Kastner A, Moyse E, Bauer S, Jourdan F, Brun G
Laboratoire de Biologie Moléculaire et Cellulaire, UMR 49 CNRS, Ecole Normale Supérieure de Lyon, France.
J Neurochem. 2000 Jun;74(6):2343-9. doi: 10.1046/j.1471-4159.2000.0742343.x.
The molecular mechanisms underlying cell cycle control in neuronal progenitors have been investigated with adult mouse olfactory epithelium as a model system. Odor receptive neurons of mammalian olfactory epithelium are short-lived and renewed in the adult by mitotic division of intrinsic neuronal progenitors. Ablation of the synaptic target, olfactory bulb, induces sequentially extensive apoptosis of sensory neurons and then stimulation of progenitor proliferation, peaking at 36 h and 4 days, respectively, postlesion. Known molecular effectors of G1 phase entry have been assessed on protein extracts of olfactory organs sampled at various postbulbectomy times in adult mice. The decay of betaIII-tubulin and olfactory marker protein levels and the rise of proliferating cell nuclear antigen (PCNA) levels, starting 1 and 3 days, respectively, postlesion, provided the kinetic frame of neuronal dynamics. Cyclin D1, cyclin E, and cyclin-dependent kinase cdk2 levels, low in olfactory organ of intact mice, increased 3 days after bulbectomy in parallel with PCNA levels; cdk4 content was initially high and unaffected by lesioning. Western blots of the known cdk inhibitors revealed proliferation-related decreases of p18, p21, and p27 from high expression in intact organs. Immunoprecipitation of cdk2 and cdk4 fractions of protein extracts at 4 days postlesion (mitotic reaction peak) versus control, followed by cyclin D1 immunoblotting, and vice versa, revealed that levels of both cyclin D1/cdk2 and cyclin D1/cdk4 complexes, as well as their kinase activities, were dramatically increased after lesion. In vivo proliferation of olfactory neuronal lineage cells thus involves functional binding of cyclin D1 with cdk2 and cdk4, with differential activation mechanisms for cdk2 and cdk4. In addition, the RT-PCR-detected cyclin D1 mRNA level remained unaffected after bulbectomy, which indicated that the cyclin D1 rise should involve posttranscriptional mechanisms in this in vivo neuronal system. These observations are discussed, along with their relevance to cell cycle control and to olfactory neuron dynamics.
以成年小鼠嗅觉上皮为模型系统，对神经祖细胞中细胞周期调控的分子机制进行了研究。哺乳动物嗅觉上皮的气味感受神经元寿命较短，在成年期通过内在神经祖细胞的有丝分裂得以更新。切除突触靶标嗅球后，依次引发感觉神经元的广泛凋亡，随后刺激祖细胞增殖，分别在损伤后36小时和4天达到峰值。在成年小鼠嗅球切除术后不同时间点采集的嗅觉器官蛋白质提取物中，对已知的G1期进入分子效应器进行了评估。βIII-微管蛋白和嗅觉标记蛋白水平的下降以及增殖细胞核抗原（PCNA）水平的上升，分别在损伤后1天和3天开始，提供了神经元动态变化的动力学框架。细胞周期蛋白D1、细胞周期蛋白E和细胞周期蛋白依赖性激酶cdk2的水平，在完整小鼠的嗅觉器官中较低，在嗅球切除术后3天与PCNA水平同时升高；cdk4含量最初较高且不受损伤影响。对已知的cdk抑制剂进行蛋白质印迹分析显示，在完整器官中高表达的p18、p21和p27与增殖相关的表达下降。在损伤后4天（有丝分裂反应峰值）与对照组相比，对蛋白质提取物的cdk2和cdk4组分进行免疫沉淀，随后进行细胞周期蛋白D1免疫印迹分析，反之亦然，结果显示细胞周期蛋白D1/cdk2和细胞周期蛋白D1/cdk4复合物的水平及其激酶活性在损伤后均显著增加。因此，嗅觉神经元谱系细胞的体内增殖涉及细胞周期蛋白D1与cdk2和cdk4的功能性结合，以及cdk2和cdk4的不同激活机制。此外，RT-PCR检测到的细胞周期蛋白D1 mRNA水平在嗅球切除术后未受影响，这表明细胞周期蛋白D1的升高应涉及该体内神经元系统中的转录后机制。对这些观察结果进行了讨论，并阐述了它们与细胞周期调控以及嗅觉神经元动态变化的相关性。
