Michael Goldstein Lab

Role of short-chain histone acylations in DNA damage response

Responses to DNA damage are important determinants of cell viability and mutagenesis. Following DNA damage induction, an intricate network of DDR signaling pathways is activated to enforce cell cycle arrest, DNA repair, and potentially cell death. The ability of a cell to detect and repair damaged DNA has a direct effect on cell survival after exposure to genotoxic stress. Importantly, many agents that are clinically used to treat cancer function by damaging DNA. Therefore, the ability of tumor cells to cope with DNA damage induced by radiation and chemotherapy affects the efficacy of these treatment modalities and disease outcomes. DNA double-strand breaks (DSBs) are the most toxic DNA lesions and are considered the major trigger of cell death induced by radiation and chemotherapy. Consequently, a specific inhibition of DSB repair in cancer cells can improve radiation response and the overall outcome in cancer patients.

Epigenetic alteration of chromatin including chromatin structure changes and post-translational modifications (PTMs) of histone proteins collectively form a complex epigenetic landscape regulating transcription, replication, and DDR. We have discovered that eviction of the H2A/H2B histone dimers is facilitated by nucleolin at the DSB sites, an event that is critical for repair of the DNA breaks by non-homologous end-joining. Additionally, multiple histone PTMs have been identified that promote recruitment of DDR factors to the break. For instance, a simultaneous methylation of H4K20 and ubiquitination of H2AK15 is required for 53BP1 recruitment and, consequently, DSB repair demonstrating how an intricate histone PTM network facilitates DDR.

Acetylation of histone lysine residues occurs frequently throughout the entire epigenome and plays an important role in transcription and DDR. A recent discovery of novel short-chain histone lysine acylations including propionylation, crotonylation succinylation and β-hydroxy-butyrylation has extended the arsenal of histone modifications. Increasing evidence suggests that these PTMs regulate cellular processes such as transcriptional activity. We are studying the role of these novel histone modifications in cellular response to radiation-induced DSBs. Our goal is to identify specific histone sites that are modified by these acylations at the DSB. Further, we are interested in dissecting the molecular mechanisms through which these modifications regulate DDR. We are employing multiple experimental techniques including laser-microirradiation, immuno-fluorescence, endonuclease-based DNA repair assays and chromatin immunoprecipitation in order to elucidate the role of these histone PTMs in DDR.

Targeting chemotherapy resistance in glioblastoma

Glioblastoma is one of the most frequent and the most aggressive primary malignant brain affecting both, pediatric and adult patients. Despite some advances in treatment, prognosis of glioblastoma remains poor with less than 5% of patients surviving 5 years after diagnosis. An adjuvant temozolomide chemotherapy combined with radiotherapy represents the standard-of-care for glioblastoma. Thus, tumor response to radiation and temozolomide is critical for patient survival. Temozolomide is an alkylating agent that induces methylation of DNA bases. Whereas, radiation directly induces DSBs, temozolomide-induced DNA lesions have to be converted into DSBs in a mismatch repair and proliferation dependent manner.

Glioblastoma is characterized by a frequent deregulation of epigenetic pathways due to mutations of histone modifiers. We are investigating epigenetic pathways that when defective result in glioblastoma resistance to the standard-of-care chemoradiotherapy. Furthermore, we are developing strategies to restore chemoradiotherapy response in these tumors by using targeted therapeutics. This investigation will contribute to the personalized medicine approach in treatment of glioblastoma by tailoring treatment based on the individual mutation profile of epigenetic modifiers and has a potential to improve outcomes of this aggressive disease.

Epigenetic modifiers promoting radiation response in cervical cancer

Radiation therapy is an important treatment modality of locally advanced cervical cancer. Thus, understanding the mechanisms of radiation response in cervical cancer is critical for development of new treatment strategies that can overcome radiation resistance and improve survival in patients. Radiation kills cancer cells by inducing toxic DNA doubles-strand breaks (DSBs).   Increasing evidence suggests that epigenetic changes at the sites of radiation-induced DSBs play a critical role in DNA repair. Identifying epigenetic pathways that are involved in radiation response in cervical cancer cells can result in development of new therapeutics that can overcome radiation resistance and improve the outcomes of the disease.

Infection with high-risk HPV subtypes, including HPV 16 and 18, is a common driver of cervical carcinogenesis, associated with over 80% of cervical cancers. The oncogenic properties of the HPV genome have been attributed to the E6 and E7 gene products that affect multiple cellular processes including cell cycle, DNA repair and epigenetic pathways such as histone acetylation and methylation. Consequently, expression of high-risk E6 and E7 genes results in an altered epigenetic landscape in HPV+ cancers.

We are using CRISPR/Cas9-based screening techniques to identify epigenetic modifiers that are involved in radiation response in HPV+ and HPV- cervical cancer. Further, we are elucidating the molecular mechanisms through which these epigenetic modifiers regulate DDR. Our ultimate goal is to identify new targets for radiosensitization that are specific for HPV+ and HPV- tumors.

DNA damage response after photon versus proton irradiation

Proton radiotherapy is an emerging treatment for medulloblastoma due to its ability to restrict radiation doses to the tumor, while sparing healthy tissues. Whereas proton radiation also kills tumor cells by generating DSBs, proton-induced DNA breaks differ from those induced by conventional γ-radiation (photon radiation). A characteristic of proton radiation is its high linear energy transfer (LET) in irradiated tissues, resulting in highly complex, clustered DNA lesions that persist for a longer period of time. Clustered DSBs require orchestration of multiple DNA processing and repair pathways, including NHEJ, HR, and base excision repair (BER) for repair and survival of proton- induced DNA damage.

We are studying the differences in molecular responses to DNA damage induced by photon versus proton radiation. These investigations will extend our knowledge of radiation biology and identify potential targets for sensitization of tumors to different types of radiotherapy.