Actin and spectrin play important roles in neurons, but their organization in axons and dendrites remains unclear. We used stochastic optical reconstruction microscopy (STORM) to study the organization of actin, spectrin and associated proteins in neurons. Actin formed ring-like structures that wrapped around the circumference of axons and evenly spaced along axonal shafts with a periodicity of ~180–190 nm. This periodic structure was not observed in dendrites, which instead contained long actin filaments running along dendritic shafts. Adducin, an actin-capping protein, colocalized with the actin rings. Spectrin exhibited periodic structures alternating with those of actin and adducin, and the distance between adjacent actin-adducin rings was comparable to the length of a spectrin tetramer. Sodium channels in axons were distributed in a periodic pattern coordinated with the underlying actin-spectrin-based cytoskeleton.

A bottom-up approach was the key to the successful fabrication of this memory. This approach both minimized the number of processing steps following deposition of the molecular monolayer, as well as protected the molecules from remaining processing steps. In the following paragraphs, we briefly describe the nanofabrication procedures utilized to construct the memory circuit. A full paper describing these procedures in more detail will be submitted for publication in the near future. form an array of top Ti NW electrodes, and the crossbar structure is complete.

Icy Adsorption The highly mobile nature of adsorbed liquid water on surfaces has made structural studies using atomic force microscopy challenging, especially for the first adsorbed layer. Moreover, scanning probe methods are hindered by the strong interaction between the probe tip and the water molecules. When graphene flakes were deposited on mica, Xu et al. (p. 1188 ; see the Perspective by Katsnelson ) found that they could use atomic force microscopy to image the first and second layers of introduced water. They consistently observed an islandike structure—0.34 nanometers in height, at low humidity—that corresponded to a single layer of ice. At higher humidity, thicker layers with a more liquid nature were observed.

By combining astigmatism imaging with a dual-objective scheme, we improved the image resolution of stochastic optical reconstruction microscopy (STORM) and obtained <10 nm lateral resolution and <20 nm axial resolution when imaging biological specimens. Using this approach, we resolved individual actin filaments in cells and revealed three-dimensional ultrastructure of the actin cytoskeleton. We observed two vertically separated layers of actin networks with distinct structural organizations in sheet-like cell protrusions.

In mammalian cells, mitochondrial dysfunction triggers the integrated stress response (ISR), in which eIF2α phosphorylation induces the transcription factor ATF4 1 - 3 . However, how mitochondrial stress is relayed to ATF4 is unknown. We found that HRI is the eIF2α kinase necessary and sufficient for this relay. In a genome-wide CRISPRi screen, we identified factors upstream of HRI: OMA1, a mitochondrial stress-activated protease, and DELE1, a little-characterized protein we found to be associated with the inner mitochondrial membrane. Mitochondrial stress stimulates OMA1-dependent cleavage of DELE1, leading to its accumulation in the cytosol, where it interacts with HRI and activates its eIF2α kinase activity. Additionally, DELE1 is required for ATF4 translation downstream of eIF2α phosphorylation. Blockade of the OMA1-DELE1-HRI pathway triggers an alternative response inducing specific molecular chaperones. Therefore, this pathway is a potential therapeutic target enabling fine-tuning of the ISR for beneficial outcomes in diseases involving mitochondrial dysfunction.

By developing a wide-field scheme for spectral measurement and implementing photoswitching, we synchronously obtained the fluorescence spectra and positions of ∼10(6) single molecules in labeled cells in minutes, which consequently enabled spectrally resolved, 'true-color' super-resolution microscopy. The method, called spectrally resolved stochastic optical reconstruction microscopy (SR-STORM), achieved cross-talk-free three-dimensional (3D) imaging for four dyes 10 nm apart in emission spectrum. Excellent resolution was obtained for every channel, and 3D localizations of all molecules were automatically aligned within one imaging path.

Recent evidence suggests that autophagy facilitates the unconventional secretion of the pro-inflammatory cytokine interleukin 1β (IL-1β). Here, we reconstituted an autophagy-regulated secretion of mature IL-1β (m-IL-1β) in non-macrophage cells. We found that cytoplasmic IL-1β associates with the autophagosome and m-IL-1β enters into the lumen of a vesicle intermediate but not into the cytoplasmic interior formed by engulfment of the autophagic membrane. In advance of secretion, m-IL-1β appears to be translocated across a membrane in an event that may require m-IL-1β to be unfolded or remain conformationally flexible and is dependent on two KFERQ-like motifs essential for the association of IL-1β with HSP90. A vesicle, possibly a precursor of the phagophore, contains translocated m-IL-1β and later turns into an autophagosome in which m-IL-1β resides within the intermembrane space of the double-membrane structure. Completion of IL-1β secretion requires Golgi reassembly and stacking proteins (GRASPs) and multi-vesicular body (MVB) formation.DOI: http://dx.doi.org/10.7554/eLife.11205.001

We report on the kinetics and ground-state thermodynamics associated with electrochemically driven molecular mechanical switching of three bistable [2]rotaxanes in acetonitrile solution, polymer electrolyte gels, and molecular-switch tunnel junctions (MSTJs). For all rotaxanes a pi-electron-deficient cyclobis(paraquat-p-phenylene) (CBPQT4+) ring component encircles one of two recognition sites within a dumbbell component. Two rotaxanes (RATTF4+ and RTTF4+) contain tetrathiafulvalene (TTF) and 1,5-dioxynaphthalene (DNP) recognition units, but different hydrophilic stoppers. For these rotaxanes, the CBPQT4+ ring encircles predominantly (>90 %) the TTF unit at equilibrium, and this equilibrium is relatively temperature independent. In the third rotaxane (RBPTTF4+), the TTF unit is replaced by a pi-extended analogue (a bispyrrolotetrathiafulvalene (BPTTF) unit), and the CBPQT4+ ring encircles almost equally both recognition sites at equilibrium. This equilibrium exhibits strong temperature dependence. These thermodynamic differences were rationalized by reference to binding constants obtained by isothermal titration calorimetry for the complexation of model guests by the CBPQT4+ host in acetonitrile. For all bistable rotaxanes, oxidation of the TTF (BPTTF) unit is accompanied by movement of the CBPQT4+ ring to the DNP site. Reduction back to TTF0 (BPTTF0) is followed by relaxation to the equilibrium distribution of translational isomers. The relaxation kinetics are strongly environmentally dependent, yet consistent with a single electromechanical-switching mechanism in acetonitrile, polymer electrolyte gels, and MSTJs. The ground-state equilibrium properties of all three bistable [2]rotaxanes were reflective of molecular structure in all environments. These results provide direct evidence for the control by molecular structure of the electronic properties exhibited by the MSTJs.