Dynamics of membrane trafficking
A yeast cell expressing fluorescently labelled endocytic proteins. The first two images show a coat protein Sla1 (green) and an actin-binding protein Abp1 (red). The last image shows both channels merged. The spots at the cell surface reveal the transient accumulation of the proteins at endocytic sites during vesicle budding
Using budding yeast as a model, the Kaksonen group wants to understand how complex molecular machineries drive vesicle trafficking.
Previous and current research
Many biological processes at the cellular level are based on complex networks of macromolecular interactions. These networks have a modular organisation, where the different modules form dynamic molecular machines that drive processes such as signalling, cell motility, cytokinesis, and vesicle trafficking. Our group’s longterm goal is to contribute to the understanding of the general principles governing the assembly and function of these supramolecular machines.
More specifically, we are interested in the formation of cargo-loaded transport vesicles, such as endocytic vesicles. The formation of the endocytic vesicle is driven by highly dynamic molecular machinery that is composed of more than 50 different protein species and of several thousand individual protein molecules. We aim to understand the processes that regulate the assembly of the endocytic machinery, the recruitment of the cargo molecules, and the selection of the location and timing of endocytic events in the cell.
Our main experimental organism is the budding yeast, Saccharomyces cerevisiae. In our studies we use quantitative live-cell imaging methods (for example particle tracking, FRAP, FCS/FCCS, high-throughput microscopy) in combination with powerful yeast genetics. We also use correlated light and electron microscopy to gain nanometre scale information about the endocytic structures, and biochemistry to characterise protein-protein and protein-lipid interactions.
Future projects and goals
We are interested in the mechanisms that initiate the assembly of the endocytic machinery and regulate the precise timing of the sequential stages of the assembly. The spatial distribution of the endocytic events is tightly coupled to the cell cycle and to the overall polarity of the cell. The spatially regulated initiation of endocytic events is critical for determining the cellular distribution of endocytosis.
We are also studying the mechanisms of selective recruitment of cargo molecules into the endocytic vesicles. The recruitment of cargo proteins is tightly regulated by a family of endocytic adaptors. We want to learn how this adaptor system integrates environmental and intracellular signals in deciding which cargoes to recruit.
Furthermore, we want to understand how actin functions to promote endocytic vesicle budding. In yeast, endocytosis is strictly dependent on actin polymerisation, but the mechanisms by which actin drives vesicle budding are not well understood. We are currently studying the molecular basis of the coupling between the actin cytoskeleton and the endocytic membrane. We have also started to investigate the evolution of the membrane-actin coupling in animals and fungi using a phylogenetic comparative approach.
The core membrane trafficking events, such as the clathrin-mediated endocytosis, are elemental cellular processes that are involved in multiple biological phenomena ranging from cell polarisation to neural plasticity. As most of the yeast trafficking proteins are widely conserved in eukaryotes, we believe that mechanisms that we unravel in yeast cells will be applicable to eukaryotes in general.