Structural basis of human TRIAP1/PRELI function in mitochondrial lipid transport and apoptosis

Mitochondria are the power generator for a cell as they are able to convert energy into usable forms. They also play important roles in the orchestrating cell growth and as well as programmed cell death. Diseases such as cancer arise when a series of unwelcome changes occur in some of these cellular processes, causing the cell to multiply out of control. Before we can identify what goes wrong in cancers it is necessary to understand exactly how our cells normally work. Mitochondrial function requires a highly coordinated supply of proteins and fat-like molecules called phospholipids. In humans two families of proteins, called TRIAP1 and PRELI, play a key role in maintaining the lipid balance in mitochondria. They interaction together in a tight complex, which promotes the transfer of important lipid precurcers from the outer mitochondrial membrane to the inner. TRIAP1/PRELI also play a role in preventing cells from entering into programmed cell death (known as apoptosis) when the cell is under stress. For example, women with breast cancer who produce a lot of TRIAP1 respond poorly to chemotherapy by virtue of their cancer cell being protected from the drug-induce death. In this proposal, we will propose to study at the atomic level how TRIAP1/PRELI perform these functions. By using nuclear magnetic resonance (NMR) and X-ray diffraction methods, the shape and flexibility of relevant protein complexes will be imaged in solution. Insight from these structural studies will shed light on the mechanisms that underlie fundamental aspects of cellular regulation. In turn, this would lead to a better understanding of how the body responds to drug treatments in cancer and may lead to new treatments ways of monitoring the progress of current treatments.

Mitochondria are dynamic organelles involved in a variety of cellular processes, such as energy. Mitochondria can also orchestrate cell death (apoptosis) via release of cytochrome c and activation of the caspase signalling cascade. Cancer cells often display misregulation in apoptosis, contributing to the generation of chemotherapy-resistant cells. Thus, mitochondria are at the centre of a plethora of cellular processes both under normal physiological circumstances and in disease.

Phospholipid composition plays an important role in mitochondrial homeostasis. Changes in lipid content and structure have been associated with mitochondrial dysfunction and a variety of diseases including ischemia, hypothyroidism, aging, and heart failure. The family proteins (Ups1-3 in yeast and PRELIs in humans) together with a novel CX9C chaperone (Mdm35 in yeast and TRIAP1 in humans) regulate the accumulation of phospholipids within mitochondrial membranes. This pathway has been shown to be important for modulating apoptotic through two independent mechanisms. One indirectly through resultant increased cardiolipin accumulation and sequestration of cytochrome C, which is required for apoptosome formation. The other is through a direct interaction with Hsp70 and APAF1 thereby preventing cytochrome C from assembling in the apoptosome. These mechanisms stall apoptosis and gives cell to the opportunity to repair DNA damage

This proposal aims to characterize the structure and interactions of human TRIAP1/PRELI using NMR and X-ray crystallography approaches. The structural and dynamic properties of TRIAP1 with PRELI proteins and Hsp70 will be probed by site-directed mutagenesis and functional assay. The outcomes of this research project will provide new insight into the biological roles for the TRIAP1/Mdm35 family of proteins, which have been implicated in mitochondrial lipid homeostasis and cytotoxic drug-resistance through the evasion of apoptosis.

The outcomes of this research project will provide new insight into the biological roles for the TRIAP1/PRELI family of proteins, which have been implicated in mitochondrial lipid homeostasis and cytotoxic drug-resistance through the evasion of apoptosis. . The applicants SM will take responsibility for maximizing impact and methods of how we will ensure that they have the opportunity to benefit from this research are described

General
We will make every effort to ensure that research is disseminated widely by the Open Access publication in high-impact journals, presentation at international research meetings, the development of new collaborations where appropriate and deposition of data into freely-accessible databases.
Pharma and wider academic community
Although this proposal describes basic science, in anticipation of findings with direct relevance to the pharmaceutical industries with an interest in the therapeutics against anti-apoptotic effects TRIAP1/PRELI. We will approach UK players in this field to gauge interest in the project outcomes. We will explore the targetability of TRIAP1/PRELI and liaise translational offices at Imperial. Where appropriate we also contribute to Industrial open-days at Imperial. All structural metadata will be freely available on-line through standard repositories and findings will be published in a timely fashion. Data will be presented at specialised international research conferences, often prior to publication.

Public sector health care professionals and research networks
We will engage with clinicians and scientists professionals through networks available through the Medical School at Imperial College and Institute of Cancer Research.

Skills, training and knowledge economy
The PDRA, technician and any undergraduate, postgraduate (PhD and MRes), or part-time students that contribute to the project will develop key interdisciplinary skills that will be extremely valuable for UK industry, contribute to the knowledge economy and increase the economic competitiveness of the UK. Training in communication will be provided to the PDRA; Imperial College has a highly active staff development programme, which includes courses on presenting science to a lay audience, developing an independent research career, grant writing and exploiting translational aspect of research.

Collaboration
The project will support a new collaborations involving SM and research studying mitochondrial function and cancer biology field. These contacts (for example with Thomas Langer (Koln) and Ernesto Yague (Cancer Biology, Imperial) will be an invaluable resource in obtaining detailed functional data to maximise the impact of our structural data.

Exploitation and Application
We will, if necessary, ensure that intellectual property opportunities are maintained through liaison through MRC's together with Imperial College's technology transfer expertise (IC INNOVATIONS Ltd).