Local structural transitions in DNA molecule upon damage and repair.
Duration: 2019 - 2022
Abnormal modifications of DNA chemical structure and composition, so-called DNA damage, may appear spontaneously or upon interaction with various damaging factors. Each cell is exposed to approximately 8000 DNA damage cases per one hour. DNA damage types might include missing bases or their chemical modifications, formation of the pyrimidine dimers, single strand breaks (SSBs), and the most dangerous for cells double strand breaks (DSBs). Unrepaired or repaired incorrectly DSBs may lead to chromosomal aberrations, mutations, or cellular death. Cells develop various DNA repair mechanisms, which have to work incessantly for the maintenance of the genomic integrity. Although the enzymology of DNA damage repair was intensively studied, many fundamental, structural aspects of DNA repair still remained elusive, such as the structures and orientations of protein complexes against DNA. Very important is the role of local conformational DNA transitions upon damage induction, or interaction with the repair proteins. DNA conformation determines the nature of interaction between DNA and the repair proteins. Therefore, local conformational transitions seem to play a key role in DNA repair processes. Apart from the induction of DNA damage and interaction with the repair proteins, the complex chromatin structure affects the susceptibility of DNA to changes of the conformation. Due to methodological limitations the local DNA conformational transition upon repair process is very poorly understood. Based on the existing knowledge, the following scientific hypothesis can be formulated: DNA conformation and chromatin integrity determine the susceptibility to DNA damage and repair. In the proposed project we are going to prove this hypothesis and complete the following project objectives: i) direct, experimental verification of the DSBs chemical structure inducted by bleomycin, ii) detailed description of the chemical bonds formed between DNA/chromatin and repair proteins, iii) understanding an influence of DNA methylation on the local DNA conformational transition, iv) development and optimization of DNA/chromatin nanoscale structural imaging in a liquid environment, v) monitoring, in real time, molecular changes of single DNA/chromatin strands upon bleomycin and introduction of repair proteins into the solution during nano-spectroscopic mapping, vi) finding molecular markers of cellular response to bleomycin treatment, especially related to synthesis of the repair proteins.