Characterization of the human NEK7 interactome suggests catalytic and regulatory properties distinct from those of NEK6.
Characterization of the human NEK7 interactome suggests catalytic and regulatory properties distinct from those of NEK6.
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DOI：
10.1021/pr500437x
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发表时间：
2014-09-05
影响因子：
4.4
通讯作者：
Kobarg, Joerg
中科院分区：
文献类型：
--
作者：
de Souza, Edmarcia Elisa;Meirelles, Gabriela Vaz;Godoy, Barbara Biatriz;Perez, Anna Marina;Costa Smetana, Juliana Helena;Doxsey, Stephen J.;McComb, Mark E.;Costello, Catherine E.;Whelan, Stephen A.;Kobarg, Joerg
人NEK7是细胞分裂的调节因子，在哺乳动物细胞的生长和存活中发挥重要作用。人类 NEK6 和 NEK7 密切相关，由保守的 C 端催化结构域和非保守且无序的 N 端调节结构域组成，对于介导与各自蛋白质的相互作用至关重要。在这里，为了更好地了解 NEK7 细胞功能，我们通过两种筛选方法来表征 NEK7 相互作用组：一种使用酵母双杂交系统，另一种基于免疫沉淀，然后进行质谱分析。这些方法导致鉴定出 61 个 NEK7 相互作用因子，这些相互作用因子有助于细胞分裂等多种生物过程。结合酵母双杂交筛选的额外相互作用和磷酸化测定，我们验证了 CC2D1A、TUBB2B、MNAT1 和 NEK9 蛋白作为潜在的 NEK7 相互作用子和底物。值得注意的是，内源性 RGS2、TUBB、MNAT1、NEK9 和 PLEKHA8 与 NEK7 一起定位在整个细胞周期的关键位点，尤其是在有丝分裂和胞质分裂期间。此外，我们获得的证据表明，除了 NEK9 之外，紧密相关的激酶 NEK6 和 NEK7 不具有共同的相互作用子，并且根据其 N 端和 C 端区域以不同的方式显示不同的蛋白质相互作用模式。总之，我们的工作首次展示了全面的 NEK7 相互作用组，结合功能性体外和体内测定，表明 NEK7 是一种多功能激酶，在不同的细胞过程中与细胞分裂信号传导相一致，且独立于 NEK6。
Human NEK7 is a regulator of cell division and plays an important role in growth and survival of mammalian cells. Human NEK6 and NEK7 are closely related, consisting of a conserved C-terminal catalytic domain and a nonconserved and disordered N-terminal regulatory domain, crucial to mediate the interactions with their respective proteins. Here, in order to better understand NEK7 cellular functions, we characterize the NEK7 interactome by two screening approaches: one using a yeast two-hybrid system and the other based on immunoprecipitation followed by mass spectrometry analysis. These approaches led to the identification of 61 NEK7 interactors that contribute to a variety of biological processes, including cell division. Combining additional interaction and phosphorylation assays from yeast two-hybrid screens, we validated CC2D1A, TUBB2B, MNAT1, and NEK9 proteins as potential NEK7 interactors and substrates. Notably, endogenous RGS2, TUBB, MNAT1, NEK9, and PLEKHA8 localized with NEK7 at key sites throughout the cell cycle, especially during mitosis and cytokinesis. Furthermore, we obtained evidence that the closely related kinases NEK6 and NEK7 do not share common interactors, with the exception of NEK9, and display different modes of protein interaction, depending on their N- and C-terminal regions, in distinct fashions. In summary, our work shows for the first time a comprehensive NEK7 interactome that, combined with functional in vitro and in vivo assays, suggests that NEK7 is a multifunctional kinase acting in different cellular processes in concert with cell division signaling and independently of NEK6.
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影响因子：
8
作者：
Hegde, Muralidhar L.;Hazra, Tapas K.;Mitra, Sankar
通讯作者：
Mitra, Sankar
影响因子：
2.6
作者：
Feige, E;Motro, B
通讯作者：
Motro, B
影响因子：
4
作者：
Kim, Sunghwan;Kim, Sungjae;Rhee, Kunsoo
通讯作者：
Rhee, Kunsoo
影响因子：
4.8
作者：
Belham, C;Roig, J;Avruch, J
通讯作者：
Avruch, J
DOI：
10.1016/s0006-291x(03)00049-4
发表时间：
2003-02-21
影响因子：
3.1
作者：
Minoguchi, S;Minoguchi, M;Yoshimura, A
通讯作者：
Yoshimura, A