Department of Molecular Physiology and Biophysics, Vanderbilt-Kennedy Center, Center for Molecular Neuroscience, Vanderbilt University School of Medicine, Nashville, Tennessee, United States of America.
PLoS One. 2012;7(2):e31554. doi: 10.1371/journal.pone.0031554. Epub 2012 Feb 13.
Mechanisms underlying age-dependent changes of dendritic spines on striatal medium spiny neurons are poorly understood. Spinophilin is an F-actin- and protein phosphatase 1 (PP1)-binding protein that targets PP1 to multiple downstream effectors to modulate dendritic spine morphology and function. We found that calcium/calmodulin-dependent protein kinase II (CaMKII) directly and indirectly associates with N- and C-terminal domains of spinophilin, but F-actin can displace CaMKII from the N-terminal domain. Spinophilin co-localizes PP1 with CaMKII on the F-actin cytoskeleton in heterologous cells, and spinophilin co-localizes with synaptic CaMKII in neuronal cultures. Thr286 autophosphorylation enhances the binding of CaMKII to spinophilin in vitro and in vivo. Although there is no change in total levels of Thr286 autophosphorylation, maturation from postnatal day 21 into adulthood robustly enhances the levels of CaMKII that co-immunoprecipitate with spinophilin from mouse striatal extracts. Moreover, N- and C-terminal domain fragments of spinophilin bind more CaMKII from adult vs. postnatal day 21 striatal lysates. Total levels of other proteins that interact with C-terminal domains of spinophilin decrease during maturation, perhaps reducing competition for CaMKII binding to the C-terminal domain. In contrast, total levels of α-internexin and binding of α-internexin to the spinophilin N-terminal domain increases with maturation, perhaps bridging an indirect interaction with CaMKII. Moreover, there is an increase in the levels of myosin Va, α-internexin, spinophilin, and PP1 in striatal CaMKII immune complexes isolated from adult and aged mice compared to those from postnatal day 21. These changes in spinophilin/CaMKII interactomes may contribute to changes in striatal dendritic spine density, morphology, and function during normal postnatal maturation and aging.
纹状体中型多棘神经元树突棘随年龄变化的机制尚不清楚。突触丝磷蛋白是一种 F-肌动蛋白和蛋白磷酸酶 1（PP1）结合蛋白，它将 PP1 靶向多个下游效应物，以调节树突棘的形态和功能。我们发现钙/钙调蛋白依赖性蛋白激酶 II（CaMKII）直接和间接地与突触丝磷蛋白的 N-和 C-末端结构域结合，但 F-肌动蛋白可以将 CaMKII 从 N-末端结构域中置换出来。在异源细胞中，突触丝磷蛋白将 PP1 与 CaMKII 共定位在 F-肌动蛋白细胞骨架上，在神经元培养物中，突触丝磷蛋白与突触 CaMKII 共定位。Thr286 自身磷酸化增强了 CaMKII 与突触丝磷蛋白在体外和体内的结合。尽管 Thr286 自身磷酸化的总水平没有变化，但从出生后第 21 天到成年期的成熟过程显著增强了从小鼠纹状体提取物中免疫共沉淀的与突触丝磷蛋白共免疫的 CaMKII 水平。此外，与来自成年纹状体裂解物的相比，突触丝磷蛋白的 N-和 C-末端结构域片段与更多的 CaMKII 结合。与突触丝磷蛋白 C-末端结构域相互作用的其他蛋白质的总水平在成熟过程中降低，这可能减少了与 C-末端结构域结合的 CaMKII 的竞争。相比之下，α-中间丝蛋白的总水平和与突触丝磷蛋白 N-末端结构域的结合在成熟过程中增加，这可能通过间接与 CaMKII 桥接。此外，与来自出生后第 21 天的小鼠相比，从成年和老年小鼠纹状体 CaMKII 免疫复合物中分离出的肌球蛋白 Va、α-中间丝蛋白、突触丝磷蛋白和 PP1 的水平增加。这些突触丝磷蛋白/CaMKII 相互作用组的变化可能导致正常出生后成熟和衰老过程中纹状体树突棘密度、形态和功能的变化。
