POSTTRANSLATIONAL MODIFICATIONS OF NERVE CYTOSKELETAL PROTEINS IN EXPERIMENTAL DIABETES
POSTTRANSLATIONAL MODIFICATIONS OF NERVE CYTOSKELETAL PROTEINS IN EXPERIMENTAL DIABETES
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DOI：
10.1007/bf02780555
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发表时间：
1992-06-01
影响因子：
5.1
通讯作者：
CASSON, IF
中科院分区：
文献类型：
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作者：
MCLEAN, WG;PEKINER, C;CASSON, IF
已知实验性糖尿病大鼠周围神经轴突转运受损。由于轴突转运依赖于神经元细胞骨架的完整性，我们研究了实验性糖尿病大鼠脑和神经细胞骨架蛋白的改变方式。采用链脲佐菌素（STZ）致糖尿病大鼠。6周后，移除坐骨神经、脊髓和大脑，用于制备神经丝、微管和细胞骨架蛋白的粗制物。脑微管蛋白和周围神经微管蛋白的非酶糖基化程度通过h -3-硼氢化钠孵育，然后在二维聚丙烯酰胺凝胶上分离，并对分离的蛋白质进行亲和层析。两周的糖尿病大鼠和非糖尿病大鼠的脑微管蛋白的非酶糖基化没有差异。微管蛋白在微管中的组装也不受糖尿病状态的影响。另一方面，糖尿病患者2周后，坐骨神经微管蛋白的非酶糖基化显著增加。我们还从2周和6周糖尿病大鼠的脑细胞骨架蛋白制备中发现脑肌动蛋白的电泳迁移率发生了改变。另一种新的多肽被证明具有比肌动蛋白稍强的酸性等电点，可以用抗肌动蛋白抗体进行免疫染色。同样的多肽可以通过纯化的肌动蛋白与葡萄糖在体外孵育产生，从而确定它是非酶糖基化的产物。这些结果与糖尿病患者的临床研究数据进行了讨论，其中我们发现血小板肌动蛋白糖化增加。stz -糖尿病也导致体内脊髓神经丝蛋白磷酸化在糖尿病6周期间增加。当P-32- atp体外培养纯化的神经丝蛋白时，这种过度磷酸化以及神经丝相关蛋白激酶活性降低导致P-32掺入减少。我们的综合数据显示，在实验性糖尿病患者中，神经元细胞骨架蛋白的一些翻译后修饰可能导致轴突转运的改变和随后的神经功能障碍。
Axonal transport is known to be impaired in peripheral nerve of experimentally diabetic rats. As axonal transport is dependent on the integrity of the neuronal cytoskeleton, we have studied the way in which rat brain and nerve cytoskeletal proteins are altered in experimental diabetes. Rats were made diabetic by injection of streptozotocin (STZ). Up to six weeks later, sciatic nerves, spinal cords, and brains were removed and used to prepare neurofilaments, microtubules, and a crude preparation of cytoskeletal proteins. The extent of nonenzymatic glycation of brain microtubule proteins and peripheral nerve tubulin was assessed by incubation with H-3-sodium borohydride followed by separation on two-dimensional polyacrylamide gels and affinity chromatography of the separated proteins. There was no difference in the nonenzymatic glycation of brain microtubule proteins from two-week diabetic and nondiabetic rats. Nor was the assembly of microtubule proteins into microtubules affected by the diabetic state. On the other hand, there was a significant increase in nonenzymatic glycation of sciatic nerve tubulin after 2 weeks of diabetes. We also identified an altered electrophoretic mobility of brain actin from a cytoskeletal protein preparation from brain of 2 week and 6 week diabetic rats. An additional novel polypeptide was demonstrated with a slightly more acidic isoelectric point than actin that could be immunostained with anti-actin antibodies. The same polypeptide could be produced by incubation of purified actin with glucose in vitro, thus identifying it as a product of nonenzymatic glycation. These results are discussed in relation to data from a clinical study of diabetic patients in which we identified increased glycation of platelet actin. STZ-diabetes also led to an increase in the phosphorylation of spinal cord neurofilament proteins in vivo during 6 weeks of diabetes. This hyperphosphorylation along with a reduced activity of a neurofilament-associated protein kinase led to a reduced incorporation of P-32 into purified neurofilament proteins when they were incubated with P-32-ATP in vitro. Our combined data show a number of posttranslation modifications of neuronal cytoskeletal proteins that may contribute to the altered axonal transport and subsequent nerve dysfunction in experimental diabetes.