Coordinated rearrangements of cytoskeletal structures are the principal source of forces that govern cell and tissue morphogenesis. However, unlike for actin-based mechanical forces, our knowledge about the contribution of forces originating from other cytoskeletal components remains scarce. Here, we establish microtubules as central components of cell mechanics during tissue morphogenesis. We find that individual cells are mechanically autonomous during early Drosophila wing epithelium development. Each cell contains a polarized apical non-centrosomal microtubule cytoskeleton that bears compressive forces, whereby acute elimination of microtubule-based forces leads to cell shortening. We further establish that the Fat planar cell polarity (Ft-PCP) signalling pathway couples microtubules at adherens junctions (AJs) and patterns microtubule-based forces across a tissue via polarized transcellular stability, thus revealing a molecular mechanism bridging single cell and tissue mechanics. Together, these results provide a physical basis to explain how global patterning of microtubules controls cell mechanics to coordinate collective cell behaviour during tissue remodelling. These results also offer alternative paradigms towards the interplay of contractile and protrusive cytoskeletal forces at the single cell and tissue levels.
We present the self-assembly of redox-responsive polymer nanocontainers comprising a cyclodextrin vesicle core and a thin reductively cleavable polymer shell anchored via host-guest recognition on the vesicle surface. The nanocontainers are of uniform size, show high stability, and selectively respond to a mild reductive trigger as revealed by dynamic light scattering, transmission electron microscopy, atomic force microscopy, a quantitative thiol assay, and fluorescence spectroscopy. Live cell imaging experiments demonstrate a specific redox-responsive release and cytoplasmic delivery of encapsulated hydrophilic payloads, such as the pH-probe pyranine, and the fungal toxin phalloidin. Our results show the high potential of these stimulus-responsive nanocontainers for cell biological applications requiring a controlled delivery.
Cell fate determination during development often requires morphogen transport from producing to distant responding cells. Hedgehog (Hh) morphogens present a challenge to this concept, as all Hhs are synthesized as terminally lipidated molecules that form insoluble clusters at the surface of producing cells. While several proposed Hh transport modes tie directly into these unusual properties, the crucial step of Hh relay from producing cells to receptors on remote responding cells remains unresolved. Using wing development in Drosophila melanogaster as a model, we show that Hh relay and direct patterning of the 3–4 intervein region strictly depend on proteolytic removal of lipidated N-terminal membrane anchors. Site-directed modification of the N-terminal Hh processing site selectively eliminated the entire 3–4 intervein region, and additional targeted removal of N-palmitate restored its formation. Hence, palmitoylated membrane anchors restrict morphogen spread until site-specific processing switches membrane-bound Hh into bioactive forms with specific patterning functions.
Gold nanoparticles (AuNPs) are subjects of broad interest in scientific community due to their promising physicochemical properties. Herein we report the facile and controlled light-mediated preparation of gold nanoparticles through a Norrish type I reaction of photoactive polymers. These carefully designed polymers act as reagents for the photochemical reduction of gold ions, as well as stabilizers for the in situ generated AuNPs. Manipulating the length and composition of the photoactive polymers allows for control of AuNP size. Nanoparticle diameter can be controlled from 1.5 nm to 9.6 nm.
Cryogenic microscopy methods have gained increasing popularity, as they offer an unaltered view on the architecture of biological specimens. As a prerequisite, samples must be handled under cryogenic conditions below their recrys-tallization temperature, and contamination during sample transfer and handling must be prevented. We present a high-vacuum cryo-transfer system that streamlines the entire handling of frozen-hydrated samples from the vitrification process to low temperature imaging for scanning transmission electron microscopy and transmission electron microscopy. A template for cryo-electron microscopy and multimodal cryo-imaging approaches with numerous sample transfer steps is presented.
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