|| Sexual reproduction is almost universal in eukaryotic life and involves the fusion of male and female haploid gametes into a diploid cell. In spite of the importance of this biological process, very little is known about the molecular mechanisms governing the plasma membrane fusion reaction between gametes. A candidate fusion protein has been only recently identified: the ancient gamete plasma membrane protein HAP2. However, the detailed protein-protein and protein-lipid interactions involved in this process are not yet clear.
This project aims at reconstituting the fusion protein machinery in models of the plasma membrane (e.g., giant unilamellar vesicles or GUVs). In these controlled and reproducible systems, it will be possible to isolate and characterize the fusion process, from a quantitative point of view. Microfluidic technology will be used to bring two GUVs in close proximity and to controllably add or remove external triggers (e.g. ions or fusion inhibitors). Cutting- edge fluorescence microscopy techniques will be used to quantify the formation of cis (i.e., within a single lipid membrane) and trans (i.e., across two interacting membranes) HAP2 complexes. Fluorescence lifetime imaging microscopy (or FLIM) will be used to quality the degree of membrane fusion. One of the aims of this project will be to determine the multimerization state of pre- and post-fusion complexes. Finally, the same combination of advanced experimental approaches will be applied to clarify HAP2 interactions directly in living cells. This project will be carried out in the context of an international collaboration between University of Potsdam, MPIKG, Technion (Haifa) and University of Hannover.