Interference of benzo[a]pyrene diol epoxide-deoxyguanosine adducts in a GC box with binding of the transcription factor Sp1

Abstract

Previous studies indicated that DNA adducts formed by the carcinogenic diol epoxide 7r,8t-dihydroxy-9t,10t-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE) can increase the affinity of the transcription factor Sp1 for DNA sequences that are not normally specific binding sites. Whether adducts that form in the normal binding site, the GC box sequence, increase the affinity of Sp1 for the modified GC-box was not determined. Starting with a 23-nt sequence that contains two natural GC box sequences, site-specifically modified oligonucleotides were prepared with a single(+)-BPDE-deoxyguanosine adduct at one of three positions: the center of each GC-box or in between the two boxes. Four modified oligonucleotides were studied, two derived from cis addition of BPDE to the exocyclic amino group and two from trans addition. For three of these site-specifically modified oligonucleotides, there was a diminution in Sp1 affinity, whereas Sp1 binding to the fourth modified oligonucleotide was abolished. Furthermore, random modification of the oligonucleotide to a level of about 1 BPDE adduct per fragment slightly decreased the affinity for Sp1, and no evidence was found for a subpopulation of molecules with high affinity. These findings suggest that BPDE modification of the GC box does not lead to an increased affinity for Sp1. This is consistent with a model in which a BPDE-induced bend in the DNA mimics the conformation of the normal GC box:Sp1 complex, leading to high-affinity binding of Sp1 to non-Gc box sites.

High-affinity binding of the cell cycle–regulated transcription factors E2F1 and E2F4 to benzo[a]pyrene diol epoxide–DNA adducts

“…The central portions of the pseudo-palindromic E2F binding sites in the E 101 fragment are fairly rich in dG residues and would therefore be expected to be good targets for modification by BPDE. However, for two other transcription factors, Sp1 [22] and AP-1 [43], site-specific BPDE-DNA adducts located in the natural target sequences actually blocked binding. Similarly, the presence of a bulky ultraviolet light-induced photoproduct in the consensus sites for several transcription factors, including E2F, was shown to abolish specific binding [44], and distamycin blocks specific binding of homeobox proteins to their cognate DNA sequences [20].…”

“…In fact, the introduction of a single BPDE adduct at the central dG residue of a GC box target sequence can abolish Sp1 binding [22]. We have suggested that the introduction of a kink or "hinge" in DNA by a BPDE adduct may be important in inducing illegitimate binding sites for Sp1 [18].…”

The stereochemistry of benzo[a]pyrene‐2′‐deoxyguanosine adducts affects DNA methylation by SssI and HhaI DNA methyltransferases

“…In the case of DNA transcription, T7 RNA polymerase is blocked more efficiently by the (+)‐ trans ‐B[ a ]P‐ N 2 ‐dG adduct than by the (+)‐ cis ‐B[ a ]P‐ N 2 ‐dG adduct [17]. The binding of the transcription factor Sp 1 to B[ a ]PDE‐modified DNA is highly dependent on the B[ a ]P‐ N 2 ‐dG conformation [18], whereas no apparent differences in the binding affinities of the Ap 1 transcription factor to DNA containing different stereoisomeric B[ a ]P‐ N 2 ‐dG adducts was observed [19]. The B[ a ]P‐DNA adducts also affect the function of human topoisomerase I by alteration of DNA cleavage patterns [20].…”

The Antileukemia Drug 2-Chloro-2′-deoxyadenosine: An Intrinsic Transcriptional Antagonist

“…On the other hand, the presence of random nitrogen mustard (bis[2-chloroethyl]methylamine)-induced alkylations of guanines within a promoter region reduced the amount of elongated transcripts by Escherichia coli RNA polymerase (Gray et al, 1991). Moreover, numerous forms of DNA damage have been shown to prevent specific transcription factor binding interactions with their consensus sequences including 8-oxoguanine and benzo[a]pyrene diol epoxide (BPDE)-modified AP-1 binding sites (Parsian et al, 2002;Persson et al, 1996), BPDE-modified Sp-1 binding sequence (MacLeod et al, 1996), and nitrogen mustard-damaged Sp-1 and AP-2 consensus sites (Chen et al, 1999).…”