Conservation and evolvability in regulatory networks: the evolution of ribosomal regulation in yeast

Abstract

Transcriptional modules of coregulated genes play a key role in regulatory networks. Comparative studies show that modules of coexpressed genes are conserved across taxa. However, little is known about the mechanisms underlying the evolution of module regulation. Here, we explore the evolution of cis-regulatory programs associated with conserved modules by integrating expression profiles for two yeast species and sequence data for a total of 17 fungal genomes. We show that although the cis-elements accompanying certain conserved modules are strictly conserved, those of other conserved modules are remarkably diverged. In particular, we infer the evolutionary history of the regulatory program governing ribosomal modules. We show how a cis-element emerged concurrently in dozens of promoters of ribosomal protein genes, followed by the loss of a more ancient cis-element. We suggest that this formation of an intermediate redundant regulatory program allows conserved transcriptional modules to gradually switch from one regulatory mechanism to another while maintaining their functionality. Our work provides a general framework for the study of the dynamics of promoter evolution at the level of transcriptional modules and may help in understanding the evolvability and increased redundancy of transcriptional regulation in higher organisms.

Fig. 1.

Conserved transcriptional modules in S. pombe and S. cerevisiae and their associated cis-elements. Shown are the S. cerevisiae and S. pombe modules for the six key conserved modules we identified, together with the cis-elements enriched in the promoters of these modules' genes. For each module, the profile shows the module genes (rows) induced (red) and repressed (green) across different experiments (columns). Rectangles indicate the orthologous genes, their number, and the P value of their cooccurrence. The enriched cis-elements associated with each module are shown in the sequence logo above or below it. (a) S phase module, associated with the conserved Mlu1 cell cycle box element (ACGCGT, bound by orthologous MBF complexes in both species), and an S. cerevisiae-specific element. (b) Respiration module, associated with the conserved HAP2345 site (CCAATCA, bound by the orthologous Hap2345 and Php2-5 complexes). (c) Amino acid metabolism module, associated with the conserved GCN4 site (TGACTCA; Supporting Materials and Methods, note 2). (d) Ribosomal proteins module associated with RAP1 (TACATCCGTACAT) and IFHL sites (TCCGCCTAG) in S. cerevisiae and with a Homol-D box (TGTGACTG) and a Homol-E site (ACCCTACCCTA) in S. pombe. (e) Stress module associated with the STRE site (AGGGG) in S. cerevisiae and the CRE site (ACGTCA) in S. pombe. (f) Ribosome biogenesis module, associated with the conserved element RRPE (AAAAATTTT) and the S. cerevisiae-specific PAC element (GCGATGAG).

Fig. 2.

Evolution of the regulatory mechanisms in the highly conserved module of ribosomal proteins. Phylogenetic cis-profile of the RP module. A schematic phylogenetic tree (branches are not drawn to scale) representing the known phylogeny (14) of the 17 analyzed species is shown, together with the sequence logos of the main cis-elements enriched in each module's promoters, grouped into three distinct types (colored boxes): RAP1 (orange), IFHL (blue), and Homol-D (red). The total number of genes in each POM is shown in parentheses, and the number of genes that contain each motif is indicated as well. Although the RP module is phenotypically extremely conserved, the phylogenetic cis-profile reveals a gradual switch from a Homol-D-dominated mechanism to a RAP1-controlled one, beginning before the speciation of A. gossypii. Concomitantly, the IFHL site underwent gradual sequence divergence and possible dimerization or domain duplication of the corresponding transcription factor.

Fig. 3.

Mechanisms for evolutionary change in the regulation of ribosomal proteins. (a) Rap1p sequence evolution. A scaled schematic representation of Rap1p sequences is shown for eight species and the human protein. Colored ovals indicate the presence and position of BCRT (orange), Myb (DNA binding, pink), silencing (olive), and TA (dark green) domains. The DNA binding Myb domain is present in all species, but the transactivation domain is apparent only in those species that harbor the RAP1 motif in their RP module genes (S. cerevisiae, S. castellii, K. waltii, A. gossypii, and all of the intermediate species; data not shown). The TA domain is absent from all species lacking the RAP1 element in RP promoters, including C. albicans, N. crassa, A. nidulans, and S. pombe. A Rap1p ortholog cannot be identified in Y. lipolytica, and no significant homology was found to the TA domain for the D. hansenii Rap1p (data not shown). (b) The Homol-D-RAP1 cis-regulatory module. Shown is a scaled schematic representation of the 35 promoters of the A. gossypii RP genes with the highest scoring Homol-D elements. Colored bars indicate the Homol-D (red) and RAP1 (orange) sites. The two sites are extremely close, with the RAP1 trailing the Homol-D site by 2-6 bp, indicating a possible interaction between their corresponding transcription factors.

Fig. 4.

Alternative modes for the evolution of the regulation of transcriptional modules. Each panel shows a distinct scenario of the inferred evolution of an ancestral regulatory program (Upper) into programs observed in 2 or more extant species (Lower). For each module, a schematic representative promoter is shown (black line) along with cis-elements (boxes) and transcription factors (ovals). Ancestral conserved sites and proteins are in light yellow, and innovations and divergences are in bright yellow or red. (a) Conservation of both the cis-element and trans-factors [e.g., the S phase (a2) and respiration (a1) modules]. (b) A gradual divergence of binding site sequence (e.g., the IFHL site in the RP module). (c) Augmentation of an existing program by the emergence of a new site along an ancestral one [e.g., the RRPE and PAC sites in the ribosomal biogenesis (RB) module]. (d) Abridgement of an augmented program by binding site loss (e.g., the loss of the TC site in the RB module). (e) Switching of the transcription factor while maintaining the same cis-element (e.g., the AA metabolism module). (f and g) Full switching of a program from one cis-element to another (e.g., the stress and the RP modules). In some cases (f), this can occur by a combination of augmentation and abridgement.