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Endogenous DNA lesions can inhibit the binding of the AP-1 (c-Jun) transcription factor
Daniel K Rogstad1, Pingfang Liu, Artur Burdzy
1Department of Biochemistry and Microbiology, Loma Linda University School of Medicine, Loma Linda, CA 92350, USA.
Abstract:
The repair of DNA damage, caused by both endogenous and exogenous sources, is necessary to remove lesions that either miscode or block DNA or RNA polymerases. We propose that damage also must be repaired to maintain sequence-specific DNA-protein interactions. In this paper, we have systematically studied two lesions that interfere with one important DNA landmark, the thymine methyl group. Oxidation of the thymine methyl group in DNA generates 5-hydroxymethyluracil (HmU) whereas the misincorporation of dUMP into DNA generates uracil (U), replacing the methyl group with a hydrogen. Both substitutions are shown to inhibit binding of the AP-1 (c-Jun) transcription factor. The energy cost of the perturbation, approximately 0.4 kcal/mol, is similar in magnitude for both U and HmU substitutions and is additive when multiple substitutions are present. A third lesion, substitution of the central C:G base pair of the AP-1 DNA binding domain with the pro-mutagenic U:G mispair, unexpectedly increases AP-1 binding, allowing the transcription factor to interfere with uracil DNA glycosylase activity. Our results support the hypothesis that an additional role for DNA repair systems is to maintain the integrity of sequence-specific DNA-protein interactions, a role of particular importance in long-lived organisms.
Insights
DNA repair is crucial for maintaining protein interactions, not just preventing coding errors. Lesions like uracil and 5-hydroxymethyluracil disrupt transcription factor binding, highlighting repair
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- DNA damage from internal and external sources necessitates repair to prevent polymerase obstruction and miscoding.
- Sequence-specific DNA-protein interactions are vital for cellular regulation and must be preserved.
- The thymine methyl group is a key DNA feature targeted by specific types of damage.
Purpose of the Study:
- To investigate how DNA damage, specifically alterations to the thymine methyl group, affects DNA-protein binding.
- To determine the energetic impact of uracil and 5-hydroxymethyluracil substitutions on transcription factor binding.
- To explore the consequences of a uracil:guanine mispair within a DNA binding site.
Main Methods:
- Systematic study of DNA lesions affecting the thymine methyl group.
- Analysis of AP-1 (c-Jun) transcription factor binding to damaged DNA substrates.
- Thermodynamic analysis to quantify the energy cost of DNA-protein interaction perturbations.
Main Results:
- Both uracil (U) and 5-hydroxymethyluracil (HmU) substitutions inhibit AP-1 binding with a similar, additive energy cost (~0.4 kcal/mol).
- A U:G mispair substitution within the AP-1 binding site unexpectedly enhanced binding.
- Enhanced binding by the U:G mispair allowed AP-1 to interfere with uracil DNA glycosylase activity.
Conclusions:
- DNA repair systems play a critical role in maintaining sequence-specific DNA-protein interactions.
- Preserving these interactions is particularly important for the function of DNA repair in long-lived organisms.
- Damage can paradoxically alter protein binding, underscoring the complexity of DNA repair's role.