Role of bromodomain containing proteins in the DNA damage response

Gong, Fade
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Chromatin-based DNA damage response (DDR) mechanisms are fundamental for preventing genome and epigenome instability, which are hallmarks of cancer. How chromatin promotes genome-epigenome integrity in response to DNA damage is a critical question. Chromatin acetylation is a key signaling event involved in detecting, signaling and repairing DNA damage. The bromodomain (BRD) containing protein is the primary reader of acetylation. Thus, BRD proteins represent attractive candidates for reading damaged chromatin to mediate genome-epigenome integrity. In the first part of this project, I performed a screen to analyze the dynamics of BRD protein at DNA damage sites. I identified one-third of BRD proteins relocalized upon DNA damage, a phenomenon common to DNA damage factors. In the second part of my thesis work, I functionally studied the BRD protein ZMYND8 in a novel transcription-dependent DNA damage recognition pathway. Upon DNA damage specifically within actively transcribing chromatin, ZMYND8 is recruited through its BRD to TIP60 mediated H4 acetylations. ZMYND8 associates with the NuRD complex and promotes its accumulation at damage sites to facilitate transcriptional repression and promote repair by homologous recombination (HR). To investigate mechanisms regulating this novel ZMYND8-NuRD pathway, I performed another screen to check the recruitment of ZMYND8 interacting factors as well as their effects on ZMYND8 recruitment. I identified the H3K4me3 specific histone demethylase KDM5A is a key upstream regulator of this transcription-dependent DNA damage recognition pathway. Upon DNA damage, KDM5A mediates the removal of H3K4me3 around active chromatin near damage sites, which is an essential step to facilitate recruitment of ZMYND8 and NuRD complex to DNA damage. Similar to ZMYND8 and NuRD, depletion of KDM5A also impairs damage induced transcriptional silencing and DNA double-strand break (DSB) repair by homologous recombination (HR). The DDR is not only a dynamic process focusing that regulates protein factor interaction at DNA damage sites, but also promotes transcriptional changes of some genes upon DNA damage. In another part of this project, I screened the functional role of BRD proteins in regulating transcription in response to different types of damage. I identified two novel p53 target genes SP110 and SP140. In response to treatment with the DNA damaging chemotherapeutic agent, Doxorubicin, in U2OS cells, SP110 and SP140 are upregulated in a p53 dependent manner. In summary, this study provides a comprehensive view for BRD reader proteins in promoting the DDR within acetylated chromatin to preserve genome-epigenome stability.