Modulation of the stathmin-like microtubule destabilizing activity of RB3, a neuron-specific member of the SCG10 family, by its N-terminal domain
Modulation of the stathmin-like microtubule destabilizing activity of RB3, a neuron-specific member of the SCG10 family, by its N-terminal domain
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DOI：
10.1074/jbc.m313693200
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发表时间：
2004-05-28
影响因子：
4.8
通讯作者：
Mori, N
中科院分区：
文献类型：
--
作者：
Nakao, C;Itoh, TJ;Mori, N
Rb3是SCG10/stathmin家族蛋白的神经元特异性同源物，具有独特的N端膜相关结构域和C末端的stathmin样结构域，促进微管(MT)突变和/或微管蛋白的隔离。在此，我们研究了RB3的N-末端亚域对MT动力学的调节作用。首先，我们在体外测定了全长(Rb3-f)和短截型(Rb3-S)Rb3对MT聚合的影响。Rb3-S在N端缺失了1-75个氨基酸，留下了所谓的Stathmin样结构域，由76-217个残基组成。虽然RB3-f和RB3-S都具有MT解聚活性，但RB3-f的解聚活性较差。Rb3-f与微管蛋白的结合亲和力也较低。用暗视野显微镜直接观察单个MT的动力学表明，RB3-S减慢了MT的延长速度，增加了灾难，减少了救援。这种作用与Stathmin/oncoProtein 18的作用几乎相同。另一方面，在较低浓度的RB3-f作用下，MT的延伸率增加。此外，RB3-f还显示出比对照组更高的救援频率，并以剂量依赖的方式表示灾难。Rb3-f的功能表明，全长Rb3不仅具有Stathmin样MT失稳活性，而且具有MT相关蛋白样MT稳定化活性。在体外，这些活性的平衡可能以浓度依赖的方式改变。RB3独特的N-末端结构域在MT动力学中的这种有趣的调节作用将有助于神经元形态发生的生理调节。
RB3 is a neuron-specific homologue of the SCG10/stathmin family proteins, possessing a unique N-terminal membrane-associated domain and the stathmin-like domain at the C terminus, which promotes microtubule (MT) catastrophe and/or tubulin sequestering. We examined herein the contribution of the N-terminal subdomain of RB3 to the regulation of MT dynamics. To begin with, we determined the effects of full-length (RB3-f) and short truncated (RB3-s) forms of RB3 on the polymerization of MT in vitro. RB3-s had a deletion of amino acids 1-75 from the N terminus, leaving the so-called stathmin-like domain, consisting of residues 76-217. Although both RB3-f and RB3-s exhibited MT-depolymerizing activity, RB3-f was less effective. The binding affinity for tubulin was also lower in RB3-f. Direct observation of the dynamics of individual MTs using dark field microscopy revealed that RB3-s slowed MT elongation velocity, increased catastrophes, and reduced rescues. This effect is almost identical to that by stathmin/oncoprotein 18. On the other hand, the MT elongation rate increased at lower concentrations of RB3-f. In addition, RB3-f, indicated higher rescue frequency than control as well as the catastrophe in a dose-dependent manner. The functionality of RB3-f indicated that full-length RB3 has not only stathmin-like MT destabilizing activity but also MT-associated protein-like MT stabilizing activity. Possibly, the balance of these activities is altered in a concentration-dependent manner in vitro. This interesting regulatory role of the unique N-terminal domain of RB3 in MT dynamics would contribute to the physiological regulation of neuronal morphogenesis.