Tail-anchored protein biogenesis: defining the ATP dependent route

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Living things are made up of cells, and cells need membranes to provide barriers inside the cell and to protect them from the outside world. Whilst the biological membranes that form these barriers are incredibly complex and diverse in nature, they are generally characterised by a lipid bilayer that is studded with many proteins. Cells must normally replenish and renew their membranes by making new ones, and a key part of this process is the insertion or stitching of new proteins into the lipid bilayer. In more complex cells, such as those that make up our body, this integration of membrane proteins is one of the key functions of a specialised compartment known as the endoplasmic reticulum. As is the case for many biological processes, it has become clear that there are different ways of integrating a protein into the lipid bilayer of the endoplasmic reticulum. One mechanism is now fairly well understood and can be described as the 'classical' route for membrane proteins synthesis at the endoplasmic reticulum. Whilst the precise details of this classical pathway remain to be fully determined, we know most of the cellular factors and machineries that mediate it. In contrast, a second quite different pathway has been far less well studied so we know very little about the cellular components that are responsible for it or exactly what they do. This second pathway is used to make an important class of molecules called tail-anchored membrane proteins that are characteristic of more complex cells such as our own. The aim of this project is to identify the cellular machines that are responsible for putting tail-anchored proteins into the membrane of the endoplasmic reticulum. We will make use of the fact that we can now replicate this second pathway in a test tube, allowing us to compare how well different combinations of cellular components can carry out tail-anchored protein insertion. We are particularly interested in looking at the contribution of cellular machines known as molecular chaperones, since we have good reason to believe they play a major role in putting tail-anchored proteins into the endoplasmic reticulum. There are many different chaperones, and our work will tell us which particular chaperones are important for the second pathway and what exactly it is that they do during this process.