Mechanism, functions and conservation of SHRED, a novel pathway regulating protein quality control
Final Report Abstract
Each cell in a living organism contains billions of protein molecules. Proteins help to carry out the many activities of a cell and are essential for a cell to survive and grow. However, proteins can only fulfil their tasks when they have their correct three-dimensional shape. If a protein is unable to reach or keep its correct shape, it cannot perform its tasks. Such faulty proteins may even interfere with the proper functioning of other proteins and in this way become toxic. Cells have evolved ways to recognize and destroy faulty and potentially dangerous proteins. The recognition of faulty proteins is achieved by a special class of proteins called ubiquitin ligases, which attach a small protein called ubiquitin to proteins that need to be eliminated and thereby mark these proteins for destruction by a large protein complex called the proteasome. The process of recognizing and eliminating faulty proteins is called protein quality control. In earlier work, we had studied protein quality control in baker’s yeast, which uses similar ways as human cells to ensure the proper shape of their proteins. Baker’s yeast is easy to grow and manipulate and the study of baker’s yeast therefore is a useful approach to learn more about the biology of our own species. We had found that the activity of the ubiquitin ligase Ubr1 is controlled by a small protein called Roq1. The abundance of Roq1 in the cell strongly increases during certain stress conditions that put many other proteins at risk of losing their correct shape. Roq1 then binds to Ubr1 and reprograms it to better detect faulty proteins. Exactly how this reprograming works was unclear. However, given that Ubr1 is also present in human cells and is important for proper cell function, we aimed, in this project, to elucidate how Roq1 acts on Ubr1 to direct it towards faulty proteins. We first isolated Roq1 and Ubr1, combined them in the test tube with properly shaped or faulty proteins and observed that Roq1 indeed promotes the Ubr1-mediated attachment of ubiquitin specifically to the faulty proteins. We then identified features in Roq1 that allowed it to control Ubr1 and found that just two small regions of Roq1 were needed. These two regions enable Roq1 to stably bind to Ubr1 and alter its activity. To find out if the binding of Roq1 may change the shape of Ubr1 in such a way that it becomes better at recognizing faulty proteins, we employed a technique called cryo-electron microscopy. These studies are still ongoing and may yield a precise understanding of how Roq1 affects the activity of Ubr1. Furthermore, they may inspire ways to use the relevant regions Roq1 to build artificial regulators of Ubr1 that can be used to manipulate Ubr1 activity at will. As a result, it may eventually become possible to enhance Ubr1’s ability to remove faulty proteins. This may be medically valuable, for example to help slow the death of neurons during aging.
