Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.1.1.37 (DNA methyltransferase)
 4,983 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have recently reported that following depletion of O6-methylguanine DNA methyltransferase (MGMT) activity by acute streptozotocin (STZ) treatment to sensitize innately chloroethylnitrosourea (CENU)-resistant HT-29 cells, the eventual repletion of activity occurs with no concommitant alterations in steady-state MGMT mRNA levels. This suggestion of a potentially stable transcript prompted studies to define the relative contributions of MGMT mRNA stability and transcription to cellular MGMT expression. Northern analysis of MGMT mRNA in actinomycin D-treated HT-29, MR-1 and A2182 cells, ranging in relative MGMT expression from high to low respectively, demonstrates relatively long MGMT mRNA half-lives of > 10-12 h. Cell lines with low and moderate levels of MGMT mRNA appear to have longer mRNA half-lives than those with high levels. Run-on transcription in nuclei isolated from cells with low to moderate MGMT mRNA levels demonstrates undetectable basal MGMT transcription rates. Collectively these data suggest that a very low transcription rate, coupled with a stable mRNA molecule, might result in the translation of pre-existing mRNA molecules. This translation may be responsible for the gradual recovery of MGMT and CENU resistance over 24 h following MGMT depletion.
Carcinogenesis 1995 Sep
PMID:The role of mRNA stability and transcription in O6-methylguanine DNA methyltransferase (MGMT) expression in Mer+ human tumor cells. 755 86

Our previous studies of Bacillus subtilis showed that the genes responsible for the adaptive response to DNA alkylation were organized as a divergent regulon, in contrast to scattered operons in Escherichia coli ada regulon. To study the generality and diversity of gene organization, several species and strains of Bacillus were examined for the responsiveness to DNA alkylation. B. cereus cells exhibited the highest resistance to MNNG treatment. When the cells were grown in the presence of MNNG, 3-methyladenine DNA glycosylase and two species of DNA methyltransferase were induced as in B. subtilis 168 cells. B. licheniformis 749 and B. amyloliquefaciens H cells exhibited a partial response that manifested itself as the induction of one species of DNA methyltransferase. On the other hand, B. thuringiensis var. Tohokuensis, B. megaterium KMT, and B. subtilis W23 cells were totally deficient in this response, and were hypersensitive to alkylating agents. To determine the cause of this deficiency in strain W23, we examined the genomic structure of the corresponding region where three genes (alkA, adaA, and adaB) were located in 168. No homologues for the three genes were detected in W23 DNA by Southern hybridization. Two genes (glmS and ndhF) flanking the adaptive response regulon in 168 were also present in W23. A sequence of about 2750 bp that carried the entire regulon in 168 was replaced with a sequence of about 250 bp that was unique to W23. At the ends of the conserved segments, palindromic sequences corresponding to the transcriptional termination sites of the adaB and glmS genes were observed. The regulon in 168 could be artificially replaced by the W23 sequence, and be regained through DNA-mediated transformation.
Mutat Res 1995 Sep
PMID:Diverse capacities for the adaptive response to DNA alkylation in Bacillus species and strains. 756 65

Differences in the methylation patterns of male and female gamete DNA are likely to be involved in genomic imprinting. However, little is known of the mechanisms that regulate de novo methylation and demethylation during gametogenesis. We report here that the well-characterized M(r) 190,000 form of DNA methyltransferase (the only known form) is present in isolated mitotic, meiotic, and postmeiotic male germ cells, with the exception of meiotic pachytene spermatocytes, where the protein is undetectable by immunoblot analysis and a novel 6.2-kb DNA methyltransferase transcript is present. Whereas replication and methylation are coupled in somatic cells, the presence of DNA methyltransferase in postreplicative male germ cells is consistent with previously observed de novo methylation events in these cells. Immunofluorescence experiments revealed that DNA methyltransferase is localized to the nuclei of male germ cells, with a subset of spermatogonia and postreplicative leptotene/zygotene spermatocytes displaying prominent nuclear foci that are strongly enriched in DNA methyltransferase. The data suggest that down-regulation of DNA methyltransferase expression during the pachytene stage of meiosis utilizes an mechanism that is associated with the production of a larger mRNA, and that de novo methylation in leptotene/zygotene spermatocytes may take place in spatially restricted nuclear domains that are enriched in DNA methyltransferase.
Biol Reprod 1995 Sep
PMID:Regulated synthesis and localization of DNA methyltransferase during spermatogenesis. 757 80

To assess the possibility that two conserved amino acids (glutamine 90 and asparagine 137) in O6-methylguanine-DNA methyltransferase (MGMT) are involved in protein-substrate contact and/or discrimination between favored and non-favored substrates, families of proteins mutant at these two sites were expressed in alkyltransferase-deficient bacteria and analyzed for stability, ability to repair O6-methylguanine (MG)-containing DNA, and ability to differentially repair a preferred (MG-containing DNA) versus a non-preferred (free base MG) substrate. All seven proteins mutant at glutamine 90 (except a proline mutant) were stable in bacteria and repaired MG-containing DNA (> 50% of wild-type levels). A representative glutamine 90 mutant protein was not, however, significantly different from the wild-type protein in the preferential repair of MG-containing DNA versus MG free base. Of eight proteins mutant at asparagine 137, only glutamine and serine mutants repaired MG-containing DNA to any degree (8.5% and 0.8% of wild-type respectively) and only the glutamine mutant protein was detectable in bacterial sonicates by Western blot analysis. Alanine and leucine mutant alkyltransferases, inactive and unstable as non-fusion proteins, could, however, be stably expressed in bacteria as glutathione S-transferase fusion proteins, although the proteins were still inactive in repair. These results suggest that while glutamine 90 has no direct role in MG-DNA methyltransferase-mediated repair or free base/lesioned DNA substrate specificity, asparagine 137 is important in both the stability and activity of the protein and may contribute to the formation or function of the active site of the protein.
Carcinogenesis 1994 Sep
PMID:The role of two conserved amino acids, glutamine 90 and asparagine 137, in O6-methylguanine-DNA methyltransferase stability, activity and substrate specificity. 792 83

EcoP15I DNA methyltransferase (Mtase) recognizes the asymmeteric sequence CAGCAG and catalyzes the transfer of a methyl group from S-adenosyl-L-methionine to the second adenine residue. We have investigated the DNA binding properties of EcoP15I DNA Mtase using gel mobility shift assays. EcoP15I DNA Mtase binds approximately threefold more tightly to DNA containing its recognition sequence, CAGCAG, than to non-specific sequences in the absence or presence of cofactors. Interestingly, in the presence of ATP the discrimination between specific and non-specific sequences increases significantly. These results suggest for the first time a role for ATP in DNA recognition by type III restriction-modification enzymes. In addition, we have shown that bromodeoxyuridine-containing oligonucleotides form complexes with EcoP15I DNA Mtase that are crosslinked upon irradiation. More importantly, we have shown that the crosslink site is at the site of DNA binding, since it can be suppressed by an excess of unmodified oligonucleotide. EcoP15I DNA Mtase exhibited Michaelis-Menten kinetics with both unmodified and bromodeoxyuridine-substituted DNA, with a higher specificity constant for the latter. Furthermore, gel mobility shift assays showed that proteolyzed EcoP15I DNA Mtase formed a specific complex with DNA, which had similar mobility as the native protein-DNA complex. Taken together these results form the basis for a detailed structure-function analysis of EcoP15I DNA Mtase.
J Mol Biol 1994 Sep 30
PMID:Interaction of EcoP15I DNA methyltransferase with oligonucleotides containing the asymmetric sequence 5'-CAGCAG-3'. 793 97

This study was undertaken to ascertain the importance of prolonged depletion of O6-methylguanine DNA methyltransferase (MGMT) activity, following O6-benzylguanine (BG) and streptozotocin (STZ) exposure, in reversing 1,3 bis(2-chloroethyl)-1-nitrosourea (BCNU) resistance in vitro. We evaluated BCNU-induced cytotoxicity and measured the temporal recovery of MGMT activity in human colon carcinoma HT-29 cells following treatment with BG, STZ, or the combination of BG and STZ. The pretreatment regimens which provided the greatest potentiation of BCNU cytotoxicity were those exhibiting the greatest temporal inhibition of MGMT activity. The combination of BG (10 microM) and STZ (1.0 mM) produced sustained inhibition of MGMT activity through 24 h and potentiated BCNU cytotoxicity by at least one log greater than either agent alone. Similarly, BG (10-100 microM) produced marked reductions in MGMT activity and increased BCNU cytotoxicity in a dose-dependent fashion. A 100-microM dose of BG inhibited MGMT activity for 48 h and potentiated BCNU induced cell kill by 3 logs greater than BCNU alone. In addition, we observed that during the period of sustained inhibition of MGMT activity, no changes in the steady-state MGMT mRNA levels occurred. We conclude that prolonged inhibition of MGMT activity is an important determinant in reversing BCNU resistance and that chemotherapeutic regimens targeting the inactivation of MGMT activity should be optimized such that MGMT activity is depleted for at least 24 h following BCNU administration.
Cancer Res 1993 Sep 15
PMID:Prolonged depletion of O6-methylguanine DNA methyltransferase activity following exposure to O6-benzylguanine with or without streptozotocin enhances 1,3-bis(2-chloroethyl)-1-nitrosourea sensitivity in vitro. 836 24

The human O6-methylguanine DNA methyltransferase (MGMT) repairs O6-methylguanine (O6-MG) in DNA at a much lower rate than the Escherichia coli Ada protein, and only MGMT repairs the altered base, O6-benzylguanine (O6-BG). The diversity in DNA repair properties between MGMT and Ada may be a result of divergent amino acid sequences outside their common proline-cysteine-histidine-arginine-valine (PCHRV) acceptor site. One notable sequence difference is an MGMT 28-amino acid carboxyl-terminal tail which is highly conserved among all mammalian alkyltransferases. The role of this tail sequence in substrate specificity was assessed by expressing full-length MGMT and Ada proteins, and mutant MGMT proteins lacking either 10 or 28 amino acids from the carboxyl terminus, as GST fusion proteins in alkyltransferase-deficient E. coli cells, and comparing rates of repair of O6-MG containing DNA and O6-BG by these fusion proteins at 4 degrees C and 37 degrees C. The MGMT carboxyl-terminal tail was not required for repair of O6-MG in DNA at 37 degrees C although the deletion of this tail sequence reversibly inhibited the ability of MGMT to repair O6-MG in DNA at 4 degrees C. Therefore, the absence of this region affects the ability of the protein to repair O6-MG in DNA at lower temperatures. Furthermore, removal of the tail sequence from MGMT decreased the rate of O6-BG repair 5-fold. We conclude that the 28-amino acid carboxyl-terminal MGMT tail, while not required for activity, modulates the rate of MGMT repair at reduced temperatures and plays a role in substrate specificity.
J Biol Chem 1993 Sep 15
PMID:The role of the carboxyl-terminal tail in human O6-methylguanine DNA methyltransferase substrate specificity and temperature sensitivity. 836 18

In Bacillus subtilis, the adaptive response to DNA alkylation depends on the ada operon, which consists of the adaA and adaB genes, which encode methylphosphotriester DNA methyltransferase (AdaA protein) and O6-methylguanine DNA methyltransferase (AdaB protein), respectively. A structural gene (alkA) that encodes 3-methyladenine DNA glycosylase was found upstream of the ada operon, but in the opposite orientation. This cluster of genes was mapped at about 235 kb from the SfiI recognition site near the origin of replication in the physical map of the B. subtilis chromosome. Disruption of the alkA gene sensitized cells to N-propyl-N'-nitro-N-nitrosoguanidine, while its overproduction rendered cells highly resistant to N-propyl-N'-nitro-N-nitrosoguanidine, indicating that lethal DNA damage produced by bulky alkylating agents was effectively counteracted by AlkA glycosylase. Transcription of the alkA gene was induced by treating adaA+ cells with methylating agents concurrent with transcription of the ada operon. This was accomplished by using methylated AdaA protein bound to a 30-bp segment in the middle of the 100-bp sequence between the transcriptional start sites of the alkA gene and ada operon. Thus, in this organism, the adaptive response to DNA alkylation is achieved by autologous activation of a divergent regulon composed of the genes for a DNA glycosylase and two species of DNA alkyltransferase.
J Bacteriol 1993 Sep
PMID:Bacillus subtilis alkA gene encoding inducible 3-methyladenine DNA glycosylase is adjacent to the ada operon. 837 46

Normal mammalian development requires a diploid combination of both haploid parental genomes. Uniparental disomy for certain segments of specific chromosomes results in aberrant development or prenatal lethality, indicating that the parental genomes have undergone modifications during gametogenesis. These modifications result in parent-of-origin specific expression for some genes, a phenomenon called genomic imprinting. Recent work with DNA methyltransferase deficient mice showed that differential methylation is the probable basis of the imprinted character of several genes. Screening for endogenous imprinted loci using restriction landmark genomic scanning with methylation sensitive enzymes (RLGS-M) identified eight imprinted RLGS (Irigs) candidate loci. Molecular analysis of the genomic region of one of the loci (Irigs2) resulted in the discovery of the paternally imprinted U2afbp-rs gene within a previously identified imprinted region on mouse chromosome 11 (refs 5, 7). This paper describes the characterisation of a novel imprinted RLGS-M locus, Irigs3, on mouse chromosome 9 (ref. 6). Within this locus we identified the Grf1 (also called Cdc25Mm) gene, which is homologous to the RAS-specific guanine nucleotide exchange factor gene, CDC25, in Saccharomyces cerevisiae. Grf1 is located about 30 kb downstream of the methylation imprinted site, identified by RLGS-M, and shows paternal allele specific expression in mouse brain, stomach and heart. Our results indicate that imprinting may have a role in regulating mitogenic signal transduction pathways during growth and development.
Nat Genet 1996 Sep
PMID:Identification of Grf1 on mouse chromosome 9 as an imprinted gene by RLGS-M. 878 30

We have characterized the inhibition exerted by histone H1 on the activity of human placenta DNA (cytosine-5-)-methyltransferase. Our experiments demonstrate that the extent of inhibition depends on the DNA base composition, AT-rich substrates being more severely affected than GC-rich substrates and CpG-rich islands. With bacterial SssI methylase, the effect is completely reversed since its activity on AT-rich substrates undergoes a 4-5-fold stimulation upon the addition of H1. Poly(L-lysine) mimicks H1 effects, suggesting an essential role of lysine residues in both the inhibitory and stimulatory effects of H1. By comparison of the different behaviors of the two enzymes, the inhibitory effect over the eukaryotic enzyme might be accounted for by hypothesizing a competition between minor groove-binding motifs (SPKK-like) present in placenta methylase as well as in histone H1.
Biochemistry 1996 Sep 10
PMID:Different effects of histone H1 on de novo DNA methylation in vitro depend on both the DNA base composition and the DNA methyltransferase. 879 46


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