Regulation of brain functions by Ca^<2+>/calmodulin-dependent protein kinase II activity

钙调素依赖性蛋白激酶II(CaMKII)是中枢神经系统中含量最丰富的蛋白激酶之一，也是细胞内钙离子对各种刺激作出反应的关键介质。CaMKII参与多种神经功能，包括突触可塑性和高级脑功能。然而，CaMKII的哪一种蛋白功能，即蛋白激酶活性、钙调素结合能力或与其他蛋白相互作用的多聚体结构，是负责这种神经元功能的，还有待确定。为了明确CaMKII蛋白激酶活性及其底物蛋白磷酸化的作用，我们用灭活的CaMKIIα亚单位通过用Arg-42取代Lys-42来改造敲入小鼠。正如预期的那样，在纯合子突变体中，CaMKII活性选择性降低，而CaMKII亚单位蛋白水平与野生型对照相当。原位杂交显示CaMKII亚单位α的正常表达模式，包括在纯合子突变中CaMKII mRNA树突状定位。急性海马片的电生理分析和培养的海马神经元细胞生物学研究表明，纯合子突变体在海马区突触可塑性异常，但基础突触传递正常。此外，在这些小鼠中也观察到了海马体依赖的学习缺陷。这些结果表明，CaMKIIα的蛋白激酶活性及其底物蛋白磷酸化是维持正常海马功能所必需的。这种功能缺陷的CaMKIIα突变小鼠将成为寻找活性依赖突触可塑性的分子细节的有力工具。

Ca^<2+>/calmodulin-dependent protein kinase II (CaMKII) is one of the most abundant protein kinase in the central nervous system and a key mediator of intracellular Ca^<2+> in response to various stimuli. CaMKII is involved in many neuronal functions including synaptic plasticity and higher brain functions. However, which protein function of CaMKII, i.e., protein kinase activity, calmodulin-binding capacity or multimeric structure interacting with other proteins, is responsible for such neuronal functions remains to be determined yet. To clarify specifically the role of protein kinase activity of CaMKII and its substrate protein phosphorylation, we engineered knock-in mice with the inactivated a subunit of CaMKII by replacing Lys-42 with Arg-42. As expected, CaMKII activity was selectively reduced, while CaMKII subunit protein levels were comparable to those of wild type controls in homozygous mutants. In situ hybridization revealed normal expression patterns of CaMKII subunit mRNAs, including dendritic localization of CaMKIIα mRNA in homozygous mutants. Electrophysiological analysis using acute hippocampal slices and cell biological study using cultured hippocampal neurons revealed abnormal synaptic plasticity but normal basal synaptic transmission in hippocampus in homozygous mutants. In addition, hippocampus-dependent learning deficit was also observed in these mice. These results indicate that protein kinase activity of CaMKIIα and its substrate protein phosphorylation are essential for maintaining normal hippocampal functions. Such a functionally deficient CaMKIIα mutant mouse will serve as a powerful tool in search for molecular details for activity-dependent synaptic plasticity.