Aletta J M
Department of Pharmacology and Toxicology, University at Buffalo, School of Medicine, New York 14214-3000, USA.
J Neurobiol. 1996 Dec;31(4):461-75. doi: 10.1002/(SICI)1097-4695(199612)31:4<461::AID-NEU6>3.0.CO;2-7.
Nerve growth factor (NGF) produces both rapid and delayed cellular responses that are involved in neuronal differentiation. Neurite formation, a conspicuous delayed response, is accompanied by phosphorylation of beta-tubulin in PC12 cells. The present work provides further characterization of the phospho form of beta-tubulin in this neuronal model system with regard to isotype, cellular localization, and the circumstances that favor its formation. The results indicate that neuron-specific type III beta-tubulin (beta III-tubulin) is selectively affected during neurite formation. This phosphorylation occurs relatively late in the NGF signal transduction cascade and increases progressively with increasing duration of NGF treatment concomitant with more extensive neurite growth. The subcellular distribution of beta III-tubulin is not markedly different from that of total tubulin, but the phosphorylated protein is uniquely associated with microtubules that are calcium and cold labile. Although NGF is capable of inducing phosphorylation of beta III-tubulin, it is not necessarily sufficient. Based on experiments that employ either nonpermissive substrate conditions or microtubule-depolymerizing drugs, this phosphorylation requires neurite outgrowth. Direct measurements of the phospho form in neurites versus cell bodies by means of a microculture system indicate that phosphorylated beta III-tubulin is enriched in neurites. The enrichment of phospho-beta III-tubulin in calcium- and cold-labile polymer within neurites and its near absence in nonneurite bearing, NGF-treated cells suggests a role for this posttranslationally modified protein in the regulation of dynamic microtubules involved in neurite formation.
神经生长因子（NGF）可产生快速和延迟的细胞反应，这些反应参与神经元分化。神经突形成是一种明显的延迟反应，在PC12细胞中伴随着β-微管蛋白的磷酸化。本研究进一步描述了在这个神经元模型系统中，β-微管蛋白磷酸化形式在亚型、细胞定位以及有利于其形成的环境方面的特征。结果表明，在神经突形成过程中，神经元特异性III型β-微管蛋白（βIII-微管蛋白）受到选择性影响。这种磷酸化在NGF信号转导级联反应中发生得相对较晚，并随着NGF处理时间的延长而逐渐增加，同时伴随着更广泛的神经突生长。βIII-微管蛋白的亚细胞分布与总微管蛋白的分布没有明显差异，但磷酸化蛋白与对钙和冷敏感的微管独特相关。虽然NGF能够诱导βIII-微管蛋白的磷酸化，但这不一定足够。基于使用非允许底物条件或微管解聚药物的实验，这种磷酸化需要神经突生长。通过微培养系统对神经突与细胞体中的磷酸化形式进行直接测量表明，磷酸化的βIII-微管蛋白在神经突中富集。磷酸化βIII-微管蛋白在神经突中对钙和冷敏感的聚合物中富集，而在未形成神经突的NGF处理细胞中几乎不存在，这表明这种翻译后修饰的蛋白在调节参与神经突形成的动态微管中起作用。
