Organization and evolution of the Cyp2 gene cluster on mouse chromosome 7, and comparison with the syntenic human cluster - Toxicogenomics: article

Environmental Health Perspectives, Nov 15, 2003 by Haoyi Wang, Kyle M. Donley, Diane S. Keeney, Susan M.G. Hoffman

Genes from the cytochrome P450 (CY-P) superfamily encode a diverse group of monooxygenases that play important roles in both endogenous processes and in the metabolism of exogenous compounds, including most drugs. A cluster of Cyp2 genes on mouse chromosome 7 was mapped and analyzed in detail and compared with the homologous cluster on human chromosome 19. The mouse cluster includes 22 loci from the same six CYP2 subfamilies--Cyp2a, Cyp2b, Cyp2f, Cyp2g, Cyp2s, Cyp2t--that are found in the human cluster. Twelve of these loci are functional genes, and 10 are pseudogenes. Parts of the mouse and human gene clusters are similarly arranged, but the data indicate that a significantly different series of duplication events created the modern gene organizations in the two species. The comparison of the mouse and human clusters provides new insights into the evolution of gene families, whereas the detailed analysis provides background information that should be informative for future studies on the expression, regulation, and function of specific mouse Cyp2 genes. Key words: Cyp2a, Cyp2b, Cyp2f, Cyp2g, Cyp2s, Cyp2t, cytochrome P450, gene family, gene duplication. Environ Health Perspect 111:1835-1842 (2003). doi: 10.1289/txg.6546 available via http://dx.doi.org/[Online 24 September 2003]

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Genes from the ancient cytochrome P450 (CYP) superfamily, which encode a large and diverse group of heme-thiolate monooxygenases, are present in the genomes of almost all species examined to date. Mammalian CYP enzymes have been particularly well studied because they detoxify or activate a wide range of environmental and ingested compounds, including many drugs (Nelson et al. 1996), and they play important roles in many endogenous processes such as the metabolism of fatty acids, steroids, and eicosanoids (Nebert and Russell 2002). The CYP superfamily is also an excellent group for studying the evolutionary mechanisms that create gene families; previous studies of" this group have provided clear examples of the molecular processes, such as tandem duplication and gene conversion, that are considered to be most important in gene family evolution (Fernandez-Salguero et al. 1995; Gonzalez and Nebert 1990).

The CYP superfamily of genes has been divided hierarchically into families and subfamilies on the basis of sequence similarity, and there is a standardized nomenclature incorporating this hierarchy (Nelson et al. 1993, 1996). Families are designated by adding a number to the root "CYP" ("Cyp" in mouse, e.g., Cyp2), and subfamilies are indicated by a letter (e.g., Cyp2a). Genes within a subfamily are numbered in order of discovery, regardless of species, and pseudogenes (both partial and full-length) are named by' adding "ps" to the related mouse gene ("P" for other species) or by adding independent numbers if no true genes are highly related. In general this system appears to reflect evolutionary relationships among the loci (Lewis et al. 1998), with different CYP gene families found in different major taxonomic groups. In vertebrates, larger families are divided into subfamilies that are typically scattered across genomes, but multiple loci from within a subfamily are usually physically clustered together on a single chromosome (Nelson et al. 1996). This pattern has been interpreted as reflecting the creation of most new CYP loci by tandem duplication (Nelson et al. 1993) so that recently duplicated and therefore highly similar loci remain in tandem clusters, whereas older duplications have been broken up over evolutionary time by chromosomal rearrangements.

In this era of genome projects, it would seem relatively simple to analyze the organization and evolution of CYP gene clusters, based on available assembled sequences. However, even though both the human and mouse genome projects have now produced huge stockpiles of sequence information (Waterson et al. 2002), additional study is typically required for highly duplicated portions of mammalian genomes such as gene family clusters (Eichler 1998). Computer-based assemblies cannot by themselves accommodate the complexities presented by clusters of closely related genes, and both polymerase chain reaction (PCR)-based and hybridization-based analyses are confounded by the high levels of sequence similarity between paralogous loci. Mouse and human are estimated to have 190 and 115 CYP loci (including pseudogenes), respectively, some of which have very similar sequences (Hoffman and Keeney 2002). Thus, the study of these gene clusters has been best served by combining genomic sequencing with fine-scale physical mapping based on analyses of cloned DNA (Hoffman et al. 2001).

Detailed physical mapping using genomic clones proved to be an extremely fruitful approach for understanding one human gene cluster, the CYP2 cluster on chromosome 19 (Hoffman et al. 2001), which includes loci from the CYP2A, 2B, 2F, 2G, 2S, and 2T subfamilies (hereafter the CYP2A-T cluster). Extending this map-based approach to the mouse, the most important model organism for genetic studies in mammals, will allow researchers to better study the expression and variation of these genes. Until all individual genes and their related pseudogenes from a cluster have been analyzed in a species, it is nearly impossible to develop PCR primers that are sufficiently locus-specific to use for accurate genotyping, cloning into expression vectors, or development of knockout mice. This study is intended to provide a practical basis for future studies of CYP gene expression, as well as to make a contribution to our understanding of the mechanisms underlying gene family evolution.