In the long axon of a neuron, cargo transport between the cell body and terminal synaptic region are mainly supported by the motor proteins kinesin and dynein, which are nano-sized drivers. Synaptic materials packed as cargos are anterogradely transported to the synaptic region by kinesin, whereas materials accumulated at the axon terminals are returned to the cell body by dynein. Extreme value analysis, typically used for disaster prevention in our society, was applied to analyze the velocity of kinesin and dynein nanosized drivers to disclose their physical properties in living cells.
Many photothermal materials have been developed in recent years to achieve efficient photothermal therapy. Although larger-sized materials are preferred to ensure a better photothermal performance, upsizing hampers the cellular uptake of these materials. To overcome this dilemma, we proposed an active control system to manipulate the assembly of magnetic composite particles (MCPs) at the cellular level. Herein, MCPs of different sizes (small, medium, and large) were synthesized, and their surfaces were modified with a polymer. These MCPs were then cultured with HeLa cells, and their uptake and mobility were investigated. Small-sized MCPs (d v = 206 nm) and medium-sized MCPs (d v = 312 nm) could be introduced into HeLa cells. However, a smaller amount of the large-sized MCPs was introduced as compared to the other MCPs. Using a direct current magnetic field [DCMF (B = 150 mT)], medium-sized MCPs were observed to quickly assemble in cells; however, redispersal did not occur after the DCMF was turned off. To improve the particle dispersibility, polyethylene glycol and polyethyleneimine (PEI) were used to modify the medium-sized MCPs. Coating the MCPs with PEI (a polymer molecular weight of 25,000 or 270,000) improved their dispersibility in phosphate-buffered saline and cellular uptake. In addition, the redispersal of assembled PEI-coated MCPs was observed after the DCMF was turned off. The photothermal conversion efficiency of the MCPs was also improved using the DCMF. Consequently, the PEI-coated MCPs were first introduced into cells in a dispersed state, and their assembly was induced at the subcellular scale by applying a DCMF, and the assembly was spontaneously redispersed by switching off the DCMF. Such a remote control could improve the cytotoxicity of MCPs under near-infrared laser irradiation.
Zika virus infection can result in profound fetal deficits, such as microcephaly and blindness. Here we identify Kif11/Kinesin-5, as a cellular target of Zika protease with both in vitro and in vivo assays. We show that soluble Zika NS2-3 protease chimera targets several sites within the motor domain of HsEg5 either when free in solution or rigor-bound to microtubules. We tested two different forms of the protease in human cells. First, we find that soluble Zika protease chimera is cytotoxic and eventually leads to cell death. Second, the native integral ERmembrane-associated protease is not only better tolerated by human cells, but also imparts unusual mitotic spindle positioning defects in cultured cells and a prolonged metaphase delay. Our data suggests a model whereby interactions at the spatially restricted ER-mitotic spindle interface bring the protease into the proximity of the pool of mitotic spindle-associated HsEg5. The resulting phenotype is distinct from the monopolar phenotype that typically results from downregulation of HsEg5 due to genetic knock-down or small molecule inhibition. Our results support the idea that not all knockouts are the same; nonuniform down-regulation of protein function can result in unanticipated phenotypes. We postulate that the mitotic spindle mobility phenotype that we observe can prematurely terminate cell lineages that are dependent on asymmetric cell division, such as neuronal lineages. Additionally, our data raise questions about novel inter-organelle communication between the mitotic apparatus and the surrounding reticulate ER network. Finally, given that Kif11 is a monogenic haploinsufficient locus linked to microcephaly, we propose that a potential direct connection between this kinesin and the protease may be a cornerstone in the subsequent pathogenesis of microcephaly.
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