The evolution of fungal metabolic pathways

Abstract

Fungi contain a remarkable range of metabolic pathways, sometimes encoded by gene clusters, enabling them to digest most organic matter and synthesize an array of potent small molecules. Although metabolism is fundamental to the fungal lifestyle, we still know little about how major evolutionary processes, such as gene duplication (GD) and horizontal gene transfer (HGT), have interacted with clustered and non-clustered fungal metabolic pathways to give rise to this metabolic versatility. We examined the synteny and evolutionary history of 247,202 fungal genes encoding enzymes that catalyze 875 distinct metabolic reactions from 130 pathways in 208 diverse genomes. We found that gene clustering varied greatly with respect to metabolic category and lineage; for example, clustered genes in Saccharomycotina yeasts were overrepresented in nucleotide metabolism, whereas clustered genes in Pezizomycotina were more common in lipid and amino acid metabolism. The effects of both GD and HGT were more pronounced in clustered genes than in their non-clustered counterparts and were differentially distributed across fungal lineages; specifically, GD, which was an order of magnitude more abundant than HGT, was most frequently observed in Agaricomycetes, whereas HGT was much more prevalent in Pezizomycotina. The effect of HGT in some Pezizomycotina was particularly strong; for example, we identified 111 HGT events associated with the 15 Aspergillus genomes, which sharply contrasts with the 60 HGT events detected for the 48 genomes from the entire Saccharomycotina subphylum. Finally, the impact of GD within a metabolic category was typically consistent across all fungal lineages, whereas the impact of HGT was variable. These results indicate that GD is the dominant process underlying fungal metabolic diversity, whereas HGT is episodic and acts in a category- or lineage-specific manner. Both processes have a greater impact on clustered genes, suggesting that metabolic gene clusters represent hotspots for the generation of fungal metabolic diversity.

Figure 1. Variation in gene clustering, GD, and HGT across the fungal phylogeny.

From top to bottom, the four box-and-whisker plots correspond to number of ECgenes per genome, percentage of clustered ECgenes per genome, percentage of horizontally transferred ECgenes per genome, and percentage of duplicated ECgenes per genome. The bottom and top of each box first and third quartiles (the 25th and 75th percentiles), respectively. The lower whisker extends from the box bottom to the lowest value within 1.5 * IQR (Inter-Quartile Range, defined as the distance between the first and third quartiles) of the first quartile. The upper whisker extends from the box top to the highest value that is within 1.5 * IQR of the third quartile. Data beyond the end of the whiskers are outliers and plotted as points. Numbers in parentheses after the lineages' names indicate numbers of genomes in each lineage; the numbers of genomes used from each lineage are also reflected by the widths of their branch triangles on the fungal species phylogeny shown at the bottom of the figure.

Figure 2. Over/underrepresentation of KEGG metabolic categories across three major fungal lineages.

From top to bottom, the box-and-whisker plots correspond to number ECgenes per genome, number of clustered ECgenes per genome, number of transferred ECgenes per genome, and number of duplicated genes per genome. Agaricomycetes boxes are colored blue, Saccharomycotina boxes are colored red, and Pezizomycotina boxes green. Box-and-whisker convention is as described in Figure 1. Up arrows under boxes indicate overrepresentation, and down arrows indicate underrepresentation of the corresponding metabolic category in the corresponding lineage. Significance of differential representation was estimated using a two-tailed Fisher's exact test using a Benjamini & Hochberg adjusted P value≤0.05 to account for multiple testing (Table S4).

Figure 3. The episodic occurrence of HGT across the fungal species phylogeny.

Numbers in parentheses indicate the number of HGT events and the number of genomes downstream of the collapsed nodes, respectively. Some clades have been collapsed for clarity; see Figure S2 for a depiction of the entire species phylogeny. The thickness and color of each branch corresponds to number of ECgenes transferred to each branch, adjusted by the number of genomes in the case of collapsed clades.

Figure 4. The association between gene innovation and gene clustering across three major fungal lineages.

Percentage of non-clustered (blue bars) and clustered ECgenes (red bars) inferred to have undergone GD (top) and HGT (bottom). Asterisks (*) indicate statistically significant differences determined using a Benjamini & Hochberg adjusted P value≤0.05 in a two-tailed Fisher's exact test (Table S6).

Figure 5. The fungal metabolic network of interactions between gene clustering and two major sources of gene innovation (GD and HGT).

Nodes of the metabolic network correspond to KEGG compounds. Thick edges of the metabolic network correspond to EC numbers from clustered ECgenes in one or more fungal species, whereas thin edges to EC numbers whose genes show no history of gene clustering. Colored edges correspond to EC numbers whose ECgenes have undergone HGT and GD (red), GD (blue), or show no history of GD or HGT (black). Note that none of the EC numbers in our dataset were affected by HGT alone. Pathway map created using iPATH2.0 .