engleski

Mechanisms of topoisomerase 1 DNA-protein crosslink repair

DNA-protein crosslinks (DPCs) consist of proteins that are irreversibly covalently bound to DNA and can be resolved by different repair pathways. Repair of DPCs is mainly studied at the cellular and biochemical level, while almost nothing is known at the level of organism. One of the most abundant DPCs in cells are Topoisomerase 1-DPCs (TOP1-DPCs). Trapped TOP1 interferes with all DNA transactions (replication, transcription, repair) and can cause double-strand breaks. In cancer therapy, many drugs have been developed that induce TOP1-DPCs and cause cell death. In humans, a mutation in the active site of TDP1 causes SCAN1 syndrome, a neurological disorder characterized by slowly progressive cerebellar ataxia in late childhood. Main repair proteins involved in TOP1- DPC repair are Tyrosyl DNA phosphodiesterase 1 (TDP1) and the protease SPRTN. Upstream SPRTN proteolysis of TOP1-DPC is considered to be a prerequisite for the action of TDP1, which hydrolyzes the covalent bond between tyrosine in the TOP1 peptide residue and the 3' end of DNA via its phosphodiesterase activity. To investigate whether SPRTN and TDP1 act within the same pathway at the organismal level, as suggested by previous in vitro studies, and to determine the dominant pathway for TOP1 DPC repair, we created a TDP1- deficient zebrafish line. Although the mutant embryos did not accumulate total DPCs, they had significantly higher amounts of TOP1 DPCs compared with wild-type embryos. In addition, even higher accumulation of TOP1 DPCs was observed after embryos were treated with camptothecin (CPT), a model TOP1- DPC inducer. We are currently investigating the interplay with SPRTN in TOP1-DPC repair by silencing SPRTN in TDP1-deficient embryos. This study and a novel TDP1 animal model will reveal the role of TDP1 in DPC repair at the level of organism and help us better understand diseases and cancer treatments related to the DPC repair pathway.

DNA, phosphodiesterase. TOP1, TDP1, camptothecin, SCAN1 syndrome, SPRTN

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