Grant: Discovery Grants Program - Individual

Recent research has revealed that the nucleolus, traditionally associated with ribosome production, plays a crucial role in stress responses, cell fate decisions, and the cellular reaction to DNA damage. This phenomenon, termed "nucleolar stress," involves disruptions of the nucleolar structure and functions due to factors such as impaired ribosomal RNA (rRNA) synthesis. Nucleolar stress affects various signaling pathways, including Mdm2-p53, NF-?B, and HIF-1a, making the nucleolus a key stress sensor and signaling hub. Nucleolar proteins, such as NPM and nucleostemin (NS), dynamically shuttle between the nucleolus and nucleoplasm, coordinating their functions. However, under nucleolar stress, these shuttling patterns can be disrupted, causing these proteins to be released from the nucleolus and trapping non-nucleolar proteins with it. DNA damage, in particular, severely impacts the nucleolus, which monitors genomic DNA stability. The highly transcribed rDNA regions in proliferating cells are especially vulnerable to DNA damage. DNA double-strand breaks and other types of damage can hinder Pol I transcription, leading to disrupted nucleolar dynamics and nucleolar stress. Additionally, various DNA repair proteins are localized within the nucleolus, such as PARP1, ATM, ATR, and Rad9B. These proteins play essential roles in DNA repair, further illustrating the intricate interplay between DNA damage and nucleolar homeostasis. Despite the dense nature of nucleoli, they are amenable to isolation and purification. Mass spectrometry-based proteomic studies have revealed a dynamic and diverse nucleolar proteome that extends beyond ribosome biogenesis-related proteins. This dynamic nature has sparked interest in comprehensively mapping the human nucleolar proteome, especially under different conditions. The research project's hypothesis posits that DNA damage-induced nucleolar stress redistributes nucleolar proteins to other cellular compartments, activating stress-related signaling pathways and influencing cell fate decisions. The project's objectives are as follows: 1.Utilize Data Independent Acquisition (DIA)-based mass spectrometry to profile the nucleolar proteome and understand post-translational modifications. 2.Investigate changes in nucleolar content under various DNA damage-related stress conditions using quantitative proteomics methods. 3.Employ proximity labeling techniques to track protein trafficking from nucleoli to other cellular compartments under different stress conditions using mass spectrometry-based proteomics. The significance of this program lies in gaining insights into the nucleolar proteome and its changes during stress conditions. This research aims to enhance our understanding of nucleolar function, stress response mechanisms, and cell fate determination, contributing to the broader field of cell biology and nucleolar biology.