Role of Protein N-Myristoylation in Driving DNA Damage Response

PROJECT SUMMARY/ABSTRACT The DNA damage response (DDR) is crucial for maintaining genome stability, with its dysregulation implicated in various diseases. Post-translational modifications (PTMs) orchestrate DDR, influencing vital signaling pathways for genomic stability. N-myristoylation, catalyzed by N-myristoyl transferase (NMT) 1 and 2, has emerged as a pivotal PTM that involves a range of biological processes. Despite its potential significance, a knowledge gap persists regarding the role of N-myristoylation in DDR. My preliminary data reveal a link between reduced N-myristoylation expression and heightened genomic instability. Utilizing quantitative proteomics, 14 N- myristoylated candidates within key DDR proteins were identified, with four overlapping the interactome of NMT1 and NMT2. Validation studies confirmed N-myristoylation on KU70 and RPA70, the key proteins in the double- strand break repair. These findings form the basis for the hypothesis that N-myristoylation actively propels DDR pathways by influencing the dynamics and functions of key proteins pivotal to DDR regulation. This project systematically investigates the role of N-myristoylation in DDR. Aim 1 delves into the association of N-myristoylation, modulated by NMT1 and NMT2, with genomic stability and DDR. This includes assessing the impact of N-myristoylation on DNA repair mechanisms and dissecting the interactome of NMT1 and NMT2 in response to various DNA damage agents. Aim 2 focuses on the detailed characterization of N-myristoylation on KU70, including the identification of specific N-myristoylation sites, evaluation of their impact on protein localization and stability, elucidation of functional consequences in DNA repair, and investigation of regulatory mechanisms. Transitioning into the R00 phase in Aim 3, I will build upon the insights gained during the K99 phase, leveraging them to validate and characterize additional N-myristoylated DDR candidates identified through proteomic screening. This phase aims to develop therapeutic implications based on a comprehensive understanding of N-myristoylation's regulatory role in DDR, enhancing the translational potential of the research. The successful completion of this work holds the promise of significantly advancing our understanding of the intricate regulatory networks governed by N-myristoylation in DDR. Beyond scientific contributions, this award provides comprehensive training in both my technical and professional skills under the guidance of experts in my mentoring committee, fostering proficiency in the DNA repair field and disease therapy development, thereby preparing me for the journey of establishing and running a successful independent laboratory.