Mechanistic insight into a novel TIF-IA-NF-kB nucleolar stress response pathway and elucidation of its role in senescence

This proposal focuses on a structure inside the cell called the nucleolus. This structure is dysfunctional in many common disease, including Alzheimer's, Parkinsons disease and cancer. It is also known to play a role in aging. If the environment of a cell changes, for example if nutrients are decreased or the cell is exposed to toxic agents, nucleoli respond and send signals to other parts of the cell to change the rate of growth/death and many other cellular processes. However, how nucleoli sense changes in the environment, and the signals they send to the other parts of the cell, remain poorly understood.

We have recently published a new method by which nucleoli respond to stress that involves degradation of a critical nucleolar protein called TIF-IA. We have also demonstrated that this degradation causes an increase in nucleolar size and simulates a protein in another part of the cell called NF-kB. Since increased nucleolar size and NF-kB stimulation are connected with ageing, we looked to see if TIF-IA-NF-kB may be important in this process using a model called oncogene-induced senescence (OIS). Here we aim to gain full understanding of how stresses degrade TIF-IA. We have exciting preliminary data to suggest that this degradation occurs by a novel mechanisms and so, this will be the focus of these studies. We also aim to identify the signals that link changed levels of TIF-IA to altered nucleolar size and NF-kB stimulation. Again we have preliminary data that has revealed some candidates that may be involved in this process. Finally, and importantly, we aim to confirm the role of TIF-IA-NF-kB in senescence.

Given that nucleolar dysfunction and NF-kB pathway activation are both markers of ageing and contribute to poor health, these studies have the very real potential of increasing our understanding of this process and identifying biomarkers that could be used to identify diseased or ageing cells. They may also open up avenues for a new class of drugs that act by targeting both these pathways simultaneously.

The nucleolus is a highly dynamic nuclear organelle which acts as a critical stress sensor and coordinates downstream responses to stress such as metabolism, differentiation, senescence and apoptosis. Perturbations in nucleolar function are associated with many common diseases and nucleolar size and number are emerging as key markers of longevity. Indeed, proper dynamic control of nucleolar activity is crucial for maintaining tissue homeostasis and health. However, understanding of the regulatory networks that control the activity of nucleoli, and those that link altered function to changes in cell physiology, are poorly understood. In this lab, we have recently identified a novel nucleolar stress response pathway which involves degradation of the critical PolI complex component, TIF-IA and subsequently, increased nucleolar size and activation of NF-kB signalling. We have shown this TIF-IA-NF-kB pathway is important in regulating cell growth/death and our preliminary data suggest it also has a role in senescence Here we aim to: 1. Elucidate the mechanisms that regulate stress-mediated degradation of TIF-IA. Our preliminary data suggest for the first time that the protein is degraded in an autophagy dependent manner and so, studies will focus on this mechanism. 2. Identify the signalling networks that link TIF-IA degradation to increased nucleolar size and activation of the NF-kB pathway. 3. Fully establish the role of TIF-IA-NF-kB signalling in senescence. The nucleolar phenotype we observe in response to stress parallels that observed in ageing cells, which also show hyperactive NF-kB. The three complementary aims outlined in this proposal will provide a deep understanding of how environmental changes can alter nucleolar function through TIF-IA, and how nucleoli signal to the NF-kB pathway. This could have significant implications for our understanding of the aetiology of ageing and identify biomarkers of this process and therapeutic targets.

The beneficiaries of this research will be:
1. Academia: As outlined in the academic beneficiaries section.
2. Pharmaceutical companies: Uncontrolled NF-kB activity is a contributory factor in a number of common diseases associated with aging. Therefore, pharmaceutical companies are extremely interested in developments that may lead to novel methods to inhibit this activity. Similarly, the nucleolus is dysfunctional in a number of age-related pathologies and is emerging as a potential drug target. The work outlined in this project will benefit the development of agents that inhibit these pathways in 2 ways: Firstly, we aim to develop a high throughput phenotypic assay to simultaneously measure NF-kB activity and nucleolar size . Here we aim to use the assay to test candidate genes but going forward, this assay would be ideal for drug screens to identify agents that act on NF-kB through altered nucleolar function. Secondly, the mechanistic studies outlined have the very real potential of identifying novel targets for drugs that act on both these pathways simultaneously. This will be especially interesting to companies developing agents to combat cancer and neurological disorders and could have a major impact on the economy and societal health.
3. Patients and health professionals: We will use aspirin as a model stress inducer. Identifying the mechanisms by which this agent inhibits nucleolar function may have an immediate impact on colorectal cancer patients as it may reveal biomarkers of response in clinical trials of chemopreventative agents. In the long term, it may lead to the development of novel agents that could be used to prevent this disease in high risk patients.
4. General public: Media publicity of grant awards and published work will raise awareness of aspirin prevention of cancer and may prompt the general public to question their risk.