Quantitative trait locus analysis reveals two intragenic sites that influence O6-alkylguanine-DNA alkyltransferase activity in peripheral blood mononuclear cells.
The repair of specific types of DNA alkylation damage by O6-alkylguanine-DNA alkyltransferase (MGMT) is a major mechanism of resistance to the carcinogenic and chemotherapeutic effects of certain alkylating agents. MGMT expression levels vary widely between individuals but the underlying causes of this variability are not known. To address this, we used an expressed single nucleotide polymorphism (SNP) and demonstrated that the MGMT alleles are frequently expressed at different levels in peripheral blood mononuclear cells (PBMC). This suggests that there is a genetic component of inter-allelic variation of MGMT levels that maps close to or within the MGMT locus. We then used quantitative trait locus (QTL) analysis using intragenic SNPs and found that there are at least two sites influencing inter-individual variation in PBMC MGMT activity. One is characterized by an SNP at the 3' end of the first intron and the second by two SNPs in the last exon. The latter are in perfect disequilibrium and both result in amino acid substitutions-one of them, Ile143Val, affecting an amino acid close to the Cys145 residue at the active site of MGMT. Using in vitro assays, we further showed that while the Val143 variant did not affect the activity of the protein on methylated DNA substrate, it was more resistant to inactivation by the MGMT pseudosubstrate, O6-(4-bromothenyl)guanine. These findings suggest that further investigations of the potential epidemiological and clinical significance of inherited differences in MGMT expression and activity are warranted.
Margison
Geoffrey P
GP
Cancer Research-UK Carcinogenesis Group, Paterson Institute for Cancer Research, Christie Hospital NHS Trust, Manchester, UK. gmargison@picr.man.ac.uk
Heighway
Jim
J
Pearson
Steven
S
McGown
Gail
G
Thorncroft
Mary R
MR
Watson
Amanda J
AJ
Harrison
Kathryn L
KL
Lewis
Sarah J
SJ
Rohde
Klaus
K
Barber
Philip V
PV
O'Donnell
Paul
P
Povey
Andrew C
AC
Santibáñez-Koref
Mauro F
MF
D001483:Base Sequence; D003062:Codon / Q000235:genetics; D017931:DNA Primers; D005091:Exons / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic; D020022:Genetic Predisposition to Disease; D005838:Genotype; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000201:enzymology*; D008175:Lung Neoplasms / Q000235:genetics; D019853:O(6)-Methylguanine-DNA Methyltransferase / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide; D040641:Quantitative Trait Loci
D003062:Codon; D017931:DNA Primers; D019853:O(6)-Methylguanine-DNA Methyltransferase