Human S-adenosylhomocysteine hydrolase: common gene sequence variation and functional genomic characterization.

Methylation plays a critical role in monoamine neurotransmitter biosynthesis and metabolism (Weinshilboum et al. 1999). Methyltransferase (MT) enzymes involved in these process include catechol O-methyltransferase (COMT (Axelrod & Cohn 1971, Axelrod & Tomchick 1958, Lachman et al. 1996), phenylethanolamine N-methyltransferase (PNMT) (Axelrod 1962a, Axelrod & Cohn 1971, Ji et al. 2005), indolethylamine N-methyltransferase (INMT) (Axelrod 1962b, Thompson et al. 1999), and hydroxyindole O-methyltransferase (HIOMT) (Axelrod et al. 1965, Donohue et al. 1993). All of these MT enzymes utilize S-adenosyl-L-methionine (AdoMet) as a methyl donor (Chiang et al. 1996), and S-adenosylhomocysteine (AdoHcy) is generated during the methyltransferase reaction (Weinshilboum 2006). AdoHcy is a competitive inhibitor of all AdoMet-dependent MT enzymes (Borchardt 1980). The ratio of AdoMet to AdoHcy is sometimes referred to as “methylation potential” (Loehrer et al. 1998). The accumulation of AdoHcy decreases this ratio, resulting in the inhibition of MT enzymes (McKeever et al. 1995a). In the brains of both human and pigs, methylation is significantly reduced with increased AdoHcy concentrations (Goggins et al. 1999, McKeever et al. 1995a, McKeever et al. 1995b). Therefore, it is important that AdoHcy be degraded in vivo to maintain methylation. S-Adenosylhomocysteine hydrolase (AHCY, EC 3.3.1.1) catalyzes the reversible hydrolysis of AdoHcy to form adenosine (Ado) and homocysteine (Hcy) (Castro et al. 2005, Palmer & Abeles 1976). Although the synthetic direction is favored thermodynamically, the reaction proceeds in the hydrolytic direction in vivo because of the rapid removal of Hcy and Ado (Cantoni 1975, Palmer & Abeles 1976, Castro et al. 2005) (Fig. 1). Reduced AHCY activity can result in elevated AdoHcy levels, with a resultant reduction in the AdoMet to AdoHcy ratio and impaired methylation (Loehrer et al. 1998). As a result, understanding the functional implications of common genetic polymorphism in the gene encoding AHCY could provide insight into individualized variation in methylation. Figure 1 The Methionine Cycle. AHCY = S-adenosylhomocysteine hydrolase; AD = adenosine deaminase; BHMT = betaine homocysteine methyltransferase; CBS = cystathione β-synthase; MAT = methionine adenosyltransferase; MTR = 5-methyltetrahydrofolate-homocysteine ... Human AHCY has 11 exons and maps to the long arm of chromosome 20. Deletion of AHCY is an embryonic lethal in mice (Miller et al. 1994). Mutations in the human AHCY gene that are associated with impaired function result in elevated AdoMet levels and even more dramatically elevated AdoHcy concentrations (Baric et al. 2005, Baric et al. 2004, Buist et al. 2006). All of these patients demonstrated severe myopathy, developmental delay and hypermethionemia. Case-control studies have reported several common AHCY sequence variants (Gellekink et al. 2004), although follow-up functional studies have generally been lacking. In the present study, we set out to resequence the AHCY gene using 240 DNA samples from four different ethnic groups, followed by functional genomic characterization of the polymorphisms observed. Specifically, we resequenced all exons and splice junctions as well as the 5′-flanking region (5′-FR) of the gene, and identified 39 single nucleotide polymorphisms (SNPs) – 28 novel, including 3 nonsynonymous SNPs. The functional implications of the nonsynonymous SNPs were then evaluated as well as the transcriptional activity of common AHCY 5′-FR haplotypes. Finally, microarray analysis was performed with the lymphoblastoid cell lines from which the resequenced DNA had been obtained to investigate the possible relationship between AHCY expression and polymorphisms in the gene. These studies represent a comprehensive attempt to link sequence variation in the human AHCY gene with function as a step toward a more complete understanding of the relationship among common AHCY genetic polymorphisms and variation in methylation.