Live cell imaging of endosomal trafficking in fungi.

Live cell imaging of endosomal trafficking in fungi.
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真菌内体运输的活细胞成像

DOI:
10.1007/978-1-4939-2309-0_24
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发表时间:
2015
影响因子:
--
通讯作者:
M. Feldbrügge
M. Feldbrügge
中科院分区:
--
文献类型:
--
作者:
S. Baumann;N. Takeshita;N. Grün;R. Fischer;M. Feldbrügge

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内体是一种多功能的膜载体,对内吞和分泌很重要。在膜运输过程中,内体运输脂质、蛋白质甚至RNA。在高度极化的细胞中,如真菌菌丝,它们在分子马达如驱动蛋白和动力蛋白的介导下,沿着微管双向穿梭。对于这些高度动态的蛋白质/膜复合物的体内研究,先进的荧光显微镜是有用的。在这一章中,我们描述了活细胞成像的内涵体在两个遥远的相关真菌模型系统,担子菌黑粉菌和子囊菌构巢曲霉。我们提供了对动态内体蛋白和RNA的活细胞成像,共定位研究的双色检测,以及用于定量的光漂白后荧光恢复(FRAP)和用于超分辨率的光激活定位显微镜(PALM)的见解。在两个充分研究的真菌模型系统中描述的这些方法适用于广泛的其他生物体。
Endosomes are multipurpose membranous carriers important for endocytosis and secretion. During membrane trafficking, endosomes transport lipids, proteins, and even RNAs. In highly polarized cells such as fungal hyphae, they shuttle bidirectionally along microtubules mediated by molecular motors like kinesins and dynein. For in vivo studies of these highly dynamic protein/membrane complexes, advanced fluorescence microscopy is instrumental. In this chapter, we describe live cell imaging of endosomes in two distantly related fungal model systems, the basidiomyceteUstilago maydisand the ascomyceteAspergillus nidulans. We provide insights into live cell imaging of dynamic endosomal proteins and RNA, dual-color detection for colocalization studies, as well as fluorescence recovery after photobleaching (FRAP) for quantification and photo-activated localization microscopy (PALM) for super-resolution. These methods described in two well-studied fungal model systems are applicable to a broad range of other organisms.
建立用于真菌基因工程的多功能金门克隆系统
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