The disassembly and reassembly of functional centrosomes in vitro.
The disassembly and reassembly of functional centrosomes in vitro.
复制标题
功能中心体的体外拆卸和重组。
DOI：
10.1073/pnas.95.16.9295
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发表时间：
1998
影响因子：
11.1
通讯作者：
Palazzo,RE
中科院分区：
文献类型：
--
作者：
Schnackenberg,BJ;Khodjakov,A;Rieder,CL;Palazzo,RE
动物细胞含有一个中心体，中心体成核并组织一系列极化的微管，这些微管在许多细胞过程中发挥作用。在大多数细胞中，中心体由两个中心粒组成，周围环绕着边界不清的中心粒周围物质“云”。最近，含有γ-微管蛋白的直径为25 nm的环状结构已被确定为分离的中心体的中心粒周围物质内可能的微管成核位点。在这里，我们证明，当Spisulacentrosomes提取与1.0 M KI，他们失去了微管成核的潜力，并出现在三维电子显微镜作为一个复杂的晶格，从12- 15纳米厚的基本纤维（S），缺乏中心粒和25纳米环。重要的是，当这些残余物在从Spisulaoocytes制备的提取物中孵育时，它们恢复了它们的25-nm环、γ-微管蛋白和微管成核潜力。这种恢复过程发生在微管，二价阳离子和核苷酸的情况下。因此，在动物中，中心体在结构上围绕KI不溶性的基于亲核试剂的“中心基质”组织，该中心基质用作微管成核所需的那些蛋白质以二价阳离子和核苷酸独立的方式直接或间接结合的支架。
Animal cells contain a single centrosome that nucleates and organizes a polarized array of microtubules which functions in many cellular processes. In most cells the centrosome is composed of two centrioles surrounded by an ill-defined “cloud” of pericentriolar material. Recently, γ-tubulin-containing 25-nm diameter ring structures have been identified as likely microtubule nucleation sites within the pericentriolar material of isolated centrosomes. Here we demonstrate that whenSpisulacentrosomes are extracted with 1.0 M KI they lose their microtubule nucleation potential and appear by three-dimensional electron microscopy as a complex lattice, built from 12- to 15-nm thick elementary fiber(s), that lack centrioles and 25-nm rings. Importantly, when these remnants are incubated in extracts prepared fromSpisulaoocytes they recover their 25-nm rings, γ-tubulin, and microtubule nucleation potential. This recovery process occurs in the absence of microtubules, divalent cations, and nucleotides. Thus, in animals the centrosome is structurally organized around a KI-insoluble filament-based “centromatrix” that serves as a scaffold to which those proteins required for microtubule nucleation bind, either directly or indirectly, in a divalent cation and nucleotide independent manner.