ N 10°20′35″ E, ca. 1080 m asl.

Figure 2

Diverse classes (categories) of secondary metabolites annotated for the selected lichen-forming fungi are given in color with the color code on the right-hand side. The total number of biosynthetic gene clusters is given on the x-axis, and the species names are on the y-axis. The following single letter code indicates families: C: Cladoniaceae, G: Graphidaceae, I: Icmadophilaceae, P: Parmeliaceae, R: Ramalinaceae, S: Stictidaceae, T: Teloschistaceae, U: Umbilicariaceae, and V: Verrucariaceae.

Figure 3

Maximum-likelihood phylogeny of ketoacyl synthase (KS) genes from Type I PKS genes as inferred by IQ-TREE. Genes from T. dissimilis are marked in red, and other Ramalinaceae genomes are in black. The putative secondary compounds clades are given in color boxes, with the largest clade assigned to melanins (green). Clades where no clear annotations could be found are in grey.