The interplay between intermediate filaments (IFs) and other cytoskeletal components is important for the integrity and motility of cells. The impact of IF assembly on other components and cell morphology is not yet fully understood. Therefore, we examined the effects of combined desmin and actin assembly on cytoskeletal network arrangement in artificial cell‐sized droplets. Fluorescently labeled desmin, with or without actin, was enclosed in droplets prepared with 1,2‐dioleoyl‐sn‐glycero‐3‐phosphoethanolamine (DOPE) using the water‐in‐oil method. Protein networks were observed using fluorescence microscopy in the presence of 150 mM KCl, 20 mM imidazole–HCl (pH 7.4), 2 mM MgCl₂, and 1 mM adenosine 5'‐triphosphate for both desmin and actin assembly. As desmin alone can assemble into filaments within seconds, desmin networks mainly localizing at the inner margins of the droplets were observed within 10 min after assembly initiation. Subsequently, deformations of droplets appeared. Furthermore, a portion of droplets formed desmin‐rich protrusions of several micrometers. Notably, actin alone rarely formed protrusions under the same conditions. When 1,2‐dioleoyl‐sn‐glycero‐3‐phosphocholine was used instead of DOPE, protrusions became less frequent. The combination of desmin and actin increased the number of deformed droplets in which the proteins were considerably colocalized. The assembly process of desmin facilitated colocalization. Atomic force microscopy failed to reveal interactions between the two filament types. These results suggest that the mechanical properties of desmin networks may influence the behavior of actin networks, as well as membrane morphology, possibly reflecting the mechanical function of desmin filaments in muscle cells.

中文翻译：

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脂质小滴中浓缩的结蛋白和肌动蛋白细胞骨架通讯。

中间丝（IF）与其他细胞骨架成分之间的相互作用对于细胞的完整性和运动性很重要。IF组装对其他组件和细胞形态的影响尚未完全了解。因此，我们检查了结合的结蛋白和肌动蛋白组装对人工细胞大小的液滴中细胞骨架网络排列的影响。使用油包水方法，将荧光标记的结蛋白（含或不含肌动蛋白）封闭在由1,2-烯丙基-sn-甘油-3-磷酸乙醇胺（DOPE）制备的液滴中。在150 mM KCl，20 mM咪唑-HCl（pH 7.4），2 mM MgCl ₂的存在下，使用荧光显微镜观察了蛋白质网络。，以及用于结蛋白和肌动蛋白装配的1 mM 5'-三磷酸腺苷。由于单独的结蛋白可以在几秒钟内组装成细丝，因此在组装开始后的10分钟内观察到了主要位于液滴内部边缘的结蛋白网络。随后，出现液滴变形。此外，一部分液滴形成了几微米的富含结蛋白的突起。值得注意的是，单独的肌动蛋白在相同条件下很少形成突起。当1,2-二油酰基使用甘油3-磷酸胆碱代替DOPE，突起的频率降低。结蛋白和肌动蛋白的组合增加了其中蛋白质被相当共定位的变形液滴的数量。desmin的组装过程促进了共本地化。原子力显微镜无法揭示两种灯丝类型之间的相互作用。这些结果表明，结蛋白网络的机械性质可能影响肌动蛋白网络的行为以及膜的形态，可能反映了结蛋白细丝在肌肉细胞中的机械功能。