Graduation Date

Fall 12-20-2019

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Programs

Medical Sciences Interdepartmental Area

First Advisor

Kishor K. Bhakat, Ph.D.

Abstract

Base excision repair (BER) pathway is required for the removal of damaged bases caused by alkylation, oxidation and ring-saturation. Human apurinic/apyrimidinic endonuclease 1 (APE1) plays a central role in BER pathway. Although repair of damaged bases by recombinant APE1 has been well investigated in vitro, how APE1 gains access to damaged bases in the context of chromatin is largely unknown. A prominent member of the histone chaperone family, FACT (Facilitates Chromatin Transcription) is thought to reorganize nucleosomes through the destabilization of multiple intra-nucleosome contacts. FACT complex is composed of two polypeptides identified as SPT16 (Suppressor of Ty 16) and SSRP1 (structure-specific recognition protein 1) that are both essential for nucleosome reorganization. Previous reports demonstrate that SPT16 is essential for transcription-coupled nucleotide excision repair (TC-NER) and homologous recombination repair (HRR). However, whether FACT complex interacts with APE1 and is involved in BER pathway remains unknown.

Here, we identified both subunits (SPT16 & SSRP1) of FACT complex as the prominent interacting partners of APE1. We show that the interaction of APE1 with FACT complex enhances upon induction of DNA damages. We demonstrate that FACT complex not only promotes the binding and subsequent acetylation of APE1 (AcAPE1), but also regulates the mobility and binding dynamics of APE1 to damage sites in chromatin. Furthermore, we found that FACT complex was required for efficient repair of DNA damages in BER pathway.

Given the prognostic significance of APE1 overexpression in various cancers, we test the translational potential of targeting FACT complex to interfere BER function in two tumor models, colon cancer and medulloblastoma, a most common brain tumor in children. We show that both APE1 and FACT levels are elevated in primary tumor tissue of colon cancer and medulloblastoma patients. A group of small molecules currently in phase II clinical trial, curaxins, is found to cause chromatin trapping effect of FACT complex. We demonstrate that curaxins inhibit FACT function and exhibits synergistic effect of tumor killing when combined with chemotherapeutic agent both in vitro and in vivo using xenograft models. Colon cancer and medulloblastoma were used as our primary tumor models.

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