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

Abstract

S‐Adenosylhomocysteine hydrolase (AHCY) is the only mammalian enzyme known to catalyze the hydrolysis of S‐adenosylhomocysteine. We have used a genotype‐to‐phenotype strategy to study this important enzyme by resequencing AHCY in 240 DNA samples from four ethnic groups. Thirty‐nine polymorphisms were identified – 28 of which were novel. Functional genomic studies for wild type AHCY and the three variant allozymes identified showed that two variant allozymes had slight, but significant decreases in enzyme activity, but with no significant differences in levels of immunoreactive protein. Luciferase reporter gene assays for common 5′‐flanking region haplotypes revealed that one haplotype with a frequency of ∼2% in Caucasian‐American subjects displayed a decreased ability to drive transcription. The variant nucleotide at 5′‐flanking region single nucleotide polymorphism (SNP) (−34) in that haplotype altered the DNA‐protein binding pattern during electrophoresis mobility shift assay. Finally, an AHCY genotype‐phenotype association study for expression in lymphoblastoid cells identified four SNPs that were associated with decreased expression. For the IVS6 (intervening sequence 6, i.e., intron 6) G56 > C SNP among those four, electrophoresis mobility shift assay showed that a C > G nucleotide change resulted in an additional shifted band. These results represent a step toward understanding the functional consequences of common genetic variation in AHCY for the regulation of neurotransmitter, drug and macromolecule methylation.

Strain‐dependent dysregulation of one‐carbon metabolism in male mice is associated with choline‐ and folate‐deficient diet‐induced liver injury

“…Another important finding of the present study is the down-regulation of the transcription factor CEBP␤ in the livers of mice fed a CFD diet and the demonstration of its critical role in the regulation of the expression of the Mat1a, Ahcy, and Mthfd1 genes. These are 3 essential genes that intimately interconnect transmethylation, transsulfuration, and folate-dependent DNA synthesis pathways in one-carbon metabolism (52)(53)(54), and the down-regulation of any of these genes may impair the entire one-carbon metabolic pathway. The significant positive correlation between the levels of CEBP␤ protein and the expression of Mat1a, Ahcy, and Mthfd1 genes suggests its direct role in their down-regulation; however, we cannot exclude the possibility that under methyl-group donor-deficient conditions single nucleotide polymorphisms existing in these genes or an altered DNA methylation pattern in the promoter region of Mat1a, Ahcy, and Mthfd1 may also contribute to their lower expression.…”

Betaine-homocysteine methyltransferase: Human liver genotype–phenotype correlation

“…We also used “1000 Genomes Project” data to impute 230 additional SNPs across BHMT and BHMT2 (see Figure 5) and identified an additional SNP, rs7700970, that also displayed a high degree of association with the two phenotypes. Since SNPs 5′ of a gene, within the gene, or 3′ of a gene can all influence transcription [26, 27], we then performed EMS assays using probes that spanned each of these SNPs, but none of the four SNPs displayed a clear “gel shift” – leaving the mechanisms responsible for the association of these SNPs with BHMT expression unresolved.…”

Human Liver Methionine Cycle:MAT1AandGNMTGene Resequencing, Functional Genomics, and Hepatic Genotype-Phenotype Correlation

“…Finally, in an attempt to begin to understand the overall regulation of the methionine and folate cycles in human liver, we performed association analyses among levels of hepatic protein for other enzymes in these pathways that had also been assayed in the same 268 liver samples used to perform the present study. Specifically, we correlated BHMT and serine hydroxymethyltransferase 1 (SHMT1) protein levels as well as the level of enzyme activity for another important methyltransferase enzyme, catechol O-methyltransferase (COMT) (Feng et al, 2009;Zhang et al, 2009;Nordgren et al, 2011) with levels of MAT1A and GNMT protein expression (Table 4). Like MAT1A and GNMT, BHMT is predominantly expressed in hepatic tissue (Li et al, 2008).…”

“…Tissue samples from only one sex were used to eliminate the possibility of confusion as a result of sex-dependent differences in enzyme protein expression. Characteristics of the patients from whom the biopsies were obtained have been described previously (Feng et al, 2009;Zhang et al, 2009;Nordgren et al, 2011). Use of these anonymized surgical biopsy samples was reviewed and approved by the Mayo Clinic Institutional Review Board.…”

“…1), including MAT1A and GNMT, for use in genotyping DNA samples from the same 268 human liver biopsy samples that we had phenotyped for level of hepatic MAT1A and GNMT protein. Specifically, a total of 768 tag SNPs for methionine and folate cycle genes, including 42 for MAT1A and 5 for GNMT, were selected using SNP genotype data from 168 unrelated HapMap white subjects (http://www.hapmap.org, data release 27/phase II ϩ III) as well as data from our resequencing studies for these genes (Shield et al, 2004;Martin et al, 2006;Feng et al, 2009;Nordgren et al, 2011) and SNP data obtained by using Illumina 550K and 510S genome-wide BeadChips (Illumina, Inc., San Diego, CA) and Coriell Institute-generated Affymetrix 6.0 genome-wide SNP genotypes for DNA from the 96 EA subjects included in the Coriell Institute Human Variation Panel (Camden, NJ), the same cell lines from which we obtained the DNA for our gene resequencing experiments. The SNPs were selected to tag across the genes and to include approximately 20 kb of flanking sequence, with r 2 Ն 0.8 and a minor allele frequency of Ն0.025.…”