Rapid Birth–Death Evolution Specific to Xenobiotic Cytochrome P450 Genes in Vertebrates
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Rapid Birth–Death Evolution Specific to Xenobiotic Cytochrome P450 Genes in Vertebrates James H. Thomas* Department of Genome Sciences, University of Washington, Seattle, Washington, United States of America Genes vary greatly in their long-term phylogenetic stability and there exists no general explanation for these differences. The cytochrome P450 (CYP450) gene superfamily is well suited to investigating this problem because it is large and well studied, and it includes both stable and unstable genes. CYP450 genes encode oxidase enzymes that function in metabolism of endogenous small molecules and in detoxification of xenobiotic compounds. Both types of enzymes have been intensively studied. My analysis of ten nearly complete vertebrate genomes indicates that each genome contains 50–80 CYP450 genes, which are about evenly divided between phylogenetically stable and unstable genes. The stable genes are characterized by few or no gene duplications or losses in species ranging from bony fish to mammals, whereas unstable genes are characterized by frequent gene duplications and losses (birth–death evolution) even among closely related species. All of the CYP450 genes that encode enzymes with known endogenous substrates are phylogenetically stable. In contrast, most of the unstable genes encode enzymes that function as xenobiotic detoxifiers. Nearly all unstable CYP450 genes in the mouse and human genomes reside in a few dense gene clusters, forming unstable gene islands that arose by recurrent local gene duplication. Evidence for positive selection in amino acid sequence is restricted to these unstable CYP450 genes, and sites of selection are associated with substrate-binding regions in the protein structure. These results can be explained by a general model in which phylogenetically stable genes have core functions in development and physiology, whereas unstable genes have accessory functions associated with unstable environmental interactions such as toxin and pathogen exposure. Unstable gene islands in vertebrates share some functional properties with bacterial genomic islands, though they arise by local gene duplication rather than horizontal gene transfer. Citation: Thomas JH (2007) Rapid birth-death evolution specific to xenobiotic cytochrome P450 genes in vertebrates. PLoS Genet 3(5): e67. doi:10.1371/journal.pgen.0030067 Introduction The human genome contains approximately 60 CYP450 genes, which encode membrane-bound oxidase enzymes that Genes in animal and plant gene families vary greatly in act on a wide variety of substrates (recent reviews include [6– their phylogenetic stability. Some genes persist as a single 8]). The CYP450 enzymes can be divided into those that act copy over a wide phylogenetic range of species, with few or on endogenous small molecules and those that act on no gene duplications or losses on different evolving lineages. xenobiotic compounds. The endogenous-substrate enzymes Other genes undergo frequent duplication and loss in a function in biosynthesis or catabolism of steroids, sterols, process called birth-death evolution [1,2]. Gene duplication retinoids, prostaglandins, and fatty acids. The xenobiotic- and subsequent divergence of the duplicate copies underlie substrate enzymes are expressed primarily in the liver and the formation of gene families and are thought to be an epithelial tissues and function to defend against environ- important source of genetic complexity and evolutionary mental toxins and carcinogens. These xenobiotic CYP450 change [3–5]. Though many specific examples of stable and genes have been studied extensively in humans because they unstable genes have been documented, there exists no comprise one of the major (and most polymorphic) activities general explanation for their different patterns of evolution. that determine drug half-life and are thus important in To explore the evolutionary basis for these patterns, I sought pharmaceutical development [6,7]. Xenobiotic CYP450 en- a gene family with characteristics that make it amenable to zymes have also been studied extensively because in some detailed analysis and interpretation. The vertebrate cyto- chrome P450 (CYP450) gene superfamily is well suited for the Editor: Barbara J. Trask, Fred Hutchinson Cancer Research Center, United States of following reasons. First, it is a large family with both America phylogenetically stable members and unstable members. Received December 6, 2006; Accepted March 14, 2007; Published May 11, 2007 Second, the biochemical and organismal functions of most A previous version of this article appeared as an Early Online Release on March 14, genes in the family have been extensively studied in humans 2007 (doi:10.1371/journal.pgen.0030067.eor). and rodents. Third, their protein products are large and have Copyright: Ó 2007 James H. Thomas. This is an open-access article distributed a well-conserved domain content, providing abundant in- under the terms of the Creative Commons Attribution License, which permits formation for studies of molecular evolution. Finally, strong unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. interest in the family has resulted in relatively high quality annotations of gene structure in a wide range of vertebrate Abbreviations: CYP450, cytochrome P450; TBLASTN, translated BLAST nucleotide species. * To whom correspondence should be addressed. E-mail: [email protected] PLoS Genetics | www.plosgenetics.org0720 May 2007 | Volume 3 | Issue 5 | e67 Cytochrome P450 Birth-Death Evolution Author Summary 628 of these sequences, marked to indicate known or probable functions based on the human proteins. A repre- Genes vary greatly in their long-term phylogenetic stability, and sentative segment of the tree is shown in Figure 1. In the tree, there exists no general explanation for these differences. Stable CYP450 enzymes that act on endogenous substrates are genes persist as a single copy over a wide range of distantly related generally represented by a single gene in each organism, and species, whereas unstable genes undergo frequent duplication and the phylogeny of that gene approximates the known species loss in a process called birth-death evolution. The vertebrate phylogeny. In some cases there are two related proteins in cytochrome P450 (CYP450) gene superfamily includes many genes one or both fish species, presumably resulting from the that are present in a single copy in species ranging from teleost fish to mammals and other groups of genes that undergo active birth- known whole-genome duplication on the fish lineage [10,11]. death evolution across the same species. The author found that In all but one case, each orthologous group of genes is nearly all stable CYP450 genes encode enzymes known to function strongly separated on the tree from all other CYP450 genes, in the synthesis and degradation of steroid and retinoid hormones indicating that they became functionally specialized well (and related molecules). These hormones function in core devel- before the divergence of teleost fish from mammals (the opmental pathways in vertebrates. In contrast, most unstable single exception is described below). In sharp contrast, nearly CYP450 genes encode enzymes that detoxify foreign small all CYP450 enzymes that act on xenobiotic substrates are molecules (called xenobiotics—foreign biochemicals). In addition, encoded by genes that have undergone active duplication and many of the unstable CYP450 genes are subject to natural selection loss throughout vertebrate evolution, reflected in multiple to change their amino acid sequence over time (positive selection), species- and lineage-specific gene expansions. The tree probably in response to changes in xenobiotic exposure. These findings suggest that stable and unstable genes differ in their rates segment in Figure 1 includes two endogenous-substrate of birth-death evolution, because stable genes have core endoge- enzymes and one class of xenobiotic-substrate enzymes; they nous functions whereas unstable genes respond to changing aretypicalofthecompletetree(FigureS1).Table1 environmental conditions. summarizes gene numbers and variance across species for three groups of CYP450 genes. Among the endogenous-substrate ortholog groups, an cases they convert exogenous compounds into active carci- expected protein from a specific species was occasionally nogens, presumably as a negative side-effect of broad absent, usually from cow, chicken, or frog. One example is substrate specificity [9]. Approximately 22 of the human found in Figure 1 (a CYP46A1 cholesterol 24-hydroxylase CYP450 enzymes are thought to act primarily on endogenous ortholog is missing from cow). Most of these apparent gene substrates and approximately 15 other CYP450 enzymes are losses probably result from incomplete sequence assemblies thought to act primarily on xenobiotic substrates (Figure S1). or gene annotations. Such apparent gene losses were found A few of the remaining enzymes are reported to have activity most often in the cow, chicken, and frog genomes presumably toward both endogenous and xenobiotic substrates; it is as a result of relatively immature assemblies and annotations unclear whether one or the other activity (or both) is their of these genomes. In several cases, translated BLAST primary function. Finally, there are about ten CYP450 nucleotide (TBLASTN) searches of the cognate genome enzymes with no known activities, most of which have been clearly indicated the likely presence