A beta-tubulin mutation selectively uncouples nuclear division and cytokinesis in Tetrahymena thermophila.
A beta-tubulin mutation selectively uncouples nuclear division and cytokinesis in Tetrahymena thermophila.
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β-微管蛋白突变选择性地解开嗜热四膜虫的核分裂和胞质分裂。
DOI：
10.1128/ec.3.5.1217-1226.2004
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发表时间：
2004
期刊：
影响因子：
--
通讯作者：
Romero,DanielP
中科院分区：
文献类型：
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作者：
Smith,JoshuaJ;Yakisich,JSebastian;Kapler,GeoffreyM;Cole,EricS;Romero,DanielP
嗜热四膜虫（Tetrahymena thermophilia）是一种纤毛原生动物，它在一个细胞内含有两个不同的细胞核，即有丝分裂微核和无丝分裂大核。虽然微管是两个细胞核正确分裂所必需的，但大核染色体缺乏着丝粒，微管在大核分裂中的作用尚未确定。在这里，我们描述了表达突变β-微管蛋白等位基因的细胞核分裂缺陷，该突变β-微管蛋白等位基因赋予对微管稳定药物紫杉醇的超敏反应。btu 1 -1（K350 M）突变对大核分裂有很大影响，产生的细胞DNA含量变化很大，包括完全缺乏大核的细胞。BTU 1 - 1等位基因表达的蛋白质比BTU 2基因座表达的野生型蛋白质占优势。当btu 1 - 1等位基因被靶向破坏失活或表达为截短的蛋白质时，正常的大核分裂得以恢复。免疫荧光研究揭示了细长的微管结构，围绕大核，未能迁移到卵裂沟。与此相反，其他细胞质微管依赖的过程，如胞质分裂，皮质图案，和口腔器组装，似乎是不受影响的突变体。在K350 M突变体中，胞核分裂也受到干扰，产生具有延长的早期后期纺锤体构型的细胞核，这些构型在胞质分裂开始后仍然存在。K350 M突变影响微管蛋白动力学，因为大核分裂缺陷会因促进微管聚合的三种处理而加剧：（i）升高的温度，（ii）亚致死浓度的紫杉醇，和（iii）高浓度的二甲基亚砜。用3-甲基腺嘌呤或渥曼青霉素抑制磷脂酰肌醇3-激酶（PI 3-kinase）也以abtu 1 -1依赖的方式诱导无核细胞形成。相反，肌球蛋白轻链激酶抑制剂ML-7对btu 1 - 1突变株的核分裂没有影响。这些发现为微管动力学提供了新的见解，并将进化上保守的PI 3-激酶信号通路与四膜虫的核迁移和/或分裂联系起来。
The ciliated protozoanTetrahymena thermophilacontains two distinct nuclei within a single cell—the mitotic micronucleus and the amitotic macronucleus. Although microtubules are required for proper division of both nuclei, macronuclear chromosomes lack centromeres and the role of microtubules in macronuclear division has not been established. Here we describe nuclear division defects in cells expressing a mutant β-tubulin allele that confers hypersensitivity to the microtubule-stabilizing drug paclitaxel. Macronuclear division is profoundly affected by thebtu1-1(K350M) mutation, producing cells with widely variable DNA contents, including cells that lack macronuclei entirely. Protein expressed by thebtu1-1allele is dominant over wild-type protein expressed by theBTU2locus. Normal macronuclear division is restored when thebtu1-1allele is inactivated by targeted disruption or expressed as a truncated protein. Immunofluorescence studies reveal elongated microtubular structures that surround macronuclei that fail to migrate to the cleavage furrows. In contrast, other cytoplasmic microtubule-dependent processes, such as cytokinesis, cortical patterning, and oral apparatus assembly, appear to be unaffected in the mutant. Micronuclear division is also perturbed in the K350M mutant, producing nuclei with elongated early-anaphase spindle configurations that persist well after the initiation of cytokinesis. The K350M mutation affects tubulin dynamics, as the macronuclear division defect is exacerbated by three treatments that promote microtubule polymerization: (i) elevated temperatures, (ii) sublethal concentrations of paclitaxel, and (iii) high concentrations of dimethyl sulfoxide. Inhibition of phosphatidylinositol 3-kinase (PI 3-kinase) with 3-methyladenine or wortmannin also induces amacronucleate cell formation in abtu1-1-dependent manner. Conversely, the myosin light chain kinase inhibitor ML-7 has no effect on nuclear division in thebtu1-1mutant strain. These findings provide new insights into microtubule dynamics and link the evolutionarily conserved PI 3-kinase signaling pathway to nuclear migration and/or division inTetrahymena.