An amphipathic helix drives interaction of Fibrillins with plastoglobule lipid droplets

Abstract

Plastoglobule lipid droplets of chloroplasts serve complex roles affecting plant development, stress tolerance and photosynthesis. They harbor a set of approximately 42 proteins that collectively dictate plastoglobule functions. Due to the monolayer structure of plastoglobules which encompass a neutral lipid core, these proteins must associate monotopically on the plastoglobule surface. However, targeting determinants have not been identified for plastoglobule proteins, and the protein-membrane interaction mechanisms that establish the plastoglobule proteome remain unclear. Here, we demonstrate that plastoglobule-localized Fibrillins harbor an amphipathic helix at the lip of their β-barrel that is necessary for proper plastoglobule association. Molecular dynamics simulations support the specific interaction of the amphipathic helix of AtFBN1a with membranes rich in lipid packing defects which are expected to be especially prevalent on the tightly curved surface of plastoglobules. Introduction of one of the amphipathic helices into stromal-or thylakoid-localized FBNs was ineffective at redirecting the proteins to plastoglobules, likely due to endogenous protein-protein interactions that override the influence of the amphipathic helix. Proteomic analyses indicate AtFBN1a influences the plastoglobule proteome through outcompeting and recruiting specific proteins. We also demonstrate that the plastoglobule-localized FBNs, AtFBN1a and AtFBN7a, bind unsaturated fatty acids, particularly C18:1, and that elimination of the amphipathic helix suppresses fatty acid binding in AtFBN1a, but promotes fatty acid binding in AtFBN7a. Predicted amphipathic helices can be identified on two-thirds of plastoglobule proteins, indicating the use of amphipathic helices may be a general mechanism by which proteins selectively associate with plastoglobules.