Belmadani Souad, Poüs Christian, Fischmeister Rodolphe, Méry Pierre-François
Laboratoire de Cardiologie Cellulaire et Moléculaire, INSERM U-446, Université Paris-Sud, Faculté de Pharmacie, Châtenay-Malabry, France.
Mol Cell Biochem. 2004 Mar;258(1-2):35-48. doi: 10.1023/b:mcbi.0000012834.43990.b6.
Little is known about the subcellular distribution and the dynamics of tubulins in adult cardiac myocytes although both are modified during cardiac hypertrophy and heart failure. Using confocal microscopy, we examined post-translational modifications of tubulin in fully differentiated ventricular myocytes isolated from adult rat hearts, as well as in immortalized and dividing HL-1 cardiomyocytes. Detyrosinated Glu-alpha-tubulin was the most abundant post-translationally modified tubulin found in ventricular myocytes, while acetylated- and delta2-alpha-tubulins were found in lower amounts or absent. In contrast, dividing HL-1 cardiomyocytes exhibited high levels of tyrosinated or acetylated alpha-tubulins. A mild nocodazole treatment (0.1 microM, 1 h) disrupted microtubules in HL-1 myocytes, but not in adult ventricular myocytes. A stronger treatment (10 microM, 2 h) was required to disassemble tubulins in adult myocytes. Glu-alpha-tubulin containing microtubules were more resistant to nocodazole treatment in HL-1 cardiomyocytes than in ventricular myocytes. Endogenous activation of the cAMP pathway with the forskolin analog L858051 (20 microM) or the beta-adrenergic agonist isoprenaline (10 microM) disrupted the most labile microtubules in HL-1 cardiomyocytes. In contrast, isoprenaline (10 microM), cholera toxin (200 ng/ml, a G(S)-protein activator), L858051 (20 microM) or forskolin (10 microM) had no effect on the microtubule network in ventricular myocytes. In addition, intracellular Ca2+ accumulation induced either by thapsigargin (2 microM) or caffeine (10 mM) did not modify microtubule stability in ventricular myocytes. Our data demonstrate the unique stability of the microtubule network in adult cardiac myocytes. We speculate that microtubule stability is required to support cellular integrity during cardiac contraction.
尽管在心肌肥大和心力衰竭过程中微管蛋白的亚细胞分布和动力学均会发生改变，但对于成年心肌细胞中微管蛋白的亚细胞分布和动力学却知之甚少。我们使用共聚焦显微镜检查了从成年大鼠心脏分离出的完全分化的心室肌细胞以及永生化且正在分裂的HL-1心肌细胞中微管蛋白的翻译后修饰。去酪氨酸化的谷氨酸-α-微管蛋白是在心室肌细胞中发现的最丰富的翻译后修饰微管蛋白，而乙酰化和δ2-α-微管蛋白的含量较低或不存在。相比之下，正在分裂的HL-1心肌细胞表现出高水平的酪氨酸化或乙酰化α-微管蛋白。轻度的诺考达唑处理（0.1微摩尔，1小时）会破坏HL-1心肌细胞中的微管，但不会破坏成年心室肌细胞中的微管。需要更强的处理（10微摩尔，2小时）才能分解成年心肌细胞中的微管蛋白。与心室肌细胞相比，含有谷氨酸-α-微管蛋白的微管在HL-1心肌细胞中对诺考达唑处理更具抗性。用福司可林类似物L858051（20微摩尔）或β-肾上腺素能激动剂异丙肾上腺素（10微摩尔）内源性激活cAMP途径会破坏HL-1心肌细胞中最不稳定的微管。相比之下，异丙肾上腺素（10微摩尔）、霍乱毒素（200纳克/毫升，一种G(S)蛋白激活剂）、L858051（20微摩尔）或福司可林（10微摩尔）对心室肌细胞中的微管网络没有影响。此外，毒胡萝卜素（2微摩尔）或咖啡因（10毫摩尔）诱导的细胞内Ca2+积累不会改变心室肌细胞中微管的稳定性。我们的数据证明了成年心肌细胞中微管网络的独特稳定性。我们推测微管稳定性是在心脏收缩过程中维持细胞完整性所必需的。
