ＫＩＦ２６Ａの分子遺伝学的研究

KIF26A的分子遗传学研究

• 批准号: 14F04904
• 负责人: 廣川 信隆
• 金额: $ 1.47万
• 依托单位: The University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for JSPS Fellows
• 财政年份: 2014
• 资助国家: 日本
• 起止时间: 2014-04-25 至 2016-03-31
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-14F04904/
• 关键词: KIF26A; megacolon; pain response; KIFs; knock-out mice; fear conditioning

The kinesin superfamily proteins (KIFs) have been shown to transport various membranous organelles and protein complexes in a microtubule- and ATP-dependent manner (Hirokawa and Noda, 2008; Schliwa, 2002). In the previous research, we identified Kinesin superfamily protein 26A (KIF26A), KIF26A is a rather atypical member as it lacks ATPase activity. We found that KIF26A suppressed GDNF-Ret signaling by direct binding and inhibition of Grb2, an essential component of GDNF/Akt/ERK signaling (Zhou et al., Cell 139, 802-813, 2009).In this study, we have occasionally noticed the hypersensitive and prolonged pain response of Kif26a-/- mouse pups during handling, and deeply investigated its cellular and molecular mechanisms.In order to understand the molecular basis of this impairment in Ca clearance, we assessed the level of the inhibitory tyrosine phosphorylation level of PMCA by using a KIF26A knockdown system in F11 cells. As a result, the tyrosine phosphorylation level of PMCA in KD cells was significantly higher than that in control cells, which could partly explain the impairment in Ca clearance in Kif26a-/- neurons via PMCA inactivation. We investigated the levels of SFK/FAK activation. Immunocytochemistry against pFAK in Kif26a-/- neurons revealed a significantly stronger signal than that in Kif26a+/+ neurons. In other hand, immunocytochemistry against pSFK, which is a kinase further upstream to FAK, was largely unaltered. These data suggested the possibility that the SFK-to-FAK signal transduction is abnormally enhanced by KIF26A deficiency.

驱动蛋白超家族蛋白 (KIF) 已被证明能够以微管和 ATP 依赖性方式运输各种膜细胞器和蛋白质复合物（Hirokawa 和 Noda，2008；Schliwa，2002）。在前期的研究中，我们鉴定出了驱动蛋白超家族蛋白26A（KIF26A），KIF26A是一个相当非典型的成员，因为它缺乏ATP酶活性。我们发现 KIF26A 通过直接结合和抑制 Grb2（GDNF/Akt/ERK 信号传导的重要组成部分）来抑制 GDNF-Ret 信号传导（Zhou et al., Cell 139, 802-813, 2009）。在这项研究中，我们偶尔会发现注意到Kif26a-/-小鼠幼崽在处理过程中出现的过敏和长时间的疼痛反应，并深入研究其细胞和分子机制，以了解其分子基础由于 Ca 清除率受损，我们通过在 F11 细胞中使用 KIF26A 敲低系统评估了 PMCA 的抑制性酪氨酸磷酸化水平。结果，KD 细胞中 PMCA 的酪氨酸磷酸化水平显着高于对照细胞，这可以部分解释 PMCA 失活导致 Kif26a-/- 神经元中 Ca 清除率受损。我们研究了 SFK/FAK 激活水平。 Kif26a-/- 神经元中针对 pFAK 的免疫细胞化学显示，其信号明显强于 Kif26a+/+ 神经元中的信号。另一方面，针对 pSFK（FAK 更上游的激酶）的免疫细胞化学基本上没有改变。这些数据表明 SFK 至 FAK 信号转导可能因 KIF26A 缺陷而异常增强。