Rewiring of posttranscriptional RNA regulons: Puf4p in fungi as an example

Abstract

It has been increasingly clear that changes in gene regulation play important roles in physiological and phenotypic evolution. Rewiring gene-regulatory networks, i.e., alteration of the gene-regulation system for different biological functions, has been demonstrated in various species. Posttranscriptional regulons have prominent roles in coordinating gene expression in a variety of eukaryotes. In this study, using Puf4p in fungi as an example, we demonstrate that posttranscriptional regulatory networks can also be rewired during evolution. Although Puf4p is highly conserved in fungi, targets of the posttranscriptional regulon are functionally diverse among known fungal species. In the Saccharomycotina subdivision, target genes of Puf4p mostly conduct function in the nucleolus; however, in the Pezizomycotina subdivision, they are enriched in the mitochondria. Furthermore, we demonstrate different regulation efficiencies of mitochondrial function by PUF proteins in different fungal clades. Our results indicate that rewiring of posttranscription regulatory networks may be an important way of generating genetic novelties in gene regulation during evolution.

FIG. 1.

The evolution of Puf proteins in fungi. The phylogenetic tree for species in fungi was reconstructed based on ten conserved proteins (the bootstrap scores are displayed in Supplementary fig. 1, Supplementary Material online, and the abbreviations for the studied species are shown in Supplementary table 2, Supplementary Material online). The tree was consistent with the results of a previous study (Fitzpatrick et al. 2006). The color in each cell denotes the number of PUM-HD repeats for that gene in the respective species. Gray means that there was no detectable orthologous gene. The arrows indicate the WGD event and the Saccharomycotina clade, respectively.

FIG. 2.

Functional rewiring of the PUF4 posttranscriptional regulon. (A) Enrichment of P4E in each functional category of genes (column) in each fungal species (row). Colors denote the P values for the enrichment of P4E after multiple test correction, as indicated at the bottom. The phylogenetic tree of the studied fungal species is shown to the left. The Saccharomycotina (pink) and Pezizomycotina (light blue) subdivisions are indicated by arrows. Two other fungal clades, Schizosaccharomyces and Basidiomycota, are indicated as dark blue and green, respectively. Yellow lines separate fungal species into four categories based on the evolutionary pattern of the PUF4 posttranscriptional regulon. (B) The average percentage of nucleolar genes with P4E in each group of fungal species.

FIG. 3.

Deep functional divergence between Puf3p and Puf4p. (A) The phylogenetic tree of the PUM-HD domains from the Puf3p and Puf4p orthologs (bootstrap scores are presented in Supplementary fig. 3, Supplementary Material online). Glam-Puf protein is used as the out-group. The branches to Puf3p and Puf4p in each species were coded in red and blue, respectively. (B) Functional conservation of Puf3p and Puf4p orthologs in fungi. The conserved amino acids for the repeat #3 and #5 of the PUM-HD domain are displayed using ClustalX (Thompson et al. 1997). The red (C ↔ T) and blue (R ↔ C) arrows denote the conserved amino acids in Puf3p and Puf4p that are critical for different motif recognition between these two proteins. (C) Little functional overlapping between Puf3p and Puf4p. The y axis denotes the ratio of genes having both P3E and P4E over the total number of genes with P3E and/or P4E in each species (x axis).

FIG. 4.

Posttranscriptional regulation of mitochondrial genes in fungi. (A) The occurrence of P3E and P4E in mitochondrial genes. The x axis denotes each studied fungal species (the Group 4 species, which include the Saccharomycotina and Pezizomycotina subdivisions, were put in the middle of fig. 4). The y axis denotes the percentage of all Puf3p or Puf4p target genes in each genome that are dedicated to mitochondrial regulation. (B) The correlation of Puf3p and Puf4p mitochondrial regulation in fungi. All mitochondrial genes were divided into four groups based on the number of species in the Pezizomycotina subdivision that have the P4E motif. The x axis represents the group of mitochondrial genes. The y axis is the average percentage of species in the Saccharomycotina subdivision that have P3E for each orthologous mitochondrial gene group. The mean and standard deviation for each category are shown.

FIG. 5.

Evolution of the PUF posttranscriptional regulons in plant and animals. (A) The occurrence of P4E in the nucleolar genes of plant and animals. (B) The occurrence of P3E in the nucleolar genes of plant and animals. (C) The occurrence of P3E in the mitochondrial genes of plant and animals. (D) The occurrence of P4E in the mitochondrial genes of plant and animals. P4E and P3E were detected at the 1,000-bp 3′UTR regions in each species. Green bars denote the percentage of genes with the motif in the studied organelle and blue bars denote the percentage of genes with the motif at the whole-genome level.