Molecular and cellular biological study about the abnormal neurite outgrowth and protein phosphorylation in temporal lobe epilepsy

颞叶癫痫的一个形态特征是海马齿状回颗粒细胞的异常突起生长。钙调素依赖的蛋白激酶II(CaM)的某些亚型可能与这种异常的轴突生长有关。在CaM激酶II中，已从脑中鉴定出四个亚基和几个亚型。为了探讨CaM激酶II各亚型在颞叶癫痫中的病理生理作用，我们主要围绕CaM激酶II的Delta亚基(CaM Kinase II Delta)对其结构和功能进行了研究。1)制备了抗CaM激酶II Delta的抗体，并对该亚单位在脑内的定位进行了检测。2)我们从大鼠脑中获得了CaM激酶II Delta的6种异构体的cDNA。在这些异构体中，Delta3定位于NG1O8-15细胞的细胞核中。3)在NG1O8-15细胞中过表达Delta3可诱导脑源性神经营养因子(BDNF)的基因表达，从而促进特定神经元群体的存活和分化。4)我们发现在PC12细胞和NG108-15细胞中，CaM激酶II的主要亚型是Delta2。而Delta2主要定位于高尔基体。5)在NG 108-15细胞中，其他亚型定位于细胞质。因此，这些异构体可能通过微管相关蛋白2等细胞骨架蛋白的磷酸化参与轴突生长的调节。这些结果表明，每种异构体都定位于神经元中特定的位置，在神经元功能中发挥重要作用。

A morphological characteristic of temporal lobe epilepsy consists of the abnormal neurite outgrowth of the hippocampal dentate granule cells. Some isoforms of Ca^<2+>/calmodulin-dependent protein kinase II (CaM kinase II) may be related to this abnormal neurite outgrowth. Among CaM kinase II, four subunits and several isoforms have been identified from brain. In order to investigate the pathophysiological roles of each isoform of CaM kinase II in temporal lobe epilepsy, we studied about the structures and functions of CaM kinase II isoforms mainly about the delta subunit of CaM kinase II (CaM kinase IIdelta). 1) We prepared the antibody against CaM kinase IIdelta and examined the localization of the subunit in brain. 2) We obtained the cDNAs of 6 isoforms of CaM kinase IIdelta from rat brain. Among these isoforms, delta3 was localized in the nuclei in transfected NG1O8-15 cells. 3) Overexpression of delta3 in NG1O8- 15 cells induced the gene expression of brain-derived neurotrophic factor (BDNF) which promotes survival and differentiation of specific populations of neurons in the brain. 4) We found that the major isoform of CaM kinase II in PC12 cells and NG108-15 cells was delta2. And delta2 appeared to be mainly localized in Golgi's apparatus.5) Other isoforms were localized in the cytoplasm in NG 108-15 cells. Therefore, these isoforms may be involved in the regulation of neurite outgrowth via phosphorylation of cytoskeletal proteins such as microtubule-associated protein 2.These results suggest that each isoform is localized in the specific site in neurons to play some important roles in neuronal functions.