Dataset Information

Toc75-V/OEP80 is processed during translocation into chloroplasts, and the membrane-embedded form exposes its POTRA domain to the intermembrane space.

ABSTRACT: The insertion of membrane proteins requires proteinaceous complexes in the cytoplasm, the membrane, and the lumen of organelles. Most of the required complexes have been described, while the components for insertion of ?-barrel-type proteins into the outer membrane of chloroplasts remain unknown. The same holds true for the signals required for the insertion of ?-barrel-type proteins. At present, only the processing of Toc75-III, the ?-barrel-type protein of the central chloroplast translocon with an atypical signal, has been explored in detail. However, it has been debated whether Toc75-V/ outer envelope protein 80 (OEP80), a second protein of the same family, contains a signal and undergoes processing. To substantiate the hypothesis that Toc75-V/OEP80 is processed as well, we reinvestigated the processing in a protoplast-based assay as well as in native membranes. Our results confirm the existence of a cleavable segment. By protease protection and pegylation, we observed intermembrane space localization of the soluble N-terminal domain. Thus, Toc75-V contains a cleavable N-terminal signal and exposes its polypeptide transport-associated domains to the intermembrane space of plastids, where it likely interacts with its substrates.

SUBMITTER: Gross LE

PROVIDER: S-EPMC7050246 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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Toc75-V/OEP80 is processed during translocation into chloroplasts, and the membrane-embedded form exposes its POTRA domain to the intermembrane space.

Gross Lucia E LE Spies Nicole N Simm Stefan S Schleiff Enrico E

FEBS open bio 20200217 3

The insertion of membrane proteins requires proteinaceous complexes in the cytoplasm, the membrane, and the lumen of organelles. Most of the required complexes have been described, while the components for insertion of β-barrel-type proteins into the outer membrane of chloroplasts remain unknown. The same holds true for the signals required for the insertion of β-barrel-type proteins. At present, only the processing of Toc75-III, the β-barrel-type protein of the central chloroplast translocon wi ...[more]

PMID: 31953987

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Project description:Fibroblast growth factor 2 (FGF2) exits cells by direct translocation across the plasma membrane, a type I pathway of unconventional protein secretion. This process is initiated by phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2)-dependent formation of highly dynamic FGF2 oligomers at the inner plasma membrane leaflet, inducing the formation of lipidic membrane pores. Cell surface heparan sulfate chains linked to glypican-1 (GPC1) capture FGF2 at the outer plasma membrane leaflet, completing FGF2 membrane translocation into the extracellular space. While the basic steps of this pathway are well understood, the molecular mechanism by which FGF2 oligomerizes on membrane surfaces remains unclear. In the current study, we demonstrate the initial step of this process to depend on C95-C95 disulfide-bridge-mediated FGF2 dimerization on membrane surfaces, producing the building blocks for higher FGF2 oligomers that drive the formation of membrane pores. We find FGF2 with a C95A substitution to be defective in oligomerization, pore formation, and membrane translocation. Consistently, we demonstrate a C95A variant of FGF2 to be characterized by a severe secretion phenotype. By contrast, while also important for efficient FGF2 secretion from cells, a second cysteine residue on the molecular surface of FGF2 (C77) is not involved in FGF2 oligomerization. Rather, we find C77 to be part of the interaction interface through which FGF2 binds to the α1 subunit of the Na,K-ATPase, the landing platform for FGF2 at the inner plasma membrane leaflet. Using cross-linking mass spectrometry, atomistic molecular dynamics simulations combined with a machine learning analysis and cryo-electron tomography, we propose a mechanism by which disulfide-bridged FGF2 dimers bind with high avidity to PI(4,5)P2 on membrane surfaces. We further propose a tight coupling between FGF2 secretion and the formation of ternary signaling complexes on cell surfaces, hypothesizing that C95-C95-bridged FGF2 dimers are functioning as the molecular units triggering autocrine and paracrine FGF2 signaling.

Dataset Information

Toc75-V/OEP80 is processed during translocation into chloroplasts, and the membrane-embedded form exposes its POTRA domain to the intermembrane space.

Publications

Toc75-V/OEP80 is processed during translocation into chloroplasts, and the membrane-embedded form exposes its POTRA domain to the intermembrane space.

Similar Datasets

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