Suppression of microtubule dynamic instability by the plus TIP protein EB1 and its modulation by the CAP-Gly domain of p150Glued
Suppression of microtubule dynamic instability by the plus TIP protein EB1 and its modulation by the CAP-Gly domain of p150Glued
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DOI：
10.1021/bi701912g
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发表时间：
2008-01-15
期刊：
影响因子：
2.9
通讯作者：
Wilson, Leslie
中科院分区：
文献类型：
--
作者：
Manna, Tapas;Honnappa, Srinivas;Wilson, Leslie
EB 1 +TIP蛋白家族及其结合伴侣跟踪细胞中微管的生长正末端，并被认为调节其动力学。在这里，我们确定了EB 1和N-末端CAP-Gly结构域（p150 n）的主要结合伙伴之一，p150（Glued），单独和一起，在正端的动态不稳定参数的影响纯化的稳态微管。与EB 1单独，缩短率，缩短的程度，和灾难的频率被抑制在没有显着影响的增长速度或救援频率。当p150 n与EB 1共同加入时，EB 1对动力学的影响显著不同。缩短的速率和程度以及突变频率的抑制比单独使用EB 1强3-4倍。此外，EB 1-p150 n复合物增加了救援频率和微管生长的平均长度，这些参数不受单独EB 1的显著影响。类似地，EB 1的C末端尾（其是p150 n的关键结合区域）的缺失显著增加了EB 1抑制缩短动力学的能力。EB 1本身沿着微管的长度结合，1摩尔EB 1二聚体结合的量类似于12摩尔微管蛋白二聚体。在p150 n存在的情况下，大约两倍量的EB 1被募集到微管。我们的研究结果表明，EB 1的灵活的C-末端尾的失活显着改变EB 1的能力，调节微管动力学。他们进一步表明，p150（Glued）可能会激活，从而促进EB 1的招募到微管的尖端，以调节其动力学。
The EB1+TIP protein family and its binding partners track growing plus ends of microtubules in cells and are thought to regulate their dynamics. Here we determined the effects of EB1 and the N-terminal CAP-Gly domain (p 150n) of one of its major binding partners, p150(Glued), both separately and together, on the dynamic instability parameters at plus ends of purified steady-state microtubules. With EB1 alone, the shortening rate, the extent of shortening, and the catastrophe frequency were suppressed in the absence of significant effects on the growth rate or rescue frequency. The effects of EB1 on dynamics were significantly different when p150n was added together with EB1. The rate and extent of shortening and the catastrophe frequency were suppressed 3-4 times more strongly than with EB1 alone. In addition, the EB1--p150n complex increased the rescue frequency and the mean length the microtubules grew, parameters that were not significantly affected by EB1 alone. Similarly, deletion of EB1's C-terminal tail, which is a crucial binding region for p150n, significantly increased the ability of EB1 to suppress shortening dynamics. EB1 by itself bound along the length of the microtubules with I mol of EB1 dimer bound per similar to 12 mol of tubulin dimer. Approximately twice the amount of EB1 was recruited to the microtubules in the presence of p150n. Our results indicate that inactivation of EB1's flexible C-terminal tail significantly changes EB1's ability to modulate rnicrotubule dynamics. They further suggest that p150(Glued) may activate and thereby facilitate the recruitment of EB1 to the tips of microtubules to regulate their dynamics.