Evolution of a global regulator: Lrp in four orders of γ-Proteobacteria

Abstract

Background: Bacterial global regulators each regulate the expression of several hundred genes. In Escherichia coli, the top seven global regulators together control over half of all genes. Leucine-responsive regulatory protein (Lrp) is one of these top seven global regulators. Lrp orthologs are very widely distributed, among both Bacteria and Archaea. Surprisingly, even within the phylum γ-Proteobacteria (which includes E. coli), Lrp is a global regulator in some orders and a local regulator in others. This raises questions about the evolution of Lrp and, more broadly, of global regulators.

Results: We examined Lrp sequences from four bacterial orders of the γ-Proteobacteria using phylogenetic and Logo analyses. The orders studied were Enterobacteriales and Vibrionales, in which Lrp plays a global role in tested species; Pasteurellales, in which Lrp is a local regulator in the tested species; and Alteromonadales, an order closely related to the other three but in which Lrp has not yet been studied. For comparison, we analyzed the Lrp paralog AsnC, which in all tested cases is a local regulator. The Lrp and AsnC phylogenetic clusters each divided, as expected, into subclusters representing the Enterobacteriales, Vibrionales, and Pasteuralles. However the Alteromonadales did not yield coherent clusters for either Lrp or AsnC. Logo analysis revealed signatures associated with globally- vs. locally- acting Lrp orthologs, providing testable hypotheses for which portions of Lrp are responsible for a global vs. local role. These candidate regions include both ends of the Lrp polypeptide but not, interestingly, the highly-conserved helix-turn-helix motif responsible for DNA sequence specificity.

Conclusions: Lrp and AsnC have conserved sequence signatures that allow their unambiguous annotation, at least in γ-Proteobacteria. Among Lrp orthologs, specific residues correlated with global vs. local regulatory roles, and can now be tested to determine which are functionally relevant and which simply reflect divergence. In the Alteromonadales, it appears that there are different subgroups of Lrp orthologs, one of which may act globally while the other may act locally. These results suggest experiments to improve our understanding of the evolution of bacterial global regulators.

Keywords: Alteromonadales; Enterobacteriales; Pasteurellales; Phylogenomics; Transcription factors; Vibrionales.

Fig. 1

Role of Lrp superimposed on core genome phylogeny. Five orders of the γ-Proteobacteria are shown, adapted (with permission) from a maximum likelihood tree generated by Gao et al. [8], and based on the concatenated sequences of 36 highly-conserved proteins. They used both maximum parsimony (MP) and maximum likelihood (ML) approaches, and the two numbers are the proportion of the puzzling quartets (ML)/% bootstrap scores (MP) that supported the given node. For each order, the colored shading and text to the right indicates the role played by Lrp in tested species (green = global, pink = local), and the tested species are also indicated. For two orders, indicated by “?” and yellow shading, the role of Lrp has not yet, to our knowledge, been tested

Fig. 2

Phylogeny and comparison of the paralogs Lrp and AsnC. Maximum likelihood phylogeny was constructed using the a Lrp and b AsnC protein sequences. The numbers above or below the internal branches show bootstrap values (%). Color keys indicate the different orders: magenta = Enterobacteriales (Ent), orange = Vibrionales (Vib), green = Alteromandales (Alt), red = Pasteurellales (Pas). c Logo comparison of all 40 Lrp vs. all 40 AsnC sequences. Areas visually identified as showing conserved differences are shaded in cyan

Fig. 3

Comparison of Lrp orthologs grouped by order. a The ten Lrp sequences from each order were used to generate aligned Logos, in order to compare globally- (Glb at right) and locally-acting (Loc) orthologs. The orders are abbreviated: Ent = Enterobacteriales, Vib = Vibrionales, Alt = Alteromonadales, Pas = Pasteurellales. The vertical arrows indicate positions of lysine acetylation (blue, from [47]) or formation of the coregulator binding pocket (red). See text for details. b Two-sample Logo comparing the global (Ent + Vib) and local (Pas) Lrp orthologs. Letters between the lines indicate amino acid residues that are conserved in both sets, symbols above the lines are selectively enriched in the globally-acting Lrp set, and symbols below the lines are selectively enriched in the locally-acting Lrp set

Fig. 4

Phylogeny of conserved housekeeping genes. Maximum likelihood phylogeny constructed for a 16S rRNA, b RpoB, and c RecA from the four bacterial orders. Colors are as assigned for Fig. 1

Fig. 5

Visualization of residues of interest in context of Lrp 3D structure. The program VMD 1.9.2 was used to visualize half of an octameric ring of E. coli Lrp subunits (from PDB 2GQQ). VMD is developed with NIH support by the Theoretical and Computational Biophysics group at the Beckman Institute, University of Illinois at Urbana-Champaign. a-d are successive 90° rotations about the vertical axis. The topmost subunit has cyan spheres highlighting residues associated with Lrp-specific signatures (see Fig. 2c), the next subunit is shown in red without additional highlighting, the next subunit shows in orange spheres the lysines that can be acetylated (see Fig. 3a), and the bottom subunit shows in green spheres the residues associated with globally-acting Lrp orthologs (see Additional file 1: Table S1 for position numbers of all highlighted residues)