Takahashi Nobuya, Hozumi Yasukazu, Tanaka Toshiaki, Okada Masashi, Iseki Ken, Hayasaka Kiyoshi, Goto Kaoru
Department of Anatomy and Cell Biology, Yamagata University School of Medicine, 2-2-2 Iida-nishi, Yamagata, 990-9585, Japan.
Histochem Cell Biol. 2014 Nov;142(5):461-71. doi: 10.1007/s00418-014-1226-x. Epub 2014 Jun 4.
Diacylglycerol kinase (DGK) catalyzes conversion of a lipid second messenger diacylglycerol to another messenger molecule phosphatidic acid. Consequently, DGK plays a pivotal role in cellular pathophysiology by regulating the levels of these two messengers. We reported previously that DGKζ translocates from the nucleus to cytoplasm in hippocampal neurons under ischemic/hypoxic stress. In addition, we also identified nucleosome assembly protein 1 (NAP1)-like proteins NAP1L1 and NAP1L4 as novel DGKζ-interacting partners using a proteomic approach and revealed that these NAP1-like proteins induce cytoplasmic translocation of DGKζ in overexpressed cells because NAP1-like proteins associate with the nuclear localization signal of DGKζ and block its nuclear import via importin α. In the present study, we examined whether NAP1-like proteins are expressed in the brain and whether the molecular interaction of DGKζ and NAP1-like proteins would be changed in the brain after hypoxic stress. Immunohistochemistry revealed that NAP1L1 and NAP1L4 are widely expressed in neurons and glial cells in the brain with some differences. After 3 days of transient whole-body hypoxic stress, DGKζ translocated from the nucleus to cytoplasm in hippocampal pyramidal neurons, whereas NAP1-like proteins remained in the cytoplasm. Contrary to our expectations, NAP1-like proteins showed no change in their expression levels. The molecular interaction between DGKζ and NAP1-like proteins was attenuated after hypoxic stress. These results suggest that DGKζ cytoplasmic translocation in neurons under hypoxic stress is regulated by some mechanism which differs from that mediated by NAP1-like proteins.
二酰基甘油激酶（DGK）催化脂质第二信使二酰基甘油转化为另一种信使分子磷脂酸。因此，DGK通过调节这两种信使分子的水平在细胞病理生理学中发挥关键作用。我们之前报道过，在缺血/缺氧应激下，DGKζ在海马神经元中从细胞核转运至细胞质。此外，我们还通过蛋白质组学方法鉴定了核小体组装蛋白1（NAP1）样蛋白NAP1L1和NAP1L4为新的DGKζ相互作用伴侣，并揭示这些NAP1样蛋白在过表达细胞中诱导DGKζ的细胞质转运，因为NAP1样蛋白与DGKζ的核定位信号结合并通过输入蛋白α阻断其核输入。在本研究中，我们检测了NAP1样蛋白是否在脑中表达，以及缺氧应激后脑中DGKζ与NAP1样蛋白的分子相互作用是否会发生变化。免疫组织化学显示，NAP1L1和NAP1L4在脑中的神经元和胶质细胞中广泛表达，但存在一些差异。短暂全身缺氧应激3天后，DGKζ在海马锥体细胞中从细胞核转运至细胞质，而NAP1样蛋白仍留在细胞质中。与我们的预期相反，NAP1样蛋白的表达水平没有变化。缺氧应激后，DGKζ与NAP1样蛋白之间的分子相互作用减弱。这些结果表明，缺氧应激下神经元中DGKζ的细胞质转运受某种机制调控，该机制不同于由NAP1样蛋白介导的机制。
