Description
The main role of DNA molecules is information storage. DNA is continuously exposed to the endogenous and environmental damaging agents (mutagens). Unrepaired lesions might lead to aging or carcinogenesis. This DNAtraffic Project is a research project focused on structural and functional characterization of DNA damage and repair proteins in order to annotate their function in DNA repair pathways. DNA repair involves many factors in cell. The plan is to study the role of AlkB human homologues towards their binding and removal in a direct manner of the mutagen-induced lesions. However, many of the proteins involved in DNA metabolism are very well described but information about different factors involved in DNA repair are scattered in the literature. Moreover, there are still processes, for which an enzymatic activity is known or suspected to exist, but the proteins have not been characterized yet. The character of the project is strictly scientific, however its influence on medical science and human health can’t be excluded.
Summary of project results
UV-radiation is an environmental agent that can induce a variety of mutagenic and bulky DNA lesions. Other bulky lesions are caused by cisplatin, which is a widely used drug. The success of cisplatin in cancer chemotherapy derives from its ability to crosslink DNA. However, the mechanism of the cisplatin-induced drug resistance is unknown. To counteract the DNA damage, all living organisms have developed a number of highly conserved DNA repair mechanisms. AlkB proteins are known to repair DNA via direct manner. The UV-induced damage were not considered as a substrates for direct repair by AlkB proteins, however, we decided to study its function in the repair process. Also, the cisplatin-induced crosslinks were not considered as a substrates for direct repair by AlkB proteins, however, the silencing of ALKBH2 enhances the sensitivity of human lung cancer cells to cisplatin-based chemotherapy. We purified bacterial AlkB, and human ALKBH2 and ALKBH3 proteins and tested them to recognize UV-induced DNA damage and cisplatin intrastrand crosslinks. We showed that human ALKBH2 and ALKBH3 proteins can bind to UV-damaged DNA in contrast to bacterial AlkB. Next, we showed that only ALKBH2 protein in contrast to bacterial AlkB and human ALKBH3 produced the additional peak when examined by HPLC in towards to modify cisplatin damage. Having our experimental data concerning HPLC analysis of ALKBH2 on the platinium adduct we performed a docking model. It showed that the damage can bind to the active site of ALKBH2. We do believe that our results can shed new light into the field of DNA repair. The DNA repair area is of scientific interest because DNA is a target for anticancer drug design and tumor therapy. We planned to work on UV-damag,e but using as a control some other agent (cisplatin) causing the bulky lesion we also noticed that the DNA damage can be modified by ALKBH2 protein. This observation needs to be studied in details, but for sure the results will be published and accessible for community. The results of the project duration were presented by two persons during three international scientific conferences. Moreover, the results obtained will be published (2016) in two peer reviewed international journals. Moreover, the project was a chance for a female researcher to be a principal investigator and leader of an independent scientific topic. Three PhD students had a chance to improve their tasks and learn the new techniques.
Summary of bilateral results