Microtubule cytoskeleton exists in various biochemical forms in different cells due to tubulin posttranslational modifications (PTMs). Tubulin PTMs are known to affect microtubule stability, dynamics, and interaction with MAPs and motors in a specific manner, widely known as tubulin code hypothesis. At present, there exists no tool that can specifically mark tubulin PTMs in living cells, thus severely limiting our understanding of their dynamics and cellular functions. Using a yeast display library, we identified a binder against terminal tyrosine of α-tubulin, a unique PTM site. Extensive characterization validates the robustness and nonperturbing nature of our binder as tyrosination sensor, a live-cell tubulin nanobody specific towards tyrosinated microtubules. Using this sensor, we followed nocodazole-, colchicine-, and vincristine-induced depolymerization events of tyrosinated microtubules in real time and found each distinctly perturbs the microtubule polymer. Together, our work describes a novel tyrosination sensor and its potential applications to study the dynamics of microtubule and their PTM processes in living cells.

中文翻译：

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用于酪氨酸微管的基因编码活细胞传感器

由于微管蛋白翻译后修饰（PTM），微管细胞骨架以各种生化形式存在于不同细胞中。已知微管蛋白 PTM 会以特定方式影响微管稳定性、动力学以及与 MAP 和马达的相互作用，这被广泛称为微管蛋白密码假说。目前，还没有工具可以特异性标记活细胞中的微管蛋白 PTM，从而严重限制了我们对其动力学和细胞功能的理解。使用酵母展示文库，我们鉴定了针对 α-微管蛋白末端酪氨酸（独特的 PTM 位点）的结合物。广泛的表征验证了我们的结合剂作为酪氨酸传感器的稳健性和非干扰性质，酪氨酸传感器是一种针对酪氨酸微管的活细胞微管蛋白纳米抗体。使用该传感器，我们实时跟踪诺考达唑、秋水仙碱和长春新碱诱导的酪氨酸微管解聚事件，发现每种解聚事件都明显扰乱微管聚合物。我们的工作共同描述了一种新型酪氨酸化传感器及其在研究活细胞中微管动力学及其 PTM 过程方面的潜在应用。