Binding sites in mammalian genes and viral gene regulatory regions recognized by methylated DNA-binding protein

Abstract

Methylated DNA-binding protein (MDBP), a ubiquitous mammalian protein, recognizes a variety of related DNA sequences. Some of these sequences require methylation of their CpG dinucleotides for binding and others do not. We report that MDBP binds, in a DNA methylation-independent fashion, to two sites in the mouse polyomavirus enhancer, one in the enhancer of the human hepatitis B virus, and to one in the long terminal repeat of equine infectious anemia proviral DNA. We have also found a number of MDBP sites in human and rodent DNAs which bind much better to MDBP when they are methylated at CpG dinucleotides within the recognition site. These include sites at the beginning of the human genes for hypoxanthine phosphoribosyl transferase, HLA-A2, -A3, and -A25 antigens, and alpha-galactosidase A. In the case of methylation-responsive MDBP sites, changes in their methylation status during differentiation or DNA replication could help drive development by modulating transcription.

The Modulatory Effects of the Polymorphisms in GLA 5′-Untranslated Region Upon Gene Expression Are Cell-Type Specific

“…Both the c.-12(A>G) and the c.-10(C>T) SNPs are located within the GLA 5 0 UTR MDBP consensus sequence, respectively, at its first and third nucleotide positions (Zhang et al 1990), but only the c.-10(C>T) SNP seem to affect enzyme expression in vivo (Oliveira et al 2008a). The amount of aGal identified by western blot analyses of leukocyte protein extracts was significantly lower in carriers of the c.-10T allele as compared to carriers of the WT allele (Oliveira et al 2008b).…”

“…Because the GLA 5 0 UTR contains a binding site for methylated DNA-binding protein (MDBP)/regulatory factor X (RFX) transcription factor family (Zhang et al 1990;Samac et al 1998), as well as partially overlapping binding motifs for the nuclear factor kappa-B (NFkB) and E-26 (Ets) transcriptional regulatory factors (Saifudeen et al 1995;Fitzmaurice et al 1997), it has been previously suggested that protein ligands to these sites might be involved in the regulation of GLA gene expression (Saifudeen et al 1995;Fitzmaurice et al 1997). Since the SNPs c.-10(C>T) and c.-30(G>A) involve nucleotides respectively located at each of those binding sites, they might act as polymorphic modulators of GLA gene expression, with possible clinical relevance particularly in males carrying GLA sequence variants associated with high REA.…”

A mutation in the 5′ untranslated region of the human α‐galactosidase A gene in high‐activity variants inhibits specific protein binding

“…In addition to the oligonucleotides shown in Fig. 1, the following duplexes (written 5' to 3') were used, with only one strand of the sequence given and the MDBP or AP-2 sites in capital letters: EIAI, tttaGTTGCCTAGCAACatga; Py2, ggcaGTTGCCTAGCAACtaat; Etp, tcatGTCTCCATGGTAACttac (containing MDBP binding sites from the equine infectious anemia virus long terminal repeat, the polyomavirus enhancer B, and the ets-2 promoter, respectively; see [12,13] and unpublished results) and Ap-2, gggagcaaagGCCTGGGGTgca (see [14] and unpublished results).…”

“…Nuclear extracts were prepared as previously described [12] from HeLa or Raji cells grown in medium containing 10% fetal bovine serum except that the HeLa cells, where indicated, had been treated with 5/zM forskolin and 100 ng/ml of phorbol 12-myristate 13-acetate (PMA) for 1 h and then harvested. Abbreviations: ~-Gal A, a-galactosidase A; WT, wild-type; EMSA, electrophoretic mobility shift assay; kb, kilobase-pair; bp, base-pair; 5' UT, 5' untranslated; Lwt, specific protein binding site in the 5' UT region of the WT ot-GAL A gene; Lmu, mutant form of the Lwt site.…”

The kinase activity of c-Abl but not v-Abl is potentiated by direct interaction with RFXI, a protein that binds the enhancers of several viruses and cell-cycle regulated genes

“…EP is a key functional element of the hepatitis B virus (HBV) enhancer (Ostapchuk et al, 1989;Zhang et al, 1990aZhang et al, ,b, 1991Dikstein et al, 1990). A second component of the EP nucleo-protein complex is RFX1.…”