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3 Janelia Publications
Showing 1-3 of 3 resultsSmall unilamellar vesicles (SUVs) are indispensable model membranes, organelle mimics, and drug and vaccine carriers. However, the lack of robust techniques to functionalize or organize preformed SUVs limits their applications. Here we use DNA nanostructures to coat, cluster, and pattern sub-100-nm liposomes, generating distance-controlled vesicle networks, strings and dimers, among other configurations. The DNA coating also enables attachment of proteins to liposomes, and temporal control of membrane fusion driven by SNARE protein complexes. Such a convenient and versatile method of engineering premade vesicles both structurally and functionally is highly relevant to bottom-up biology and targeted delivery.
The flagella of mammalian sperm display non-planar, asymmetric beating, in contrast to the planar, symmetric beating of flagella from sea urchin sperm and unicellular organisms. The molecular basis of this difference is unclear. Here, we perform in situ cryo-electron tomography of mouse and human sperm, providing the highest-resolution structural information to date. Our subtomogram averages reveal mammalian sperm-specific protein complexes within the microtubules, the radial spokes and nexin-dynein regulatory complexes. The locations and structures of these complexes suggest potential roles in enhancing the mechanical strength of mammalian sperm axonemes and regulating dynein-based axonemal bending. Intriguingly, we find that each of the nine outer microtubule doublets is decorated with a distinct combination of sperm-specific complexes. We propose that this asymmetric distribution of proteins differentially regulates the sliding of each microtubule doublet and may underlie the asymmetric beating of mammalian sperm.
The severe acute respiratory syndrome associated coronavirus 2 (SARS-CoV-2) and SARS-CoV-1 accessory protein Orf3a colocalizes with markers of the plasma membrane, endocytic pathway, and Golgi apparatus. Some reports have led to annotation of both Orf3a proteins as a viroporin. Here we show that neither SARS-CoV-2 nor SARS-CoV-1 form functional ion conducting pores and that the conductances measured are common contaminants in overexpression and with high levels of protein in reconstitution studies. Cryo-EM structures of both SARS-CoV-2 and SARS-CoV-1 Orf3a display a narrow constriction and the presence of a basic aqueous vestibule, which would not favor cation permeation. We observe enrichment of the late endosomal marker Rab7 upon SARS-CoV-2 Orf3a overexpression, and co-immunoprecipitation with VPS39. Interestingly, SARS-CoV-1 Orf3a does not cause the same cellular phenotype as SARS-CoV-2 Orf3a and does not interact with VPS39. To explain this difference, we find that a divergent, unstructured loop of SARS-CoV-2 Orf3a facilitates its binding with VPS39, a HOPS complex tethering protein involved in late endosome and autophagosome fusion with lysosomes. We suggest that the added loop enhances SARS-CoV-2 Orf3a ability to co-opt host cellular trafficking mechanisms for viral exit or host immune evasion.