Jenny E. Hinshaw’s research while affiliated with National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health and other places

... To locate rare proteins or proteins of unknown structure, we implemented a CLEM protocol. The HEK293 cell line described above expresses, upon induction, a fluorescently labeled well-characterized dominant negative mutant of dynamin 1, Dyn1(K44A)-GFP, that blocks clathrin-mediated endocytosis [25][26][27] . Using this cell line, we tested our CLEM protocol to search for sites of arrested clathrin-mediated endocytosis and dynamin accumulation. ...

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Cryo-electron tomography pipeline for plasma membranes

... crucial [39]. OPA1 promotes the fusion of the mitochondrial inner membrane, and the proteolytic products separate the mitochondrial inner membrane from the outer membrane [39]. ...

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Macrophage-derived mitochondria-rich extracellular vesicles aggravate bone loss in periodontitis by disrupting the mitochondrial dynamics of BMSCs

... The polymerization of proteins such as actin and b-microglobulin has been reported to generate active forces in driving the transition of a flat membrane to a bud. 44,45 We used numerical simulations to capture the scaling between the force generated by a growing network of ESAT-6 fibrils and membrane shape. The polymerization of ESAT-6 results in the formation of fibrils with varying lengths (Figures 3G and 3H). ...

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Real-time visualization reveals Mycobacteriumtuberculosis ESAT-6 disrupts phagosome-like compartment via fibril-mediated vesiculation

... [38][39][40] It was shown that GlpG is able to locally readjust the membrane, thinning a portion of the lipids that embrace the protein, [38] a feature that also has been reported for a variety of membrane protein insertases. [41][42][43][44][45] It is believed that this local membrane thinning facilitates diffusion of GlpG [38,46] by minimizing the hydrophobic mismatch, as the protease scans the membrane for substrates and potentially enhancing substrate interaction. A direct correlation between membrane thickness and proteolytic activity could be established for GlpG where an optimized hydrophobic thickness of the lipid bilayer resulted in the most efficient substrate proteolysis. ...

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Maintaining the Integral Membrane Proteome: Revisiting the Functional Repertoire of Integral Membrane Proteases

... In mammals, they are called atlastins and have several isoforms (ATL1-3). These membranebound GTPases mediate the tethering and fusion of ER tubules to form the three-way junctions of the polygonal network [14][15][16][17][18][19] . ...

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The membrane curvature-inducing REEP1-4 proteins generate an ER-derived vesicular compartment

... This suggests the possibility that there is stored elastic energy in the positively curved interphase lattice that is relieved during cytokinesis to help to form the furrow as a prelude to scission. This resembles what has been proposed to happen in eukaryotic cells when the mechanical energy stored in flat clathrin lattices on the plasma membrane assists in the formation of endosomes (55,56). Despite this, the S-layer must be flexible, since in wildtype cells it completely cover the entire cell surface at all times -and completely coats small extracellular vesicles generated by Sulfolobus cells at division (34, unpublished data). ...

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The mechanics of a continuous self-assembling surface-layer aids cell division in an archaeon

... Due to the high heat capacity of the cryogen, the sample is cooled at a rapid rate, leading to efficient vitrification (Dubochet & McDowall, 1981). Additionally, samples can also be vitrified using a cryogen stream (Ravelli et al., 2020), dispensed onto a grid in minute volumes and at rapid intervals designed principally for time sensitive specimens (Dandey et al., 2020), or cryofixed during lightmicroscope imaging using a correlative light and electron microscope (CLEM) fitted with a microfluidics device (Fuest et al., 2019). Even more excitingly, protein samples can be passed through a mass spectrometer in a gaseous state and deposited on a cryo-cooled grid for cryo-EM, allowing an accurate characterisation of the applied specimen prior to imaging (Esser et al., 2024). ...

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Structural biology inside multicellular specimens using electron cryotomography

... However, it has been puzzling how cells overcome the transition from flat membrane to hemispherical domes during CME at high tension. The exact mechanism of curvature formation in CCPs is hotly debated 11,126,168,171 and whether membrane tension and presence of membrane bending proteins during CCP initiation control fate of the CCP is an open question. ...

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Mechanical Regulation of Clathrin-Mediated Endocytosis by Membrane Bending Protein Epsin

... This includes mitochondria quality control 20 and the regulation of mitochondrial transport [21][22][23][24] . Similarly, actin plays an important role in recruiting and activating dynamin family GTPases to various cell compartments 25,26 , and has been shown to act in concert with dynamin to promote cell-to-cell fusion 27 . As the ER likely acts as a platform to recruit the machinery for both mitochondrial fission and fusion, and ER-anchored INF2 and actin play crucial roles in GTPase-mediated mitochondrial fission, we hypothesize that mito-and ER-associated actin ("ER-actin") is important for GTPase-mediated mitochondrial fission and fusion. ...

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Mitochondria- and ER-associated actin are required for mitochondrial fusion

... They play critical roles in organelle homeostasis and as innate immunity effectors that restrict certain pathogens (Bui and Shaw, 2013;Haller et al., 2015). Independent of their membrane binding activity, DSPs also play an increasingly appreciated role as regulators and organizers of both the actin-and microtubulebased cytoskeleta (Strack et al., 2013;Hatch et al., 2014;Zhang et al., 2019). ...

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Open and cut: allosteric motion and membrane fission by dynamin superfamily proteins